Leptin

Leptin Raffaella Faggioni, Kenneth R Feingold and Carl Grunfeld* Metabolism Section, Department of Veterans Affairs Medical Center, Department of Medicine, University of California, San Francisco, CA 94124, USA * corresponding author tel: 415-750-2005, fax: 415-750-6927, e-mail: [email protected] DOI: 10.1006/rwcy.2001.06007.

SUMMARY Leptin is a 16 kDa protein mainly produced by adipose tissue in direct proportion to fat depot mass. Originally thought to be a satiety factor, leptin displays pleiotropic activities. Both the structure of leptin and that of its receptor suggest that leptin might be classified as a cytokine. The secondary structure of leptin has similarities to the long-chain helical cytokine family, which includes IL-6, IL-11, CNTF, and LIF, and the leptin receptor is homologous to the gp130 signal-transducing subunit of the IL-6-type cytokine receptors. Furthermore, leptin levels are acutely increased by inflammatory stimuli such as LPS and turpentine and by cytokines, indicating that leptin induction is part of the host response to inflammation. Defects in leptin production, such as observed in ob/ob mice, or in the long isoform of the leptin receptor (db/db mice and fa/fa rats) cause a complex syndrome characterized by severe obesity, infertility, and impaired immune responses.

db/db mice, which have a mutation in the db gene and display a phenotype identical to ob/ob mice, make the factor missing in ob/ob mice, but cannot respond to it. Therefore it was hypothesized that the db gene would encode for the ob receptor. In 1994 using positional cloning technique, Friedman identified the molecular defect responsible for the obesity syndrome in ob/ob mice (Zhang et al., 1994). The 16 kDa protein encoded by the ob gene was named leptin, from the Greek leptos which means thin. Leptin is produced in adipose tissue and circulating levels directly correlate with adipose tissue mass. Leptin reverses the obesity syndrome in ob/ob mice and causes decreased food intake and increased activity when administered to normal mice (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995). The leptin receptor (OB-R) was cloned shortly thereafter by virtue of its high affinity to leptin through an expression cloning strategy (Tartaglia et al., 1995). The OB-R was found to be the product of the db gene and db/db mice were shown to be resistant to leptin (Lee et al., 1996).

BACKGROUND

Alternative names

Discovery

Authors refer to the product of the ob gene as leptin, LEP or OB protein.

Forty years ago a genetic defect was identified in mice, which, if homozygous, causes a severe obese phenotype due to overeating and decreased energy expenditure (Coleman, 1978). The gene was named ob and the obese mice carrying the mutation were called ob/ob mice. Parabiotic animal experiments suggested that the ob/ob animals were unable to make a satiety factor, but could respond to such a factor from a parabiotic mate. Similar experiments suggested that

Cytokine Reference

Structure Despite the absence of sequence similarity between leptin and other long-chain helical cytokines, there is a striking structural similarity in the tertiary structure (Zhang et al., 1997). Leptin has a four-helix bundle similar to that of the long-chain helical cytokine

Copyright # 2001 Academic Press

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Raffaella Faggioni, Kenneth R Feingold and Carl Grunfeld

family, which also includes IL-6, IL-11, IL-12, LIF, G-CSF, CNTF, and oncostatin M. Furthermore, the leptin receptor is homologous to the gp130 family, the signal-transducing subunit of the IL-6-type cytokine receptors (Baumann et al., 1996).

