Different immunosuppressive mechanisms in multi‐drug‐resistant tuberculosis and non‐tuberculous mycobacteria patients

Previous studies have demonstrated that cells from both multi-drug-resistant tuberculosis (MDR-TB) and non-tuberculous mycobacteria (NTM) patients respond poorly to mycobacterial antigens in vitro. In the present study, we compared the in vitro response of cells isolated from sensitive TB (NR-TB)-, MDR-TB- and NTM-infected patients. Analysis of T cell phenotype ex vivo revealed that both MDR-TB and NTM patients present an increased percentage of CD4⁺CD25^+- forkhead box protein 3 (FoxP3)⁺ and CD4⁺CD25⁺CD127⁻ regulatory T (T_reg) cells when compared to NR-TB. Increased numbers of T_reg cells and interleukin (IL)-10 serum levels were detected in MDR-TB, whereas elevated serum transforming growth factor (TGF)-β was found in the NTM group. Cells of MDR-TB patients stimulated with early secretory antigenic target (ESAT)-6, but not purified protein derivative (PPD), showed a lower frequency of CD4⁺/interferon (IFN)-γ⁺ T cells and enhanced CD4⁺CD25⁺FoxP3⁺, CD4⁺CD25⁺CD127⁻ and CD4⁺CD25⁺IL-10⁺ T cell population. In addition, increased IL-10 secretion was observed in cultured MDR-TB cells following ESAT-6 stimulation, but not in NR-TB or NTM patients. In vitro blockade of IL-10 or IL-10Rα decreased the CD4⁺CD25⁺FoxP3⁺ frequencies induced by ESAT-6 in MDR-TB, suggesting a role of IL-10 on impaired IFN-γ responses seen in MDR-TB. Depletion of CD4⁺CD25⁺ T lymphocytes restored the capacity of MDR-TB T cells to respond to ESAT-6 in vitro, which suggests a potential role for T_reg/T regulatory 1 cells in the pathogenesis of MDR-TB. Together, our results indicate that although the similarities in chronicity, NTM- and MDR-TB-impaired antigenic responses involve different mechanisms.     

The calcineurin inhibitor tacrolimus allows the induction of functional CD4+CD25+ regulatory T cells by rabbit anti-thymocyte globulins

Rabbit anti-thymocyte globulins (rATG) induce CD4⁺CD25⁺forkhead box P3 (FoxP3⁺) regulatory T cells that control alloreactivity. In the present study, we investigated whether rATG convert T cells into functional CD4⁺CD25⁺FoxP3⁺CD127^−/low regulatory T cells in the presence of drugs that may hamper their induction and function, i.e. calcineurin inhibitors. CD25^neg T cells were stimulated with rATG or control rabbit immunoglobulin G (rIgG) in the absence and presence of tacrolimus for 24 h. Flow cytometry was performed for CD4, CD25, FoxP3 and CD127 and the function of CD25⁺ T cells was examined in suppression assays. MRNA expression profiles were composed to study the underlying mechanisms. After stimulation, the percentage CD4⁺CD25⁺FoxP3⁺CD127^−/low increased (from 2% to 30%, mean, P < 0·01) and was higher in the rATG samples than in control rIgG samples (2%, P < 0·01). Interestingly, FoxP3⁺T cells were also induced when tacrolimus was present in the rATG cultures. Blockade of the interleukin (IL)-2 pathway did not affect the frequency of rATG-induced FoxP3⁺ T cells. The rATG tacrolimus-induced CD25⁺ T cells inhibited proliferative responses of alloantigen-stimulated effector T cells as vigorously as rATG-induced and natural CD4⁺CD25⁺FoxP3⁺CD127^−/low T cells (67% ± 18% versus 69% ± 16% versus 45% ± 20%, mean ± standard error of the mean, respectively). At the mRNA-expression level, rATG-induced CD25⁺ T cells abundantly expressed IL-10, IL-27, interferon (IFN)-γ, perforin and granzyme B in contrast to natural CD25⁺ T cells (all P = 0·03), while FoxP3 was expressed at a lower level (P = 0·03). These mRNA data were confirmed in regulatory T cells from kidney transplant patients. Our findings demonstrate that tacrolimus does not negatively affect the induction, phenotype and function of CD4⁺CD25⁺ T cells, suggesting that rATG may induce regulatory T cells in patients who receive tacrolimus maintenance therapy.     

