Color Atlas of Immunology - PDF Free Download

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Author: Gerd Burmester | Antonio Pezzutto

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... Key to Symbols

Antigens

Membrane, phospholipid layer

Receptors

Thymus

TCR

Class I MHC molecule

Class II MHC molecule

Cytokine receptor

Antibody (Ab)

Lymph node

Cellular tissue

Bacteria and viruses

Arrows denoting transportation, effect, and direction

Positive effect

Negative effect

Auto-Ab

Inhibition Blockade

Complement

Signal

... Key to Symbols

Stem cell

Plasma cell

B cell

Antigenpresenting cell (APC)

T cell

Natural killer cell (NK cell)

Erythrocyte

Macrophage

Monocyte

Megakaryocyte

Neutrophil granulocyte

Thrombocytes

Basophil granulocyte

, Langerhans cell

Eosinophil granulocyte

Interdigitating dendritic cell

Basophil mast cell

Mature dendritic cell

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#?+$?%+@

CD34+

Pluripotent stem cell

Myeloid progenitor

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Lymphoid progenitor

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T-cell precursor

Monoblast

Thymus

B-cell precursor

Platelets B lymphocytes

P

h

Erythrocytes

a

g

o

c

y

t

e

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Basophils Eosinophils

Dendritic Neutrophils Monocytes cells A. Origin of cells of the immune system Innate immunity

Adaptive immunity

Infectious agents

Phagocytes

Natural killer cells

;2'<91+'.9, :$%'8%=,+-

Erythroblast

"

Generation of specific receptors

Natural killer cells Blood components Hours

Days, weeks

B. Defense mechanisms against infections CD34+

Hematopoietic stem cell

Hepatocytes Neurons C. Plasticity of stem cells

Cardiomyocytes Smooth muscle cells

Endothelial cells

Skeletal muscle cells

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Cortex

Secondary lymphoid organs

!

Spleen

T lymphocytes

Thymus

T lymphocytes

Bone marrow

B lymphocytes

9&6:0.(6$05 ;%16'1/5(2

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Erythron

B lymphocyte

"

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Liver

Lung

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Mesenteric lymph nodes

Peyer´s patches

Efferent lymphatics

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Spleen

Peripheral lymph nodes

! ?

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10('23

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Thyroid gland Large vessels

Heart

Fat and connective tissue

Cortex

20

!

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1. Position of the thymus

10

20 30 Age/years

40

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Hassall’s body

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Interlobular connective tissue (trabecula)

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B T

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T

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Follicle-associated epithelial cells Dome region

T-cell zone

Postcapillary venule Muscular mucosa

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Thymic hormone

2.

!

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4.

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Mature T cells

Location

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Mature thymocyte

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Cell

TCR

TCR

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CD3 CD7

CD1

CD8

CD7 CD2

(CD1)

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TCR CD2

CD3

Marker

TCR

CD5 CD7

Rearrangement of TCR!

Transcription of TCR! rearrangement of TCR#

Cell surface expression of TCR!,", #

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TdT enzyme

TdT, CD2, CD7

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(CD1)

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Thymus (see B)

CD2, 5, 7, TCR",#

C. Development of mature T cells

CD3 CD4 CD45RA

Naive T cell

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Memory T cell

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Prethymocytes

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A

B Dendritic cell

"

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D

Autoantigen

!

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Thymus with maturing thymic cells

Thymocyte (type)

Self-MHC binding

Autoantigen recognition

Fate

A

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_

B

Strong

_

Cell death

C

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Yes

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D

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No

Complete maturation

A. Mechanisms of T-cell selection in the thymus

Cell death

Mature immunocompetent T cell

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#$%&'' (&)&*+,-4

!, " (Chromosome 14) V!1 V!2

# (Chromosome 7q) V#1 V#2

V!n

V"n

V#3

D" J "

V#n

D#1

C"

V

J! segments

C#1

J#1

D#2

!

C!

C#2

V#14

$ (Chromosome 7p)

V1.1

V1.2

V1.3

V1.4

V1.5

V$II

V$IV Jp1 Jp J1 C$1

V$III

V1.6 V1.8 VA V1.5P V1.7

C$2

VB VII

Pseudogene

08/?12&/+1' <-3/)3*'&4

V$I

(Reversed direction of transcription)

A. T-cell receptor gene families I II I II

"

I II Deletion

Unequal chromosome switch

Inversion

B. T-cell receptor rearrangement ! (") Extracellular region

V!

# ($) S S S S

Gene segments

Transmembrane region Cytoplasmicregion

S S S S

D

0

2

J

100

13

V x D x J-combinations

104

N-sequences

104

2 x103 104

V

SS

C

#-chain

100

CHO C!

!-chain

V

100

Total number of !/# combinations

1015

D. Possible combinations of the T-cell receptor (!/#) 248 282

! chain = V - J - C # chain= V - D - J - C " chain = V - D - J - C $ chain = V - J - C

C. Configuration of the T-cell receptor

Total: Marker:

!/# 95 %

$/" 5%

66 %

<1%

CD4+ CD4– CD4–

CD8– CD8+ CD8–

33 %

25 %

<1 %

70 %

CD4+

CD8+

<1 %

<12 %

E. Distribution of !/# and $/" T cells

Weak

! " CE

<$';&%+'1&, =0.'4.8,+5

!

"

! >?

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Molecule

Cell expression

Function

Thymocytes, dendritic cells, few B cells (CD1c)

Antigenpresentation (glycolipids)

1p13

Thymocytes, all T cells, NK cells

Receptor for CD58 (LFA-1), T-cell activation

11q23 11q23 11q23 1q22 1q22

Maturing thymocytes, T cells

Signaltransduction after MHC-TCR contact

55

12p12

Thymocytes, T-helper cells, monocytes/macrophages, dendritic cells, eosinophilic granulocytes

Binding to MHC class II MHC

67

11q13

Thymocytes, all maturing T cells, few B cells

Signal transduction

40

17q25

All cells of T-cell lineage

Not known

2p12 2p1

Thymocytes, cytotoxic T-cells, NK cells (weak, CD8a)

Binding to class I MHC molecules

33

Xq26.327.1

CD4+ T cells (after activation), CD8+ T cells (subpopulation), basophils

Binds to CD40, activates B cells and dendritic cells

40

2q33

Thymocytes CD4+T cells, CD8+T cells, (subpopulation)

Ligand for CD80, CD86 (“co-stimula„ tory signal )

33

2q33

Activated T cells

Ligand for CD80, CD86 (negative regulator of T-cell activation)

Molecular weight (kDa)

#

43 – 49

Gene locus 1q22-23

CD1a b,c,d,e

$ 2m

50 CD2 %

CD3g 25 CD3d 20 CD3e 20 !-chain 16 "-chain 22

& !/" #(&) $(%) '

CD3/TCR

CD4

CD5

CD7 # $

CD8# 33 CD8$ 33

CD8

CD154 (CD40L)

CD28

CD152 (CTLA–4)

A. Human T-cell differentiation molecules

! <$';&%+'1&, =0.'4.8,+5

()*+,, !'1.@+'5

"

! " >A

A4./,8&.),' B2*.(*='&3

!

"

! CD

#$>?8=7-(?)& @&+&'-=8&.) ,./ @*99&2&.)*,)*-.

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> #$33) 4YZT>9 =<+2 &'$ 5<$$A%)&%"; =+<2 &+ &'$ (#&%I$ =+<2 *. ?(. += 5'+)5'+<.3(&%+" 53(.) ( -$#%)%I$ <+3$ %" &'$ 5<+#$))6 YZT> 2%;<(&$) &+ &'$ ",#3$,): *%"-) &+ &'$ )5$#%=%# UX@M 5<+2+&$< <$;%+": ("- #++5$<(&$) ?%&' ("+&'$< ",#3$(< *%"-%"; =(#&+< 4TN@8 #+253$A9 %" )&(<&%"; UX@M ;$"$ &<(")#<%5&%+" I%( KYT 5+3.2$<()$ UU6

5" #$%&'' !()*+,)*-.6 #7& #*8& %-423& -9 :&.& ;<=2&33*-. T -%)&%"#&%+" %) 2(-$ *$&?$$" %22$-%(&$: $(<3.: ("- 3(&$ >@#$33 (#&%I(&%+" 5<+#$))$)6 N<+@ &+@+"#+;$"$) 4#@1&0 ("- #@'*#9: ",#3$(< *%"-@ %"; 5<+&$%") 4)$$ )$#&%+" !9: ("- #.&+H%"$ ;$"$) *$#+2$ %"I+3I$- %" &'%) +<-$< += ),##$))%+"6 >'$ %"#<$()$- $A5<$))%+" += /01 -$&$<2%"("&) 4+" #$<&(%" #$33 ).)&$2)9 ("- (-'$)%+" 2+3$@ #,3$) +##,<) +"3. )$I$<(3 -(.) 3(&$<6

#$%&'' !()*+,)*-.

APC

MHC

$

'

!

!(")

!

!

$

#

&

%

PIP2

PLC

DAG

TCR IP3

p56Ick

PKC

p59fyn Ca2+

Tyrosine kinases

Ca-dependent kinases

T cell Cytoplasm

Phosphorylated proteins Gene activation regulation of transcription

Nucleus

Phosphorylated proteins

Immediate

A. T-cell activation: Signal transduction Proto-oncogenes Nuclear binding proteins Minutes Cytokines

Early

Late

c-fos

Receptors Activating antigens Enzymes, intracellular proteins Hours

IL-4 IL-5 IL-6

GM-CSF

Insulin-R IL-2R CD69

1

2

Transferrin-R

Cyclin 3

MHC Cytokines Adhesion proteins Days

30

TGF-# IL-3

ODC Actin

4

Transferrin 5

6

12

Histones 18

24

48

HLA-DR Rantes VLA-4 2

"

c-myc NF(B

NFAT-1 15

IFN-' IL-2

A4./,8&.),' B2*.(*='&3

%

CD4(CD8)

#

Ag

CD3 CD45

!

4

B. T-cell activation: Time course of gene expression

VLA-1 6

8

10

! " CE

C7(/*?&()*2 :'$(1$@2&3

!

"

! 0D

,=>
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,-. *(/ ,-0 ;&223

Antigens

Bacteria

Protozoans (e.g. helminths)

Fungi

!

Pollen

NK cell

Macrophage

!/" T cell IFN-!

IL-12

Basophils, mast cell IL-4

C7(/*?&()*2 :'$(1$@2&3

Genetic susceptibility (e.g. MHC, TCR etc.)

?

Cytokine milieu IL-12

MHC II Dendritic cell

B7-1 (CD80)

IFN-!

TCR

TCR

CD28

Intracellular killing

MHC II

CD28

TH1

IFN-

!

Dendritic cell

B7-1 (CD80)

TH0

IFN-! IL-2 IFN-! TNF-#

IL-4

"

IL-4 TH2

0 IL-1 IL-4

IL-5 Antibody

IL-4

Macrophage

B cell

A. Differentiation into TH1 and TH2 cells

IL-10 IL-4

IL-4R TCR

TH2 B. Regulation of IgE production

IL-4R

IL-13 IL-4

CD40 ligand

Ag IL-4R

sIgM

MHC + Ag

IgE

CD40 B cell

! " 0.

;<*87)$*+7& =:9*19(&$2

!

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-63$&& ?*+'@$*$292

CD34 CD19

CD38

CD19 CD20 CD38 CD10

CD117

CD10

Pro-B

Plasma cell

Intestinal submucosa

Pro-plasma cell

Follicle Pro-PC Pro-B

Plasma cell

Marginal zone

Antigens Circulating follicular B cell T-cell zone

Circulating follicular B cell

Extrafollicular B cell

Apoptosis Pre-B

„Virgin”-B

Bone marrow : primary B-cell production CD20 CD19

CD22

Peripheral blood

CD20 CD19

CD21 CD22

cyt-µ

Pre-B

Virgin-B

slgM Virgin-B

Secondary lymphatic organs: antigen-induced B-cell proliferation

CD20 CD21 CD19 CD22 CD23 CD37 slgM slgD

Circulating follicular B cell

CD20 CD21 CD19 CD22

;<*87)$*+7& =:9*19(&$2

CD22

!

"

CD39 slgM CD11c

Extrafollicular B cell

A. Development of B lymphocytes Pleural space Bone marrow

CD5+ slgM+

CD5+ B cells

– T-independent – Secrete low-affinity lgM-autoantibodies – Can renew themselves – No hypermutations CD5

CD45R CD20 CD11a

Early ontogenesis B. CD5+ B cells

Peritoneal cavity

slgM

! " >A

A9./,3&.),' 62*.(*?'&>

!

"

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! BC

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1&23*.,' %&.)&2 4&,()*-.

= Follicular dendritic cells (FDC)

Follicle mantle Primary B-blasts

Phase 1 of follicle reaction (3-4 days)

Primary lymphoid follicle Follicle mantle

– Centrocytes – FDC – T-helper cells – Macrophages

Secondary B-blasts Phase 3 of follicle reaction (months)

Light zone

– Centroblasts

Phase 2 of follicle reaction (3 weeks)

Dark zone

A. B-cell activation: the germinal center reaction

Antigen profile

Follicular B cell

Primary B-blast

– – – – – – – – –

– – – – –

CD19 CD20 CD21 CD22 CD23 CD39 CD40 CD76 slgM/D

Centroblast – – – – – – –

CD19 CD20 CD22 CD40 slgM

CD19 CD20 CD22 CD38 CD77 CD40 slg±

Centrocyte – – – – – – – –

CD10 CD19 CD20 CD22 CD37 CD38 CD40 slgM/A/G/E

Circulating memory cell

Apoptosis

Plasmablast

Intestinal mucosaplasma cell

Bone marrow plasma cell

B. Antigen profile of B cells during germinal center reaction

"

FDC

FDC

Antigen-specific B cell

A9./,3&.),' 62*.(*?'&>

Follicle mantle

!

Apoptosis Apoptosis

Dark zone of germinal center

Repeated cycles

– Proliferation

Light zone of germinal center

Mutated immunoglobulin with higher affinity binds antigen-presenting FDC and survives

– Point mutation in – V-region of H/L-chains A A C T G C

A A C C G C

T T G A C G

T T G G C G

= Amino acid switch = 10-fold increase in affinity

C. Selection of high-affinity antibodies through hypermutation in the germinal center

! " BD

B%&A5$0&-5) C.+&/+3)0=

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#$$%&'()'*%)+&=

Variable

Constant

Domain C1q binding site

Light chain

CH1 VH

Heavy chain

S

S S S S

CH2 COOH

S S

CH3

CH1

COOH VH

CH2

CL NH2

CH3

CL Fc

Antigen binding

Effector function

CDR = complementarity-determining region Fab = antigen-binding fragment Fc = crystallizable fragment

CH4 Only at IgE and IgM

VL

VL Fab

NH2

SSSS

S

Antigen binding VH = variable domain of heavy chains VL = variable domain of light chains CH/L = constant domain of heavy / light chains

A. Immunoglobulin structure N

N N

N N N ! (")

!2m #

C C IgG

CC TCR I/II

C MHC class I

N

N

NN

# # ($)

N

NN !

CC MHC class II

C CD4

C C C CD8 Poly-IgR

C CD19

C C CD22 N-CAM/ CD 56

B%&A5$0&-5) C.+&/+3)0=

CDR

!

"

B. Immunglobulin-“superfamily”

-N-terminal end

SSSS

CDR L3 90-95 CDR L1 26-32 CDR L2 49-55

Variable domain of light chain with the 3 hypervariable complementarity determinants

CDR H3 95-101

CDR L2 49-55

CDR H1 30-35

CDR H2 49-64

e n - E p i t o p e t i g A n CDR L1 26-32

C. Hypervariable regions determine the antigen specificity

CDR L3 90-95

! " DF

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5667)%8+%97+() :+
Albumin

Breast milk

Globulins

! 1 !2

"

!

# Fc receptors

IgD

IgA

1. Circulating Ig 2. Membrane2. bound Ig

IgG

A. Protein electrophoresis

VL

CL

4. Cell-bound Ig

B. Different immunoglobulin types

IgG

SS SS

V

3. Secretory Ig

IgA

IgM

IgD

Serum: Monomeric lgA

Serum: lgM

Serum: lgD

IgE

>7)=<6')&<+ #$(),(-+'/

IgM

SS

65%

SS

SS SS

SS

SS

IgG1

SS

SS

SS SS SS SS

23%

Membranebound lgM

Secretory lgA; dimer SS SS SS SS SS SS SS SS SS SS SS

SS

8%

"

J chain

S S

SS

IgG3

SS

Hinge region

Membranebound lgD

SS

IgG2

SS

Mature B lymphocyte

SS SS

Mature B lymphocyte IgG4

Basophil granulocyte

4%

Serum lg

80%

Half-life

23

13%

6%

0.1%

0.002%

6

5

3

2.5 days

C. Immunglobulin structure and features Lumen

pH 7.4

Fc#R

Binding at pH <7.0

Release at pH 7.4

D. IgG transport through intestinal epithelium

Secretory component

lgA

E. Secretion of IgA

Lumen

J chain

! " CE

!

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56- 7-"-/ 2,+ # (#76. )6'#"/ '+- (,)'.-$ ," )6+,: 9,/,9- J4 !+,&"$ FBE>1 2&").#,"'((8 ').#%- C 7-"-/ ;.,7-.6-+ S#.6 .6- ),++-/*,"$#"7 P 7-"-/= ),$- 2,+ '9#", ')#$/ 0 ., DB ,2 .6- %'+#: '3(- +-7#," ,2 # (#76. )6'#"/X B I 7-"-/ ),$- 2,+ '9#", ')#$/ DH ., 0014 C 7-"-/ '+- 3+,&76. "-W. ., I 7-"-/ $&+#"7 .6- *+,)-// ,2 GT! +-'++'"7-: 9-".4 ! *+-)&+/,+ 9UT! #/ .+'"/)+#3-$ 2+,9 ' CI /-Q&-")-@ .,7-.6-+ S#.6 .6- /-Q&-")- 2+,9 .6- ),"/.'". +-7#," ,2 .6- # (#76. )6'#" ;K#=4 56P /-Q&-")- .6-" /*(#./ ,22 2+,9 .6- *+,.-#"4

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;" #$%&'()&*(+' +0 " 9(%:* ;:&(' 2.'.3 56- ,+7'"#V'.#," ,2 $ (#76. )6'#" 7-"-/ ," )6+,: 9,/,9- JJ #/ ",. 2&((8 &"$-+/.,,$4 56-+- '+- ' "&93-+ ,2 7-"-/ 2,+ .6- ),"/.'". +-7#,"@ '"$ .6- I /-Q&-")-/ (#- $#+-).(8 #" 2+,". ,2 .6- K 7-"-/4 56-+- '+- *+-/&9'3(8 A&/. '/ 9'"8 7-"-/ 2,+ .6- $ )6'#" '/ 2,+ .6- # )6'#"4 Z-)'&/- -%-+8 6-'%8 )6'#" #/ (#"R-$ S#.6 ' # ,+ $ )6'#"@ .6-+- '+-@ .6-,+-.#)'((8@ B4J!01B ;0YB!F!01F= ., 04[!01H ;J11!D!01F= $#22-+-". #:3-'+#"7 '".#3,$8 +-),93#"'.#,"/4 56- *+,A-).-$ "&93-+ ,2 $#22-+-". $:3-'+#"7 #99&",7(,3&(#"/ #/ *+,3'3(8 /#9#('+4 56- '): .&'( "&93-+ ,2 *,//#3(- '".#3,$8 9,(-)&(-/ #/@ 6,S-%-+@ 9&)6 6#76-+4 56#/ #/ '..+#3&.'3(., /-%-+'( +-'/,"/@ /&)6 '/L ;0= 9&.'.#,"/ '. .6- GT! (-%-( $&+#"7 ,".,7-"-/#/X ;J= -++,+/ #" .6- ),&+/- ,2 $-(-.#," '"$ +-),93#"'.#," ,2 C@ G@ '"$ I 7-"-/@ $&+#"7 .6- ),&+/- ,2 S6#)6 "&)(-,.#$-/ '+- #"/-+.-$ 2+,9 GT! /-Q&-")-/ .6'. '+- ",. ",+9'((8 .+'"/)+#3-$X ;F= .6/8".6-/#/ ,2 "-S #99&",7(,3&(#"/ S#.6 -"6'")-$ '22#"#.8 $&- ., .6- /S#.)6#"7 ,2 '9#", ')#$/ %#' *,#". 9&.'.#," $&+#"7 7-+9#"'( )-": .-+ +-').#,"/ ;/-- *4 J>=4

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

4//5'+%6+756(' 2.'. #$%&'()&*(+'

n = ~50 Vn

Chromosome 14

✂

✂

VH1 VH2 VH3 5'

!

3' L1

L2

L3

Ln

D1 D2 D 3 D 4

DJ rearrangement

VH1 VH2 VH3

Dn

Cµ C% C#3 C#1 C&1 C#2 C#4 C' C&2

J1 J2 J3 J4 J5 J6

Germline DNA

Deletion

C#4

Vn

3'

5' L2

L3

Ln

D1 D2 D3 J5 J6

Transcription

Deletion

DJ-Cµ mRNA

VDJ-rearrangement

DJ-Cµ protein

VH1 VH2 5'

Cleavage of L-coded sequence

3' L1

L 2 D3 J 5 J6

C' C&2

Cµ C% C#3 C#1 C&1 C#2

Cµ C% C#3 C#1 C&1 C#2 C#4 C( C&2

VDJ-Cµ coding mRNA

µ heavy chains

A. Organization and rearrangement of immunoglobulin H genes n = ~35 V$1 V$2 V$3

V$n

5' L1

L2

L3

Ln

J$1 J$2 J$3 J$4 J$5

3'

Chromosome 2

3'

Chromosome 22

C$

B. Organization of ! light chain genes V"1

V"x Lx

"

V"n

5' L1

D5',&/.'*&6 E$('@(B6.3

L1

VpreB Ln

J"2 C"2 J"4 C"4 J"x C"x

C. Organization of " light chain genes

VDJ

s Cµ

C%

µ

s C#3

mRNA

lgM-expressing B cell D. Immunoglobulin class-switching

s C#1

Maturation V D J

C #3

C µ

C%

s

s C& 1

C# 1

#1

mRNA

lgG1 -expressing B cell

! " FH

>5,9.3&,-.' ?+),1)<'&@

!