Main activities and pathophysiological roles Leptin and the Control of Food Intake Leptin was originally proposed as a satiety factor produced by adipose tissue in proportion to fat depot mass (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995). Leptin signals the nutritional status from the periphery to the center of the brain involved in the homeostasis of energy balance. In agreement with the proposed role of leptin as a satiety factor, the absence of leptin, as observed in leptin-deficient, obese ob/ob mice, causes overeating and decreased activity. Leptin replacement normalizes food intake and increases activity, decreasing body weight to normal levels. However, the physiological role initially proposed for leptin as a satiety signal is not consistent with certain aspects of its physiologic regulation. For example, leptin levels acutely change with feeding or fasting disproportionately to the changes in fat depot (Ahima et al., 1996). Therefore, leptin cannot just be a read-out of the fat stores. Secondly, leptin treatment at physiological levels does not cause satiety, but decreases food intake in the ob/ ob to normal; higher doses of leptin are required to decrease food intake in normal animals (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995). More important, a pleiotropic role for leptin in mammalian physiology is suggested by the complex syndrome exhibited by leptin-deficient ob/ob mice and leptin receptor-deficient db/db mice. Those mice are not only obese, but they have abnormal reproductive function, hormonal abnormalities, and impaired immune function (Chehab et al., 1996; Flier, 1998; Howard et al., 1999; Faggioni et al., 1999). Leptin as a Cytokine Leptin levels are acutely increased by inflammatory stimuli such as LPS and turpentine and by cytokines, such as TNF and IL-1, indicating that leptin induction is part of the host response to inflammation (Grunfeld et al., 1996b; Sarraf et al., 1997; Faggioni et al., 1998). Furthermore, the increase in leptin production during local and systemic inflammation is absent in IL-1 deficient mice (Faggioni et al., 1998). Thus, during inflammation leptin expression is regulated in a manner

similar to the cytokine response to infection and injury. In addition, both the structure of leptin and that of its receptor suggest that leptin might be classified as a cytokine (Baumann et al., 1996; Zhang et al., 1997). Furthermore, leptin deficiency in ob/ob mice causes impaired immune responses, with lymphoid atrophy, reduced T cell function and enhanced susceptibility to infections (Chandra, 1980; Faggioni et al., 1999). Hyperresponsiveness to monocyte/macrophage-activating stimuli, i.e. LPS or TNF, is present in both genetically leptin-deficient (ob/ob) mice and physiologically leptin-deficient starved mice (Faggioni et al., 1999, 2000b; Takahashi et al., 1999). Leptin deficiency is accompanied by increased susceptibility to LPS- and TNF-induced lethality and liver injury and decreased induction of anti-inflammatory cytokines (Yang et al., 1997; Faggioni et al., 1999; Takahashi et al., 1999). Humans with genetic leptin deficiency also exhibit increased lethality from infections (Ozata et al., 1999). Leptin has been shown to have direct effects on T lymphocytes, enhancing the T helper (TH) alloproliferative response (Lord et al., 1998). Leptin polarizes TH cells toward a TH1 phenotype by enhancing INF and IL-2 and inhibiting IL-4 production. Leptin plays an important role in T cell-mediated liver toxicity in association with a regulatory effect on thymus and peripheral blood cellularity as well as on the production of two proinflammatory cytokines, TNF and IL-18 (Faggioni et al., 2000a). A diminished immune response has long been recognized as a consequence of starvation (Chandra, 1996). Thymic atrophy and decreased T lymphocyte responses, such as delayed-type hypersensitivity (DTH) reaction, are prominent features of starvation. Increased susceptibility to infection also accompanies malnutrition. Starvation also causes a significant reduction in circulating leptin levels. This decrease is now thought to be responsible for the diminished immune response during starvation (Ahima et al., 1996). Leptin administration protects mice from the lymphoid atrophy associated with starvation and reverses the inhibitory effect of starvation on the development of DTH reactions (Lord et al., 1998; Howard et al., 1999). Leptin administration is also effective in reversing the increase in LPS sensitivity caused by starvation (Faggioni et al., 2000b). Leptin and the Neuroendocrine Response to Starvation Leptin promptly signals the shift between sufficient and insufficient energy availability (Ahima et al., 1996). In fact, leptin levels fall rapidly with the onset of starvation, disproportionally to changes in adipose tissue mass. This fall in leptin levels is a signal for the

Leptin brain to initiate the adaptative responses to starvation. During the adaptative response to starvation all the nonvital activities that might increase metabolic demands are downregulated in order to put all effort into preservation of the energy stores. The endocrine changes include suppression of reproductive and thyroid function and stimulation of the hypothalamus-pituitary-adrenal (HPA) axis. Preventing the starvation-induced fall in leptin with exogenous leptin administration substantially blunts the changes in gonadal, adrenal, and thyroid axes in male mice and prevents the starvation-induced delay in ovulation in female mice (Ahima et al., 1996). When adequate caloric intake and energy stores are normal, leptin levels return to normal and its permissive role on behavioral, metabolic, and endocrine function is restored. Likewise, genetically leptin-deficient mice are not only obese, but also have some of the hormonal and metabolic disorders characteristic of early starvation, such as abnormal reproductive function, decreased thyroid hormones levels, hypercortisolemia, and decreased activity (Chehab et al., 1996; Flier, 1998). Therefore ob/ob mice seem to exist in a state of perceived starvation and as a consequence, they became obese when given free access to food.