Increased serum IL-10 in lupus patients promotes apoptosis of T cell subsets via the caspase 8 pathway initiated by Fas signaling

We sought to investigate the expression of Fas and FasL on T cell surface and caspase 8 involvement in T cell apoptosis promoted by serum IL-10 in systemic lupus erythematosus(SLE) patients.Cells and sera were obtained from 35 SLE patients.Apoptosis of T cells in patients with SLE was increased and associated with the SLE disease activity index(SLEDAI).Elevated expression of Fas and FasL on T cell surface contributed to increased apoptosis of T cells.Increased IL-10 in the sera of SLE patients was capable of inducing Fas and FasL expression on CD4~+T cell surface,promoting apoptosis of this cell subset.Decreased IL-10 serum levels and low expression of Fas were found in 5 patients of the first follow-up group after 2-month treatment.In another group with one-year treatment,the SLEDAI declined to inactive scores.Serum IL-10 was decreased significantly,and expression of Fas and FasL on T cells was also reduced.Declined apoptosis was predominant only in CD4~+T cell subset.When sera with high level of IL-10 were used to culture PBMCs from healthy controls,activated caspase 8 was elevated in CD3~+T,CD4~+T and CD8~+T cells.The study showed that serum IL-10 induced apoptosis of T cell subsets via the caspase8 pathway initiated by Fas signaling.Increased apoptosis of T cells contributes to autoantigen burden,which is pathogenic in the development of SLE.     

Imbalance of Different Types of CD4+Foxp3+ T cells in renal transplant recipients

Aims: To determine the number of CD4⁺CD25^-Foxp3⁺, CD4⁺CD25⁺Foxp3⁺ and CD4⁺CXCR5⁺Foxp3⁺ T cells in renal transplant recipients that are transplanted stable (TS), or experiencing accelerated rejection (ALR), or acute rejection (AR).

Methods: Renal transplantation was conducted in 28 patients with end-stage renal failure (ESRF). The number of peripheral CD4⁺CD25^-Foxp3⁺, CD4⁺CD25⁺Foxp3⁺, or CD4⁺CXCR5⁺Foxp3⁺ T cells and the serum levels of interleukin-10 (IL-10) were measured in pre- and post-transplant patients and these results were compared to 10 healthy controls (HC). Correlation between CD4⁺CD25⁺Foxp3⁺ and estimated glomerular filtration rate (eGFR), CD4⁺CD25^-Foxp3⁺ and serum creatinine (Cr) levels, or Cr and IL-10 levels in TS patients was also determined.

Results: The number of CD4⁺CD25^-Foxp3⁺ T cells was significantly increased in patients with ESRF, as compared to HC. Stratification analysis demonstrated that TS patients contained greater numbers of CD4⁺CD25⁺Foxp3⁺ and CD4⁺CXCR5⁺Foxp3⁺ T cells, higher levels of serum IL-10, and fewer numbers of CD4⁺CD25^-Foxp3⁺ T cells than ESRF patients. In contrast, ALR and AR patients contained fewer numbers of CD4⁺CD25⁺Foxp3⁺ and CD4⁺CXCR5⁺Foxp3⁺ T cells, greater numbers of CD4⁺CD25^-Foxp3⁺ T cells, and lower levels of serum IL-10 than ESRF patients. In TS patients, the numbers of CD4⁺CD25⁺Foxp3⁺ and CD4⁺CD25^-Foxp3⁺ T cells were positively correlated with eGFR and serum Cr levels, respectively.

Conclusion: An imbalance of different types of CD4⁺Foxp3⁺ T cells might be involved in renal transplant rejection.     