"

! AB

#$:;3<2/1;-& (&=&'/<3&,- .,9 ()**&+&,-).-)/,

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

4335,/6'/75'), C&,& ?+/951- DE<+&@@)/, pro-B

pre-B-I

pre-B-II

Immature B

Mature B

Plasma cell

!

HL A -class II (HL A -DR) CD34 CD10

Surface

lg-!/lg-"

Surface

VpreB/#5

slgM/slgD

cyt-µ

Immunoglobulins CD22 CD20 CD21 CD23 A. B-cell differentiation scheme CD34 IL-7R

H

L$

Cytoplasmic

CD19 CD22cyt

IL-7R

H

L$

DJ-rearrangement L#

CD19 CD22cyt CD10

L#

Pro-B

pre-B-I VpreB/#5

VpreB/#5 DJ-Cµ transcription

CD19 CD22cyt H

L$

CD10

H

L$

VDJ-rearrangement

"

CD10

L#

L# Pre-B-II

Prot.

Pre-B-I

VpreB/#5

DJ-Cµ

Hµ/VpreB/#5

HµL$

VL-JL- Rearrangement CD19 CD22cyt

Hµ/L$ (slgM)

CD19 CD22s

CD20

CD20

or

or

Immature B Hµ/VpreB/#5

CD19 CD22cyt

>5,9.3&,-.' ?+),1)<'&@

CD19

Hµ/L#(slgM)

Mature B slgM+

B. Immunoglobulin modulation during B-cell differentiation

CD21 CD23 HµL# (slgM)

! AA

56/10%+/*0. 742/)2&.+8

!

"

! 9:

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

+.,)0/%&#T:+/+0/%&# 520#33 (/)4," *+0,)/ )00./+- + 03#+8+(# :/)'.0, )* <=M>6 !"($ %- 1+%&39 #?:/#--#' $9 5 0#33-; $., +3-) $9 '#&'/%,%0 0#33- A=; %- + 1#1$#/ )* ,"# %./%)"1 3)*!"+5"'(/$ /#0#:,)/ *+1%396 G%H# <=IC; %, %- + $/)+' 520#33 1+/H#/; $., %,- *.&0,%)& %- &), 9#, .&'#/-,))'6 Q+/3%#/ #8%'#&0# ,"+, <=BM *.&0,%)&- +- + 3%(+&' *)/ <=W "+- &), 9#, $##& 0)&*%/1#'6 !"*$ +&' !"*+ +/# &), 520#33 3%&#+(#2-:#0%2 *%06 !"#9 +/# #?:/#--#' )& ),"#/ +&,%(#&2:/#2 -#&,%&( 0#33-; -.0" +- 1)&)09,#- +&' '#&'/%,%0 0#33-6 !"# %&,#/+0,%)& )* <=NJT<=NY 4%," ,"#%/ 3%(+&'- )& ! 0#33- A<=MN +&' YD6

Molecule

Molecular Gene locus Cell expression weight(kDa) (chromosome)

slg

150–900 14 (!-chains) 2 ("-chain) 22 (#-chain)

Ig-$(CD79a) Ig-%(CD79b)

Function

Mature B cells

Antigen binding part of B-cell antigen receptor (BCR)

NH COOH NH2 COOH

COO COOH

34 39

19q132 17q23

Pre-B cells Mature B cells

sIg-associated molecules signal transducing part of BCR

100

3q21-q27

Pre-B cells B cells of germinal center Granulocytes

Neutral endopeptidase

95

16p11.2

All B cells incl. progenitor B cells

With CD21-, CD81Leu-13 co-receptor for BCR

35–37

11q-q13

Pre-B cells Mature B cells

Ion channel subunit

140

1q32

Mature B-cells Follicular dendritic cells (FDC)

C3d/EBV-receptor (CR2) BCR-association, signal transduction

135

19q13.1

All B cells (cytoplasm) Mature B cells (surface)

B-cell adhesion molecule (B-B and B-T interaction) modulation of BCR

45

19p13.3

Mature B cells FDC Act. monocytes eosinophils

Low-affinity Fc& receptor (Fc&RII) cleavage product = B-cell growth factor

48

20q12q13.2

Pre-B cells Mature B cells Dendritic cells (DC)

Interaction with CD40 ligand (T cell) Anti-apoptosis signal

43–39

9p

All cells of B-cell lineage Macrophages

Adhesion molecule

B cells, act. monocytes

T-APC interaction (ligands for CD28/CTLA-4)

CD10 (CALLA)

CD19

CD20

CD21

CD22

NH2 CD23

COO

CD40

NH2

CD72

COOH

60 CD80/86

A. Important B-cell antigens

3q21

Dendritic cells (DC)

! 56/10%+/*0. 742/)2&.+8

;%&(4*0/* !"<+.. =/*2>+/8

"

! " 9?

B(+-1@&+/1% 60.+'.?%&)

!

"

! CD

5&%%A5&%% *+/&01'/.$+)

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*+/&01'/.$+) 2&/3&&+ 4 5&%%) 1+- !+/.9&+
CD54 (ICAM-1)

CD4 or CD8 ICOS

B7h/ICOSL

TCR CD3 ICOS

ICOSL/B7h

CD40

CD40L

MHC class I (or class II)

CD80/CD86 (B7.1/B7.2) CD106 (VCAM)

CD80/CD86 (B7.1/B7.2)

CD28/CTLA-4

CD2 CD8 or CD4

CD28/CTLA-4 CD49d

A. Molecules involved in T cell – APC interaction APC Non APC

APC

CD80/ CD86

APC

CD80/ CD86

CD28

CD152

CD80/ CD86

CD28

CD152 T

T

MHC I antigen TCR

CD80/ CD86 CD28

CD28 T

3. Self-limitation/downregulation APC

T

Anergy/Tolerance

T-cell activation

CD80/ CD86

CD40

CD28

2. Co-stimulation via CD28

B. Several signals needed for T-cell activation APC

IL12

CD154 IFN-#

T

1. Signal 1 only

"

Upregulation of CD152/CTLA-4

MHC I

B(+-1@&+/1% 60.+'.?%&)

MHC class II (or class I)

TCR

CD58

APC = B cell

T cell

CD3

CD11a/18

CD54

APC = dendritic cell

CD58 (LFA-3)

CD2

CD40L

CD40

T cell

CD 11a/18 (LFA-1)

!

TH1

CTL

Upregulation of CD154/CD40L

4. Effector cell induction

B cell IL-6

CD80/ CD86

MHC II

AgR

TCR

CD28

B7RP-1/ ICOSL

ICOS IL-4

T

IL-13

Upregulation of ICOS C. ICOS in T-cell activation

"

IL-2

T H1

!

Superantigen Variable region of ! chain (V!)

T H2

D. Superantigen stimulation

! " CE

=>*?3)$*+3& @48*78(&$0

!

"

! AB

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-3+>43& .8&&$4 /$&&0

T progenitor cell TCR rearrangement Thymus T/NK-progenitor cell

IL-7 CD33 CD7

Bone marrow

Fetal liver

Further T-cell development

CD2+/CD5+/IL-5 CytCD3

NK progenitor cell

Mature NK cell

CD34CD33CD7+ CD2+/CD5CD1CD16CD56-

T/NK

CD34CD7+ CD2+/CD5CD1CD16+/CD56+

A. Development of NK cells

Unknown carbohydrate?

NKp30,44,46

MHC I KIR3DL1

B. Target recognition by NK cells

Cw1,3,7,8 Bw4

A3, A11

HLA-E

C. Inhibitory receptors of C. NK cells

NK cell

Activation Inhibition

Cw2,4,5,6

APO-1 Fas ligand

DNAfragmentation

NKG2D MICA MICB

CD95

Target cell

KIR2DL1

KIR2DL2

NKR-P1

Apoptosis

1. Nonsecretory lysis Target cell

Formation of pores Granzyme Perforin

Degranulation

=>*?3)$*+3& @48*78(&$0

CD34

CD34+ CD33CD2+ CD7+ CD5+ CD1+ CytCD3+

!

"

DNAfragmentation Apoptosis +necrosis

2. Secretory lysis Target cell

Perforin Granzyme

NK cell

KIR3DL1 KIR3DL2 CD94 + NKG2A

DNA fragmentation Apoptosis + necrosis

Antibody against target antigen

Fc-receptor

3. ADCC (antibody-dependent cellular cytotoxicity) D. Cytolytic mechanisms D. of NK cells

! " AC

8?26'/%2,'> &=32*3<>%.

!

"

1)2)*+,%. '26 :%26=3,3* 7%>>.

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IDC

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Langerhans cell in skin/mucosa

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Dermis/ Interstitial DC

Kupffer cells

Mesangial cell

Microglia

Tissue macrophage

LN/spleen monocyte

Alveolar macrophage

Serosa macrophage

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Coated pit

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TNF-$ IL-1 IL-6 IFN-#

IL-8

TNF-$ IL-1

3. Phagocytosis, scavenger function, tissue repair

NO

Elastase Collagenase – fibrogenic stim-factor – angiogenic factors

Acute phase proteins IL10 GM-CSF

IL12 TH2

TNF-$

T H1

5. Antigen presentation 6. Immunomodulation A. Function of monocytes/macrophages

– Toxic factors – TNF proteases

– NO – H2O2 – O2 – Lysozyme – Hydrolase

4. Effector function

MolecularGene locus Cell expression weight (kDa) (chromosome)

Function

CD11a: 180 CD18: 95

16p11-13.1 21q22.3

Monocytes, !", lymphocytes (T-B), granulocytes

Cell–cell and cell–matrix adhesion, ligand for ICAM-1, ICAM-2

CD11b: 170 CD18: 95

16p11-13.1 21q22.3

Myeloid cells, NK cells

Cell–cell and cell–matrix adhesion, ligand for iC3b, C4b, ICAM-1

CD11c: 150 CD18: 95

16p11-13.1 21q22.3

Macrophages, myeloid cells

Cell–cell adhesion, ligand for iC3b, C4b, ICAM-1, LPS, fibrinogen

55

5q31

Monocytes, !", granulocytes, (B cells)

Receptor for LPS and LPS/LPB complex

50–80

1q23

Monocytes, !", NK cells, granulocytes (GPI)

Low-affinity Fc-lgG receptor

40

1q23-24

Phagocytes, B cells, platelets

Intermediateaffinity Fc-lgG receptor

72

1q21

Monocytes, !"

High affinity Fc-lgG receptor

48

20q12q13.2

Pre-B cells, mature B cells dendritic cells (DC)

Interaction with CD40 ligand (T cell), anti-apoptosis signal

CD80/86

60

3q21

B cells, act. macrophages, T-APC interaction dendritic cells (DC) (ligands for CD28/CTLA-4)

CD83

45

6p23

Mature dendritic cells, some B cells and T cells

Adhesion receptor

180

10p13

Phagocytes, lymphoid endothelium

Pattern recognition receptor

43–49

1q22-23

Thymocytes, DC

Antigen presentation (glycolipids)

110

17p13

Monocytes, !", granulocytes, basophils, LGL lymphocytes

Lysosomal and endosomal glycoprotein

Molecule CD11a LFA-1

CD18 CD11b

CR3 CD18 CD11c CR4 CD18 CD14

or CD16 (FcRIII) CD32 (FcRII) CD64 (FcRI) COOH

CD40

COOH

Mannose receptors CD1a

CD68

B: Monocytes and DC antigens

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#&(*'$)$+ ,&-- ./01-2)$/(3

CD34+cell

!

Flt3L, IL-3 M-CSF GM-CSF TNF-!

Monocyte Langerhans cell

IL-3

CD11c+ CD13+ CD33+ CD14+ CD1c+ Pre-DC-1

CD1a+

Endothelium transmigration

Thymic DC

GM-CSF IL-4

CD1c+ BDCA3+ CD11c+ CD13+ Pre-DC-2

IDC

CD40 ligation viral infection

LPS CD40 ligation

Ag uptake

HEV Interstitial DC

GCDC

CD11c+ CD123+ CD2/5/7 (+) BDCA2+ BDCA2+ BDCA4+ TLR9+

“Lymphoid” DC2

Myeloid DC 1 CD83+

– LPS – Bacterial products

A. Different DC populations Langerhanscell

Interdigitating DC

Veiled cell

Antigen HLA-DR

CD40 CD40L

T

Macrophages

IL-2 IFN-"

NK cells

Activated T cell

B. Life cycle of a dendritic cell Epithelium (Tonsil, gut)

SLC

MIP-3!

MIP-3! CCR6

Lymphatic vessel

CCR7 SLC

1. Migration to and from inflamed tissue

CCR7 MIP-3#

Mature DC

Immature DC

"

CD80/ CD86 CD28

Eosinophils

T cells

;1(*28&()2- .'$(+$0-&3

TNF-!

2. Lymph node homing

C. Recruitment of dentritic cell precursors/DC migration

! " <=

8/%&.G$%)., 5'(%*(7,$4

!

"

-0%0*6)$4 .%& #$%&'()(* +$,,4

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! HI

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#" <0,$'03$%(* 50)$%)(., 0; FGG.)/'$ #$%&'()(* +$,,4 T%',5#% 7$'4?# J( ,664'7+# 12/ ,% '"# /?,%A 67-&/4A 4%* #;#% ,% '76&+/ 64( ,%*7-# 4%',F 5#%F/$#-,),- '&.#+4%-# +4'"#+ '"4% !F-#.. 4-',;4F ',&% ,) )7+'"#+ 64'7+4',&% /',67., 0#858A !PDA LOK3 4+# 6,//,%58 9% '"#/# -&%*,',&%/A -&F/',67.4'&+( 6&.#-7.#/ 4+# %&' 7$+#57.4'#*> 9LF
#+ -.)/'.)(0%1 +2.%3$4 (% 52$%0)67$ .%& 8/%*)(0%

Resting DC

Immature DC Mature DC

++ (+) ++ +++ + +

(+) +++ ++ ++ + (+) +++ ++++

High level of MHC: Extended long life

Foreign antigen

Antigen + LPS, TNF Recycling

A. DC maturation: changes in phenotype and function CD34+ Myeloid CD11c+ progenitor CD13/33/14+

GMCSF TNF-! IL-4

BDCA3+ Mannose rec+ CD1a/b/c+ TLR2+ TLR4+

PreDC-1

CD34+ CD11CD2/7/5(+) TCR-!+

FLT3L IL-3

CD40L LPS

CD123+ BDCA2+ BDCA4+ TLR7+ TLR9+

IL-3 CD40L

Low phagocytic capacity Low IL-12 secretion Secretes IFN-!/#(IPC)

High IL-12 IFN-"

TH

High IL-12

Low IL-12 Low IL-12

– Unmethylated CpG oligo– nucleotides – – CD40 ligation – – LPS – – IFN-" $ High DC/T ratio Both DC-2 and DC-1

CTL

PreDC-2

Viral infection

High phagocytic capacity High IL-12 secretion High costimulation

DC-1

Lymphoid progenitor

TH2

DC-2

IL-4 IL-10 IFN-"

8/%&.G$%)., 5'(%*(7,$4

CD13, CD33, CD14 CD40, CD80, CD86 CD83 LAMP CCR1, CCR5, CCR6 CCR7 Phagocytosis AG presentation IL-12 secretion

Activated DC

Low level of MHC: High turnover

!

"

IL-10 TGF# PGE2 Ox40L signalling

B. Polarization of TH responses by DCs: lineage duality vs. plasticity TH

IFN-"

Mucosa

High IL-12

+ DC-1

pre-DC-1

– low costimulation – low IL-12 secretion – IL-10 secretion

TH1 Skin Low IL-12

pre-DC-2

DC-2

TH2

C. Feedback regulation of DC-1/2

Immature DC

IL-4 Tumors

Antigenspecific T-cell tolerance

D. Tolerogenic potential of immature DCs

! " HJ

>?%@-'$%/-6 A+(%)(56$:

!

"

! BC

23! 89:/$' ;<24 89:/$'=

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#$%&'() *+,-%(.-/(&% &0 /1$ 23! 4&'56$7

1.

21B C4B long 21A

C4A

21B C4B short 21A

C4A

Chromosome 6 Short arm

!

p

21B C4B short C4A

21A

C4A

21B C4B short 21A C4B short 21A

C4A

21B C4B short 21B C4B short 21A

C4A

21A C4B long 21A C4B long 21A C4B long 21A

C4A

q

C4/CYP21 gene organization (haplotypes) 2.

MHC class II D-region

C4B C4A Bf C2

TNF !"

CYP 21P

LTB

CYP 21

B

MHC class I C

A

Direction of transcription

DPB2

3. 4.

DPA2

DPB1

DPA1

DNA

B1

B6

B1

B6

B1

B2

B3

B1

B7

B8

Exon 1

DMB

DOB

B9 B5

B1

5.

DMA

DQA1 DRB2 DRB9 DQB1 DRB1 DRB3 DRA

"

DR3, 11, 12, 13, 14, 1403, 1404 (DR52 group)

B9

Exon 3

DQB3

DR15, 16, (DR51 group)

DR4, 7, 9 (DR53 group)

B9

Exon 2

DQA2

DR1, 10, 103 (DR1 group)

B9 B9

B4

DQB2

>?%@-'$%/-6 A+(%)(56$:

21B

DR8 (DR8 group) DRB gene organization (haplotypes) Exons 4, 5, 6 DRB1 gene

1 4 9 13

25

38 57 60 67 Amino acids

74

A. Genomic organization of the HLA complex

94

DRB1 polypeptide First domain

! " BD

F*6?/.(60/' @416)13'(5

!

"

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#$! %&'()*'( ,04*)0*4( /6? :'/55 ; !''('(5

Class II

!1

NH2

T-cell receptor

"1

!

Antigenic peptide

!1

CHO

!2

CHO

!2

"2

!1

!2

N

!3

NH2 COOH

NH2

CHO

COOH

COOH

"2m

"2 m

!3

C

CD8

Exosol

COOH

Class I

Cytosol

Antigen

C CD8

N

HVIII

View from above on antigen-binding part

B. Structure of an HLA class I molecules

A. HLA molecules (schematic)

HLA-B alleles

HLA-A alleles

HLA-C alleles

New nomenclature

Old nomenclature

New nomenclature

Old nomenclature

New nomenclature

Old nomenclature

A*0101 A*0201 A*0202 A*0203 A*0204 A*0205 A*0206 A*0207 A*0208 A*0209 A*0210 A*0301 A*0302 A*1101 A*1102 A*2401 A*2501 A*2601 A*2901 A*3001 A*3101 A*3201 A*3301 A*6801 A*6802 A*6901

A1 A2, A2.1 A2, A2.2F A2, A2.3 A2 A2, A2.2Y A2, A2.4a A2, A2.4b A2, A2.4c A2, A2-ZB A2, A2-LEE A3, A3.1 A3, A3.2 A11, A11E A11, A11K A24 (9) A25 (10) A26 (10) A29 (w19) A30 (w19), A30.3 A31 (w19) A32 (w19) Aw333 (w19), Aw33.1 Aw68 (28), Aw68.1 Aw68 (28), Aw68.2 Aw69 (28)

B*0701 B*0702 B*0801 B*1301 B*1302 B*1401 B*1402 B*1501 B*1801 B*2701 B*2702 B*2703 B*2704 B*2705 B*2706 B*3501 B*3502 B*3701 B*3801 B*3901 B*4001 B*4002 B*4101 B*4201 B*4401 B*4402 B*4601 B*4701 B*4901 B*5101 B*5201 B*5301 B*5701 B*5801 B*7801

B7, B7.1 B7, B7.2 B8 B13, B13.1 B13, B13.2 B14 Bw65 (14) Bw62 (15) B18 B27, 27f B27, 27e, 27K, B27.2 B27, 27d, 27J B27, 27b, 27C, B27.3 B27, 27a, 27W, B27.1 B27, 27D, B27.4 B35 B35 B37 B38 (16), B16.1 B39 (16), B16.2 Bw60 (40) B40, B40* Bw41 Bw42 B44 (12), B44.1 B44 (12), B44.2 Bw46 Bw47 B49 (21) B51 (5) Bw52 (5) Bw53 Bw57 (17) Bw58 (17) B'SNA'

Cw*0101 Cw*0201 Cw*02021 Cw*02022 Cw*0301 Cw*0501 Cw*0601 Cw*0701 Cw*0702 Cw*1101 Cw*1201 Cw*1202 Cw*1301 Cw*1401

Cw1 Cw2, Cw2.1 Cw2, Cw2.2 Cw2, Cw2.2 Cw3 Cw5 Cw6 Cw7 Cw7, JY328 Cw11 Cx52 Cb-2 CwBL18 Cb-1

C. HLA class I alleles

F*6?/.(60/' @416)13'(5

CHO

"

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!

