Claudio Procaccini

Leptin as an immunomodulator

Leptin is an adipocyte-derived hormone/cytokine that links nutritional status with neuroendocrine and immune functions. In humans, leptin influences energy homeostasis and regulates neuroendocrine function primarily in states of energy deficiency. Initially described as an antiobesity hormone, leptin has subsequently been shown also to influence basal metabolism, hematopoiesis, thermogenesis, reproduction, and angiogenesis. As a cytokine, leptin can affect thymic homeostasis and the secretion of acute-phase reactants such as interleukin-1 (IL-1) and tumor-necrosis factor-alpha (TNF-α). Leptin links nutritional status and proinflammatory T helper 1 (Th1) immune responses and the decrease in leptin plasma concentration during food deprivation leads to impaired immune function. Similar to other pro-inflammatory cytokines, leptin promotes Th1-cell differentiation and can modulate the onset and progression of autoimmune responses in several animal models of disease. Here, we review the advances and controversy for a role of leptin in the pathophysiology of immune responses and discuss novel possible therapeutic implications for leptin modulators.

Regulatory T cell proliferative potential is impaired in human autoimmune disease

Human CD4+CD25highCD127−FoxP3+ regulatory T (Treg) cells suppress immune responses in vitro and in vivo. Reduced suppressive function and/or number of peripheral Treg cells has been previously reported in autoimmune disorders. Treg cells represent the most actively replicating compartment within the CD4+ cells in vivo, but they are hyporesponsive to classical T cell receptor (TCR) stimulation in vitro, a condition that is secondary to their overactive metabolic state. Here we report that proliferation of Treg cells after TCR stimulation is impaired in subjects with relapsing-remitting multiple sclerosis (RRMS) because of altered interleukin-2 (IL-2) secretion and IL-2 receptor (IL-2R)–signal transducer and activator of transcription 5 (STAT5) signaling. This is associated with decreased expression of the forkhead box P3 (FoxP3) 44- and 47-kDa splicing forms, overactivation of S6 ribosomal protein (a downstream target of the mammalian target of rapamycin, mTOR) and altered activity of the cyclin-dependent kinase inhibitor p27 (p27kip1) and extracellular signal–related kinases 1 and 2 (ERK1/2). The impaired capacity of Treg cells to proliferate in RRMS correlates with the clinical state of the subject, where increasing disease severity is associated with a decline in Treg cell expansion. These results suggest a previously unrecognized mechanism that may account for the progressive loss of Treg cells in autoimmune disease.

Leptin neutralization interferes with pathogenic T cell autoreactivity in autoimmune encephalomyelitis.

Recent evidence has indicated that leptin, an adipocyte-secreted hormone belonging to the helical cytokine family, significantly influences immune and autoimmune responses. We investigate here the mechanisms by which in vivo abrogation of leptin effects protects SJL/J mice from proteolipid protein peptide PLP(139-151)-induced EAE, an animal model of MS. Blockade of leptin with anti-leptin Abs or with a soluble mouse leptin receptor chimera (ObR:Fc), either before or after onset of EAE, improved clinical score, slowed disease progression, reduced disease relapses, inhibited PLP(139-151)-specific T cell proliferation, and switched cytokine secretion toward a Th2/regulatory profile. This was also confirmed by induction of forkhead box p3 (Foxp3) expression in CD4 T cells in leptin-neutralized mice. Importantly, anti-leptin treatment induced a failure to downmodulate the cyclin-dependent kinase inhibitor p27 (p27) in autoreactive CD4 T cells. These effects were associated with increased tyrosine phosphorylation of both ERK1/2 and STAT6. Taken together, our data provide what we believe is a new molecular basis for leptin antagonism in EAE and envision novel strategies of leptin-based molecular targeting in the disease.

Regulatory T cells in obesity: the leptin connection

Studies to understand the pathogenesis of obesity have revealed mediators that are responsible for the control of food intake and metabolism at the hypothalamic level. However, molecular insight explaining the link between obesity and low-degree chronic inflammation remains elusive. The adipocyte-derived hormone leptin, and thereby the nutritional status, could control immune self-tolerance by affecting regulatory T (Treg) cell responsiveness and function. Furthermore, resident Treg cells, which are capable of modulating metabolism and glucose homeostasis, are abundant in adipose tissue. Here, we provide an update on recent findings relating Treg cells to obesity and discuss how the intricate network of interactions among leptin, Treg cells and adipose tissue might provide new strategies for therapeutic interventions.

