Leptin favors imbalance of antigen-specific CD4+ T-cells associated with severity of cat allergy.

Introduction: Obesity can complicate IgE-mediated allergic diseases. In the present study, we aimed to investigate the ability of obesity-related concentrations of leptin to modulate the in vitro effector and regulatory Fel d1-specific CD4+ T-cell subsets in patients allergic to cat, considered the third most common cause of respiratory allergy in humans. Methods: For this study, plasma and peripheral blood mononuclear cells (PBMC) from 30 cat-allergic patients with mild, moderate and severe respiratory symptoms were obtained. The PBMC cultures were stimulated with Fel d1 antigen (10 µg/mL) in the presence or absence of obesity-related leptin dose (50 ηg/mL). After 6 days, the levels of cytokines and IgE in the supernatants were evaluated by multiplex and ELISA, respectively. The frequency of different non-follicular (CXCR5-) and follicular (CXCR5+) Fel d1-specific CD4+ T cell subsets was determined by flow cytometry. The plasma levels of leptin and IgE anti-cat titers were evaluated by ELISA and ImmunoCAP, respectively. Results and conclusions: Fel d1 induced both IgE production and release of cytokines related to Th2, Th9 and Th17 cell phenotypes. Feld1 was more efficient in increasing the frequency of TFHIL-21- cells positive for IL-4, IL-5 and IL-13 than TFHIL-21+ cell subsets. Leptin favored the expansion Th2-like and Th9-like cells and TFHIL-21- cells positive for IL-4, IL-5 and IL-13, but reduced the proportion of conventional (Treg/Tr-1) and follicular (TFR) regulatory CD4+ T-cell subsets expressing or not CD39 marker. Finally, many of the imbalances between Fel d1-specific CD4+ T-cells were also correlated with plasma leptin and anti-Fel d1 IgE titers. In summary, hyperleptinemia should negatively impact on the severity of cat allergies by favoring the expansion of pathogenic Fel d1-specific CD4+ T-cell phenotypes and damaging the functional status of regulatory CD4+ T-cell subsets.

Leptin favors Th17/Treg cell subsets imbalance associated with allergic asthma severity.

Background: Obesity has often been associated with severe allergic asthma (AA). Here, we analyzed the frequency of different circulating CD4+T-cell subsets from lean, overweight and obese AA patients. Methods: Mononuclear cells from peripheral blood were obtained from 60 AA patients and the frequency of different CD4+T-cell subsets and type 1 regulatory B cells (Br1) was determined by cytometry. The effect of obese-related leptin dose on cytokine production and Treg cell function in AA-derived CD4+ T cell cultures was evaluated by ELISA and 3H thymidine uptake, respectively. Leptin levels were quantified in the plasma by ELISA. According to the BMI, patients were stratified as lean, overweight and obese. Results: AA severity, mainly among obese patients, was associated with an expansion of hybrid Th2/Th17 and Th17-like cells rather than classic Th2-like cells. On the other hand, the frequencies of Th1-like, Br1 cells and regulatory CD4+ T-cell subsets were lower in patients with severe AA. While percentages of the hybrid Th2/Th17 phenotype and Th17-like cells positively correlated with leptin levels, the frequencies of regulatory CD4+ T-cell subsets and Br1 cells negatively correlated with this adipokine. Interestingly, the obesity-related leptin dose not only elevated Th2 and Th17 cytokine levels, but also directly reduced the Treg function in CD4+ T cell cultures from lean AA patients. Conclusion: In summary, our results indicated that obesity might increase AA severity by favoring the expansion of Th17-like and Th2/Th17 cells and decreasing regulatory CD4+T cell subsets, being adverse effects probably mediated by leptin overproduction.

Major depression favors the expansion of Th17-like cells and decrease the proportion of CD39+Treg cell subsets in response to myelin antigen in multiple sclerosis patients.