CD4(+) CD25(-) FoxP3(+) regulatory cells are the predominant responding regulatory T cells after human rotavirus infection or vaccination in gnotobiotic pigs

The distribution and dynamic changes of CD4⁺ CD25⁺ FoxP3⁺ and CD4⁺ CD25⁻ FoxP3⁺ regulatory T (Treg) cells induced by human rotavirus (HRV) infection and vaccination were examined in neonatal gnotobiotic pigs infected with virulent HRV (VirHRV) or vaccinated with attenuated HRV (AttHRV). Subsets of gnotobiotic pigs in the AttHRV and control groups were challenged with VirHRV at post-inoculation day (PID) 28. We demonstrated that VirHRV infection or AttHRV vaccination reduced frequencies and numbers of tissue-residing Treg cells, and decreased the frequencies of interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) producing CD4⁺ CD25⁻ Treg cells in ileum, spleen and blood at PID 28. The frequencies of IL-10 and TGF-β producing CD4⁺ CD25⁻ Treg cells in all sites at PID 28 were significantly inversely correlated with the protection rate against VirHRV-caused diarrhoea (r = −1, P < 0·0001). Hence, higher frequencies of functional CD4⁺ CD25⁻ Treg cells can be an indicator for poorer protective immunity against rotavirus. Our results highlighted the importance of CD4⁺ CD25⁻ Treg cells over CD4⁺ CD25⁺ Treg cells in rotavirus infection and immunity. AttHRV vaccination (induction of immune effector responses) reduced the expansion of CD4⁺ CD25⁻ Treg cells in ileum seen in the challenged naive pigs during the acute phase of VirHRV infection and preserved normal levels of intestinal TGF-β producing Treg cells post-challenge. The reduced suppressive effect of Treg cells in AttHRV-vaccinated pigs would unleash effector/memory T-cell activation upon challenge. Preserving TGF-β producing CD4⁺ CD25⁻ Treg cells is important in maintaining homeostasis. Based on our findings, a model is proposed to depict the dynamic equilibrium course of Treg and effector T-cell responses after primary rotavirus infection/vaccination and challenge.     

Increase in tumour‐infiltrating lymphocytes with regulatory T cell immunophenotypes and reduced ζ‐chain expression in nasopharyngeal carcinoma patients

The pathological significance of the mechanisms of tumour immune-evasion and/or immunosuppression, such as loss of T cell signalling and increase in regulatory T cells (T_regs), has not been well established in the nasopharyngeal carcinoma (NPC) microenvironment. To evaluate the T_reg immunophenotypes in tumour-infiltrating lymphocytes (TILs), we performed a double-enzymatic immunostaining for detection of forkhead box P3 (FoxP3) and other markers including CD4, CD8, and CD25 on 64 NPC and 36 non-malignant nasopharyngeal (NP) paraffin-embedded tissues. Expression of CD3ζ and CD3ε was also determined. The prevalence of CD4⁺FoxP3⁺ cells in CD4⁺ T cells and the ratio of FoxP3⁺/CD8⁺ were increased significantly in NPC compared with those in NP tissues (P < 0·001 and P = 0·025 respectively). Moreover, the ratio of FoxP3⁺/CD25⁺FoxP3⁻ in NPC was significantly lower than that in NP tissues (P = 0·005), suggesting an imbalance favouring activated phenotype of T cells in NPC. A significant negative correlation between the abundance of FoxP3⁺ and CD25⁺FoxP3⁻ cells (P < 0·001) was also identified. When histological types of NPC were considered, a lower ratio of FoxP3⁺/CD25⁺FoxP3⁻ was found in non-keratinizing and undifferentiated carcinomas. Increased CD4⁺FoxP3⁺/CD4⁺ proportion and FoxP3⁺/CD8⁺ ratio were associated with keratinizing squamous cell carcinoma. A reduced expression of CD3ζ in TILs was found in 20·6% of the NPC tissues but none of the NP tissues. These data provide evidence for the imbalances of T_reg and effector T cell phenotypes and down-regulation of signal-transducing molecules in TILs, supporting their role in suppression of immune response and immune evasion of NPC.     