Glycolysis controls the induction of human regulatory T cells by modulating the expression of FOXP3 exon 2 splicing variants

Human regulatory T cells (Treg cells) that develop from conventional T cells (Tconv cells) following suboptimal stimulation via the T cell antigen receptor (TCR) (induced Treg cells (iTreg cells)) express the transcription factor Foxp3, are suppressive, and display an active proliferative and metabolic state. Here we found that the induction and suppressive function of iTreg cells tightly depended on glycolysis, which controlled Foxp3 splicing variants containing exon 2 (Foxp3-E2) through the glycolytic enzyme enolase-1. The Foxp3-E2–related suppressive activity of iTreg cells was altered in human autoimmune diseases, including multiple sclerosis and type 1 diabetes, and was associated with impaired glycolysis and signaling via interleukin 2. This link between glycolysis and Foxp3-E2 variants via enolase-1 shows a previously unknown mechanism for controlling the induction and function of Treg cells in health and in autoimmunity.

Cellular and molecular crosstalk between leptin receptor and estrogen receptor-α in breast cancer: molecular basis for a novel therapeutic setting

Obesity is associated with an increased risk of breast cancer. A number of adipocytokines are increased in obesity causing low-level chronic inflammation associated with an increased risk of tumors. The adipocytokine leptin shows profound anti-obesity and pro-inflammatory activities. We have hypothesized that in common obesity, high circulating leptin levels might contribute to an increased risk of breast cancer by affecting mammary cell proliferation and survival. Leptin exerts its activity not only through leptin receptor (LepR), but also through crosstalk with other signaling systems implicated in tumorigenesis. In this study, we focused our attention on the relationship between the leptin/LepR axis and the estrogen receptor-alpha (ERalpha). To this aim, we utilized two human breast cancer cell lines, one ERalpha-positive cell line (MCF 7) and the other ERalpha-negative cell line (MDA-MB 231). We observed that the two cell lines had a different sensitivity to recombinant leptin (rleptin): on MCF 7 cells, rleptin induced a strong phosphorylation of the signal transducer and activator of transcription (STAT) 3 and of the extracellular related kinase 1/2 pathways with an increased cell viability and proliferation associated with an increased expression of ERalpha receptor. This response was not present in the MDA-MB 231 cells. The effects induced by leptin were lost when LepR was neutralized using either a monoclonal inhibitory antibody to LepR or LepR gene-silencing siRNA. These data suggest that there is a bidirectional communication between LepR and ERalpha, and that neutralization and/or inactivation of LepR inhibits proliferation and viability of human breast cancer cell lines. This evidence was confirmed by ex vivo studies, in which we analyzed 33 patients with breast cancer at different stages of disease, and observed that there was a statistically significant correlation between the expression of LepR and ERalpha. In conclusion, this study suggests a crosstalk between LepR and ERalpha, and could envisage novel therapeutic settings aimed at targeting the LepR in breast cancers.

Leptin-Induced mTOR Activation Defines a Specific Molecular and Transcriptional Signature Controlling CD4+ Effector T Cell Responses

The sensing by T cells of metabolic and energetic changes in the microenvironment can determine the differentiation, maturation, and activation of these cells. Although it is known that mammalian target of rapamycin (mTOR) gauges nutritonal and energetic signals in the extracellular milieu, it is not known how mTOR and metabolism influence CD4+CD25−FOXP3− effector T cell (Teff) responses. In this article, we show that leptin-induced activation of mTOR, which, in turn, controls leptin production and signaling, causes a defined cellular, biochemical, and transcriptional signature that determine the outcome of Teff responses, both in vitro and in vivo. The blockade of leptin/leptin receptor signaling, induced by genetic means or by starvation, leads to impaired mTOR activity that inhibits the proliferation of Teffs in vivo. Notably, the transcriptional signature of Teffs in the presence of leptin blockade appears similar to that observed in rapamycin-treated Teffs. These results identify a novel link between nutritional status and Teff responses through the leptin–mTOR axis and define a potential target for Teff modulation in normal and pathologic conditions.