BACKGROUND: Mood disorders have been associated with risk of clinical relapses in multiple sclerosis (MS), a demyelinating disease mediated by myelin-specific T cells. OBJECTIVES: We aimed to investigate the impact of major depressive disorder (MDD) and cytokine profile of T-cells in relapsing remitting MS patients. METHODS: For our study, plasma and PBMC were obtained from 60 MS patients (30 with lifetime MDD) in remission phase. The PBMC cultures were stimulated with anti-CD3/anti-CD28 beads or myelin basic protein (MBP), and effector and regulatory T cell phenotypes were determined by flow cytometry. The cytokine levels, both in the plasma or in the supernatants collected from PBMC cultures, were quantified by Luminex. In some experiments, the effect of serotonin (5-HT) was investigated. RESULTS: Here, higher Th17-related cytokine levels in response to anti-CD3/anti-CD28 and MBP were quantified in the plasma and PBMC cultures of the MS/MDD group in comparison with MS patients. Further, elevated frequency of CD4+ and CD8+ T cells capable of producing IL-17, IL-22 and GM-CSF was observed in depressed patients. Interestingly, the percentage of myelin-specific IFN-γ+IL-17+ and IFN-γ+GM-CSF+ CD4+ T cells directly correlated with neurological disabilities. In contrast, the occurrence of MDD reduced the proportion of MBP-specific CD39+Tregs subsets. Notably, the severity of both neurological disorder and depressive symptoms inversely correlated with these Tregs. Finally, the addition of 5-HT downregulated the release of Th17-related cytokines in response to anti-CD3/anti-CD28 and myelin antigen. CONCLUSIONS: In summary, our findings suggested that recurrent major depression, by favoring imbalances of effector Th17 and Treg cell subsets, contributes to MS severity.

Human pregnancy levels of estrogen and progesterone contribute to humoral immunity by activating TFH /B cell axis.

Circulating TFH (cTFH ) cells express CXCR5, PD-1, and, when activated, ICOS, and release IL-21. According to the production of IFN-γ, IL-4, and IL-17 and expression of FoxP3, these cells are also classified as cTFH 1, cTFH 2, cTFH 17, and cTFR cells, respectively. This CD4+ T-cell subset is pivotal to efficient humoral immunity, and pregnancy appears to favor IgG production. Here, not only pregnancy amplified the in vivo production of anti-HBsAg IgG in HBV immunized women, but the frequency of cTFH cells was directly correlated with estradiol levels. In vitro, pregnancy-related dose of 17-ß-estradiol (E2) directly increased the percentage of different cTFH subsets. While E2 and progesterone (P4) increased the proportion of differentiated TFH cells derived from naïve CD4+ T-cells, only E2 amplified the release of IL-21 in those cell cultures. In addition, E2 and P4 increased the proportion of memory B cells and plasma cells, respectively. In SEB-activated B/TFH cell co-cultures, E2, in the presence of P4, increased the production of total IgG. Finally, among the hormones, P4 was stronger in upregulating the percentage of IL-10+ TFR cells. Collectively, our findings suggested that E2 and P4 cooperate in the humoral immune response by favoring the expansion of different cTFH and B cell subsets.

Different core-specific T cell subsets are expanded in chronic hepatitis C with advanced liver disease.

Chronic hepatitis C (CHC) is frequently related to liver fibrosis, and several studies have suggested that the immunological activity of HCV antigens contributes to hepatic damage. In the present study, among structural and non-structural HCV antigens, elevatedIL-1ß, IL-6, IL-17 levels were secreted by PBMC cultures obtained from CHC patients following stimulation with core antigen. Moreover, the percentage of core-specific IL-6+IL-17+(CD4+ and CD8+) T cells was significantly higher in patients with worsehepatic lesions, determined on the Metavir scale. When compared with healthy subjects, the percentage of circulating Treg cells was elevated in CHC patients, mainly among those with advanced liver fibrosis. Nevertheless, in this last group of patients, the proportion of CD39+ Treg subsets was very low. Finally, the percentage of senescent (CD57+ CD28-) and exhausted (PD-1+CD28+) core-specific T cells in CHC patients was also found to be a result of fibrotic hepatic status. In summary, imbalances between different core-specific T cell subsets are associated with liver fibrosis severity.