Thymus transplantation restores the repertoires of forkhead box protein 3 (FoxP3)+ and FoxP3− T cells in complete DiGeorge anomaly

The development of T cells with a regulatory phenotype after thymus transplantation has not been examined previously in complete DiGeorge anomaly (cDGA). Seven athymic infants with cDGA and non-maternal pretransplantation T cell clones were assessed. Pretransplantation forkhead box protein 3 (Foxp3)⁺ T cells were detected in five of the subjects. Two subjects were studied in greater depth. T cell receptor variable β chain (TCR-Vβ) expression was assessed by flow cytometry. In both subjects, pretransplantation FoxP3⁺ and total CD4⁺ T cells showed restricted TCR-Vβ expression. The development of naive T cells and diverse CD4⁺ TCR-Vβ repertoires following thymic transplantation indicated successful thymopoiesis from the thymic tissue grafts. Infants with atypical cDGA develop rashes and autoimmune phenomena before transplantation, requiring treatment with immunosuppression, which was discontinued successfully subsequent to the observed thymopoiesis. Post-transplantation, diverse TCR-Vβ family expression was also observed in FoxP3⁺ CD4⁺ T cells. Interestingly, the percentages of each of the TCR-Vβ families expressed on FoxP3⁺ and total CD4⁺ T cells differed significantly between these T lymphocyte subpopulations before transplantation. By 16 months post-transplantation, however, the percentages of expression of each TCR-Vβ family became significantly similar between FoxP3⁺ and total CD4⁺ T cells. Sequencing of TCRBV DNA confirmed the presence of clonally amplified pretransplantation FoxP3⁺ and FoxP3⁻ T cells. After thymus transplantation, increased polyclonality was observed for both FoxP3⁺ and FoxP3⁻ cells, and pretransplantation FoxP3⁺ and FoxP3⁻ clonotypes essentially disappeared. Thus, post-transplantation thymic function was associated with the development of a diverse repertoire of FoxP3⁺ T cells in cDGA, corresponding with immunological and clinical recovery.     

Critical stoichiometric ratio of CD4+ CD25+ FoxP3+ regulatory T cells and CD4+ CD25− responder T cells influence immunosuppression in patients with B‐cell acute lymphoblastic leukaemia

Regulatory T (Treg) cells act to suppress activation of the immune system and thereby maintain immunological homeostasis and tolerance to self-antigens. The frequency and suppressing activity of Treg cells in general are high in different malignancies. We wanted to identify the role and regulation of CD4⁺ CD25⁺ FoxP3⁺ Treg cells in B-cell acute lymphoblastic leukaemia (B-ALL). We have included patients at diagnosis (n = 54), patients in clinical remission (n = 32) and normal healthy individuals (n = 35). These diagnosed patients demonstrated a lower number of CD4⁺ CD25⁺ cells co-expressing a higher level of FoxP3, interleukin-10, transforming growth factor-β and CD152/CTLA-4 than the normal population. Treg cells from patients showed a higher suppressive capability on CD4⁺ CD25⁻ responder T (Tresp) cells than normal. The frequency and immunosuppressive potential of CD4⁺ CD25⁺ FoxP3⁺ Treg cells became high with the progression of malignancy in B-ALL. Relative distribution of Tresp and Treg cells was only ˜5 : 1 in B-ALL but ˜35 : 1 in normal healthy individuals, further confirming the elevated immunosuppression in patients. A co-culture study at these definite ex vivo ratios, indicated that Treg cells from B-ALL patients exhibited higher immunosuppression than Treg cells from normal healthy individuals. After chemotherapy using the MCP841 protocol, the frequency of CD4⁺ CD25⁺ cells was gradually enhanced with the reduction of FoxP3, interleukin-10 positivity corresponded with disease presentation, indicating reduced immunosuppression. Taken together, our study indicated that the CD4⁺ CD25⁺ FoxP3⁺ Treg cells played an important role in immunosuppression, resulting in a positive disease-correlation in these patients. To the best of our knowledge, this is the first detailed report on the frequency, regulation and functionality of Treg cells in B-ALL.     