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Old nomenclature

DRB1-alleles

1)23-4'25-& 6782(89&'*

DRB1-alleles

"

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New Old nomenclature nomenclature

DRB1*0101

DR1, Dw1

DRB1*1304

DRw6, DRw13

DRB1*0102

DR1, Dw20

DRB1*1305

DRw6, DRw13

DRB1*0103

DR' BR', Dw' BON'

DRB1*1401

DRw6, DRw14, Dw9, Drw6b

DRB1*1501

DR2, DRw15, Dw2

DRB1*1402

DRw6, DRw14, Dw16

DRB1*1502

DR2, DRw15, Dw12

DRB1*1403

DRw6, DRw14

DRB1*1601

DR2, DRw16, Dw21

DRB1*1404

DRw6, DRw6b.2

DRB1*1602

DR2, DRw16, Dw22

DRB1*1405

DRw6, DRw14

DRB1*0301

DR3, DRw17, Dw3

DRB1*0701

DR7, Dw17

DRB1*0302

DR3, DRw18,

DRB1*0702

DR7, Dw'DB1'

DRB1*0401

DR4, Dw4

DRB1*0801

DRw8, Dw8.1

DRB1*0402

DR4, Dw10

DRB1*08021

DRw8, Dw8.2

DRB1*0403

DR4, Dw13, 13.1

DRB1*08022

DRw8, Dw8.2

DRB1*0404

DR4, Dw14, 14.1

DRB1*08031

DRw8, Dw8.3

DRB1*0405

DR4, Dw15

DRB1*08032

DRw8, Dw8.3

DRB1*0406

DR4, Dw'KT2'

DRB1*0804

DRw8

DRB1*0407

DR4, Dw13, 13.2

DRB1*09011

DR9, Dw23

DRB1*0408

DR4, Dw14, Dw14.2

DRB1*09012

DR9, Dw23

DRB1*0409

DR4

DRB1*1001

DRw10

DRB1*0410

DR4

DRB1*0411

DR4

DRB1*1101

DR5, DRw11, Dw5, DRw11.1

Other DRB alleles

DRB1*1102

DR5, DRw11, DRw11.2

DRB3*0101

DRw52a, DW24

DRB1*1103

DR5, DRw11, DRw11.3

DRB3*0201

DRw52b, Dw25

DRB1*1104

DR5, DRw11

DRB3*0202

DRw52b, Dw25

DRB1*1201

DR5, DRw12, Dw'DB6'

DRB3*0301

DRw52c, Dw26

DRB1*1202

DR5, DRw12, DRw12b

DRB4*0101

DRw53

DRB1*1301

DRw6, DRw13, Dw18, DRw6a

DRB5*0101

DR2, DRw15, Dw2

DRB1*1302

DRw6, DRw13, Dw19, DRw6c

DRB5*0102

DR2, DRw15, Dw12

DRB1*1303

DRw6, DRw13, Dw'HAG'

DRB5*0201

DR2, DRw16, Dw21

DRB1*1304

DRw6, DRw13

DRB5*0202

DR2, DRw16, Dw22

A. HLA-DR, HLA-DQ, and HLA-DP alleles in the HLA system (class II alleles)

New Old nomenclature nomenclature

New Old nomenclature nomenclature

DQA1 alleles

DPA1 alleles

DQA1*0101

DQA 1.1, 1.9

DPA1*0101

LB14/LB24, DPA1

DQA1*0102

DQA 1.2, 1.19, 1.AZH

DPA1*0102

pSBa-318

DQA1*0103

DQA 1.3, 1.18, DRw8-Dqw1

DPA1*0103

DPw4a1

DQA1*0201

DQA 2, 3.7

DPA1*0201

DPA2, pDAa13B

DQA1*03011

DQA 3, 3.1, 3.2

DQA1*03012

DQA 3, 3.1, 3.2, DR9-DQw3

DQA1*0302

DQA 3, 3.1, 3.2, DR9-DQw3

DPB1 alleles DPB1*0101

DPw1, DPB1, DPw1a

DQA1*0401

DQA 4.2, 3.8

DPB1*0201

DPw2, DPB2.1

DQA1*0501

DQA 4.1, 2

DPB1*02011

DPw2, DPB2.1

DQA1*05011

DQA 4.1, 2

DPB1*02012

DPw2, DPB2.1

DQA1*05012

DQA 4.1, 2

DPB1*0202

DPw2, DPB2.2

DQA1*05013

DQA 4.1, 2

DPB1*0301

DPw3, DPB3

DQA1*0601

DQA 4.3

DPB1*0401

DPw4, DPB4.1, DPw4a

DPB1*0402

DPw4, DPB4.2, DPw4b

DQB1 alleles

DPB1*0501

DPw5, DPB5

DQB1*0501

DQw5 (w1), DQB 1.1, DRw10-DQw1.1

DPB1*0601

DPw6, DPB6

DQB1*0502

DQw5 (w1), DQB 1.2, 1.21

DPB1*0801

DPB8

DQB1*05031

DQw5 (w1), DQB 1.3, 1.9, 13.1

DPB1*0901

DPB9

DQB1*05032

DQw5 (w1), DQB 1.3, 1.9, 13.2

DPB1*1001

DPB10

DQB1*0504

DQB 1.9

DPB1*1101

DPB11

DQB1*0601

DQw6 (w1), DQB 1.4, 1.12

DPB1*1301

DPB13

DQB1*0602

DQw6 (w1), DQB 1.5, 1.2

DPB1*1401

DPB14

DQB1*0603

DQw6 (w1), DQB 1.6, 1.18

DPB1*1501

DPB15

DQB1*0604

DQw6 (w1), DQB 1.7, 1.19

DPB1*1601

DPB16

DQB1*0605

DQw6 (w1), DQB 1.8, 1.19b

DPB1*1701

DPB17

DQB1*0201

DQw2, DQB 2

DPB1*1801

DPB18

DQB1*0301

DQw7 (w3), DQB 3.1

DPB1*1901

DPB19

DQB1*0302

DQw8 (w3), DQB 3.2

DQB1*03031

DQw9 (w3), DQB 3.3

DQB1*03032

DQw9 (w3), DQB 3.3

DQB1*1401

DQw4, DQB 4.1, Wa

DQB1*0402

DQw4, DQB 4.2, Wa

DR4

B. HLA-DR, DQ, and DP alleles in the HLA system (continued)

DW4 DW10 DW13 DW14 DW15 DW"KT2"

DRB1*0401 DRB1*0402 DRB1*0403 DRB1*0407 DRB1*0404 DRB1*0408 DRB1*0405 DRB1*0406

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

#$% %&'(( ))*/,-,./,.0 !.012,. 34,(,.0'015. Extracellular protein

!

Coat proteins Endocytosis Early endosome

Golgi MHC class II compartment

Transport vesicles

Ii

Endogenous proteins ER

HLA-DM

mRNA

A. MHC class II-dependant antigen processing

Anchor AA

P1

P4

P6

P9

N

HVIII

B. Anchor AA in MHC II peptides

Inhibitory AA

P1

Phe, Ile, Leu, Met, Val, Trp, Tyr

P4

Asp, Met, Gln, Ser Gly, Lys, Pro, Arg, Trp, Tyr

P6

Ser, Thr, Val

P9

Ser Asp, Gln, Leu, Asn, Pro

Gln, Phe, Gly, His, Lys, Leu, Met, Tyr

AB./'9,.0'& 341.61-&,(

Calnexin Terminal lysosome

"

C. Allele-specific motifs in DRB1*0401-binding peptides

IL-2

CD4

B

CD4

IL-4 IL-6

IFN-!

TNF-"

NO H2O2 O2

D. T-cell activation via MHC II

Antibody production

! CC

;1*-,6'*.,< #$)*&)=<'(

!

"

! >?

23! 45(.'6 7829 45(.'6:

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

829 9<,(( @A-'='*-'*. !*.)+'* #$'('*.,.)%*

Subunits

Ubiquitinated protein

!

Exocytosis via Golgi

! " " !

Ubiquitin

TAP Erp57 20S

19S

26S proteasome

"2m

Tapasin

Enzymatic digestion Single AA

Unfolded MHC I chain

Calreticulin

A. Processing and presentation of endogenous antigens

P2

P9

P9=Valine, Tyrosine

COO

NH3+

HLA-A*0201 P2=Leucine

Pockets in ! chain of MHC I

HLA-A3:

P2=Leucine P9=Lysine

HLA-B7:

P2=Leucine P9=Proline

MAPPQVLAFGLLLAAATATFAAAQEECVLENY KLAVNCFVNNNRQCQCTSVGAQNTVICSKL AAKCLVMKAEMNGSKLGRRAKPEGALQNND GLYDPDCDESGLFKAKQCNGTSTCWCVNTA GVRRTDKDTEITCSERVRTYWIIIELKHKAREK PYDSKSLRTALQKEITTRYGLDPKFITSILYENN VITIDLVQNSQQKTQNDVDIADVAYYVEKDV KGESLFSHKKMDLTVNGEQLDLDPGQTLIYY VDEKAPEFSMQGLKAGVIAVIVVVVIAVVAGI VVLVISRKKRMAKIEKAEIKEMGEMHRELNA

;1*-,6'*.,< #$)*&)=<'(

19S

"

C. HLA-A2-binding epitopes of a 314 AA protein

B. Binding motifs VPYGSFKHV

Non-APC cells

••TGSTA VPYGSFKHV DTRLQ•••

Lack of presentation US2 US11

KHVDTRLQ Virus infected cells – tumor cells

APC, IFN-#-treated cells

D. Immunoproteasome

E. Immune escape mechanisms

Thymic medulla dendritic cell Self antigen

Non-APC cell

CD4+/CD8+ Thymocyte

Self-antigen

Apoptosis Clonal deletion

F. Central tolerance to self antigens

Apoptosis/ anergy

Lack of co-stimulation

G. Peripheral tolerance to self-antigens

! " >B

96):.%()*.' ;3,)+,&'(8

!

"

! <=

27( #$%&'(%()* 418*(%

!" #$%&'(%()* !+*,-.*,$)

/" 01*,+ 2(3%,).' 4(56()+(

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!+*,-.*,$) .): >??(+*$38

C3

Antigen C1r C1s

Immune complex

Contact with microorganisms

C1q C1s

C1s C4

Spontaneous hydrolysis

!

C4b

C2a

C2 C3a C3b C2b Factor D

Factor B Ba

Amplification loop

C4b2a = C3 convertase of the classical pathway

C3b

Bb

Properdin

C3bBb = C3 convertase of the alternative pathway

P C3

C3b C3a

C4b2a3b = C5 convertase of the classical pathway

C3bBbP C3b C2a

C4b

Cell

1. Classical pathway A. Complement activation C5

C5a

C3bBb3b = C5 convertase of the alternative pathway

C3b C3bBb

Cell

2. Alternative pathway C6

C7

96):.%()*.' ;3,)+,&'(8

C4a

"

C8

C5 convertase C5b

C9

C9

C5b67

(Up to 14 molecules)

C5b678

Inhibition of the stable C9 bond by CD59 (MAC-inhibitor)

CD59 Unstable C9 bond

Stable C9 bond

B. Lytic terminal sequence

Poly-C9 membrane attack complex (MAC)

! " <@

:,*@45'*&4- #$)*()6-'/

"

#

=>' 0%56-'5'*& ?2/&'5

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0" 8)%-%.)(4- 9++'(&/ %+ 0%56-'5'*&; <55,*%-%.)(4- 9++'(&/

" AB

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!

!

3'.,-4&)%* 4*@ 9++'(&/

C2 C4b

C2a

C4b

C2a

C4b

C4b

FI

DAF

1. Blockage of binding

C3b

FI

CR1

2. Dissociation of C2a and C4b, 2. enhanced by DAF or CR 1

"

C3c C3dg

CR1

3. Cleavage of C4b and C3b by CR1/FI

Anaphylatoxins

1. Contraction of 1. smooth muscles

2. Increased permeability of blood vessles

3. Degranulation 3. of basophils

:,*@45'*&4- #$)*()6-'/

A. Regulation of complement effects: protection of autologous cells

4. Chemotaxis, release 4. of O2-radicals and 4. lysosomal enzymes

C3a

+

+

+

-

C4a

(+)

(+)

(+)

-

C5a

++++

++++

+

++++

#

B. Biological effects of complement factors: inflammatory effects C3b/C3d/iC3b receptor

1. Antimicrobial effects APC

2. Clearing of immune complexes FDC

Killer cell

FcR

MHC

T cell

3. Cell adhesion

C3b CR1

C3d CR2

FcR

B C3b CR1

Target cell

B

B cell

B CR2

4. B-cell stimulation

C. Biological effects of complement: immunological effects

C3d Memory B cell

" AC

A2*/$?)*%$1 #3.*-.51),

!

"

=**$%) =??2*.%@

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6" #$%%)3* 4)-'(*.%.'* 4)-)5%'3,

! BC

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PAMPs

Bacterial DNA Mycobacteria Gram +ve bacteria Gram -ve bacteria

Yeasts

PRR

– CpG

Toll-like-receptor 9

– LPS

Scavenger receptors, LBP, CD14, Toll-like receptors 4

– Lipoproteins

Toll-like-receptor 2

– Peptidoglycans – Lipoarabinomannan

CD14, Toll-like-receptor 2 CD1, Toll-like-receptor 2

– Mannan

Mannose receptor Mannose-binding protein

– Zymosan

Mannose receptor, !-glucan receptors, Toll-like receptor 2

A. Pathogen-Associated Molecular Patterns and Pattern Recognition Receptors Bacteria

C3

Mannosebinding lectin

“Acute phase” MASP1

C3b Complement activation

MASP2

C3a

1. Secreted pattern recognition receptors Mannose receptor MHC II

Lysosome

Scavenger receptor

Presentation

! A2*/$?)*%$1 #3.*-.51),

#$%&'()*+$,,'-.$%)/ 0'1)-21$3 #$%%)3*, $*/ #$%%)3* 4)-'(*.%.'* 4)-)5%'3,

"

2. Endocytic pattern recognition receptors

MD-2

MyD88 TLR4

Bacterial lipopolysaccharide

CD14

Adapter proteins

P IRAK TRAF 6

Nucleus

NF"B

Activation of inflammatory and immune response genes

MAPK

3. Signaling pattern recognition receptors B. Pattern recognition receptors LPS

I"B

IL-1, IL-12

Toll-like receptor MHC II

Endocytic PRR

CD80/86

Clonal expansion

TCR

CD28

Quiescent T cell

C. Dendritic cells as a link between innate and adaptive immunity

Help

! BD

@%/+0?$/*09 A3./(.B9$-

!

"

#$%&'()*$ :.;30*.'/

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#$%&'()*$ !+,$-.'/ 0/+ :.;30*.'/

!

Endothelium

Glycoprotein PSGL-1

Rolling L-selectin

Activation !4"7 !4"1 integrin integrin

7TMR

Arrest

! L" 2 integrin

! M" 2 integrin

ICAM-2

ICAM-1

!4"7 integrin*

!4"1 integrin*

Chemokines

P-selectin E-selectin

PNAd

MAdCAM-1 VCAM-1

MAdCAM-1 VCAM-1 *activated

A. Leukocyte adhesion and extravasation T cell CCR 7

HEV

SLC

CXCR5

Lymphoid follicle

BLC

T cell CCR 7

ELC

B cell FDC, stromal cells

c

@%/+0?$/*09 A3./(.B9$-

Tethering

"

DC, MØ

C. B-cell migration

B. T-cell migration

IL-8

MCP-1

RANTES MIP-1! TNF-!

CCR1 CCR2 CCR5 (CCR6) Immature DC CXCR1

D. DC migration into inflamed tissue

CCR7 CXCR4

Mature DC

Migration to lymph nodes

! CE

!

31.4(5(6-215 7889,& :&241,-'8' 1,; #(5&+1,2& !"#$$#%&'()*+ ') ,)$#(-* %*'(-#*+ &%* .#%/ ') '(++0# /%1%-#2 30&4 $#+5)*+#+ %$# 6*)7* %+ !"#$%&$'&()(*()" %$+,)(-'&8 74(&4 %$# /("(/#/ (*') ,)0$ '95#+2 :95# ;<;;; $#%&'()*+ %$# 1#/(%'#/ =9 %*'(=)/(#+8 74#$#%+ '95# ;> $#%&? '()*+ %$# 1#/(%'#/ =9 : &#..+2

<9,;18&,.15 3+-,2-%5&'

!" #$%&' () *$%&+'&,'-.-/-.$ 0&12.-(,'

"

! ==

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*$%&+'&,'-.-/-.$ 0&12.-(,'

Food

Pollen

Dust mite

Fc!R

Antigen

Antigen Second antigen contact Increased permeability of blood vessels

Bronchial constriction

Insect venom

Reexposure to antigen

Mucus secretion

!

Urticaria

IgE

Penicillin

Basophil/mast cell

Mother RhD-

Antigen

Fetus RhD+ R'

S

CH3 CH3

O

C

Complement

Anti-BM antibodies

1st pregnancy

Immune complex

N COO

Post partum

Degranulation, mediator release

Immune complex deposit

antiRhD

C3a C5a

Pulmonary bleeding

Lysis

2nd pregnancy

Hapten

Hapten – carrier complex

Langerhans cell

"

Enzyme release, cell damage

Nephritis

Type II : Cytotoxic antibody reactions

<9,;18&,.15 3+-,2-%5&'

Type I: Immediate hypersensitivity

Type III: Immune complex reaction

Lymph nodes

Hapten/antigen reexposure

Epidermis

Dermis

Induction of antigen-specific T cells Type IV: Delayed type hypersensitivity (DTH ) Lymphatic

A. Types of hypersensitivity reactions

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Adult mouse, strain A

Embryonic mouse, strain B

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Dendritic cell + antigen Tolerance to A

Mature T cells of strain B

T cell activation against A A. Induction of T-cell tolerance by antigen-presenting dendritic cells Mature T cell

Auto-peptide/MHC not accessible

Foreign antigens/recognizable autoantigens

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T -cell receptor Co-receptor

Ignorance innocence

Elimination, deletion, loss by consumption

Anergy

TCR/ co-receptor modulation

Regulatory cells

B. Peripheral mechanisms of tolerance induction Fertilized egg

Microinjection of foreign gene

Implantation of egg in syngenic mouse

Own gene product Foreign gene product

Tolerance

C. Transgenic mice

Transgenic offspring

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TC Resting T cells Islet cell Antigenic peptide

TH

Cytotoxicity

APC Signal

Signal

Activated T cells

A. Induction of an autoimmune response by virally induced T-cell activation Antigen Foreign part Autoantigenic part

Surface immunoglobulin

Autologous cell

Virus Auto antigen

Viral antigen

T cell Processing B cell

Autoantibodies

C. Induction of an autoimmune reaction C. by molecular mimicry Target cell and virus

HLA class II antigen presentation

Viral antigen

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T cell that recognizes viral Ag

Help T-cell receptor

IFN-! HLA class II antigen

T cell

Autoantigen Cell damage

B. Induction of autoantibodies by T-cell help after autoantibody-mediated antigen presentation

T cell that recognizes auto-Ag

D. Induction of an autoimmune response after viral infection by aberrant MHC class II antigens

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Organ-specific diseases

Non-organspecific diseases

Brain MS

Brain SLE

Cartilage Polychondritis

APC

TH

B

T-helper/ inducer cell Macrophage TR Presentation of autoantigens

Regulatory cell

Autoaggressive effector functions

Cartilage

Kidney SLE, Wegeners’ granulomatosis

Intestine Crohn’s disease, ulcerative colitis Bone marrow Autoimmune hemolytic anemia, ITP Testes

C. Sequestered antigens Disease

Muscle/skin Dermatomyositis

Liver PBC, autoimmune hepatitis Pancreas Juvenile diabetes

d er e es t

qu Hidden antigens An tig en s

Se

Lung Scleroderma, MCTD, Wegeners’ granulomatosis

Stomach Pernicious anemia

TZ

A. Induction of autoimmunity by loss A. of regulatory mechanisms Lens

Nose Wegener’s granulomatosis

Thyroid gland Hashimoto’s thyroiditis, primary myxedema, thyrotoxicosis (Graves’ disease)

Allele*

Skin Pemphigus

Joints Rheumatoid arthritis Skin Scleroderma, SLE

B. Organ-specific and non-organB. specific autoimmune diseases Frequency (%) Patients

Controls

Behçet’s disease

B5

41

10

6,3

Ankylosing spondylitis Reiter’s syndrome

B27

90

9

87,4

Acute anterior uveitis Subacute thyroiditis Psoriasis vulgaris Dermatitis herpetiformis Celiac sprue Graves’ disease Diabetes mellitus type I Myasthenia gravis Systemic lupus erythematosus Idiopathic membr. nephropathy Narcolepsy Multiple sclerosis Rheumatoid arthritis Hashimoto’s thyroiditis Pernicious anemia Juvenile chronic arthritis

Relative risk

B27

79

9

37,0

B27

52

9

10,4

B35

70

15

13,7

Cw6

87

33

13,3

DR3

85

26

15,4

DR3

79

26

56

26

10,8 3,7

DR3 DR3 and/or DR4 DR3

91

57

7,9

50

26

2,5

DR3

70

26

5,8

DR3

75

26

DR2

100

25

12,0 N.D.

DR2

59

25

4,1

DR4

50

19

4,2

DR5

19

6

3,2

DR5

25

6

5,4

DRw8

23

8

3,6

D. Associations between diseases and the HLA system (*old nomenclature)

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#" =<05/):,/*$* /% !2/2,/,$) 8&77* I&/.*4 +2)2')$.$; 20)$20+'.-,#$"/.*" 7LD8 .$ "A2)$"- +' '0" ("## $&/3+("9 D(+1"*4"/ /"("2')/$ ? +*- "*-)')A.* /"("2')/ !IN] (+* 5.*- ') LD9 :0" 1.'/)*"('.* /"("2')/ ())2"/+'"$ 6.'0 '0" '0/)%5)$2)*-.* 7:DL8 /"("2')/ !IGH ') 5.*:DL9 :/.44"/.*4 )3 '0"$" /"("2')/$ .*-&("$ +('.* /"+//+*4"%"*'; #"+-.*4 ') .*'"/*+#.E+'.)* )3 +2)2')'.( ("##$9 :0" /"#"+$" )3 2/).*3#+%%+')/, (,')B.*"$ )/ (0"%)B.*"$ .$ -)6*/"4&#+'"-9

!2/2,/*$*

Swelling of the cell, damage to organelles

Apoptosis signal

Chromatin condensation

Inflammation

Necrosis

Cell shrinkage, zeiosis

Lysis – organelles destroyed chromatin destroyed

Chromatin margination

Segmentation of the nucleus, DNA fragmentation

Apoptotic bodies

Phagocytosis: no inflammation

A. Differences between necrosis and apoptosis Oxidants Apo-1/FAS antigen CD95 Glucocorticoid receptor

Calcium

T/Bantigen receptor

Active caspases

Mitochondrion Cytochrome c release

i

O2, H2O2 OH, NO

Bax

Ceramide

Loss of cellcell contact Hyperthermia

DNA damage (UV light, irradiation)

-p53 -nur 77 -c-myc -ICE

-bcl-2 -bcl-xL -Rb

Activation of further caspases, apoptosis

C. Caspase activation by mitochondria Apoptotic cell TSP

Apoptosis promoting genes

Vitronectin receptor

-bax -bad -bcl-xS

CD36

PS Scavenger CD14 receptor-A

Actin polymerization Actin

Survival B. Regulation of apoptosis

"

Granzyme Deprivation of growth factor

Effector genes

Apoptosisinhibiting genes

Apaf-1 pro-caspase 9

>6(10?&(,07 ='$()$27&*

– Ischemia – Hyperthermia – Hypothermia – Physical or chemical damage – Trauma

!

Actin

Anti-inflammatory response TNF-"

IL-1!

MCP-1

IL-8

Apoptosis D. Phagocytosis of apoptotic cells

! " @B

D/(0)/.0)C !--'%,/.%017

!