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binding protein  (C/EBP) (Miller et al., 1996). In contrast, thiazoladinedione agonists for peroxisome proliferator-activated receptor (PPAR ) transcription factor suppress leptin expression in vitro and in vivo in rodents, and this may involve, at least in part, a functional antagonism between C/EBP and PPAR on the leptin promoter (Hollenberg et al., 1997). The leptin promoter has also been shown to be transactivated by adipocyte determination differentiation dependent factor 1 (ADD1)/ sterol regulatory element binding protein 1 (SREP1), a transcription factor responsive to insulin (Kim et al., 1998).

Cells and tissues that express the gene

Accession numbers

White adipose tissue is the major site of leptin gene expression (Zhang et al., 1994). Constitutive leptin mRNA has also been detected in placenta trophoblasts and amnion cells and in a cultured human choriocarcinoma cell line, BeWo cells (Masuzaki et al., 1997). Leptin mRNA is also selectively transcribed in specific areas of rat brain and pituitary, and in a rat glioblastoma cell line (Morash et al., 1999). Leptin gene expression is present in a number of tissues in the fetal mouse, such as bone and cartilage (Hoggard et al., 1997). Leptin messenger RNA has been detected in rat gastric epithelium and in the glands of the gastric fundic mucosa (Bado et al., 1998).

GenBank: Human: U18915 Mouse: U18812

PROTEIN

GENE AND GENE REGULATION

Chromosome location The human leptin gene exists as a single copy gene located on chromosome 7g31 (Green et al., 1995).

Relevant linkages The human ob gene consists of three exons and two introns and spans about 18 kb, encoding a 3.5 kb cDNA (Gong et al., 1996).

Regulatory sites and corresponding transcription factors The ob gene promoter is positively regulated through a functional binding site for CCAAT/enhancer

Accession numbers Human: AAA60470 Mouse: AAA64564

Sequence See Figure 1.

Description of protein The structure reveals a four-helix bundle similar to that of the long-chain helical cytokine family (Zhang et al., 1997). The N-terminal region of leptin has been shown to be essential for both its biological and receptor-binding activities. The amino acid sequence of the C-terminal loop structure is also important for

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Raffaella Faggioni, Kenneth R Feingold and Carl Grunfeld Figure 1 Amino acid sequence for human and mouse leptin.

enhancing these actions, whereas the C-terminal disulfide bond is not needed (Imagawa et al., 1998).

CELLULAR SOURCES AND TISSUE EXPRESSION

Cellular sources that produce White adipose tissue is the major site of leptin secretion (Zhang et al., 1994). Another source of leptin is placenta. Leptin is produced by primary cultured human amnion cells. Leptin production has been detected in a cultured human choriocarcinoma cell line, BeWo cells (Masuzaki et al., 1997). Leptin protein has been shown in specific areas of rat brain and pituitary, and in a rat glioblastoma cell line (Morash et al., 1999). Cells of the rat gastric epithelium and of the glands of the gastric fundic mucosa are immunoreactive for leptin (Bado et al., 1998).

Eliciting and inhibitory stimuli, including exogenous and endogenous modulators Insulin increases leptin expression and levels (Saladin et al., 1995; Leroy et al., 1996). Falling insulin levels may be a key regulatory signal for the suppression of leptin expression with starvation (Schwartz et al., 1997). On the other hand, leptin can inhibit insulin gene expression and production and modulates insulin sensitivity (Kulkarni et al., 1997; Zhao et al., 1998; Shimomura et al., 1999). Glucocorticoids at high doses are positive regulators of leptin expression and levels (De Vos et al., 1995; Murakami et al., 1995; Slieker et al., 1996). Interestingly, a regulatory loop exists between the hypothalamus±pituitary±adrenal (HPA) axis and circulating leptin. In mice, adrenalectomy decreases basal leptin levels and corticosterone replacement therapy restores circulating leptin to physiological levels (Spinedi et al., 1998). Furthermore, leptin deficiency, as observed in ob/ob mice, results in