Ribavirin modulates the conversion of human CD4+ CD25− T cell to CD4+ CD25+ FOXP3+ T cell via suppressing interleukin‐10‐producing regulatory T cell

Because regulatory T (Treg) cells play an important role in modulating the immune system response against both endogenous and exogenous antigens, their control is critical to establish immunotherapy against autoimmune disorders, chronic viral infections and tumours. Ribavirin (RBV), an antiviral reagent used with interferon, is known to polarize the T helper (Th) 1/2 cell balance toward Th1 cells. Although the immunoregulatory mechanisms of RBV are not fully understood, it has been expected that RBV would affect T reg cells to modulate the Th1/2 cell balance. To confirm this hypothesis, we investigated whether RBV modulates the inhibitory activity of human peripheral CD4⁺ CD25⁺ CD127⁻ T cells in vitro. CD4⁺ CD25⁺ CD127⁻ T cells pre-incubated with RBV lose their ability to inhibit the proliferation of CD4⁺ CD25⁻ T cells. Expression of Forkhead box P3 (FOXP3) in CD4⁺ CD25⁻ T cells was down-modulated when they were incubated with CD4⁺ CD25⁺ CD127⁻ T cells pre-incubated with RBV without down-modulating CD45RO on their surface. In addition, transwell assays and cytokine-neutralizing assays revealed that this effect depended mainly on the inhibition of interleukin-10 (IL-10) produced from CD4⁺ CD25⁺ CD127⁻ T cells. These results indicated that RBV might inhibit the conversion of CD4⁺ CD25⁻ FOXP3⁻ naive T cells into CD4⁺ CD25⁺ FOXP3⁺ adaptive Treg cells by down-modulating the IL-10-producing Treg 1 cells to prevent these effector T cells from entering anergy and to maintain Th1 cell activity. Taken together, our findings suggest that RBV would be useful for both elimination of long-term viral infections such as hepatitis C virus infection and for up-regulation of tumour-specific cellular immune responses to prevent carcinogenesis, especially hepatocellular carcinoma.     

The severity of systemic lupus erythematosus negatively correlates with the increasing number of CD4+CD25(high)FoxP3+ regulatory T cells during repeated plasmapheresis treatments of patients

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by increased pathologic autoantibody production. A decrease in the number of CD4+CD25(high)FoxP3+ regulatory T cells can play a key role in the loss of tolerance to self antigens. Our aim was to determine the absolute number of peripheral CD4+CD25(high)FoxP3+ T cells in 44 patients with SLE, furthermore, to measure the changes in the number of CD+CD25(high)FoxP3+ T cells in 5 patients with severe SLE treated with repeated plasmapheresis for 4-6 days in comparison to the changes in the activity of disease (SLEDAI). Percent of CD4+CD25(high)FoxP3+ T cells were measured by flow cytometry. The absolute number of peripheral CD4+CD25(high)FoxP3+ T cells was significantly decreased in the 44 patients with SLE compared to the healthy controls n = 32 (0.012 +/- 0.006 vs. 0.038 +/- 0.017 G/L, p < 0.05). In the 5 patients with severe SLE the repeated plasmapheresis treatments increased the peripheral number of CD4+CD25(high)FoxP3+ T cells. As the number of CD4+CD25(high)FoxP3+ T cells increased during the treatment, the activity of disease (the value of SLE activity index) decreased. In the peripheral blood of SLE patients not only the ratio was decreased (as it was published earlier) but also the absolute number of these regulatory T cells. The repeated plasmapheresis treatments of SLE patients induced a significant increase in the number of peripheral CD4+CD25(high)FoxP3+ T cells in parallel to the decrease in the values of SLEDAI (the activity of disease). This phenomenon is, among others, possibly due to the elimination of interpheron-alpha and lymphocytotoxic antibodies during plasmapheresis.     