Serum leptin levels correlate negatively with the capacity of vitamin D to modulate the in vitro cytokines production by CD4+ T cells in asthmatic patients.

Both obesity and low vitamin D levels have been associated with allergic asthma (AA) severity. In the present study, severity of AA was associated with obesity but to the in vitro IgE production. In those patients, higher levels of IL-5, IL-6 and IL-17 were quantified in CD4+ T-cell cultures as compared with patients with mild and moderate AA. In addition, the lowest IL-10 levels were detected in the cell cultures from patients with a worse prognosis. Interestingly, the occurrence of AA elevates the plasma levels of leptin, and this adipokine was positively correlated with the release of IL-5, IL-6 and IL-17, but inversely correlated with IL-10 production, by CD4+ T-cells from patients. In AA-derived CD4+ T-cell cultures, 1,25(OH)2D3 was less efficient at inhibiting IL-5, IL-6 and IL-17 production, and up regulating IL-10 release, as those from healthy subjects. Interestingly, the in vitro immunomodulatory effects of vitamin D were inversely correlated with serum leptin levels. In summary, our findings suggested that obesity, probably due to the overproduction of leptin, negatively impacts AA as it favors imbalance between Th2/Th17 and regulatory phenotypes. The deleterious effects of leptin may also be due to its ability to counter-regulate the immunosuppressive effects of vitamin D.

TLR-2 and TLR-4 agonists favor expansion of CD4+ T cell subsets implicated in the severity of neuromyelitis optica spectrum disorders.

BACKGROUND: High frequency of circulating Th17 cell subsets expressing TLR2, TLR4 and TLR9 was observed in Neuromyelitis optica spectrum disorder (NMOSD) patients, a severe humoral autoimmune disease of the central nervous system. Our objective was to evaluate the direct effects of different TLR ligands on CD4+ T-cells form those patients. METHODS: CD4+ T-cell cultures from NMOSD and healthy individuals were stimulated with different TLR ligands and the cell proliferation and cytokine profile was analyzed by [3H] TdR up take and ELISA/ cytometry, respectively. The plasma levels of CD14 were determined by ELISA. RESULTS: Here, Pam3C (TLR2) and LPS (TLR4) induced significant cell proliferation and IL-6, IL-17 and IL-21 production by CD4+ T-cells from NMOSD. Additionally, while both TLR ligands were more potent in favoring the expansion of TFH-like cells, Pam3C reduced the frequency of IL-10-secreting FoxP3+and FoxP3- CD4+ T-cells. With regard to disease severity, the levels of IL-6, IL-17 and IL-21 produced by CD4+ T-cells, as well as the frequency of TFH-like cells, in response to TLR2 and TLR4 agonists were positively correlated with neurological disabilities and the occurrence of new acute relapses during follow up. Finally, circulating levels of CD14, an indirect marker of microbial translocation, were positively correlated with IL-6, IL-17 and IL-21 release by Pam3C- and LPS-activated CD4+ T-cells. CONCLUSIONS: In summary, our data suggest that microbial antigens may affect NMOSD outcomes by favoring an imbalance between Th17 and TFH-like cells and regulatory T cell subsets.

The expansion of circulating IL-6 and IL-17-secreting follicular helper T cells is associated with neurological disabilities in neuromyelitis optica spectrum disorders.

Due to their function in assisting B cells, TFH cells may be involved in the production of pathogenic IgG in neuromyelitis optica spectrum disorder (NMOSD). In the present study, the proportion of IL-6+ and IL-17+ TFH cell subsets was higher in NMOSD patients than healthy individuals. The frequency of both TFH cell subsets were directly associated with disease activity. By contrast, NMOSD patients with a higher proportion of IL-10+ TFH cell subsets showed a lower neurological disabilities score. In summary, all findings suggest that expansion of peripheral IL-6+ and IL-17+ TFH cells may be involved in the severity of NMOSD.

Serotonin decreases the production of Th1/Th17 cytokines and elevates the frequency of regulatory CD4+ T-cell subsets in multiple sclerosis patients.