chronic HPA axis activation, which is reversed by leptin treatment (Ahima et al., 1998). In addition, leptin administration substantially prevents the activation of the HPA axis in response to stress or fasting (Ahima et al., 1996; Heiman et al., 1997). Negative regulators include -adrenergic agonists and cAMP (Slieker et al., 1996; Trayhurn et al., 1996). Interactions exist between leptin and cytokines. Proinflammatory cytokines increase leptin levels, whereas leptin regulates the production of several pro- and anti-inflammatory cytokines. In vivo, leptin levels are acutely increased by TNF and IL-1 (Grunfeld et al., 1996b; Sarraf et al., 1997). In vitro, leptin has been shown to modulate cytokine production by macrophages and T cells (see Leptin regulation of cytokine production). Leptin production and gene expression in BeWo cells, a human trophoblastic cell line, are increased by treatment with phorbol myristate acetate (PMA). The PMA-induced increase in leptin production is completely suppressed by H7 and staurosporine, both of which are inhibitors of protein kinase C (Yura et al., 1998).

RECEPTOR UTILIZATION The leptin receptor (OB-R) is related to class I cytokine receptors, which include gp130, the common signal transducing component for the IL-6 related family of cytokines (Baumann et al., 1996). Several alternatively spliced isoforms of OB-R have been cloned (Fei et al., 1997). The weight-regulating effects of leptin are mediated through the OB-Rb form in the hypothalamus (Vaisse et al., 1996). The OB-Rb isoform is also present in the kidney, where mediates the clearance of leptin from the circulation. The short isoform (OB-Ra) is the predominant OB-R mRNA found in most tissues and cells, including kidney, lung, liver, spleen, and macrophages (Tartaglia et al., 1995). Leptin, a relatively large protein that would ordinarily be inaccessible to the brain, is transported through the blood±brain barrier via a saturable transport system (Golden et al., 1997). OB-Ra is highly expressed in the choiroid plexus

Leptin where it might function as a transporter across the blood±brain barrier (Tartaglia et al., 1995). Leptin circulates both in bound and free form (Sinha et al., 1996). The OB-Re isoform is a soluble receptor (Li et al., 1998). The OB-Rb isoform contains a full-length cytosolic domain that includes binding motifs required for the activation of the Janus kinase (JAK)/signal transduction and activators of transcription (STAT) signaling pathways and has been shown to have signaling capabilities of IL-6 type cytokine receptors (Vaisse et al., 1996). It has also been demonstrated that leptin can activate the mitogen-activated protein kinase (MAP) signal transduction pathway in a variety of in vitro systems (Takahashi et al., 1997; Tanabe et al., 1997). Leptin induces expression of SOCS (suppressor of cytokine signaling)-3 mRNA in the hypothalamus (Bjorbaek et al., 1999). SOCS-3 is a member of a new family of cytokine-inducible inhibitors of signaling that has recently been identified. Members of the cytokine superfamily including leptin, IL-6, interferons, and LIF, induce transcription of SOCS genes in vivo and in vitro, and when expressed in cell lines, SOCS proteins inhibit signaling and biological activities of cytokines. Therefore, SOCS proteins are thought to function as inducible intracellular negative regulators of cytokine signal transduction. Accordingly, transfection data suggest that SOCS-3 is an inhibitor of leptin signaling (Bjorbaek et al., 1999).

IN VITRO ACTIVITIES

In vitro findings Leptin Regulation of Cytokine Production Leptin has been shown to potentiate LPS-stimulated production of TNF, IL-6, and IL-12 in peritoneal macrophages (Loffreda et al., 1998; Santos-Alvarez et al., 1999). In addition, leptin induces IL-1Ra production and upregulates the IL-1Ra induction by LPS in RAW cells (Faggioni et al., 1999). Leptin can enhance phagocytic function of murine peritoneal and bone marrow macrophages of both Leishmania and Candida parasilopsis (Gainsford et al., 1996; Loffreda et al., 1998). Furthermore, leptin regulates the balance of TH1/ TH2 cytokines. Leptin enhances the alloproliferative response of peripheral blood lymphocytes by provoking a strong proliferative response by both nave and memory T cells. Moreover, leptin has been shown to increase IL-2 and IFN while inhibiting IL-4 production by T cells (Lord et al., 1998).