Cyclosporin but not everolimus inhibits chemokine receptor expression on CD4+ T cell subsets circulating in the peripheral blood of renal transplant recipients

The peripheral chemokine receptors chemokine receptor 3 (CXCR3) and CC chemokine receptor 5 (CCR5) have been reported to be associated with allograft rejection. The impact of the expression of immunosuppressive drugs on peripherally circulating CD4⁺ T cell subsets after renal transplantion is unknown. Expression of CXCR3 and CCR5 was investigated by flow cytometry in 20 renal allograft recipients participating in a prospective, randomized trial ({"type":"clinical-trial","attrs":{"text":"NCT00514514","term_id":"NCT00514514"}}NCT00514514). Initial immunosuppression consisted of basiliximab, cyclosporin A (CsA), mycophenolate sodium and corticosteroids. After 3 months, patients were treated either with CsA, mycophenolate sodium (MPA) plus corticosteroids (n = 6), CsA and everolimus plus corticosteroids (n = 8) or CsA-free (CsA_free) receiving everolimus, MPA and corticosteroids (n = 6). After initial reduction of CD4⁺forkhead box protein 3 (FoxP3)⁺ and CD4⁺CD25^hiFoxP3⁺ regulatory T cells (T_regs) (P < 0·05; P < 0·01), 3-month post-transplant percentages of T_regs were reconstituted in CsA_free and CsA_lo arms compared to CsA_reg 12 months post transplant. Expression of CCR5 and CXCR3 on CD4⁺FoxP3⁺ and CD4⁺FoxP3^- T cells 12 months post transplant was increased in CsA_freeversus CsA_reg. Increase in CCR5⁺CXCR3⁺ co-expressing CD4⁺FoxP3^- cells between 3 and 12 months correlated negatively with the glomerular filtration rate (GFR) slope/year [modification of diet in renal disease (MDRD); r = −0·59, P < 0·01]. CsA, but not everolimus, inhibits both T_reg development and expression of CXCR3 and CCR5 on CD4⁺ T cell subsets. Increase in CCR5⁺CXCR3⁺ co-expressing CD4⁺FoxP3^- T cells is associated with early loss in allograft function.     

Reduction of forkhead box P3 levels in CD4+CD25high T cells in patients with new-onset systemic lupus erythematosus

The aim of this study was to quantify and evaluate the forkhead box P3 (FoxP3) expression regulatory T cells in new-onset systemic lupus erythematosus (SLE) patients before and after treatment. Forty-four newly diagnosed and untreated SLE patients, including 24 with active disease (SLEDAI > or = 10) and 20 with inactive disease (SLEDAI < 5), were enrolled in this study. Twenty-one age- and sex-matched healthy volunteers were also included as controls. Peripheral blood samples were collected and mononuclear cells isolated. The expression of CD25 and FoxP3 in CD4(+) T cells were analysed with flow cytometry. CD4(+)CD25(+) (3.95-13.04%) and CD4(+)CD25(high) (0.04-1.34%) T cells in peripheral blood in untreated patients with new-onset active lupus were significantly lower than that in the patients with inactive lupus (7.27-24.48%, P < 0.05 and 0.14-3.07% P < 0.01 respectively) and that in healthy controls (5.84-14.84%, P < 0.05). Interestingly, the decrease in CD4(+)CD25(high) T cells was restored significantly in patients with active lupus after corticosteroid treatment. There was, however, a significantly higher percentage of CD4(+)FoxP3(+) T cells in patients with active (5.30-23.00%) and inactive (7.46-17.38%) new-onset lupus patients compared with healthy control subjects (2.51-12.94%) (P < 0.01). Intriguingly, CD25 expression in CD4(+)FoxP3(+) T cells in patients with active lupus (25.24-62.47%) was significantly lower than that in those patients with inactive lupus (30.35-75.25%, P < 0.05) and healthy controls (54.83-86.38%, P < 0.01). Most strikingly, the levels of FoxP3 expression determined by mean fluorescence intensity in CD4(+)CD25(high) cells in patients with active SLE were significantly down-regulated compared with healthy subjects (130 +/- 22 versus 162 +/- 21, P = 0.012). CD4(+)CD25(high) T cells are low in new-onset patients with active SLE and restored after treatment. Despite that the percentage of CD4(+)FoxP3(+) T cells appear high, the levels of FoxP3 expression in CD4(+)CD25(high) T cells are down-regulated in untreated lupus patients. There is a disproportional expression between CD25(high) and FoxP3(+) in new-onset patients with active SLE.     