"

!1.%*$1B!1.%(0&C =1.$)/,.%017

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2" +)$,%-%./.%01 6$/,.%017 %1 8'3%& +9/7$

! EF

10"2%3%4#'"5, @* (*'%1#* .(0&7# %. &7($#/ %* ( $-B#''# (*/ (77)<#/ ') "#($' <%'+ (* #>$#.. )6 (* (*'%.#"-0 $)*'(%*%*1 '+# $)""#.&)*/%*1 (*'%2)/34 =)7-27# %00-*# $)0&7#>#. 6)"04 5+# $+(*1# %* '-"2%/%'3 %* '+# $-B#''# %. 0#(9 .-"#/ 23 &+)')0#'"34 5) /#'#"0%*# '+# #*/ &)%*': '+# %*$"#(.# %* (2.)"&'%)* <%'+%* ( $#"9

'(%* '%0# %. -.#/ (. '+# 0#(.-"# )6 (*'%1#* $)*9 $#*'"('%)*4 6#&7#/)4#'"5, 5+%. '#$+*%,-# %. (7.) 2(.#/ )* '+# "#($'%)* &"%*$%&7# )6 %00-*# $)0&7#> 6)"0('%)* /-# ') (*'%1#*I(*'%2)/3 %*'#"($'%)*4 5+# $-B#''# %. %""(/%('#/ <%'+ 7(.#" 7%1+': <+%$+ .$(''#". 6")0 '+# %00-*# $)0&7#>#.4 J#*.#. ("# -.#/ ') 6)$-. '+# .$(''#"#/ 7%1+' )*') ( &+)')0#'#"4 K.%*1 '+# 0#(.-"#/ .%1*(7.: '+# -*D*)<* (*'%1#* $)*$#*'"('%)* $(* '+#* 2# /#'#"0%*#/ 6")0 ( .'(*/("/ $-"B#4

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

:$>%1%.%017 /1& +)$,%-%./.%01 G$,91%H3$7

!

Immune complex formation with molecular antigens

Size of immune complexes (precipitate mass)

Precipitation Ag surplus

Equivalence range

Ab surplus

Ag

Immune complex formation with antigenic particles (e g. erythrocytes, latex particles)

Ab

Agglutination A. Heidelberg curve

B. Precipitation and agglutination

Turbidimetry (Extraction measurement)

Laser/ quartz lamp

Photodetector Cuvette Scattered light detector

D/(0)/.0)C !--'%,/.%017

Ag : Ab ratio

"

Nephelometry (Scattered light measurement)

Gel with antibodies in solution Spot for antigen to be tested Precipitate rings (equivalence range) d

Ag-concentration C

C. Precipitation techniques in fluid phase

! C ~ d2 4

Standard curve

d

Relationship between diameter d of precipitate ring and antigen concentration D. Simple radial immunodiffusion (RID) according to Mancini

! " EE

D$+)7$6)78 !>>'&4$6&)/1

!

"

! EF

!/6&A,/C!/6&+)%8 -/6,7$46&)/1

!" #$%&$' ()*+', -..*/)%&00*1&)/ 23*456,7')/8 9,65)%:

(" #)4?,6 <',467)>5)7,1&1 2<',467)>5)7@ ,1&1 &/ !/6&+)%8@4)/6$&/&/A B,':

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

!" #$%& #'4$"%50'( +"#%,'"& 9%,1+#' #$1*0,$ + ,'2 4*"#+%"%", +" +"#%)*-. #*3+1- +" +"*-'6 :$%& 2'+-& #* #$' /*19+#%*" */ 2*",( 1*4<'#? &$+8'- 81'4%8%#+#'& =1*4<'#&> #$+# 4+" )' ;%&0+? 2%K'- ). &#+%"%",6 :$%& 9'#$*- 8'19%#& 50+"#%? #+#%*" */ #$' +"#%,'" 4*"4'"#1+#%*" ). 4*98+1? %", #$' #'&# &8'4%9'" #* + &%902#+"'*0&2. 9'+&01'- &#+"-+1-6

;" <',467)&..*/)%&00*1&)/ 2=)*/6,74*77,/6 <',467)>5)7,1&1: @$'" + &+982' %& +-A0&#'- #* + ,%;'" 8B ='6,6( 8B C6D /*1 #$' #'&# +"#%,'"E 'F#1+4#+)2' "042'+1 +"#%,'"&( GHI>( #$' +"#%,'" +"- +"#%)*-. 9%? ,1+#' %" *88*&%#' -%1'4#%*"& #$1*0,$ #$' ,'2 -0' #* -%//'1'"4'& %" #$'%1 '2'4#1%4 4$+1,'&6 !990"' 4*982'F'& /*19 -01%", #$' #1+"&9%? ,1+#%*" 81*4'&& %/ #$' 4*11'&8*"-%", +"#%)*-. %& 81'&'"# %" #$' 8+#%'"#J& &'1096 :$' %990"' 4*982'F'& /*19 2%"'& */ 81'4%8%#+#%*" #$+# 4+" )' ;%&0+2%K'- ). &#+%"%",6

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

G,45/&H*,1 )0 <',467)>5)7,1&1

Anti-!, "

Anti-!, "

y

x

Anti-!, "

y

x

y

x

x

y

x

y

x

y

Antigen

!

!

!

"

!

!"

or:

!"

!"

or:

"

" Identity x=y

No identity x#y

Partial identity x y

A. Radial double diffusion (Ouchterlony) Principle Anode

Cathode

Ab Ag Anti-ENA (anti-ENA) (ENA) B. Transmigration electrophoresis ( countercurrent electrophoresis ) Start Anode

Serum

Cathode

Electrophoresis Alb

!2

!1

"

ENA

D$+)7$6)78 !>>'&4$6&)/1

Possible precipitation patterns of anti-!, "antiserum with mixtures of the antigens !, " in spots x and y

!

"

$

Antiserum/diffusion in gel

A

n t

i

s e r

u m

Precipitation Immunoelectrophoresis

C. Immunoelectrophoresis (time course of procedure )

Ag

Cathode

Ag

Anode Precipitation line

D. Electrophoresis in antibody-containing gel (rocket electrophoresis)

! " EI

?)6*>)&*>< !44$'-)&'*(0

!

"

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Before: chemical binding

Agglutination of Ag-loaded test erythrocytes in the presence of specific antibodies in patient serum

Ag

!

Sheep erythrocyte

1. Hemagglutination

IgG Agglutination = positive test reaction IgM-RF in patient serum

Latex particle

2. Latex agglutination in rheumatoid factor screening A. Agglutination techniques Principle: competition between test system(Ag + Ab) and indicator system (erythrocytes loaded with test Ab) for complement Complement Ag

+

Pre-incubation

Indicator system

Ab Test system Complement Ag (exogenously added) Ab (patient serum)

Test system Positive result (complement bound by antigenantibody complex)

(test erythrocytes, loaded with test Ab)

?)6*>)&*>< !44$'-)&'*(0

RF (IgM anti-IgG)

"

No hemolysis

Negative result (complement available for lysis of test erythrocytes

Hemolysis

Complement

Test system (no specific Ab present)

B. Complement-binding reaction (CBR)

+ Test system

Indicator system

! " @C

*53025402& !GG=+A54+0$1

!

"

!$4+D($F!$4+30-& .$4(25A4+0$1

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Indicator-H (colored) + H2O

Indicator (no color) + substrate (H2O2)

!

Color reaction

E

E

E

E

Anti-mouse antibody Monoclonal Ig

E

E

Antigen

Detection of specific antibodies directed against an antigen by enzyme-labeled secondary antibodies ( conjugate )

*53025402& !GG=+A54+0$1

Polyclonal Ig (e.g. rabbitanti- xy)

Sandwich ELISA for sensitive antigen detection (e.g. cytokines)

E

Antibody

Antigen

Secondary antibody, enzyme linked

A. Enzyme-linked immunosorbent assay (ELISA)

Serum + tracer

High titer: low radioactivity

E

C1q

* * *

Test Ab

Measurement of radioactivity

Patient antibodies

*

Ig Anti-human Ig with subsequent color reaction

MW (kDa)

*

*

"

Antigen at solid phase Principle: antibody in patient serum competes with radioactively labeled test antibody

B. C1q solid-phase ELISA 1. SDS-PAGE

Low titer: high radioactivity

C. Radioimmunoassay (classical method) 2. Transfer PAG

3. Staining

NC

100

AP

80 50 41 31 10

Mouse Ab against human Ig Human Ab against B. burgdorferi

D. Immunoblotting (Western blot)

AP

Alkaline phosphatase B. burgdorferi proteins

Immunoblot

! " HK

<8:&'84&'3 !==$-*84-&+)

!

"

!+4-7(+9!+4-:&;3 /+4('8*4-&+)

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/,,%+&0$%&'()*(+*(

Energy (nm)

400 500 555 600

Fluorescing substrate

UV radiation

FITC excitation

FITC fluorescence

(Invisible)

Exciter filter 450490 nm

520560 nm Bandpass filter

Energy decrease Green (visible)

Fluorescence

700

300

1. Generation

400 500 ![nm]

600

2. Absorption/emission2. spectrum of FITC

A. Fluorescence Fluorescing antibody

FITC-

1. Direct immunofluorescence

PE-conjugated antibody

Fluorescing anti-Ig antibody (anti-mouse)

4. Intracytoplasmic 4. fluorescence

3. Indirect immunofluorescence B. Immunfluorescence Photomultiplier 530 – 550 nm (green)

Lightsensitive diode

Forward scatter

Granulocytes

Lymphocytes Forward scatter

2. Separation of cell fractions

C. Flow cytometry Negative control No fluorescing cells

Anti-CD3 antibody Negative Positive

Fluorescence intensity

1. Histograms D. Flow cytometry: Histograms

Fluorescence intensity

CD3+ CD4-

CD3+ CD4+

CD3CD4-

CD3CD4+

CD3-FlTC

75% positive cells

Cell count

Cell count

Monocytes

"

Side scatter

Side scatter

560 – 580 nm (red)

1. Principle

3. Fluorescence microscope

2. Double fluorescence

Primary antibody (mouse anti-human)

Cells in suspension

Dichroic mirror >510 nm

Light

<8:&'84&'3 !==$-*84-&+)

365

!

CD4-PE

2. Dot plot, two-color analysis

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E.g. lymph node or spleen or N2 tumor tissue Shock freeze Surgical specimen

– Heat – Formamidase – Alkaline solutions

!

Rupture of H-bonds: single-strand DNA

Thin sections ( 4-8 µm) on microscope slides

Cryostat sections

Fluorochromelabeled complementary DNA-probe

1. Preparation of samples 1. With DNA probes 3 Streptavidin/peroxidase 2 Antibody B (rabbit anti-mouse, biotinconjugated )

S POX

1 Antibody (murine)

+ Chromogenic substrate (DAB: diaminobenzidine)

Section with antigen

RNA

+ Complementary RNA probe, digoxigeninlabeled Hybridization

Mild fixation F

F

F

F

F

2. Biotin–avidin/peroxidase staining Alkaline phosphatase

+ FITC-labeled antidigoxigenin Ab

RNA-specific fluorescence

3 AP–AAP complex (mouse Ab)

+ Chromogenic substrate

1 Primary antibody (mouse)

2 Bridging antibody (rabbit)

2. With RNA probes

E/8'3/+'37 !>>,)./+)'&*

Hybridization

"

B. Fluorescence in-situ hybridization B. methods Chromosome 8 Chromosome 21

Chromosome 8qChromosome 21q+

3. APAAP staining

ETO gene

AML 1 gene

t(8:21)

4. Detection of 5. Staining of the tumor cells in follicle mantle the bone marrow with CD22 Abs A. Immunohistological staining

Normal cell

t(8:21) cell

Interphase cytogenetics C. Example of FISH staining in 8:21 C. translocation

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1/&& #$%&'()%* 9/.5*)E-/$

Centrifuge

!

Mononuclear cell fraction Ficoll Ficoll (density: 1.077)

Granulocytes, erythrocytes, platelets

A. Isolation of mononuclear cells from peripheral blood

Non-T fraction (B lymphocytes, monocytes

Sheep erythrocytes Centrifuge Mononuclear cells

Ficoll

Binding of sheep erythrocytes to T cells: rosette formation

T lymphocytes, sheep erythrocytes

B. Separation of T and B lymphocytes: Rosette formation Antibody labeled with iron beads

Antigenbearing cells adhere

Antigennegative cells

1. Panning method

Antigenbearing cells attracted by magnet

Antigennegative cells

Magnet

Plastic surface coated with antibodies

2. Immunomagnetic separation

:'@%0'(%0; !44&).'()%*$

Diluted blood

"

C. Antibody-mediated separation of cell fractions Cells in suspension Vibrating cuvette

Photomultiplier for side-scattered light

Bandpass filter

Dichroic mirrors

Laser

–

+ +

+ Positive cells

–

Photomultiplier for light 530–550 nm (green)

Photomultiplier for light 560–580 nm (red) Light-sensitive photodiode

Forward scatter

– Negative cells

D. Cell separation by flow cytometry

! " DF

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Stimulus

T0

Stimulus

4

12

24 h

!

CD69 [Ca2+]i

CD71

GM-CSF IL-2, IL-4 IFN-!

HLA-DR

H-thymidine

5% CO2

4

6

8

10 min

2. Flow cytometry [Ca2+]i

3

37˚C

2

3. Expression of 3. activation antigens Dist. H2O

= Radioactive

Glass fiber filter captures cells

16 h, 37 ˚C, 5%CO2 Cell division: DNA synthesis incorporation of 3H-thymidine in new DNA strands

Scintillation fluid Vacuum

"-counter – Mononuclear cells – Stimulus (lectins, antigens) – Growth factors B. Proliferation assay

Waste

='>(/'$(/< !??0%#'$%(),

0

1. Cytokine measurement 1. (ELISA) A. Activation assays

48 h

CD25

"

cpm (counts per minute)

Proximal 1 Tetanus

Proteus 8

2 Diphtheria

Trichophyton 7

3 Streptococcus

Candida 6 Control 5 (glycerin)

C. T-cell function in vivo: Mérieux multitest

4 Tuberculin Distal

! " @C

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!

"

! AB

3$22;2'& >::;%)(6

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!%(),$%-4/$0)+)0 1$4(4

+ Irradiated autologous MNC + Antigen +IL-2 4-7 days

Cell growth 7 days

37°C

37°C

5% CO2

5% CO2

Limiting dilution 1000 cells/well 100 cells/well 10 cells/well 1 cell/well 0.3 cell/well

!

Repeat 3 – 4x

– Mononuclear cells (MNC) – Antigen – Human AB serum

+ Antigen + IL-2

Medium

?'@*&'(*&6 !//2)0'()*%4

A. Generatiion of antigen-specific T cell clones No effector cells = background Effector/target ratio1 : 1 Effector/target ratio 5 : 1

Washing Binds to intracellular proteins

Effector/target ratio 20 : 1

+ Triton

Maximal cell lysis cpm probe – cpm background cpm maximum – cpm background

x 100

"

B. Cytotoxicity assay: Chromium release assay Target cell + 3H-thymidine

Labeled DNA

1. Dist. H2O

Target cell apoptosis

Fragmented DNA

2.

!-counter

Lysis in %

Filter

3.

Effector cell

waste

C. Cytotoxicity assay: Jam test

Only intact DNA sticks to filter

cpm without effector cell

cpm probe x 100

cpm without effector cells 4.

cpm = counts per min

! " AC

(1?801/807 !@@4:21/:8.<

!

"

! AB

63445410 #>>5.:/7

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!<<17 *0823C503< D80 6E1012/30:F:.; !./:;3.&<@32:D:2 , 2344<

Coating Ab

Over night

24–28 h

Wash

Antigen-presenting cells, antigen, T cells Detection Ab

1h

!

E

E E

Wash

Wash

Chromogenic substrate

(1?801/807 !@@4:21/:8.<

E

A. IFN-" ELISPOT ?

!-CD4PerCP

!-CD8AP 2

Surface staining Intracellular cytokines Wash + fixate with PFA

!-IL-4PE

Permeabilize with saponin

!-IFN-" FITC

CD-8-APC

"

IL-4-PE

Intracellular staining CD4-PerCP

B. Intracellular IFN-" staining

IFN-"-FITC

CD45

Restimulation, IFN-" production for 45 min

Bispecific Ab, !-CD45 ˜!-IFN-" PE

FACS sorting

!-IFN-"-Ab PE

PE

MACS sorting

!-PE bead

C. Cytokine secretion assay

PE

! " AG

<,4-0,)-06 !=='*(,)*-.>

!

"

89:-0,' ;::9.*)6

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! ?@

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A&>)> -1 #$%&'' B9.()*-.

Antigen

Marker T0

Stimulus

4

24 h

!

CD69

[Ca2+]i

CD71

CD23 0

Cross-linkage of slg

Activated B cell

2

4

6

( )

( )

8 Min

Increase in intracytoplasmic calcium

Expression of activation antigen

<,4-0,)-06 !=='*(,)*-.>

Anti-IgM antibody, SAC

( )

CD25

A. B-cell activation H-thymidine incorporation (cpm)

IL-2 IL-14 (BCGF) IL-4 sCD 23

CD40 ligand, anti-CD40 antibody

30 20 10

3

Activated B cell

40

Anti-IgM

IL-14

B. B-cell proliferation

SAC

Anti-IgM Anti-IgM + + IL-14 anti-CD40

Sheep erythrocytes coated with anti-Ig serum + Complement

+

"

Hemolytic plaques

B

Agarose gel

1. Inverse plaque-forming cell assay E E

E

Washing

Washing

+

+

E

Ab-producing B cells

E

E

Antigencoated Petri dish

E

E

Enzyme-linked anti-Ig antibodies

2. ELISPOT test C. B-cell differentiation: Antibody secretion

Gel with chromogenic substrate

EE E EE

EE E EE

! " ?C

>4?$)4&$)2 !@@,-74&-$*5

!

"

! AB

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!*4,2&-74, C(7'*-;%(5 Slot

!

15.8 kb 13.1 kb 9.0 kb 7.1 kb 6.5 kb 2.9 kb 1.0 kb b

c

DNA transfer filter

Autoradiography film or enzymatic color reaction

labeled DNA probe xy

>4?$)4&$)2 !@@,-74&-$*5

a

Agarose gel after gel electrophoresis 75 V; 60 mA A. Southern blotting

28S 18S 12S 5S a

b

c

RNA transfer filter labeled cDNA-probe xy

Region to amplify Denaturation to DNA single strands Hybridization with strandspecific primers

Second heat denaturation, hybridization with primers and synthesis of complementary strands

!-gt-11 left arm

!-gt-11 right arm

PCR mit

Streptavidincoated particle

Millions of copies of DNA product

C. Polymerase chain reaction

Biotin-conjugated primers Nonconjugated primers

Separation of single strands by streptavidin-linked magnetic beads C

G

Sequence analysis

PCR cycle repeated 20–35 times

"

Insert

A

Primer sequence

Autoradiography film or enzyme color reaction

D. DNA sequencing

T

Direction 5'-3'

Agarose gel after gel electrophoresis 75 V; 60 mA B. Northern blotting

3'ddTACGGAGCTGATTA.... 3'ddACGGAGCTGATTA.... 3'ddCGGAGCTGATTA.... 3'ddGGAGCTGATTA.... 3'ddGAGCTGATTA.... 3'ddAGCTGATTA.... 3'ddGCTGATTA.... 3'ddCTGATTA.... 3'ddTGATTA....

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Pyoderma

Meningitis

Albumin

Infections of upper and lower airways Sepsis

Xq21

X-linked recessive

! 1 !2 "

Fc# receptor IgG1 + IgG3

C1q

Ag

Opsonization, phagocytosis, Ab-mediated cellular cytotoxicity Absence increased susceptibility to bacterial infections

IgG2

Half life (t 1 2 ):

Not able to activate complement

of IgG1 = 23 days t1

IgG4

Produced from 2nd year or life on; Ab against bacterial polysaccharides, teichoic acid Absence increased susceptibility to infections

S. pneumoniae, H. influenzae, N. meningitidis

Complement Dangerous when other subclasses cascade are absent

2

~12 days

IgG3

Dangerous when IgG1 is absent because of short half-life of IgG3

1. Selective IgG subclass defects/properties of IgG subclasses Mutations in the gene for CD40 ligand

TCR

2. Hyper-IgM syndrome B. Dysgammaglobulinemia pre-B Arrest in pre-B phase Impaired TH regulation Defective glycosylation of Ig

MHC

IgM or IgD

#

Class switch

Xq26

Pro-B

Ag

5%&'&:,% 8--0'.%.+3

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#-fraction is missing

Serum protein electrophoresis

A. Bruton’s agammaglobulinemia Bacterium

"

CD40 ligand

CD40

Virgin-B Circulating B cell

Auto-Ab Association with HLA-A1, B8, DR3, see SLE-association Plasma cell

C. CVID: Common variable immunodeficiency – possible causes

IgG or IgA

Recurrent pneumonia

Bronchiectasis Gastrointestinal symptoms: spruelike celiac disease, diarrhea, malabsorption

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TCR, " chain

IgG, heavy chain

RAG I, II-genes

A. Forms of SCID 1. 22q11

2.

Eustachian tube Pharyngeal tonsil

3. 4. 5.

X-linked recessive

"

!

IL-2-receptor Clinical picture Thymic hypoplasia

Facial abnormalities Hypoparathyroidism

Thymus

Embryonic developemental disorder of 3rd and 4th pharyngeal pouches

#

Point mutationXq13

Frequent infections

Parathormone Lower

Parathyroid Upper glands Ultimobranchial body (C cells)

Lateral pharynx

cCa2+

Aortic arch malformation

cp i

Hypocalcemic seizures

B. Di George syndrome Autos. recessive, heterogenous group

Ionizing radiation #, ", !, X

1. Oculo-1. cutaneous 1. telangi1. ectasia 2. Cerebellar 2. ataxia

Chromosomal instability

Breakages and translocations in chromosome 14 (gene loci for TCR and Ig)

#

Impossible to repair DNA damage

Thymidine dimer

3. Dysmorphism

';+,+18; .))/,(;(62

V+D+J+C

"

Caution during radiodiagnostics!

Clinical triad

C. Ataxia telangiectasia Xp11.22

1.Frequent infections

Thrombocytes

CD43 (Sialoglycoproteins)

T cells Bundle formation of actin filaments in cytoskeleton

D. Wiskott – Aldrich syndrome

Impaired function

2. Eczema 2. constitution 3. Thrombo3. cytopenic 3. purpura: 3. epistaxis; 3. petechial 3. skin bleeding; 3. melena

Clinical triad

1 2

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4" 56*7(&89:(;&36( +2$7/1<*

! BCD

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./&$0)1-2'(- ?*%(-(*$-(*3

Glucose-6- P -dehydrogenase

Hexose monophosphate pathway

Glucose-6- P

Ribulose-6- P + CO2

NADP+

Xp21

NADPH + H+

NADPH oxidase

e-

g1 22

Staphylococcus, Klebsiella, Serratia

eO2 (radical)

O2

Cytochrome b558

Pronasal Septic abscesses in lung, intestine, bones, liver pyoderma A. Infantile septic granulocytosis

Chemotaxis B. Chediak – Higashi syndrome CD18-expression

! "

CD18 CD11b

! "

Homo- a logies with WF and FB granulocyte surface CD18 CD11c

Abscesses in lymph nodes

Streptococcus, Pneumococcus, H. influenzae

Neurological symptoms

Intracellular bactericidal function (degranulation)

Partial oculocutaneous albinism

Mannose

Clinical outcome

5)($(-&) #<<0$1)1;2

Defect

CD18 CD11a

!