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Proliferative and Anti-apoptotic Activities Leptin acts on murine hematopoiesis. In bone marrow cells from normal mice, leptin induces granulocyte± macrophage colony formation in a dose-dependent manner. Similar to IL-6, leptin stimulates the proliferation of murine myelocytic progenitor cells and synergizes with stem cell factor in the proliferation of primitive hematopoietic progenitors (Umemoto et al., 1997). In transfection experiments, the long form of the OB-R has been shown to be capable of signaling for cell survival and proliferation in the murine IL-3dependent bone marrow-derived Ba/F3 cell line and for the differentiation of leukemic M1 cells into macrophages (Gainsford et al., 1996). Importantly, leptin also increases the proliferation of CD34+ stem cells from human umbilical cord blood and murine fetal liver stem cells (flASK cells), indicating a role for leptin as a hematopoietic regulator (Bennett et al., 1996). Dexamethasone induces apoptosis of murine thymocytes. Leptin protects thymocytes from steroidinduced apoptosis in vitro (Howard et al., 1999). Leptin has been shown to induce cell proliferation of a wide spectrum of cell types. It can stimulate proliferation of primary cultures of murine tracheal epithelial cells and human lung squamous cell line (SQ5), as well as mouse embryonic cell line (C3H10T1/2) (Takahashi et al., 1997; Tsuchiya et al., 1999). Leptin induces proliferation of pancreatic cell line MIN6 through activation of MAP kinase (Tanabe et al., 1997). In addition, leptin protects against apoptosis of cells induced by fatty acids through maintenance of Bcl-2 expression (Shimabukuro et al., 1998). Effect on Insulin Secretion and Insulin Activities Leptin has a direct effect on insulin secretion. An inhibitory effect of mouse leptin on insulin secretion was observed in both human and rat islets (Kulkarni et al., 1997). Leptin suppresses insulin release stimulated with glucose in mouse ( TC6) and rat (RIN5HA and RINm5F) insulinoma cell lines (Kulkarni et al., 1997). In pancreatic cells (rat pancreatic islet and a cell line, HIT-T15) leptin inhibits glucose- and glucagon-like peptide 1-stimulated insulin secretion, via phosphatidylinositol (PI) 3-kinase-dependent activation of cyclic nucleotide phosphodiesterase 3B (PDE3B) and subsequent suppression of cAMP levels (Zhao et al., 1998). In human hepatic cells (HepG2) leptin causes attenuation of several insulin-induced activities, including tyrosine phosphorylation of the insulin receptor substrate 1 (IRS-1), association of the adapter molecule growth factor receptor-bound protein 2 with IRS-1, and downregulation of gluconeogenesis. In contrast,

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Raffaella Faggioni, Kenneth R Feingold and Carl Grunfeld

leptin increased the activity of IRS-1-associated phosphatidylinositol 3-kinase (Cohen et al., 1996). These findings raise the possibility that high leptin levels in obesity could directly contribute to diabetes and insulin resistance in vivo. To date there are no data demonstrating such activities of leptin in humans.

IN VIVO BIOLOGICAL ACTIVITIES OF LIGANDS IN ANIMAL MODELS

Knockout mouse phenotypes Leptin-deficient ob/ob mice exhibit a complex phenotype characterized not only by obesity, but also by multiple hormonal and metabolic disorders, including infertility and dysfunctional adrenal and thyroid axes (Coleman, 1978). Ob/ob mice have abnormal immune and inflammatory responses (Faggioni et al., 1999; Howard et al., 1999). In accordance with the proliferative activity of leptin on stem cell populations, a deficit in hematopoiesis is observed in ob/ob mice (Howard et al., 1999). They have lymphoid atrophy accompanied with alterations in the number of circulating lymphocytes and monocytes. The ability of leptin to protect against thymic atrophy probably involves a direct anti-apoptotic mechanism. Ob/ob mice exhibit defective cell-mediated immunity, as they have an impaired delayed-type hypersensitivity reaction (DTH) (Chandra, 1980). Furthermore, ob/ob mice are protected from liver damage in models of T cell-mediated hepatitis associated with reduced induction of TNF and IL-18 (Faggioni et al., 2000a). Exogenous leptin replacement restored the responsiveness of ob/ob mice to ConA and normalized their lymphocyte and monocyte populations (Faggioni et al., 2000a; Howard et al., 1999). Therefore, leptin acts as a regulator of T cell-mediated inflammation in vivo. In contrast, increased sensitivity to proinflammatory monocyte/macrophage-activating stimuli, particularly LPS and TNF, is observed in ob/ob mice (Faggioni et al., 1999; Takahashi et al., 1999). However, in addition to reduced thymic and circulating lymphocytes, a 4-fold increase in the number of circulating monocytes is present in ob/ob mice (Faggioni et al., 2000a). Therefore, it is likely that the absence of leptin will lead to reduced sensitivity to T cellactivating stimuli and enhanced responses to monocyte activators. Interestingly, it has recently been shown that prevention of lymphocyte apoptosis is associated with improved survival in a murine model of sepsis, suggesting a critical role of the lymphocyte in resolving severe infection (Hotchkiss et al., 1999).