Leptin Modulates the Survival of Autoreactive CD4+ T Cells through the Nutrient/Energy-Sensing Mammalian Target of Rapamycin Signaling Pathway

Chronic inflammation can associate with autoreactive immune responses, including CD4+ T cell responses to self-Ags. In this paper, we show that the adipocyte-derived proinflammatory hormone leptin can affect the survival and proliferation of autoreactive CD4+ T cells in experimental autoimmune encephalomyelitis, an animal model of human multiple sclerosis. We found that myelin olygodendrocyte glycoprotein peptide 35–55 (MOG35–55)-specific CD4+ T cells from C57BL/6J wild-type mice could not transfer experimental autoimmune encephalomyelitis into leptin-deficient ob/ob mice. Such a finding was associated with a reduced proliferation of the transferred MOG35–55-reactive CD4+ T cells, which had a reduced degradation of the cyclin-dependent kinase inhibitor p27kip1 and ERK1/2 phosphorylation. The transferred cells displayed reduced Th1/Th17 responses and reduced delayed-type hypersensitivity. Moreover, MOG35–55-reactive CD4+ T cells in ob/ob mice underwent apoptosis that associated with a downmodulation of Bcl-2. Similar results were observed in transgenic AND-TCR- mice carrying a TCR specific for the pigeon cytochrome c 88–104 peptide. These molecular events reveal a reduced activity of the nutrient/energy-sensing AKT/mammalian target of rapamycin pathway, which can be restored in vivo by exogenous leptin replacement. These results may help to explain a link between chronic inflammation and autoimmune T cell reactivity.

The intricate interface between immune and metabolic regulation: a role for leptin in the pathogenesis of multiple sclerosis?

Over the last few years, a series of molecules known to play a function in metabolism has also been shown to play an important role in the regulation of the immune response. In this context, the adipocyte‐derived hormone leptin has been shown to regulate the immune response in normal as well as in pathological conditions. More specifically, it has been shown that conditions of reduced leptin production (i.e., genetic leptin deficiency, anorexia nervosa, malnutrition) are associated with increased susceptibility to infections. Conversely, immune‐mediated disorders such as autoimmune disorders are associated with increased secretion of leptin and production of proinflammatory, pathogenic cytokines. Leptin could represent the “missing link” among immune response, metabolic function, and nutritional status. Indeed, more recently, leptin‐deficient mice have been shown to be resistant to a series of experimentally induced autoimmune disorders including experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. Normal wild‐type mice show increased secretion of leptin in serum upon EAE induction, and brain inflammatory infiltrates stain positive for leptin. Finally, leptin neutralization with leptin antagonists improves the EAE course by profoundly altering intracellular signaling of myelin‐reactive T cells and increasing the number of regulatory forkhead/winged helix transcription factor 3+CD4+ T cells. These data suggest that leptin can be considered as a link among immune tolerance, metabolic state, and autoimmunity and that strategies aimed at interfering with the leptin axis could represent innovative, therapeutic tools for autoimmune disorders.

Leptin in autoimmune diseases

The past twenty years of research on leptin has provided crucial information on the link between metabolic state and immune system function. Adipocytes influence not only the endocrine system but also the immune response, through several cytokine-like mediators known as adipokines, which include leptin. Initially described as an antiobesity hormone, leptin has subsequently been shown also to influence hematopoiesis, thermogenesis, reproduction, angiogenesis, and more importantly immune homeostasis. As a cytokine, leptin can affect thymic homeostasis and the secretion of acute-phase reactants such as interleukin-1 (IL-1) and tumor-necrosis factor-alpha (TNF-α). Leptin links nutritional status and proinflammatory T helper 1 (Th1) immune responses and the decrease in leptin plasma concentration during food deprivation leads to impaired immune function. Conversely, elevated circulating leptin levels in obesity appear to contribute to the low-grade inflammatory background which makes obese individuals more susceptible to increased risk of developing cardiovascular diseases, diabetes, or degenerative disease including autoimmunity and cancer. In this review, we provide an overview of recent advances on the role of leptin in the pathogenesis of several autoimmune disorders that may be of particular relevance in the modulation of the autoimmune attack through metabolic-based therapeutic approaches.