"

Endothelium

Fucose

Granulocyte Transmigration

Adhesion Rolling

! " Sgp50

Type 1

LFA-1

CR-3

C3dg-R

L-selectin

Type 2

E-selectin

LewisX ligand

C. Leukocyte adhesion deficiency 17q21

H2O2 + Cl-

defect D. Myeloperoxidase deficiency

MPO

OCl- + H2O

Bactericidal activity

C. albicans

! " BCE

#7%&%/37 F669&*7*;0

!

"

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#*6C7-6-&) ,-.%/%-&/%-5

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#*6C7-6-&) C+*)-%&5

,-.%/%-&/0E355*/%3)-@ 63&%.-5)3)%*&5

!"#!$

%&'( )*+,-./0 /.1-02/+.3 4-5,5( ).-67+0+0089 3-)3/3:

!;( <=( <"

>*+,-./0 /.1-02/+.3( ,9+7-?69+.-)@?/2/3

!A

B.1-02/+.3( -3)-0/899* C* !"#$$"%#& 3))5 4,+.+0+00/( 7-./.,+0+00/:D 309-?+E802*9*

!F;( !F$( &
H/.,/I/2/3( E-98*-E E-0/E682/+. +1 2@- 67C/9/089 0+?E( ?-06??-.2 3-)3/3

#*6C7-6-&) ,-.%/%-&/%-5 3&@ ,-.-/)5

C1 inhibitor

!

Plasminogen

C1 XI

C4

Prekallikrein

C2

Plasmin

C1qrs XII C2b

Complement activation

Coagulation

“Kinin-like” High molecular C2b-fragment kininogen

Kallikrein

Fibrinolysis

Bradykinin

Local permeability of blood vessels , edema A. C1 inhibitor deficiency

P

G

M

G

M M C I

H2C C O

CH O

C O C O

CH2

Phosphatidylinositol

P

C3

"

Bb

C2b

Ba

C3a C3b C3b

B B

C3b I

C3c B. Paroxysmal nocturnal B. hemoglobinuria (PNH)

P

H

C3b C4b

GPI-anchor

G

G

E.g. DAF z.B. LFA-3 z.B. N-CAM z.B. erythr. z.B. AChE

Carbohydrate chain

Protein C-terminal

#7%&%/37 F669&*7*;0

C4b

C3dg

C. Positive feedback loop dysfunction

! " $GI

782-2'/8 >..&-)8)4;

!

"

! ?@A

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5>1 #$%&'$&%( /-0 9(:82'/$2)-

LTR

gag

Increases infectiosity

pol Reverse transcriptase Protease Ribonuclease

Long terminal repeats

Enables budding

vif

Viral surface proteins mediate CD4 binding and membrane fusion

vpu

vpr

nef

env tat rev

LTR

Regulator of expression of structural proteins

Weak, strong transcription activator

gp 120 gp 41 RNA p7-nucleocapsid Lipid membran p24-capsid p17-matrix

Structural proteins Enzymes Regulatory proteins Other proteins

782-2'/8 >..&-)8)4;

Nucleocapsid nuclear proteins

A. Structure of the genome and the virion gp 41

HIV

Hematopoietic stem cell

2. gp 120

1. Binding 1. to CD4 1. molecule

Chemokine receptor CXCR-4 or CCR-5

HIV

Reverse transcriptase transforms RNA into DNA

?

Change in conformation (V3-loop)

CD4+ T cell

B. Binding to the host cell

!

Monocyte

CD4+ T cell

Langerhans cell

"

Macrophage

D. Cells susceptible to HIV

Viral DNA

Production of viral RNA and proteins Integration in cell genome Protease

Integrase

Binding to CD4, membrane fusion

Release of virion contents into the cell

RER Production of integral membrane proteins (gp 41, gp 120)

C. Replication cycle of HIV in the host cell

Budding of virus particles

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Acute phase

Symptom-free interval Helper T cells

ARC

AIDS

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Cytotoxic T cells

Anti-HIV antibodies HIV particles

Months

1

2

3

4

5 6 Time (years)

7

8

9

10

11

12

Infection – Sexual intercourse – Blood products – Mother child – Infected needles

Immune response – Acute infection, often unapparent – Symptom-free carrier

– Anti-HIV antibodies – Immune cells that recognize HIV – Cellular cytotoxicity – Viral suppression

– Viral mutation – Viral replication

– New mutations – Increased viral replication

– New immune response – Humoral and cellular adjustment to new epitopes

– Virus activated by - other infections ? – Migration into brain ?

– Viral suppression

– Loss of immune cells - CD4+ T-cells <400/µL

– Insufficientimmune response

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– Viral remains active

ARC B. Immune response AIDS Opportunistic infections Pneumocystis carinii Toxoplasmosis Mycobacteria Herpes viruses: HSV, CMV, EBV

Pneumocystis carinii infection

C. AIDS

Neurological complications Acute meningoencephalitis

Vacuolar myelopathy

Peripheral neuropathy

Chronic AIDSencephalopathy

HIV-associated tumors Kaposi’s sarcoma Malignant lymphomas: NHL

Kaposi’s sarcoma

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Patient serum

Anti-human-Ig + enzyme

Chromagic substrate

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Well coated with HIV antigen

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A. Diagnosis of HIV infection by ELISA a) Blocking of catalytic center

H

H3C

N

N

Produced DNA

b) Incorporation N3 of wrong nucleoside, strand breakage

Reverse transcriptase

Translation

HO O

Nevirapin

O

N

H

O

N

N

N

Azidothymidine (AZT) O

CH3

c) Blocking of catalytic center because of similarity with cleavage site H H

Viral proteins

Protease

O

Saquinavir

1. Antiviral drugs

O

N

N

O

N

N NH2

H

OH O

N

H

H

MHC I–peptide complex

Binding to MHC I molecules

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Translation

Cellular Immune response

Endogenous processing

Humoral

Plasmid Free proteins

2. Pure DNA as vaccine HIV-provirus

Antigen, mitogen

IL-16

Activated CD8+ T cell

3. Interleukins B. Treatment strategies

? Infected CD4+ T cell

TCR MIP-1!/", RANTES

Signal transduction

Virus production Chemokine receptor

CD4

Noninfected

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Blood group A: ~ 42% Antibodies against B

Blood group B: ~ 14% Antibodies against A

Blood group AB: ~ 6%

B antigen

A antigen

Blood group O: ~ 38%

No antibodies against A or B

!

Antibodies against A and B

A antigen

No A or B antigens

B antigen

Bacteria, pollen

Deletion of clones producing antibodies against own erythrocyte antigens

Rich in A, B antigens IgM-antibodies against A, B, AB

Newborn B. Development of A and B antibodies

Glu

Ceramide

Gal

GlcNAc

Gal

Precursor substance (paragloboside)

Erythrocyte

GPDfucose

L-fucosyltransferase (H-gene active in 99.9% of population)

Fuc

Chromosome 9 Ceramide

Glu

Gal

GlcNAc

Gal

8-121:6- ;**<2+-+3&

A. The ABO blood group system

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H-antigen B A 0 Active A gene (galactosaminyltransferase) N-Acetylgalactosamine

Blood group O

Blood group A

A antigen

Unmodified H antigen

C. Development of the ABO antigens

Active B gene (galactosyltransferase)

Both A and B genes active

Galactose

A antigen + B antigen

B antigen

O gene (silent allele)

Blood group B

Blood group AB

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Locus 1

Locus 2

2 possible alleles: D or d (inactive)

4 possible alleles: Ce, cE, CE, ce cE

CE

Most common Frequency genotypes

Ce D

ce

DCe/DCe

18%

DCe/dce

35%

DCe/DcE

13%

DcE/dce

12%

DcE/DcE

2%

Dce/dce

2%

dce/dce

15%

Immunogenicity

Phenotype

D

+++

c Rh+

E C e

Rh-

(+)

A. Rhesus system

1st Contact D antigen, immunogenic ++++

1. Pregnancy Fetus Rh+

Reexposure to < 0,5 ml Rh+ erythrocytes

Fetus Rh+

Mother RhEntry of Rh+ erythrocytes during labor

Rh-

1 ml Rh+ erythrocytes

2. Pregnancy

IgM-antiD production in 15%

Mother produces IgM Ab against D antigen

Immediate IgG production

Lysis of fetal erythrocytes

Entry of a few Rh+ erythrocytes sufficient IgG production of anti-D Ab

2. Hemolytic disease of the newborn

250 ml Rh+ erythrocytes

IgM-Anti-D production in 80%

Immediately after delivery anti-D immunoglobulins

Fetus Rh+ Mother Rh-

1. Anti-D immune response

4-/0/92- <++'0.-.3)

Chromosome 1

!

"

Lysis of Rh+ erythrocytes: no immune stimulation

3. Prophylaxis by Rh immunization

B. Alloimmunization against Rh antigens Kell system = K and k Phenotype Frequency

Erythrocyte membrane

Duffy system = Fya and Fyb Phenotype

Frequency (USA)

K- k+

91%

K+ k+

8.8%

Fya+b-

17%

9%

K+ k-

0.2%

Fya+b+

49%

1%

K immunogenic after Fya-b+ – pregnancy – infection Fya-b– transfusion C. Other erythrocyte antigens

34%

22%

<1%

68%

Whites

Blacks

Fyglycoprotein

Plasmodium vivax

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– Free hemo- globin – immune - complex -formation – DIC – nephro- pathy – hemo- globinuria

Complement binding Ab (IgM)

Complement receptor Complement

RES

!

Fc-receptor

Phagocytosis

2. Extravascular hemolysis

1. Intravascular hemolysis

RBCs sensitized with Ab or complement

Immunization with human serum

Test erythrocytes with known antigen profile

+ Patient + serum

+ Coombs + serum + Coombs + serum

or Polyspecific Coombs serum: anti-IgG, antiIgM, anti -complement 1. Production of Coombs serum

Agglutination

8/)()%'/ =++5(,/,30

A. Mechanisms of hemolysis

"

3. Indirect Coombs test

2. Direct Coombs test

B. Detection of erythrocyte antibodies: Antiglobulin test (Coombs test) Severe

IgM-Ab: agglutination in suspension = complete Ab

Coombstest +/-

Hemolysis

No agglutination = incomplete Ab (IgG) or complement only + Coombs serum

lgG only IgG+C3 rare high-affinity antibodies

None Agglutination by anti-C3

Coombstest +++

Agglutination by anti-IgG

C. Complete vs. incomplete antibodies

0

101

102

103

104

Antibody molecules/erythrocyte

D. Hemolysis and antibody affinity

105

! " @@B

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Idiopathic

Clonal transformation – Hodgkin’s disease CLL, NHL

B cell

APC Infections – Myco- plasms – Viruses

CH3 C CH2 C COOH NH2 !-methyldopa-type Drugs

T-helper

!

Regulatory T cell

Collagen vascular diseases – SLE

Alteration of T-cell regulation: – lymphoproliferative disease (AILD) – thymoma

A. Pathogenesis of autoimmune hemolytic anemia

Fc-mediated phagocytosis +20°C

+37°C

Spleen

Macrophage

Warm antibodies = IgG, rarely IgM, IgA B. Warm antibodies

Anemia jaundice

Immunosuppressive therapy:

Spleen irradiation

LDH Haptoglobin Urobilinogen C. Clinical features

"

Cyclophosphamide Azathioprine

Hepatosplenomegaly

Hyperplasia of bone marrow, erythropoiesis

91,*,3$1 >//.*'1'(2

Plasmocytoma

Cyclosporine Anti-CD20 antibody Prednisone: 20% success

Splenectomy Saturation of Fc-receptors by high-dose i.v. immunglobulins

1. Reduced Ab production 2. Reduced2. phagocytosis

D. Treatment of warm antibody-induced autoimmune hemolysis

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Infections – Mycoplasma pneumonia – Infectious mononucleosis – Cytomegaly – Listeriosis

– Idiopathic (chron. cold agglutinin disease)

Polyclonal agglutinins

Monoclonal agglutinins

!

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Predominantly anti-I (adult erythrocytes)

Anti-I (adult erythrocytes)

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50%

Ab binding

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Complement activity

- Acrocyanosis - Livedo - reticularis - Trophic - disorders

Hemolysis

0% 0

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3. Changes in blood chemistry

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CD20 antibody

0%

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+4°C

+20°C

+30°C

Cold antibodies = IgM, rarely IgG

Kupffer cell B. Mechanisms of hemolysis

C3-receptormediated phagocytosis C. Therapy

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2/&(&1-/ 7''#(%/%=;

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Viral infections

Drugs. Caution with heparin !!

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SLE

FcRII

gpIIIa

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Chronic Myeloid Leukemia (CML) Acute Myeloid Leukemia (AML)

Acute Lymphatic Leukemia (ALL))

M0 = Undifferentiated AML M1 = AML without maturation M2 = AML with maturation M3 = Promyelocytic leukemia M4 = Myelomonocytic leukemia M5 = Monoblastic/monocytic leukemia M6 = Erythroleukemia M7 = Megakaryocytic leukemia

CFU-B

undifferentiated ALL T-ALL B-ALL

Pluripotent stem cell

CFUGEMM

CFU-Eo

CFU-GM

B

T

A. Hematopoiesis and origin of leukemia MPO

HLA-DR

CD34

CD117

CD13

CD14

CD15

CD33

CD61

CD64

+/-

+/-

+

+/-

+/-

+/+

+/-

-

+/-

-

-

+/-

-

+/+ +/-

+/-

-

+/+/-

-

-

+ + +

+/+/-

+

-

+/-

+

+ +

-

+ +

-

+/-

+

M0 M1 M2 M3 M4

+ -/+

+

+

-/+

+/-

+

M5 M7

-

+

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-/+

+

-

+/-

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= positive for glycophorin -A

+ +/+/-

+/-

-

B. Immunophenotypic features of acute myeloid leukemias TdT

CD1a

CD2

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slg

Pro-T-ALL

+

-

-

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-

-

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Pre-T-ALL

+

-

+

+

+/-

-

-

-

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+

+

+

+/-

+/-

+/-

-

-

-

-

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+/-

-

+

-

+

-

-

-

-

-

Pro-B-ALL

+

-

-

-

-

+

+

-

-

Pre-B-ALL

+

-

-

-

+/-

+

+

+

-

Mature B-ALL

-

-

-

-

+/-

+

+

-

+

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C. Immunophenotypic features of acute lymphatic leukemias Lymphatic leukemias

Myeloid leukemias t (8;21)

M2

t (9;22)

Ph'+-ALL

inv (16), del (16q)

M4Eo

t (4;11)

newborn -ALL or biphenotypic ALL

t (15;17)

M3

t (8;14), (2;8), (8;22)

B-ALL

t (11;17)

M3-like

t (11;14), (1;14)

T-ALL

del (11) (q22-23)

M5, M4

t (1;19)

pre-B-ALL

t (9;11), t (11;19)

M5, M4

t (5;14)

B-ALL

monosomy/del 7 & 5

M1, M2, M5

del 9, t (9;n..)

hyperleuk., extramedull. manifestations

D. Cytogenetic features of acute myeloid and lymphocytic leukemias

! " @AC

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"

B-lymphoblastic lymphoma

1. CLL, PLL, small-cell lymphocytic lymphomas

B-lymphocytic lymphoma

2. Lymphoplasmacytoid lymphoma (immunocyt.)

Lymphoplasmacytic immunocytoma

3. Mantle cell lymphoma

Centrocytic lymphoma Centrocytoid subtype of centroblastic lymphoma

4. Follicular center lymphoma, follicular, grade I – III diffuse (predominantly small cell) (preliminary)

Centroblastic-centrocytic lymphoma Follicular, diffuse

5. Marginal zone-B-cell lymphoma, extranodal (MALT-type, +/- monocytoid B cells), nodal (+/- monocytoid B cells)

Monocytoid lymphoma, including marginal zone lymphoma

6. Marginal zone lymphoma of the spleen (preliminary) 7. Hairy cell leukemia

Hairy cell leukemia

8. Plasmocytoma/myeloma

Plasmocytic lymphoma

9. Diffuse large cell B-cell lymphoma (subtype:

Centroblastic lymphoma, B-immunoblastic Lymphoma, large cell anaplastic Ki1+ lymphoma

primary mediastinal large cell B-cell lymphoma)

10. Burkitt’s lymphoma

Burkitt’s-lymphoma

11. High-malignancy B-cell lymphoma, Burkitt-like

Centroblastic, immunoblastic lymphoma

T-Cell and Natural Killer-Cell Lymphomas I. Precursor T-cell neoplasias Precursor T-lymphoblastic lymphoma /leukemia

T-lymphoblastic lymphoma

II. Peripheral T-cell and NK-cell neoplasias 1. T-CLL /T-PLL (prolymphocytic leukemia)

T-lymphocytic lymphoma, CLL-type, PLL-type

2. Large cell granular lymphocyte leukemia a) T-cell type; b) NK-cell type

T-lymphocytic lymphoma, CLL-type

3. Mycosis fungoides/Sézary syndrome

Mycosis fungoides, Sézary syndrome

4. Peripheral T-cell lymphomas, subcutanous panniculitic lymphoma (preliminary), hepatosplenic !-"-lymphoma (preliminary)

T-zone lymphoma, lymphoepitelioid lymphoma, Pleomorphic small/large cell lymphoma, T-immunoblastic lymphoma

5. Angioimmunoblastic T-cell lymphoma

Angioimmunoblastic lymphoma (AILD, LgX)

/0%4%2.0 @++B4'0'7*

II. Peripheral B-cell neoplasia

#

6. Angiocentric lymphoma 7. Intestinal T-cell lymphoma (+/- enteropathy) 8. Adult T-cell lymphoma/leukemia, HTLV1+

Pleomorphic HTLV1+ lymphoma

9. Anaplastic large cell lymphoma (T and null)

Anaplastic large cell T-cell lymphoma (Ki1+)

Hodgkin’s Lymphoma I. II. III. IV. V.

Lymphocyte-predominante type Nodular-sclerosis type Mixed cellularity type Lymphocyte depletion type Lymphocyte-rich classic type

Nodular sclerosis type Mixed cellularity type Lymphocyte-depletion type Lymphocyte-predominante type

1. REAL classification 2. Kiel classification A. Lymphoma entities: REAL classification vs. Kiel classification

" # CDC

3/+*+,$/ @77A*'/'(6

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1'.(E+*F2 9+2)$2)

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T cell

CD30 CD15 CD25 CD71 HLA-DR (CD20) (CD2) (CD3) (CD4)

EBV

? B cell

Activated B cell

Monocyte/ dendritic cell

VH genes ReedSternberg cell

?

!

Cytokine secretion

Lymphocyte predominant type ca.12%

Nodular sclerosis type ca. 46%

Mixed cellularity type ca. 31%

Lymphocyte-depletion type ca. 10%

B. Histological classification B-symptoms:

Liver

–Fever > 38°C –Weight loss- > 10% –Night sweats

Bone marrow/ bone

–Anemia –Leukocytosis –Thrombocytosis –Eosinophilia

Lung, pleura, pericardium –Pruritus – Itching –“Alcohol pain” –Bone pain

Upper mantle field Inverted Y

1. Radiotherapy E. Therapy

Erythrocyte sedimentation

Extranodal manifestations

– Adriamycin – Bleomycin – Vinblastine – Dacarbazine

BEACOPP High-dose chemotherapy

"

Signs of inflammation

Skin

C. Symptoms

3/+*+,$/ @77A*'/'(6

A. Pathogenetic model

D. Laboratory findings CD3

– Cyclophosphamide – Oncovin (Vincristine) – Procarbacine – Prednisone

E = etoposide Autologous stem cell transplantation

2. Polychemotherapy

CD30

T cell

Bispecific antibody

3. Experimental approaches

Hodgkin’s cell

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Thymus

TdT+

CD7+ CD3+ CD2+/CD5+/CD1a+/-

1. T-CLL (<1%)

–Mediastinum –Bone marrow –Blood –Lymph nodes

CD7+ CD2+ CD3+ CD5+ CD4+(65%)

Bone marrow

;25@5&32 A..'@)2)>-

A. Precursor T-lymphoblastic lymphoma/leukemia

CD4+8+(25%)

Circulating peripheral cell

" Peripheral CD4+ T-cell Extrafollicular CD30+ blasts

CD3+ CD7+ CD8+/CD103+

! BCF

HTLV-1-infection

CD2+ CD3+ CD5+ CD4+ CD25+

CD30+ CD25+/CD3+/+ EMA+/HTLV1 genome

7. Intestinal 7. T-cell lymphoma (<1%) B. Peripheral T-cell neoplasms

9. Anaplastic large-cell 9. lymphoma ALCL (~2.5%)

8. Adult T-cell lymphoma/ 8. leukemia (<1%)

*+;%22 ,-./0).3(

CD2+ CD3+ CD8+ CD16+ CD57+/TCRab+

2b) Large granular 2b. lymphocyte leukemia 2b. NK-cell type 2b. (<1%) CD2+ CD8+/CD16+ CD56+/CD57+/-

Circulating peripheral CD8+ T cell / NK cell

3) Mycosis fungoides/ 3. Sézary syndrome (<1%)

CD2+ CD3+ CD5+ CD7+(35%) CD4+(95%)

Peripheral epidermotrope CD4+ T cell

;25@5&32 A..'@)2)>-

2a) Large granular 2a) lymphocyte leukemia 2a) T-cell type (<1%)

!

" Peripheral T cell in different phases of maturation ?

?

NK or T cell?