It is therefore possible that the significant decrease in the number of lymphocytes which occurs in septic and endotoxic shock, will impact multiple facets of the immunological response and may lead to uncontrolled inflammatory response and death. The profound lymphopenia of ob/ob mice might, therefore, substantially contribute to their increased susceptibility to infection and inflammation.

Pharmacological effects Effect of Leptin on Food Intake Chronic peripheral administration of leptin to ob/ob mice has been shown to lower their body weight, percentage body fat, food intake, and serum concentrations of glucose and insulin (Halaas et al., 1995; Pelleymounter et al., 1995). In addition, metabolic rate, body temperature, and activity levels were increased by this treatment. None of these parameters was altered beyond the level observed in lean controls, suggesting that leptin normalized the metabolic status of the ob/ob mice. Lean animals injected with leptin had a smaller weight loss throughout the 28-day study and showed no changes in any of the metabolic parameters. The central route of administration of leptin is more effective than the peripheral route in reducing food intake and body weight in ob/ob and dietinduced obese mice, indicating the brain as the main target for effect of leptin on food intake (Campfield et al., 1995). The Neuroendocrine Response of Starvation Leptin levels fall during starvation disproportionally to the decrease in fat depot. Preventing the starvationinduced fall in leptin with exogenous leptin substantially blunts the changes in gonadal, adrenal, and thyroid axes in male mice, and prevents the starvation-induced delay in ovulation in female mice. In contrast, leptin repletion during this period of starvation has little or no effect on body weight, blood glucose, or ketones (Ahima et al., 1996). The Immunosuppression of Starvation Starvation suppresses immunity, particularly T lymphocyte responses, and decreases resistance to infection. Prevention of the fasting-induced fall in the level of leptin by administering exogenous recombinant leptin reverses the suppressive effects of acute starvation on cell-mediated immunity. In addition, administration of leptin to starved mice protected mice from starvation-induced thymic atrophy (Lord et al., 1998). Furthermore, administration of leptin to

Leptin starved mice markedly reversed their increased susceptibility to both LPS and TNF toxicity (Faggioni et al., 2000b). Correction of the ob Phenotype Chronic leptin treatment of leptin-deficient ob/ob reverses their neuroendocrine and metabolic abnormalities (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995). Noteworthy, leptin restores the responsiveness of ob/ob mice to T cellactivating stimuli, mainly by increasing thymic cellularity (Howard et al., 1999). The increased susceptibility to LPS- and TNF-induced lethality observed in ob/ob mice is also reversed by leptin treatment (Faggioni et al., 1999; Takahashi et al., 1999).

Interactions with cytokine network The induction of leptin during the host response to infection and inflammation is mediated by release of the cytokines IL-1 and TNF. During local and systemic inflammation caused by injection of turpentine or LPS in mice, IL-1 has been shown to play an essential role in the induction of leptin (Faggioni et al., 1998). Leptin levels are increased in mice during bacterial peritonitis, and blocking the TNF response blunts the increase (Moshyedi et al., 1998). Conversely, leptin upregulates LPS-induced phagocytosis and proinflammatory cytokine expression (TNF, IL-6, IL-12) in ex vivomacrophages from mice (Loffreda et al., 1998; Santos-Alvarez et al., 1999). Leptin actions in the brain appear to depend on IL1 . Luheshi and coworkers showed that leptin increases levels of IL-1 in the hypothalamus of normal rats. The effect of leptin on fever and food intake is abolished by IL-1 receptor antagonist (IL1Ra) and is absent in mice lacking the main IL-1 receptor (80 kDa, R1) responsible for IL-1 actions (Luheshi et al., 1999).