Excessive levels of proinflammatory cytokines in the CNS are associated with reduced serotonin (5-HT) synthesis, a neurotransmitter with diverse immune effects. In this study, we evaluated the ability of exogenous 5-HT to modulate the T-cell behavior of patients with MS, a demyelinating autoimmune disease mediated by Th1 and Th17 cytokines. Here, 5-HT attenuated, in vitro, T-cell proliferation and Th1 and Th17 cytokines production in cell cultures from MS patients. Additionally, 5-HT reduced IFN-γ and IL-17 release by CD8+ T cells. By contrast, 5-HT increased IL-10 production by CD4+ T cells from MS patients. A more accurate analysis of these IL-10-secreting CD4+ T cells revealed that 5-HT favors the expansion of FoxP3+ CD39+ regulatory T cells (Tregs) and type 1 regulatory T cells. Notably, this neurotransmitter also elevated the frequency of Treg17 cells, a novel regulatory T-cell subset. The effect of 5-HT in upregulating CD39+ Treg and Treg17 cells was inversely correlated with the number of active brain lesions. Finally, in addition to directly reducing cytokine production by purified Th1 and Th17 cells, 5-HT enhanced in vitro Treg function. In summary, our data suggest that serotonin may play a protective role in the pathogenesis of MS.

Expansion of IL-6+ Th17-like cells expressing TLRs correlates with microbial translocation and neurological disabilities in NMOSD patients.

Different microbial antigens, by signaling through toll-like receptors (TLR), may contribute to Th17-mediated autoimmune diseases, such as neuromyelitis optica spectrum disorder (NMOSD). The objective of this study was to determine the proportion of different Th17-like cell subsets that express TLR in NMOSD patients. For this study, the frequency of different Th17 cell subsets expressing TLR subsets in healthy individuals (n=20) and NMOSD patients (n=20) was evaluated by cytometry. The peripheral levels of soluble CD14 (sCD14) and cytokines were determined by ELISA. Our results demonstrated that the proportion of peripheral CD4+ T cells expressing TLR2, 4 and 9 was significantly higher in NMOSD samples than in healthy subjects. In NMOSD, these cells are CD28+PD-1-CD57- and produce elevated levels of IL-17. Among different TLRs+ Th17-like subsets, the proportion of those that co-express IL-17 and IL-6 was significantly higher in NMOSD patients, which was positively correlated with sCD14 levels and EDSS score. By contrast, the percentage of TLRs+Treg17 cells (IL-10+IL-17+) was negatively related to sCD14 and the severity of NMOSD. In conclusion, the expansion of peripheral IL-6-producing TLR+ Th17-like cells in NMOSD patients was associated with both bacterial translocation and disease severity.

The proportion of different interleukin-17-producing T-cell subsets is associated with liver fibrosis in chronic hepatitis C.

Studies have suggested the pivotal role of T helper type 1 (Th1) -related cytokines on the outcome of hepatitis C virus (HCV) infection. Nevertheless, the role of different interleukin-17 (IL-17) -secreting T cells on chronic hepatitis C (CHC) is less clear. Here, the in vivo IL-1ß, IL-6, and IL-17 levels were positively correlated with both alanine transaminase (ALT) levels and hepatic lesions. When compared with the control group, CHC patients showed a lower proportion of IL-17-secreting (CD4+ and CD8+ ) T cells capable of simultaneously producing IL-21. Moreover, the percentage of IL-10-secreting Th17 cells was also lower in CHC patients. Notably, advanced liver lesions were observed among those patients with lower percentage levels of IL-17-producing T cells positive for IL-21, interferon-γ (IFN-γ) and IL-10. In contrast, the severity of hepatic damage was associated with peripheral single IL-17+ T cells. The percentage of IL-17+ IL-21- IFN-γ+ (CD4+ and CD8+ ) T-cell phenotypes was positively associated with plasma CD14 levels. Finally, elevated levels of circulating CD14 were detected among CHC patients with extensive liver damage. In summary, although preliminary, our results suggest that a balance between different IL-17-producing T cells, associated with peripheral levels of CD14, may be a progress marker for liver disease in chronically HCV-infected patients.