Peripheral T cell

Cytokine+++ CD2+ CD5+/CD7+/CD3+/+ CD56+

6. Angiocentric 6. lymphoma (~1%)

CD3+/CD2+/CD5+/CD4+/-

5. Angioimmunoblastic 5. T-cell lymphoma (~1%)

CD3+/CD2+/CD5+/CD7-/+ CD4>CD8

4. Peripheral T-cell 4. lymphoma (~10%) a) Medium cell b) Medium and large cell c) Large cell d) Lymphoepithelioid e) Hepatosplenic e) !" lymphoma f) Subcutaneous pannif) culitic lymphoma

! " BCG

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TdT

CD10+/CD19+ CD22 cyt CD79a

–Bone marrow –Blood –Lymph nodes

Bone marrow

,-090&3- ://'9)-)7;

A. Precursor B-lymphoblastic lymphoma/leukemia

"

11. High-malignancy Burkitt-like lymphoma 11. (~2.5%) CD19+ CD20+ CD22+ Slg+/CD10+/-

1. Chronic lymphatic 1. leukemia / prolympho1. cyte leukemia / lympho1. cytic lymphoma (~7%) CD19+ CD20+ CD22 cyt CD5+ CD23 + SlgM (+) SlgD (+)

Deletions in chromosome 13 trisomy 12 CD19+ CD20+ CD22 cyt

PANB+

Recirculating CD5+-CD23+ B cell

Translocation of bcl-2 in 30%

Peripheral B cell in different phases of differentiation ?

EBV infection

Translocation of c-myc gene t(8;14), t(2;8), t(8;22) CD19+ CD20+ CD22+ SlgM+ CD10++

!
10. Burkitt’s lymphoma (<1%) B. Peripheral B-cell neoplasms

CD19+ CD20+ CD22++ SlgM+ CD11c+ CD25+ CD103+

7. Hairy cell leukemia (<1%)

CD19+ CD20+ CD22+ SlgM+ cyt lg +/CD43-/+

6. Marginal zone lymphoma 6.of the spleen (<1%)

2. Lymphoplasmocytoid 2. lymphoma/immuno2. cytoma (~2%) CD19+ CD20+ CD22 cyt SlgM + SlgD -/+ cit lg+

t(9;14) deletions in chromosomes 13, 11, 17

Peripheral B cell maturing into plasma cell

3. Mantle cell lymphoma (~6%)

4. Follicle center lymphoma 4. (~25%)

CD19+ CD20+ CD22(+) CD5+ CD10 +/SlgM+

CD10+/CD19+ CD20+ CD22 cyt SlgM,G,A+

t(11;14) rearranged bcl-1 gene (cyclin)

t(14;18) overexpression of bcl-2 proteins

CD5+–CD23- cell of follicular mantle

Germinal center cells

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Marginal zone of lymph nodes

Translocation of: bcl-2 (30%) bcl-6 (30%) CD19+ CD20+ CD22+ CD10-/+ CD30-/+ Slg+/-

9. Diffuse large-cell 9. lymphoma (~30%)

Marginal zone of mucosaassociated lymphoid tissue

Antigen stimulation e.g., Helicobacter pylori CD19+ CD20+ CD22+ SlgM+ cyt lg +/CD43-/+

5b) Nodal marginal zone 5b. lymphoma (monocytoid 5b. lymphoma) (~2%)

CD19+ CD20+ CD22+

SlgM,G+ cyt lg +/CD43-/+

5a) Extranodal marginal 5a. zone lymphoma 5a. (MALT-Type) (~8%)

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TH

TR Bacteria, antigen mixture

Poyclonal B-cell stimulation

Malignant transformation

Hypergammaglobulinemia

Monoclonal B-cell proliferation

Defective T-cell regulation

Monoclonal gammopathy

A. Polyclonal vs. monoclonal gamma-globulin increase TS

Bone marrow TH

Hepatosplenomegaly Lymphadenopathy

Neurological symptoms

Blood Impaired vision, blindness

CD19 CD20

=%2(2')% A..1($%$<&

“spike” (M-component)

"

Congestive heart failure

cyt/IgM++

Bleeding

Lymphoplasmacytoid B cells

Viscosity

B. Waldenström’s macroglobulinemia

Overproduction of heavy chains

C. Heavy chain disease

! chains

Lymphadenopathy, rarely osteolytic lesions (Franklin’s disease)

" chains

Malabsorption, diarrhea, intestinal lymphadenopathy

µ chains

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Stromal cell

Immunglobulins

Large amounts of monoclonal proteins

Plasma viscosity (esp. IgA)

Hyperviscosity syndrome

Cytokines: TNF

IL-6

Autocrine production

Plasma cell

IL-1

(osteoclast-activating factors)

Unbalanced synthesis of light chains

Suppression of normal Ig-production

Suppression of hematopoiesis

Activation of osteoclasts

!/"-chains in urine (Bence Jones proteinuria)

Decrease of normal antibodies

Bone marrow

Osteolysis

Cytopenias

Hypercalcemia

Nephropathy

Frequent infections

83,*,/$3 9..2*'3'(5

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A. Pathogenesis and pathomechanisms of multiple myeloma

B. Localization B. of osteolysis

Histology

Buckshot skull

Cytology

Osteolysis, decalcification

C. Bone marrow findings

D. Radiographic findings

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Collect serum

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B. Determination of cryoglobulins in the blood

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Rheumatoid factor activity

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IgA + IgG

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Immune complex precipitation

IgG + IgG

IgM + IgG

Polyclonal

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Polyclonal

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Polyneuropathy

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– Infections - (HCV, HBV, HIV, syphilis, - borreliosis) – Essential cryoglobulins – Connective tissue diseases

Vasculitis

Arthralgia

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C. Classification of cryoglobulinemia

Nephritis 3. Type III (mixed polyclonal cryoglobulins)

Purpura

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!012',3'+,+

- Immunglobulin - light chain - Protein AA - !2-Microglobulin - Transthyretin - ! Protein A4 - Cystatin - Procalcitonin - "-Natriuretic - peptide

1000nm

10 nm

!

Degradation of proteins or precursors

A. Pathogenesis

Polymerization

Amyloid fibrils

Polymerized polypeptides

Waldenström’s macroglobulinemia

Monoclonal Ig light chain surplus

Chronic infections

Hepatocytes

Serum amyloid A = SAA

Amyloid P component (serum amyloid P = SAP)

AL-fibrils

Chronic inflammations

B. AL amyloidosis B. (primary, nonhereditary amyloidosis)

Accumulation: AA fibrils

C. AA amyloidosis (secondary amyloidosis) Accumulation of AA fibrils in tissue Dementia, vasculopathy

Cells of the immune system

Hepatocytes

!2-microglobulin

Glomerular filtration

Macroglossia

Cardiomegaly, congestive heart failure

Hepatomegaly

Tubular absorption

Adrenal gland insufficiency

Reduced filtration in longterm renal failure

!2-m fibrils

D. AH amyloidosis C. (hemodialysis-associated)

Peripheral and autonomous neuropathy: polyneuropathy, incontinence, impotence

"

Bronchitis, diffuse or nodular infiltration of the lung

Glomerular damage, proteinuria, azotemia

Splenomegaly

Carpal tunnel syndrome

Hypomotility, malabsorption, obstruction

E. Clinical features

/2,*,9$2 >00<*'2'(1

IL-1, IL-6,TNF-" Myeloma, plasma cell dyscrasia

! " ?@B

75)()%45 2..-(&5&'6

!

"

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;$*$%*)&( 4(3 23$(*)+)%4*)&( &+ ,-.&/ !(*)'$(0

Myeloma cell

!

Cloning

Spleen cell Immunization with tumor cells

Spleen removal

Fusion (+PEG) Hybrid cell

HAT-sensitive mutant

Reactivity test against tumor cells

Culture in HAT medium

Hybridoma clones

1. Production of monoclonal antibodies against tumor antigens E.g., mutated protein

Tumor cell

CD8

ER

Tumorspecific T-cell

Proteasome MHC I

TCR

"

Tumorspecific peptide

TAP 2. Antigen recognition by T cells

A. Recognition of tumor antigens Tumor peptide Tumor cell (e.g., HLA-A2)

MHC I

CD8

CD8

TCR

T cell clone

DNA extraction

Tumor cell

T cell clone TCR

Tumor peptide

Fractionation by HPLC columns

DNA fractionation

75)()%45 2..-(&5&'6

Myeloma cell line

Acid elution of MHC-bound peptides

Insertion of DNA fragments in vectors Insert in HLA-A2+ cells T-cell clone only lyses cells with tumor antigen-coding DNA 1. Identification of genes

B. Identification of tumor antigens

Peptide loading of cell line with empty MHC I molecules T-cell clone

Lysis by T-cell clone 2. Identification of peptides

! " 898

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!

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/%%$(, 1-234, 5,263(*-%- &> #$%&' !()*+,(Normal Gene Products Fetal antigens Fetal liver Fetal bowel

Normal melanocyte

Tyr

Tyrosinase peptide

Tyr

Tyr Tyr

Clonal antigens B cell T cell

Tyr

Melanoma cell

Tyr

Carcinoembryonic antigen (CEA)

!-Fetoprotein (AFP)

Hepatic/testicular cancer AFP+++

Bowel cancer CEA+++

mIg

B-cell lymphoma

p21 t 9;22

ABL

Viral proteins

Point mutations

22

BCR

T-cell lymphoma

Idiotypic antibody orTCR sequences

Mutated or Abnormal Proteins Chromosomal translocations 9

TCR

Clonal proliferation

p53

p16

HTLV1

Cell cycle-regulated proteins BCRABL

Infected T cell

Point mutations

BCR ABL Fusion gene

Adult T-cell leukemia (ATL)

BCR-ABL protein

Fusion peptide = tumor specific

Normal peptide

A. Tumor antigens

Tumorspecific peptide

Tumor cells: MHC I expression absent or reduced

2. Downregulation of MHC molecules

1. Tumor peptide does not bind to MHC

CD8 Empty MHC I molecule

Defective TAP 3. Defective antigen-processing machinery

TCR

T-cell

T-cell anergy, tolerance

CD28 4. Absence of co-stimulatory molecules IL-10, TGF-", prostaglandins

Tyr Tyr

Tyr

"

Viral peptide

Normal cells: express MHC I molecules

TAP

:8*(*238 /%%$(&8&+9

Differentiation antigens overexpressed in tumor cells

!

Tumor antigen no longer synthesized/ expressed

5. Antigen modulation

B. Immune recognition escape mechanisms

T-cell FAS ligand FAS 6. T-cell-inhibitiory molecules, cytokines

T-cell anergy/ apoptosis

! " ;
9:0$0'&: 1))2$+:+=3

!

"

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1))2$+*%(<&/(2*0' 6*<&*(=0(. A1B

Addition of adjuvants: BCG, Corynebacterium parvum

!

Interferon-!

– Chronic - myelocytic - leukemia – Plasmocytoma – Lymphoma – Melanoma – Renal cell - carcinoma

Interleukin-2

– – –

Postoperative injection (s.c.)

Tumor removal, tumor cell isolation

IL-2

Insert cytokine-encoding genes into tumor cells (retroviruses, adenoviruses, physical methods)

IL-7 IL-4 GM-CSF

TNF-!

Renal cell carcinoma Melanoma Leukemias

– local perfusion of - sarcomas/melanomas

2. Cytokine therapy

1. Vaccination with adjuvants, cytokine-producing cells

A. Enhancement of nonspecific immunity Irradiation

B7 gene B7 CD28

Tumor removal, tumor cell isolation

Tumor cell lysate CD80 gene

Postoperative injection s.c., c.v.

Tolerance

CD4

APC

KSQSSALRK

B. Induction of a specific T-cell response

T-cell activation

Phagocytosis of tumor proteins

Tumor cell lysate

1. Vaccination strategies

"

2. Direct antigen presentation

CD80+ tumor cells

Defined tumor antigen/ tumor peptide

9:0$0'&: 1))2$+:+=3

s.c. or c.v. administration of cytokines

Autologous APC

Pulse with purified antigen, peptide

CD8 T-cell activation

3. Indirect antigen presentation

! " >??

5=*+*(;= >11'+%=%.9

!

"

! ?@A

0'1%$ >11'+%=%.9

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>11'+%):/$;4/')*( B)$;)/.*/3 C>>D

Tumor peptide

MHC-I

TCR

Cloning the DNA sequence of TCR from tumor-infiltrating lymphocytes

Gene transfer in lymphocytes, expansion of T cells with tumor -reactive TCR

DNA expression in vectors

Adoptive immune therapy

!

A. Production of tumor-specific TCR by gene modification Human Fc

1.

Murine Fab

Anti-APO-1 Fas

Fc + part of Fab = human

1.

APO-1 Fas Murine monoclonal antibody (MAb) 2.

“Humanized antibody”

Monospecific antibodies

Only variable sequence is murine CD3

Pore formation

Tumor cell

Bispecific antibodies

Activation of the complement cascade Variable part of antibody

E.g., !-chain of TCR

Chimeric T-cell receptor

Only variable part of antibody

Transfection of T cells

Necrosis, apoptosis

Antibody-dependent cellular cytotoxicity (ADCC)

Genetically produced antibody derivatives Toxin

Tumor antigen

Radioisotope

Murine MAb

Ab therapy

Immunotoxins - immunoconjugates

B. Antibody therapies

3.

Fc-receptor

Perforins, granzymes

4.

"

Bispecific antibody

Fusion of hybridomas (quadromas)

Fv protein

2.

Tumor-associated antigen

T cell

3.

Antibody imitates the natural ligand of a molecule: “agonistic effect”

5=*+*(;= >11'+%=%.9

Apoptosis

C. Mechanisms

4.

Production of human anti-mouse antibodies (HAMA), which imitate the antigen

! " ?@E

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!"#$%&'#$(#()*$ *+ ,-(*'*.*-% /*$0 1#""*23405#(*&*)0()6 7(05 80''% Bone marrow ~850 ml BM blood

0.5% CD34+ cells

!

CD34+

Bone marrow (BM) aspiration Peripheral blood ~350 ml leukapheresis product

A. Stem cell harvesting

CD34+ Stem cell graft

Leukapheresis BM-aspiration leukapheresis

Cryoconser-196ºC vation

Cytokine stimulation

Stem cell reinfusion Reconstitution by graft

mG-CSF GM-CSF

20000

"

Myeloablative therapy ± radiotherapy

Leukocytes

5000

8')$)6#' >55-$*'*.B

0.1% CD34+ cells

Aplasia 10–20 days

1000 0

B. Mobilization of hematopoietic stem cells and transplantation schedule – Immune defects – Hemophilia – Immune therapy of cancer – Gene modification - of stem cells

– Acute leukemias – Hodgkin’s lymphoma – Non -Hodgkin’s lymphoma – Breast cancer – Germ cell tumors

1. Enhancement of the chemotherapeutic effect

2. Genetic and experimental therapy

C. Indications Avidin

Cobalt– samarium magnet

MAb

Biotin CD34

Enrichment of CD34+ cells ~90% purity

Stem cell

D. Purging of the autograft

Bead with iron core CD34

MAb

CD34+

CD34+

Depletion of contaminating tumor cells

! " CDF

8)+&+0%) 966>&,),.B

!

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Search for an HLAcompatible donor within the family

Alternatives

!

– CML Cord blood

– AML – Aplastic anemia – SCID

Unrelated donor

– PNH – Fanconi’s anemia – Thalassemia

Patient

Myeloablative therapy

– Solid tumors?

A. Allogeneic BM transplantation Hemorrhagic cystitis

Interstitial, toxic pneumopathy

Veno- occlusive disease (VOD)

Infertility

1. Chemotherapy-induced complications

Bleeding

HSV, CMV

Aspergillus, Candida

Coccus, Pseudomonas

2. Aplasia-induced complications Host APC

TNF-! IL-1 IL-6

Chemotherapy

Irradiation

B. Indications

Graft

Donor lymphocytes

CTLs

8)+&+0%) 966>&,),.B

– Aggresive lymphomas?

"

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NK cell

3. Immunological complications: GVHD C. Complications of allogeneic transplantation BM donor

Decresed risk of GVHD , increased risk of relapse T-cell depletion

D. T-cell depletion and graft-versus-leukemia effect

Reinfusion of T/NK cells after 4,8,12 weeks Anti-leukemia effect

! " CDC

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Identical twins

Autologous

Syngeneic

Allogeneic

Xenogeneic

A. Transplantation types

Organ donor’s written consent Irreversible absence of spontaneous breathing

24 h Isoelectric EEG

2. Conditions

- ABO blood group - HLA test: DR>B>A>C - Test for cross-reactivity - HIV exclusion, CMV status

2 x angiography: absence of cerebral blood flow 1.

30 min

1. Dissociated brain death

Approval of family members

2.

3. Testing

B. Criteria for organ transplantation Vena cava

Aorta Lamellar keratoplastic surgery Ureter

Pelvic vessels

Bladder

Perforating keratoplastic surgery

1. Kidney C. Transplantable organs: Examples

2. Cornea

Complement regulators

Plasmapheresis

Knock Out For antigens For preformed antibodies

DAF (CD55) MCP (CD46) MIRL (CD59)

Transgenic pig

D. Xenogeneic transplantation

Ciliary body Iris Lens

"

Anterior eye chamber Epithelium Endothelium Stroma

Soluble inhibitory factors of complement lysis CD35

!-Galactosyl compounds

Host

0)+&+=%) ;55<&,),3-

Irreversible areflexia (pupils, cornea)

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Fc"R

C3b

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ADCC

C3a, C5a

Cell activation

Chemotaxis

Mast cells Histamine, serotonin

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1. Humoral rejection Processing (indirect)

Rejection

APC

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#$%&%'($ )*(+,-*. /0 12*,3(+/%4 !-+2-%+%.

Articular

Extra-articular

!

Episcleritis

Pericarditis

Elbow

Pleuritis

Hand

#$%&%'($ ?33,&/$/@A

Cervical spine

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Rheumatoid nodules

Knees

Vasculitis

Feet A. Clinical manifestations of rheumatoid arthritis

! " 5:;

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Normal

!

Pathological Enlarged layer of lining cells

Intra-articular space Normal layer of lining cells

CD8+ T cells

Connective tissue

Increased vascularization

A. Cellular structure of the synovium

1. Synovial membrane with lymphoid follicle-like structure B cells T cells

Bone Cartilage

1. Synovitis Erosion

Synovial fluid

#%202+'% 944&0,%,B/

CD4+ T cells

"

Plasma cells Granulocytes

Pannus NK cells

Lining cell layer

Dendritic cells

Synovium

Macrophages/ monocytes Endothelial cells 2. RA: histology B. Cells in the arthritic joint

3. Activated macrophages: electron micrograph

4. Cellular composition

! " CDF

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Amino acid

1. Class II 1. antigen 1. (view from above)

DRB1

57

69

67

70

71

74

86

DR4Dw4 DR4Dw10 DR4Dw13 DR4Dw14 DR4Dw15 DR1 DR6Dw16

Asp – – – Ser Ile Leu

Glu – – – – – –

Leu Ile – – – – –

Gln Asp – – – – –

Lys Glu Arg Arg Arg Arg Arg

Ala – Glu – – – –

Gly Val Val Val – Ala Ala

!

2. Variable amino acid position in the sequence of DR

Postcapillary venules

Synovial proliferation

T cells B cells

Arthritogenic peptide

Immunoglobulins Rheumatoid factor Antigen presentation by synovial cells

Synovitis Pannus formation

Immune complexes

:,*8*&2, ;11$8.,.7-

A. Susceptibility to rheumatoid arthritis

"

Complement activation

Cytokines

Bone and joint damage

B. Pathogenesis of rheumatoid arthritis Unknown antigen

Pannus tissue GM-CSF

Synovial tissue

FGF IL-1 T cells

Macrophages

C. Induction of rheumatoid arthritis

TNF-!

Chondrocytes Osteoclasts

! " ?@?

93(2(&*3 @..-2/3/71

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F*'$/7%2%:(: /G ,$%-.*'/(+ !5'$5('(: =@@>

PGE2, neopterin

CD4

CD14 DR DP Enhanced phagocytosis

DQ

TGF-!, FGF IL-10

!

CD68

TNF-", IL-1

Enhanced chemotaxis

Fc receptor

1. Synovial macrophage

2. Giant cell

93(2(&*3 @..-2/3/71

A. Activated rheumatoid synovial macrophage

3. Stellate synovial cell

B. Activated cells of the rheumatoid synovial membrane

2. IgG rheumatoid factor ( also IgA and IgE rheumatoid factor )

1. IgM rheumatoid factor

C. Rheumatoid factors (anti-globulins) Allo- or autoantigen?

Synovial membrane

"

Intra-articular space

+ APC

Polyclonal B-cell activation

AutoImmune antibodies complexes (RF)

Complement activation

C3a,b

Vasculitis

C5a

D. Impaired B-cell regulation

Cytokines + inflammatory mediators Tissue and cartilage destruction

Cartilage

! " CDH

#$',')%$ D119,+$+=/

!

"

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29?0,'$0 #>8+,') !8*>8'*'&

Juvenile chronic arthritis (JCA) Polyarticular 40%

Systemic 10%

!

Oligoarticular 50%

RF-negative (early onset)

Early onset type I ANA+ (EOPA)

RF-positive (late onset)

Late onset HLA-B27+ type II (LOPA)

A. Subtypes Fever (up to 41°C) Exanthema

Lymphadenopathy

Polyserositis Organ enlargement

f:m 2:3 Exanthema

Arthritis (polyarticular)

Lab. findings: ESR , CRP leukocytes (shift to the left) anemia

B. Systemic form ( Still’s disease) Chronic iridocyclitis

Arthritis (few joints)

f:m 9:1

Lab. findings: ANA+ 50% RF neg.

Seronegative

Seropositive

Age of onset 2–5 years

Age of onset >10 years

Mandibular involvement (micrognathia) Symmetric polyarthritis (epiphysial growth defect)

f:m 4:1

Lab. findings: unspecific (ANA+ 25%)

Symmetric destructive arthritis

Lab. findings: RF+ ANA+ 75% DR4+

"

C. Polyarticular forms Uveitis

Sacroiliitis

f:m 1:9

Lab. findings: HLA-, B27+ ANA-, RF neg.

Iridocyclitis D. Early childhood oligoarthritis

#$',')%$ D119,+$+=/

( Still’s disease)

E. Juvenile spondyloarthropathy

! " EFH

!

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!