PATHOPHYSIOLOGICAL ROLES IN NORMAL HUMANS AND DISEASE STATES AND DIAGNOSTIC UTILITY

Role in experiments of nature and disease states Obesity Leptin deficiency as a cause of human obesity appears to be extremely rare (Montague et al., 1997;

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Ozata et al., 1999). One of two very obese consanguineous cousins who have congenital leptin deficiency due to an inactivating mutation of the leptin gene has recently been treated with leptin. Administration of leptin to one of these obese children ameliorates hyperphagia, promoted weight loss while preserving lean mass, and may have had a permissive role in the onset of puberty (Farooqi et al., 1999). More commonly, obese humans are not leptin deficient and plasma leptin levels are elevated in most overweight individuals (Maffei et al., 1995). The results of the first clinical trial show that some study participants given leptin lost more weight than controls (Heymsfield et al., 1999). The differences were statistically significant, however, only in obese subjects given the two highest leptin doses. Weight loss was small and did not result in loss of obesity. Infectious and Inflammatory Diseases Acute illness: Leptin levels are elevated in patients with sepsis (Torpy et al., 1998). Interestingly, there is a positive correlation between leptin levels and survival (Bornstein et al., 1998; Arnalich et al., 1999). In addition, in humans genetic leptin deficiency is associated with high mortality due to infections; 7 out of 11 obese members of the family with consanguineous homozygous leptin deficiency died of infection in childhood (Ozata et al., 1999). The following results have been noted in animal models:  Leptin levels are transiently induced by LPS in rodents with endotoxic shock (Grunfeld et al., 1996b).  Polymycrobic sepsis (cecal ligation and puncture, CLP): leptin levels are elevated (Moshyedi et al., 1998).  Sterile abscess: turpentine injection increases leptin expression and secretion (Faggioni et al., 1998).  Leptin deficiency is associated with increased susceptibility to endotoxic shock (Faggioni et al., 1999). Chronic illness: Leptin levels are significantly lower in patients with AIDS compared with normal subjects, related to body mass index (Grunfeld et al., 1996a). Leptin concentrations are similar in the inflammatory bowel disease and control groups (Ballinger et al., 1998).

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Raffaella Faggioni, Kenneth R Feingold and Carl Grunfeld

IN THERAPY

Preclinical ± How does it affect disease models in animals? Obesity Leptin administration is effective in normalizing the metabolic, endocrine, reproductive, and immune abnormalities of ob/ob mice (Campfield et al., 1995; Halaas et al., 1995; Pelleymounter et al., 1995). In mice, diet-induced obesity produces resistance to peripheral leptin treatment. Interestingly, those mice retained sensitivity to centrally administered leptin (Van Heek et al., 1997). Inflammation Leptin deficiency is associated with enhanced susceptibility to LPS, as assessed by increased liver injury and lethality (Yang et al., 1997; Faggioni et al., 1999). Leptin administration reverses the increased sensitivity to LPS lethality observed in ob/ob mice (Faggioni et al., 1999). Starvation Leptin levels drop dramatically with the onset of starvation. Leptin administration during the period of starvation effectively prevents the neuroendocrine response to starvation (Ahima et al., 1996). The adaptative response to starvation includes changes in gonadal, adrenal, and thyroid axes in male mice and delay in ovulation in female mice, all of which are affected by leptin treatment. In addition, leptin administration during fasting protects mice from the lymphoid atrophy and reverses the inhibitory effect of starvation on the development of a delayed-type hypersensitivity reaction (Lord et al., 1998; Howard et al., 1999). Furthermore, leptin administration markedly reversed the increased sensitivity to LPS caused by starvation (Faggioni et al., 2000b).

Toxicity Leptin can cause redness and swelling at the site of injection, severe enough to cause subjects to drop out of the study early (Heymsfield et al., 1999).

Clinical results The results of the first clinical trial show that some study participants given leptin lost more weight than controls (Heymsfield et al., 1999). The differences

were statistically significant, however, only in obese subjects given the two highest leptin doses.

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