Axial and peripheral arthritis

#$%&%'($ GAA,&2$23@

Ankylosis

Enteritis/ diarrhea

Balanitis/ urethritis

Enthesiopathy

Keratoderma blenorrhagicum

"

A. Clinical manifestations of spondyloarthropathies Genetic

Environmental

- HLA - gender - ethnic - factors

Ankylosing spondylitis

95%

Reiter’s syndrome (classic form)

80%

Undifferentiated spondyloarthropathy

70%

Arthritis with IBD

25%

Psoriatic arthritis

25%

Infections

Disease

Cytokine genes

Genetic modifier Treatment Clinical outcome

B. Pathogenesis of spondyloarthropathy

Reactive arthritis

Normal population

20–80%

8–10%

C. Frequency of HLA-B27 antigen

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Antigen determinant

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HLA-B27

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CD4+ T cell

Own gene product Human HLA-B27

C. Promiscuous B27 hypothesis

Arthritis Psoriasis Colitis

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Diet

Allopurinol Secondary

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Benzbromarone Decreased urate elimination

O

7 mg/dl

Gout osteoarthropathy Olecranon bursitis Renal damage

Urate pool

Uric acid

Colchicine

Macrophage

Urate crystals IL-1

Chemotaxis

Inflammatory reactions

Soft tissue tophi on hands

Granulocyte

Knee joint (gonagra)

Piercing

Release of enzymes and mediators

Phagocytosis 1. Pathogenesis and therapy

2. Manifestations

A. Gout Uveitis

Cauliflower ear

Saddle nose

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First metatarsophalangeal joint (podagra)

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Primary

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3 Genital ulcerations

Oligoarthritis, esp. lower limbs

Clinical triad

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6. Anti-centromere antibody pattern

7. PM-Scl pattern

8. Anti-spindle-apparatus antibody pattern

9. Anti-coilin antibody pattern

10. Anti-PCNA antibody pattern

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80% common symptoms (fever, fatigue)

A. Clinical manifestations of systemic lupus erythematosus (SLE)

70% vasculitis/ skin involvement

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Histones

Sm-antigen

DNA

nRNP

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La (SS-B)

Ro (SS-A)

Nuclear antigens

Aberrant expression of autoantigens

Apoptosis

Cytoskeleton filament

Apoptotic blebs

Cell membrane protein

Nucleosomes

Autoantibodies against phospholipids Thrombosis, miscarriages, cerebral infarctions

Blood vessel

Antibodies against cells

Ro (SS-A)

Anemia

Cytoplasmic antigens

ThromboLeukopenia penia

Polyclonal B-cell activation

Impaired regulation B-cell help B-cells

Genetic susceptibility

APC

Epitope spreading

Phagocytosis Antibodies against subcellular antigens (DNA, RNP etc.)

Tissue lesion: release of autoantigens

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Immune complex

Vasculitis

Deposition in the kidney Deposition of small immune complexes

Granulocyte

Glomerular basal membrane

Large immune complexes

Complement receptor Fc-receptor Platelet aggregation

Endothelial cells

Epithelial cell

Immune complexes and glomerulonephritis

A. Pathogenesis of systemic lupus erythematosus

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IL-1

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Collagen synthesis

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Shortened tongue frenulum

Myocardial fibrosis

Interstitial lung fibrosis

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Microstomia

Occlusion of blood vessels

Esophageal hypomotility Malabsorption, impaired motility

AutoAb

Sclerodactyly

Centromere (CREST) Topoisomerase I

ANA

Pitting scars on fingertips

SLE Polymyositis

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Coronary angiopathy

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Scleroderma

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Sjögren’s syndrome/RA

Dermatomyositis Sclerodactyly

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Calcinosis

Esophageal hypomotility

Telangiectasias

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Raynaud’s syndrome

Pulmonary involvement Impaired esophageal motility

Hand swelling, sclerodactyly

Skin symptoms

Polyarthralgia polyarthritis Association with PBC

Anti-U1-RNP-Ab

ANA

highly positive

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T cells

Viruses (herpes, retroviruses) IL-1

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Sicca symptoms

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Lymphocyte infiltration

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Expression of HLA class I antigens

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Cytotoxic T cell

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Muscle fiber

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Arteriole

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Vein

Schönlein-Henoch purpura and essential cryoglobulinemia

Panarteritis nodosa and Kawasaki’s disease Temporal arteritis and Takayasu’s arteritis

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B//50% 12345-6*68%3 208 @&-(2'*%'6*63 A&8&32

ANCA

!1-antitrypsin PR3-!1antitrypsin

!

PR3-ANCA-IC

PR3 TNF-! LFA-1

LFA-1 LFA-1

ICAM-1

ICAM-1

LFA-1

ICAM-1

ICAM-1

Endothelial cells (EC)

1. Resting PMN and EC

2. PMN and EC preactivated

3. PMN adherent to EC

4. PMN activated by ANCA

A. Vasculitis development theory e.g., in Wegener’s granulomatosis

2. C-ANCA

c-ANCA

3. Saddle nose Bulbar protrusion Otitis Sinusitis Swelling of parotid gland Pulmonary infiltration CNS granulomas Episcleritis Rhinitis, ulcers Subglottic stenosis Tracheal/ bronchial stenosis Glomerulonephritis Myalgias/myositis Arthralgias Leukocytoclastic vasculitis Polyneuropathy

4. Vasculitis of the toes

Sinusitis

BP diastolic >90

Allergic rhinitis

Encephalomalacia (juvenile stroke)

Pulmonary infiltration

Ulcers in stomach and intestine

Carditis Eosinophilc gastroenteritis

Testicular pain

Interstitial nephritis

Livedo reticularis

Myalgias Arthralgias

Myalgias

Purpura

Arthralgias

Polyneuropathy

Painful cutanous nodules

Skin nodes Skin necrosis

1. Clinical findings B. Wegener’s granulomatosis C. Churg-Strauss syndrome

"

Coronary insufficiency

Asthma

IgE Eosinophilia

=-60642- B//50&-&:(

IL-8, IL-1

HBsAg

Polyneuropathy

D. Polyarteritis nodosa

! " CDF

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#$%&' ()** !+')+$'$,

Temporal arteritis Scalp skin necrosis

Disease onset within 2 weeks

Jaw claudication

Initial ESR increase of >40 mm in 1 hour

Pulmonary vasculitis (rare)

Morning stiffness >1 hour

Coronary insufficiency (rare) Cerebral infarction via extracranial vessels (rare) Ocular findings – Loss of vision – Eye muscle-paresis

ESR CRP

Aching shoulders and/ or bilateral stiffness

Polymyalgia of pelvis/ shoulder (bilateral)

1. Clinical findings in temporal arteritis/ polymyalgia rheumatica

Age over 65 years Depression and/or weight loss Bilateral tenderness on palpation of upper arm 2. Diagnostic criteria of polymyalgia rheumatica Patient over 50 years at first manifestation Newly occurrent headache Clinical findings in temporal arteries: tenderness on palpation, pulselessness highly increased ESR positive arterial biopsy 3. Diagnostic criteria of temporal arteritis

4. Histology of Takayasu’s arteritis

6. Histology of occluded temporal artery

7. Temporal arteritis

(*$&$7%* ;//5&1*1<4

Headache

!

"

5. Branch stenosis in Takayasu’s arteritis

8. Head skin ulcer

A. Clinical features of giant cell arteritis: Takayasu’s and temporal arteritis

! " =>>

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Neutral proteases

Histamine

Histamine, neutral proteases, prostaglandin D2, leukotriene C4 Mast cell

Blood vessel Vasodilatation

Skin lesion

Permeability

A. Pathogenesis

Aspirin kg

Dermographism

Pressure Cold

Heat Light

Drugs

Bridging of IgE molecules

X-ray contrast agents

IgE

Aquagenic X-ray Adrenergic stress

Cholinergic sweating Food

1. Physical urticaria

Preservatives

Hypersensitivity, histamine

Sensitization, Min. degranulation

2. Immunological urticaria

Mediators

Edematous swelling

>24h

Hyperpigmentation

Healing with residual pigmentation

C1q Vasculitis

"

3. Pseudoallergic urticaria

Rubbing

Urticaria

38,*,9$8 :55;*'8'(1

Edema

Immune complexes C3

4. Urticarial vasculitis B. Triggers and manifestations

Complement factors

Mastocytoma

5. Urticaria pigmentosa

Spontaneous regression in 90%

! " <=?

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Toxic event

Allergen

Acute toxic dermatitis

Cumulative toxic dermatitis

Allergic contact dermatitis

– strong event – only once – e. g., sunburn

– weak event – repeatedly – e. g., frequent hand washing or desinfection

– non-toxic event – sensibilization causes type IV allergy – e. g., nickel

A. Causes of dermatitis Nickel Lymphocyte infiltration

Leather, chromate

Vasodilation and dermal edema

Intraepidermal edema

Hair dyes with p-phenylene diamine

#$',')%$ 6778,+$+09

Toxic event

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Rubber compounds Antibiotics, neomycin

B. Contact allergens Epidermis

Edema

T cell

Light

Inflammatory Endothelium: infiltrate adhesion molecules Extravasation

Sensitization Langerhans cell

Migration Lymph nodes

Allergen

Repeated antigen exposure

TNF-! IL-1 Keratinocytes

a) Sensitization C. Pathogenesis

TNF-! IL-1

b) Induction of eczema

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Genetic factors Metabolic disorders Immunological disorders

Fc!RII on monocytes

IgEimmune complexes

IgE

Allergic rhinitis

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IgE

Allergic asthma Eczema on flexor surfaces

TH2

IL-4

B cell

Fc!R Food

Frequent infections

Skin irritations Psychological factors

TH1

Mast cells, basophil granulocytes, releasability

2. Pathogenesis

1. Trigger factors

A. Atopic dermatitis (neurodermatitis)

73&6&',3 8++06$3$92

Environmental factors

" Systemic manifestation Complement Deposition of immune complexes in vessel

IgA nephropathy Petechias Urticaria Papules Hemorrhage Necrosis

Blood vessel changes B. Leukocytoclastic vasculitis

Symmetric manifestation

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Psoriatic arthritis

Immune dysregulation IgA , ANA IL-8

Latent psoriasis

Exogenous

b c

HIV

Provocation UV

Ca2+

Manifest psoriasis Epidermal hyperregeneration

a

Endogenous

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Impaired keratinization IL-8, LTB4, EGF, TGF-! keratinocytes

Sites of predilection

Genetic predisposition HLA-DR7, -Cw6, -B13

!

Skin scaling C2H5OH

Inflammatory reaction

Autoactivation

1. Pathogenesis 2. Symptoms

A. Psoriasis Desmoplakin

"

Cell membrane

Autoantibodies Cytoskeleton

Normal skin

Desmoglein

Acantholysis

Desmosome Tonofilaments Autoantibodies

Bullous pemphigoid Epidermolysis bullosa acquisita Herpetiform dermatitis

2. Subepidermal blister formation B. Bullous skin diseases

Laminin BP-I, -II Collagen VII Reticulin

Collagen IV

Basal cell Hemidesmosomes Lamina lucida Lamina densa

Basement membrane

1. Pemphigus

Dermal connective tissue

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Vitamin B12

Chief cell

Pepsinogen

Autoantibodies

HCL

Maldigestion/ malabsorption

Achlorhydria

Parietal cells

Intrinsic factor

Vit. B12 deficiency

Intrinsic factor Chief cell

Posterior funiculi

Parietal cell Atrophic mucosa

"

Pyramidaltract

Pernicious anemia:

Fundus

Hyperchromic, macrocytic anemia

Hypersegmentarted granulocytes

Funicular myelosis

A. Chronic atrophic gastritis type A Nutritional fats

Macrophages with PAS+ phagosomes

Hyperpigmentation

Fever of unknown origin Arthritis

Heart involvement Intestinal lumen

Intestinal Lymph epithelium lymphatic blockade

Steatorrhea

Malabsorption

#6('()-6 =>>9'&6&?1

Corpus

#

Blood vessel

Weight loss

B. Whipple’s disease

Diarrhea

Tropheryma whippeli

Steatorrhea

Gliadin from food

Transglutaminase

Cross-linked gliadin

Normal intestinal mucosa

TH2 TNF-!

TH1

Fibroblasts Celiac atrophy

C. Gluten-sensitive enteropathy

Mucosal transformation and destruction

IgA Autoantibodies

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HLA-DR1 -DQw5 Assoc. with smoking Segmental

Shorter breast feeding period

HLA-DR2

Refined carbohydrate

Associated with Autoimmune IgA-nephritis hepatitis

Discontinuous

Primary sclerosing cholangitis

continuous

!

Ulcers

Transmural

superficial

Bladder fistula Diarrhea Inflammatory conglomerate tumor

Typical sites of manifestation

Typical pattern

Typical sites of manifestation

A. Crohn’s disease IgA switch

Pseudo-polyps Flattening of mucosa

Bloody mucous diarrhea

Typical pattern

B. Ulcerative colitis

IgG

Chemotaxis Activation of complement cascade

C3a

Chemotaxis adhesion molecules

C5a

Granulocytes Eosinophils Eicosanoids

Macrophages Degranulation

#0+'+1-0 5667'%0%89

Fissural

"

Sulfasalazine, 5-ASA, steroids Leukotriene LTB4

PGE2, PGF2, thromboxane

Lipid peroxidation DNA damage protease inhibitors

Vasodilation edema pain Invasion of foreign antigens into intestinal lumen

Histamine, serotonin, bradykinin lysomal enzymes, O2 radicals

Invasion of foreign antigens into intestinal lumen

Tissue damage

C. inflammatory reactions in the intestinal wall

IL-12

T B

IgG2 IFN-!

D. Pathological mechanisms

IL-5

T H1

ANCA

IgG1 TNF-" IL-2 Crohn’s disease

TH2

IgG3

Ulcerative colitis

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!#$%&''#() ?&@)0 ;&-),-)-

Periportal field with artery, periportal vein, and bile duct Central vein

Criteria [ ] : HLA-DR3, -DR4 Associated with rheumatoid arthritis, glomerulonephritis, ulcerative colitis, Crohn’s disease Hashimoto’s disease Clinical findings:

Lymphocyte infiltration

Lobe of liver Lab. findings:

Jaundice

Fever

Sjögren’s syndrome

!

Histology: chronic hepatitis

Hypergammaglobulinemia

Inflammatory infiltrates in periportal field

Hepatosplenomegaly

Itching Nausea

Cryoglobulinemia Diarrhea

Transaminases GOT, GPT cholestase: bilirubin, !-GT, AP Autoantibodies LKM 1

cytochrome P450 IID6, in autoimmune hepatitis

LKM 2

cytochrome P450 II C9, in drug-induced hepatitis

LKM 3

UDP-glucuronyltransferase in chron. hepatitis D

ANA Ab

asialoglycoprotein receptor, smooth muscles

A. Autoimmune hepatitis Bile duct

Hepatocytes

Lymphocyte infiltration

> 40 years

Associated with Symptoms: Sjögren’s syndrome melanin (50%), scleroderma, CREST, rheumaHypertoid arthritis, pigmenHashimoto’s tation of the thyroiditis skin B. Primary biliary cirrhosis

32&(&8,2 <''#(%2%91

Vasculitis

"

Micronodular cirrhosis Cholestase, maldigestion Jaundice

Xanthomas

bilirubin , !-GT , alkaline phosphatase serum cholesterol

AMA (antimitochondrial autoantibodies)

Cholestasis, malabsorption bilirubin , !-GT , AP > 40 years Associated with HLA-B8, DR3 ulcerative colitis in 50% C. Primary sclerosing cholangitis

Thickening and stenosis of bile ducts ANCA

! " =>A

47'%'(37 :;;<%.7.19

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Chromosome 5

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House mite

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Increased IgEsynthesis

R

A. Genetic predisposition

Animal hair Animal dung

SO2

Feathers

O3

Mold

Cold air

Nicotine

Stress

Drugs Chemicals

Physical activity

B. Trigger factors

Mucus T cell MBP ECP

Proteases PAF histamine leukotrienes

IL-4 IL-10

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TH2 IL-4 IL-6

IL-13

IgEsynthesis Mast cell

1. Sensitization C. Pathogenesis Grass Grains

47'%'(37 :;;<%.7.19

Reexposure

APC

Smooth muscle contraction Glands

2. Reexposure, early reaction

3. Late reaction

IgE-loaded basophils

Alder Hazel Birch Mold Mites

Histamine

-Sneezing -Rhinorrhea -Contralateral -hypersecretion

Nasal allergen exposure

Animal allergens Occupational allergens D. Allergic rhinitis

Edema of mucosal epithelium

Prick test

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Unknown pathogen Monocytes chemotaxis, activation

Mycobacteria? Viruses?

!

Hematogenic spread??

Alveolus

Inhalation of pathogens

Lymphocytic (CD4) alveolitis

Noncaseating granuloma

Fever, general symptoms

CNS involvement

Calcium ACE neopterin

Parotitis

CD4/CD8 ratio "-globulins

Lung involvement 90%

Uveitis Neuropathy Mediastinal lymphadenopathy

3. BAL

Cardiac involvement Spleen infiltration

Liver infiltration Skin lesions Loefgren’s syndrome:

4. Chest radiograph

fever, polyarthritis, erythema nodosum, biliary lymphadenopathy

Bone lesions

Inhaled noxae

Immune complexes

T cells

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2. Clinical manifestations A. Sarcoidosis

R

;2(4(&$2 ,3314'2'<5

1. Pathogenic model

IGF-1 PDGF TGF-!

Monocyte chemotaxis and activation

Interstitial infiltration, fibrosis Type I Granulocytic alveolitis

Alveolar macrophage

IL-8, leukotrienes

5. T-cell infiltrate in the liver

Granulocyte chemotaxis

B. Idiopathic pulmonary fibrosis

Pulmonary fibrosis

Type II pneumocytes

Honeycomb lung

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Farmer’s lung

Hay Actinomycetes

Bird-breeder’s lung

Animals

Animal-derived proteins

Airconditioning

Air conditioner lung

Compost lung Compost

Aspergillus

Polyurethane foam

Isocyanates

Chemical worker’s lung

R

1. Most common clinical syndromes Early granulocytic alveolitis

R IL-8, chemokines

C3

Alternative complement activation pathway C3a

C3b

4–12 h

Lymphocytic alveolitis (CD8+), interstitial mononuclear infiltration, granuloma formation

5+'('*2+ 6778(/+/-3

Aureobasidium p. Actinomycetes

"

48–72 h

2. Immunological pathogenesis 4–8 h

Inhalation

a) Acute course

Chronic exposure

b) Chronic course

Therapy with steroids

12–24 h

Fever, coughing, dyspnea

Weakness anorexia weight loss

Dyspnea while resting

Impaired ventilation

3. Clinical outcomes A. Extrinsic allergic alveolitis

4. Chest radiograph

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Secretory proteins M. tuberculosis

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Minimal lesions

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Parietal epithelium

Periorbital edema

Immune complex in capillaries Endothelial cell Glomerular basement membrane (GBM) Podocyte with foot processes

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Pleural effusion

Hypertension

Ascites

1. Anatomy Hypoalbuminemia

Proteinuria Urine sediment; hyaline and granular casts

Edema 2a. Immune complex deposition

2b. Anti-GBM Ab

2c. Anti-epithelial cell Ab

1. Clinical symptoms

Children Adults Membranous glomerulonephritis

Direct damage (C5b-C9)

Secondary T cell migration

5%

20%

Lipoid nephrosis minimal change GN

60%

10%

Focal segmental glomerulosclerosis

10%

10%

Membranoproliferative glomerulonephritis

10%

5%

Proliferative GN (focal, lgA....)

10%

15%

5%

40%

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Proteinuria Cytotoxicity

Hypertension Oliguria

C5a Hematuria

Platelet aggregation

Neutrophil activation

Monocyte activation

Proteases, eicosanoids, NO, cytokines, growth factors

3. Mediators of glomerular damage A. Mechanisms

Mesangial cell activation

1. Clinical symptoms

Urine sediment; Red blood cell casts

Postinfectious GN Rapidly progressive GN IgA nephropathy

2. Causes of the nephritic syndrome C. Nephritic syndrome

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7(0&,/8(0%,.*/$1$1$9 B?C

Loss of digital extensions Albuminuria T-cell cytokines

Lipid aggregation in tubular cells Nephrotic syndrome

Fusion, loss of foot processes

A. Minimal change glomerulonephritis (GN) HIV-infection, heroin abuse

Deposition of IgM, fibrin

Secondary to other GN

Loss of foot processes

Electron microscopy Damage to epithelial cell

Poor response to corticosteroids

Mesangial proliferation

B. Focal segmental glomerulosclerosis Idiopathic >80% (genetic predisposition)

Segmental sclerosis Thickening of basement membrane

Infections Carcinomas

Poor response to corticosteroids

Nonselective proteinuria

SLE Drugs Gold, mercury

M

Deposition of proteins, lipids, fibrin

Secondary to compensatory hypertrophy Idiopathic

Good response to corticosteroids, good prognosis

Loss of foot processess

40% - Progressive course with renal failure

C. Membranous glomerulonephritis Type I Hepatitis B, C SLE Infections? Subendothelial IgG, complement deposition

Type II Ab against C3-convertase Complement activation, consumption Intramembranous C3 deposition

D. Membranoproliferative GN (MPGN)

)($%$5'( ?&&8%0(0+2

Podocyte

!

"

Granular IgG deposits Nephrotic syndrome acute nephritis

40% - Progressive renal failure

Tramtrack effect

30% - Partial renal failure 30% - Persistent nephrotic syndrome EM: Dense deposits

! " @@D

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Immune complex deposits

Common symptoms, hematuria

No specific therapy

!

Hypercellularity in all glomeruli Children

Adults

> 80% Healing

~50% Healing

A. Postinfectious (acute proliferative) GN Type I Anti-basement membrane Ab

with/without pulmonary bleeding

Type II With immune

complexes: idiopathic, systemic diseases, postinfectious, SchönleinHenoch

Renal insufficiency in 90%

Aggressive immunosuppression: high-dose steroids cyclophosphamide

Type III Pauci-immune: idiopathic, Wegener’s, PAN

B. Rapidly progressive GN Genetic predisposition, infections

Crescent Infection of airways

No specific therapy

Hematuria, kidney pain

Increased IgA synthesis

Children

IgA immune complexes

Linear IgG deposition

Mesangial IgA deposition, mesangial proliferation

High Slowly healing rate progressive IgA deposits Medications

? Infections

Rehydration Steroids??

Fever, eosinophilia, exanthema (25%)

Drugs

Renal damage (hematuria, proteinuria, leukocyturia)

Interstitial deposition

Granuloma formation

"

Adults

C. IgA immune complexes Idiopathic

=0'('+10 >55,($0$<;

Hypercellularity

Interstitial edema, kidney enlargement

D. Tubulointerstitial nephritis

Renal insufficiency (avoid dehydration) Eosinophil infiltrates

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TSH TSH receptor (TSHR) TPO

I2

I2 I 2

Target

Ab

Thyroglobulin (Tg)

Tg-Ab

Microsomal antigen, thyroidperoxidase TPO

TPO-Ab

Decreased iodination of Tg

TSI

Thyroid stimulating immunoglobulin: hyperthyroidism (Graves’ disease)

TBII

TSH-binding inhibitory immunoglobulin: hyper- or hypothyroidism

TSBI

Thyroid-stimulation blocking immunoglobulin: hypothyroidism

T3 I2

TSHReceptor

T4

I2

T4 T3 Thyroglobulin

A. Autoantigens of thyroid

B. Most important autoantibodies APC

Exophthalmos

TH2

Goiter Heat intolerance

Palpitations Tachycardia IL-4, IL-6

Viral antigen “mimicry” of TSHR?

Diarrhea

Exophthalmus

Tremor Weight loss HLADR3+ HLAB8+

Effect of antibody

B cell

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1. Clinical symptoms

TSHR

2. Immunopathogenesis

C. Graves’ disease ?

HLA DR5

Activation of CD4 cells

IFN-! TNF IL-2

HLA DR3 CD8

Thin hair

CT scan of the eye muscles 3. Endocrine orbitopathy

Goiter (often small) Macroglossia Deep, hoarse voice

Damage to thyroid cells Release of autoantigen

TPO and Tg Ab

D. Immunopathogenesis of Hashimoto’s thyroiditis

Bradycardia

Hashimoto (histology)

Cold intolerance Hyporeflexia

Myxedema E. Clinical signs of hypothyroidism

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Ketoacidosis Polydipsia Polyphagia Polyuria

1. Early symptoms

2. Late complications

P1

Risk DRB1-0405 DQB1-0201 DQB1-0302 DQB1-0502

P11

Binding of peptides with negative charge at position 9 (Asp/Glu)

P1

Protection

P11

Aspartate at AA 57

TH2

#-islet

IL-4 IL-10

Mass of #-cells

Disease signs

5 10 20 15 25 Years after disease onset

Intrainsulitis TH1

Infection Molecular e.g. Coxmimicry sackie virus of GAD

TCR

Binding of peptides with positive charge at position 9 (Ser/Gly/ALA)

B. Genetic control

A. Clinical manifestations

Peri-insulitis

IFN-! TNF-" IL-12

TH1

Serine/Ala at AA 57

DRB1-0403 DQB1-0301 DQB1-0602

TCR

Anti-GADimmune response

Release of other autoantigens (e.g. insulin)

APC

#$%&%'($ 5::9&.$.8;

Microangiopathy Stroke Retinopathy Cataract, glaucoma Hypertension Ischemic cardiopathy Glomerulosclerosis Autonomous neuropathy Peripheral vasculopathy Gangrene

!

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Local immune response

C. Hypothetical pathogenesis Genetic defects

Adrenal cortex

21-OH

Parathyroid gland

Calciumsensor

Autoantibodies Interstitial with Leydig multiple cells reactivity

Molecular mimicry

17"-OH/ p450scc

Liver

Cytochrome p 450IID6

Gastric parietal cell

Intrinsic factor

Antigens in steroid-producing cells D. Autoimmune polyglandular syndrome (APS)

Type I: juvenile form

Adrenocortical insufficiency Hypoparathyroidism Candidiasis Hypogonadism, chronic hepatits, pernicious anemia less frequently

Type II:

Adrenocortical insufficiency Autoimmune thyropathy

Type III:

Autoimmune thyropathy Hypogonadism Ovarian insufficiency

! " @AC

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Minor criteria

Major criteria

Fever

(Pan-)carditis

!

Erythema marginatum Arteriole

Chorea (uncontrollable movements)

High ESR

CRP

3. Histopathological picture

Subcutaneous nodules Arthritis

1. Primary manifestations Cross reactivity: Capsule Cell wall

Cytoplasm

Protein antigens = M Groupspecific carbohydrates Mucopeptide Cytoplasmic membrane

myosin, tropomyosin, synovia, cartilage, brain

Streptococcal pharyngitis

Lymph nodes/tonsils

Valves Neurons of subthalamic and caudate nuclei

2. Streptococcal antigens

Valvular vegetations

Aschoff’s bodies

Fibrinous pericarditis

A. Rheumatic fever

4. Pathogenesis

1 week– 4 months after myocardial infarction

After cardiothoracic surgery

57*:*+(7 >''&:272?1

Long PR interval

"

Cytotoxic reaction Bacteria CD8

Spirochetes

MHC II CD4

Parasites

Release of myocardial antigens

Rickettsia Fungi

Infection

Viruses

B. Myocarditis

Expression of microbial antigens, adhesionmolecules

Antisarcolemmal Ab Crossreactive antibodies

Antimyosin Ab

C. Dressler’s syndrome

! " @AA

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#8(1*&(6 D+(6%$0*0

1. Early lesion

2. Late lesion

!

3. MRI

Myelin proteins: MBP PLP MOG MAG

Genetic predisposition (DR15, DQ6) + noxious agent (Virus? Mimicry?)

<(*-*+,( =778-$($)9

A. Morphological and histopathological findings Reduced Phagomyelin synthesis, cytosis apoptosis

IL-1, TNF-!, NO, H2O2

Encephalitis

Release of myelin antigens Diapedesis of autoreactive T cells

T cell transfer

Macrophage

Astrocytes

IL-2 TNF-! IFN-"

Microglia

Disease transfer

CD4

TH1

Activation

TH2

B. Experimental autoimmune encephalitis (EAE)

C. Immunopathogenetic mechanisms

Reduced signal transmission

Increased signal transmission Stimulating neuron Inhibitory neuron

Stimulating neuron

Myelin auto-Ab

IL-4 IL-10

1. Treatment of relapsing-remitting MS Corticosteroids, IFN-#, polypeptides (glutiramer), azathioprine, intravenous immunoglobulins

2. Treatment of primary 2. progressive MS Impaired vision Paresis Ataxia

D. Clinical features

Paresthesia Tonic contractions

Lhermitte’s syndrome

"

Methotrexate, cyclophosphamide, (IFN-#), cyclosporine

E. Treatment approaches

! " ABE

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!$:1'(:%D10/E2,0%':,0 ?%4,'4,4

Ascending Dysphagia symmetrical muscle weakness

Respiratory failure vegetative disorders

1. Clinical features

Neutralizing antibodies

Antigen release

Campylobacter jejuni, EBV/CMV Molecular mimicry

Autoreactive T cells

APC CD4+ T cell

TH2

TH1

Blocking of Fc-receptors

a) Plasmapheresis

Antiglutamate receptor Ab

Uncontrollable epileptic seizures Physiological

mV +20

Na+

Na+

Na+ Glu

Na+

0

Proteases NO O2

2. Pathogenesis

A. Guillain-Barré syndrome

Na+ Na+ Na+ Na+

TNF-!

b) High-dose i.v. b) immunglobulins

3. Therapy

Tumors (malignant lymphomas)

Central neuron

Hu antigen Ri antigen

Neuroblastoma

Na+

Breast carcinoma Peripheral neuron

Hu antigen

Na+ Na+

Ovarial carcinoma Purkinje cell

Yo antigen

-90 B. Rasmussen’s encephalitis

"

Lung carcinoma

Anti-glutamate R3 Ab

Na+

;&%(%7'& @22$(1&1>/

Surgery

!

C. Paraneoplastic neurological syndromes

! " ABF

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mV 0

Diplopia Chewing problems

Nasal voice Dysphagia

!

-90

Decremental response to repetitive stimulation mV 0

a) Ocular form

b) Generalized form

-90

Improvement after administration of acetylcholinesterase inhibitor

c) Electrophysiology

1. Clinical features of myasthenia gravis Defective T cell interactions

Antibody production

c) Thymoma-associated MG Complementmediated lysis Na+

Expression of AChR/titin in tumor tissue

Myogenous cells in thymus HLA-DR3 HLA-B8

Na+

+ APC

CD4

>:+*+<%: ?119*5:5;$

Failure of respiratory musculature

"

Reduced AChR count

a) Neuromuscular a) junction

b) Changes b) in MG

2. Pathogenesis A. Myasthenia gravis (MG)

d) Thymitisd) associated d) MG

Dry mouth Lung cancer Weakness of shoulder and pelvic muscles Micturition disturbances

B. Lambert–Eaton syndrome

Crossreactivity of tumor cells with presynaptic calcium channels

Reduced ACh release 0

Low baseline potential

-90

Serial stimulation

! " @AC

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Sclera

Ciliary body

Conjunctiva

Iris

Intraocular inflammation

Extraocular inflammation

!#$%&'( $#A 4$%*&1+#+525

Lens Vitreous body

Cornea

Retina Choroid

!

Retina

Choroid Sclera

1. Direct antigen invasion a) Loss of protection

2. Hematogenic spread of germs or deposition of preformed immune complexes

b) Perforating wound

Autoantigen

Foreign antigen

B. Immunological pathomechanisms Photoreceptor

"

4. Release of lens antigens induces autoimmune reaction against different eye antigens

3. Modification of autoantigens, molecular mimicry

Immunization with IRBP

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A. Anatomy of the eye

APC

TH1

CD4

IL-12 Lymphocyte transfer to a healthy animal

Uveitis

C. Experimental autoimmune uveoretinitis (EAU)

Uveitis

! " >?B

25).)(-5 >004.%5%68

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1. Causes B. Conjunctivitis

Pollen, grass, blossoms

2. Dry eye

IgE-bound antigens

Binding to mast cells

3. Sjögren’s syndrome

Histamine release

Vernal conjunctivitis

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Cornea

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3. Neovascu3. larization

4. Epithelial lesion Corneal ulcer

3. Episcleritis

4. Scleritis

D. Pathology of the cornea Retina Choroid Sclera

Viruses (HSV, HZV), fungus, parasites Idiopathic form 50%

Episclera

Rheumatoid arthritis, Sjögren’s syndrome, panarteritis, SLE, Wegener’s granulomatosis

1. Normal sclera

2. Etiology

E. Pathology of the sclera

Necrotizing scleritis

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50 % Idiopathic HLA-B27 associated Spondyloarthropathy associated

Molecular mimicry

Heterochromic uveitis (Fuchs’ disease)

Immune complex deposition

Behçet’s disease, JRA, sarcoidosis

Iridocyclitis

HLA-B27 Polyclonal B-cell activation

Herpes infection, yersiniosis, borreliosis Iritis Cyclitis

Iritis

A. Anterior uveitis Peripheral Idiopathic retinitis

70 %

Antigen?

T-cell activation

Vitritis

Sarcoidosis

Snow balls Snow banking

Lyme borreliosis

Cystoid macular edema

Multiple sclerosis

IFN-! HEV

Antigen presentation T-cell proliferation

B. Intermediate uveitis Syphilis Tuberculosis Borreliosis Leprosy Whipple’s disease Brucellosis

Endothelium

CMV EBV HSV VZV HIV

Histoplasmosis

Toxoplasmosis

PAN Behçet’s disease Sarcoidosis

Vitreous cells Birdshot retinopathy ApMPPE

?

Sepiginous choroiditis

19'#'809 -..6#(9(?@

Posterior cyclitis

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C. Posterior uveitis: Causes/associations Congenital/acquired

Retinal vasculitis

Vitritis Retinitis/papillitis Inflammation of anterior chamber

Granulomatous papillitis

Destruction of neural fibers

Retinal infiltrates + pigmented scars D. Toxoplasmosis

Scar focus

Hypersensitivity to histoplasma Ag

F. Histoplasmosis

Choroidal infiltrates

HLA-A29: RR=224!! Binding of S-antigen-peptides?

Subretinal neovascularization, bleeding Peripheral + peripapillary scars

E. Sarcoidosis

Granulomatous intra-/subretinal inflammation Macular lesion

Pigment defects G. Birdshot chororetinopathy

! " ABE

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Sympathetic ophthalmia

Behçet’s disease

Vogt-Koyanagi-Harada syndrome

Phacogenic uveitis

Sarcoidosis

Multifocal chorioiditis and panuveitis

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A. Panuveitis: Causes and associations

Obliterative vasculitis of the retina

B. Uveitis with Behçet’s disease

Hypopyon

Neuritis, accompanying vasculitis

Bilateral panuveitis

EBV??

CNS symptoms Meningism Headache Pleocytosis

Vitritis Chorioretinal lesions (fundus) Scar formation in macula

Hearing loss, tinnitus Vitiligo, alopecia

Choroid detachment

C. Vogt-Koyanagi-Harada Syndrome HLA-A11

Perforating wound

D. Multifocal choroiditis and panuveitis Anti-S-100 Anti-IRBP

CD8

Injured eye

Attack on healthy eye

CD8 CD4

TNF

SPA

Right eye: scar

Left eye: beginning iritis

E. Sympathetic ophthalmia RA

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Granuloma formation

CD8

Müller cells

Chronic outcome with recurrent disease

Choroid lesions

-A)%)0$A G??&%/A/8;

IL-8

SLE

SJS

JRA

Wegener

PAN

Behçet

Sarcoidosis

Conjunctivitis sicca Keratitis, corneal ulcer Episcleritis, scleritis Anterior uveitis Vasculitis Choroiditis, panuveitis

F. Ocular manifestations of systemic diseases

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Spermatocyte peptide antigen

Infertile due to auto-Ab

Acrosomal antigen

Oocyte 1. Antigens

Zona pellucida antigens, ZPI ZPII ZPIII

Infertile due to auto-Ab or iso-Ab (antispermatocyte)

2. Auto- or isoimmunity

Prevention of sperm exposure for 6 to 12 months

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Intrauterine insemination Immunosuppression In vitro fertilization (intracytoplasmic sperm injection) 3. Treatment options

TH0

T cell NK cell Progesterone Fetus: paternal MHCantigens = foreign

PIBF TNF IL-10 Trophoblast cell derived factor HLA-G

TH2 IL-4 IL-10 IgG1

APC FAS-L

NK cell

Trophoblast FAS

CD94 Placenta

Trophoblast cell

Trophoblast

Inhibitory signal

T cell

Apoptotic cell

B. Maternofetal tolerance Transient hypothyroidism

Transient myasthenia

Transient hyperthyroidism

’s d G r av e

I TP

Discoid skin change

A

SS-A Ab

SL E, S j ögr en

SS-B Ab

AV-block Bradycardia

Transient neonatal lupus syndrome

C. Neonatal autoimmune syndromes

Anticardiolipin Ab

AC

Day 1 to 7 after birth

~20% of pregnancies of myasthenic mothers

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Anti Ib/IX Ab

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Difficulty in swallowing and breathing

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Cytopenia

Stroke, thrombosis

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Basic immunization

Booster vaccinations Measles mumps rubella

H. influenzae, diphtheria, pertussis, tetanus

Tuberculosis (Still performed in some countries)

Rubella vaccination for girls

Poliomyelitisoral vaccination

BCG i.cutan.

DT

Poliomyelitis

PPD-test

DPT

3

15

18

6

Age in months

10

11–14

14

Age in years

A. Vaccination calendar in childhood Pathogen

Indications

HBV

– Hospital staff, Short – Dialysis patients, fever reaction – Family members of HBV-carriers – prostitutes – Homosexuals – Drug users – Travel in endemic regions

recombinant HBsAg-vaccine

Influenza

Very ill patients > 60 y

killed vaccine

Side effects

Contraindications – Pregnancy – Known allergy

Allergic reactions (rare)

– Acute diseases – Egg albumin allergy

Rabies

– After exposure (animal bite) – Before exposure in veterinarians, forest-workers, hunters, etc.

Fever, lymph node swelling

Before exposure: allergy to neomycin and tetracycline in vaccine

Pneumococci

– Splenectomy – Sickle cell anemia

Mild local reaction

Ongoing pneumococcal infection, chronic purulent diseases

killed vaccine

dead vaccine capsular polysaccharides in 14 serotypes

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Birth

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Low grade fever

Yellow fever

Low grade fever

Typhoid fever

No problems

Allergies

killed vaccine live vaccine attenuated D17-strain live vaccine

B. Optional vaccinations

– Pregnancy – Egg albumin allergy

Use killed vaccine in high-risk patients

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=(B 74**)%(.

+

MHC class II

Processing by APC

A. Synthetic peptides

Strong B-cell response weak T-cell response

E. coli Plasmid with gene for vaccine and with antibodyresistant genes

Selection of plasmid-bearing E. coli by antibiotics production and export of vaccine into supernatant

Centrifugation and purification

Recombinant vaccine

B. Recombinant proteins

"

Bacille CalmetteGuérin (BCG)

Plasmid

Vaccination Recombinant vaccine alone is ineffective

Cloning into suitable carrier

69)%)*49 >22;%191?$

Synthetic peptide

Antigen

!

Protected by T cells

C. Recombinant vaccine strains

Restriction endonuclease Wild-type virus: highly virulent

Single strand-specific nuclease S1

Ligase

Isolation of genome Virus receptor

Reduced replication – but strong stimulation of immune response

CD8 MHC class I

D. Deletion mutants

Low virulence

! " @AC

>-*%*;,- /114%$-$52

"

/114%( <=,)1,;$-$52

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#$%&'()$*+,- !%'*.*%0-,11,'$)2 3)45& ,%+ :-4;$;$)'*;$*+&

Physiological stimulus

Inflammatory stimulus IL-1

"

Phospholipids

NSAIDs

Phospholipase A2

Glucocorticoids

COO– CH3

Arachidonate COX 2

Prostaglandin H

Glucocorticoids

Lipoxygenase Leukotriene A4

(indirect effects)

Prostacyclin

Prostaglandins

Leukotriene B4 chemotaxis

Thromboxane A2

Peripheral resistance Stomach protection Renal function Hyperalgesia Bronchodilatation

Inflammation: Vasodilation Mediators Proteases

Leukotriene C4, D4 bronchoconstriction

Florid ulcers Bronchial asthma 2. Contraindications

!

Impaired bloodcell formation Pregnancy, nursing

1. Mechanisms A. Nonsteroidal anti-inflammatory drugs (NSAIDs) Myelopoiesis MHC class II Fc-receptors IL-1, IL-6, TNF-" prostaglandin

CH 2 OH CO HO

OH

O

Lymphocytes via redistribution IL-2 production IL-2 effect

>-*%*;,- /114%$-$52

COX 1

#

IL-2

Second messenger

hsp 90

hsp

90

NF!B

?

Cation transport

Membrane stability Activation capacity

mRNA

mRNA

ACE, lipocortin, endonuclease

Phospholipase A2, COX2, receptor expresion, cytokines

1. Mechanisms B. Glucocorticoids

HO

Free radicals

CH2OH CO OH

O

Adhesion to endothelium Eosinophils Basophils Neutrophils

Permeability of blood vessels Adhesion molecules IL-1 Prostaglandins

Fibroblast proliferation Fibronectin, COX 1 prostaglandins

2. Effects on the immune system

" # ?@@

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!#$%&'$()*+%$',A ./0+*12*,12(&%3'A 45+6(,(+(7%#'A (#3 9*+3

Methotrexate

Dihydrofolate reductase

!

Thymidine synthesis T

Folate

FH4 (tetrahydrofolate)

FH2 SH

Feedback

N

A

6-Mercaptopurine Ribose

C1

Purine biosynthesis

N

G

Wrong base

P

1

Methotrexate 5 10

Inhibition of expression

A. Antimetabolites Cyclophosphamide

Immune modulation Structures

Liver

H

Cl

CH2

CH2

Cl

CH2

CH2

OH

N P

O

Cross-linkage

O

B. Cyclophosphamide

Cell death

.+%#%0(+ :&&5#*+*=/

N

IN

Azathioprine

"

COOH OH

H N

SO2

N

N

Au Sulfapyridine

5-ASA

?

Inhibition

Side effects – – – –

Headache Dizziness Fever Allergic reaction

C. Sulfasalazine

T cells

NK cells

– Chemotaxis – Inflammation mediators – Enzymes

SH Au HS D. Gold

! " >?B

#&+,+%1& 8665,'&':$

!

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MHC class II (Auto-)antigen

TCR

!

Tacrolimus (FK506)

Cyclosporin A FKbinding protein

IL-3

PKC"

Ca2+

IL-4

IFN-! IL-8

Calcineurin

P P

NFAT

Cytokine production

IL-2 effect

NFAT Promoter Cytosol NFAT NFAT

Rapamycin

Tacrolimus Cyclosporin A

IL-2 gene

A. Cyclosporin A/FK506

Nucleus

Effect on cellular immunity

"

Gln, HCO3, ATP

IMP Mycophenolate

#&+,+%1& 8665,'&':$

Cyclophilin

IL-1

NAD+

IMP-DH

NADH XMP Glu + ADP

GMP synthase

Leflunomide

Gln + ATP

DHO-DH

GMP Guanosine 5 diphosphatefucose N-Glycosylation of glycoproteins Adhesion molecules

B. Mycophenolate

GTP, dGTP DNA/RNA synthesis

Dihydroorotate FMNH2

Adhesion molecules

FMN Orotate

GTPcyclohydrolase 1

RNA/DNA synthesis

Tetrahydrobiopterin Redox reactions NO formation

UMP -Pool Antibodies

TTP

C. Leflunomide

UTP

CTP

! " ;
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Abs of single specificity produced

+

Murine part

Fusion

Myeloma cell

(Nonsecretory)

Human part

Hybridoma cell

1. Production

2. Reduction of antigenicity

TNF-!

Anti-TNF antibody

TNF-!

T cell Blood vessels

Migration

Synovitis

Cartilage/bone

– proliferation – enhanced adhesion

– activation – chemotaxis – IL-8 production

– proliferation – activation – pannus formation

– degradation – activation – faulty repair

rhs-TNF-R: Fc fusion protein

Joint changes of RA

"

a) Inhibition of TNF-! effects by anti-TNF Ab (e.g., in RA) Anti-ICAM-1 Ab

Complementmediated lysis ADCC phagocytosis

Shedding !

"

TCR

LFA-1

CD4

:'*%*&(' 4556%$'$91

B cell

Antigen

!

ICAM-1

LFA-1 ICAM-1

Modulation Apoptosis

b) Anti-CD4 antibodies

Adhesion

c) Anti-ICAM-1 Ab

Co-stimulatory signal

A. Monoclonal antibodies Antigen phagocytosis

Antigen presentation

B-cell activity

Neutralization of pathogenic Ab

Overloading of MPS

Ag Ag

Fc#R Ag CR

B. Polyclonal antibodies

Polyclonal antibodies

Phagocytosis

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