Higher frequencies of circulating ICOS+, IL-21+ T follicular helper cells and plasma cells in patients with new-onset membranous nephropathy

Background: T follicular helper (TFH) cells and B cells are known to regulate humoral immune responses. This study is aimed at examining the putative contribution of different subsets of circulating of TFH cells and B cells to membranous nephropathy (MN).

Methods: A total of 45?MN patients and 19 healthy controls (HCs) were examined for the number of TFH cells and B cells by flow cytometry. The level of 24-h urinary protein and eGFR were calculated, and the level of serum cytokines was examined. The potential association among these measures was analyzed.

Results: Compared to the HCs, MN patients had significantly higher numbers of circulating CD4⁺CXCR5⁺, CD4⁺CXCR5⁺ICOS⁺, CD4⁺CXCR5⁺CD154⁺, CD4⁺CXCR5⁺IL-21⁺, and CD4⁺CXCR5⁺CD28⁺ TFH cells, as well as IgD⁺CD27^?CD19⁺ and CD138⁺CD19⁺ B cells. However, the number of IgD⁺CD27⁺CD19⁺ B cells was significantly lower in MN patients than in the HC. The levels of serum IL-21, IL-2, IL-4, IL-10, IL-17A, and IFN-γ were significantly higher in MN patients than in the HC. Furthermore, the numbers of CD4⁺CXCR5⁺, CD4⁺CXCR5⁺ICOS⁺, CD4⁺CXCR5⁺CD154⁺, CD4⁺CXCR5⁺IL-21⁺, CD4⁺CXCR5⁺CD28⁺ TFH cells, CD138⁺CD19⁺ B cells, and the level of sera IL-21 were negatively correlated with the values of eGFR, but positively correlated with the levels of 24-h urinary proteins. Following treatment, the numbers of CD4⁺CXCR5⁺, CD4⁺CXCR5⁺ICOS⁺, CD4⁺CXCR5⁺CD154⁺, CD4⁺CXCR5⁺IL-21⁺, CD4⁺CXCR5⁺CD28⁺ TFH cells, CD138⁺CD19⁺ B cells, and the levels of IL-21 were significantly reduced. In contrast, IL-4 and IL-10 levels were noticeably increased after treatment.

Conclusions: Data suggest that activated TFH and plasma cells may contribute to the pathogenesis of MN.     

HLA DRB1*1503 allelic haplotype predominance and associated immunodysregulation in systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a chronic, relapsing, and remitting disease affecting primarily African American females of child bearing age. Familial aggregation of this disease suggests that at least part of the susceptibility for this disease is genetic, although environmental and hormonal influences are also likely to play a role. Early studies of genetic susceptibility to SLE revealed several of the major histocompatibility complex molecules, namely HLA DR, to be linked to SLE. Meta-analysis of genome scans has yielded loci significant for lupus patients, one of which includes the MHC region.Regulatory T cells are immunoregulatory cells that modulate activated immune cells. These cells play a large role in homeostasis of the immune responses and maintenance of immunologic tolerance, i.e., prevention of autoimmunity. Decreased numbers of regulatory T cells have been described in many autoimmune diseases, including systemic lupus erythematosus.Autoantibody production in systemic lupus erythematosus and the resulting immune complex formation and complex deposition into tissues are arguably the central core of immune dysregulation leading to disease manifestations and symptoms. Inability of the immune system to recognize and inhibit autoreactive immune cells in this particular autoimmune disease may be the result of inappropriate numbers and function of regulatory T cells.This study aims to characterize the immune cell population in patients from our community suffering from systemic lupus erythematosus and to prove that these patients exhibit a unique cellular profile compared to healthy age, race and gender matched control subjects. Surprisingly, our findings demonstrate that patients from the local Mississippi area exhibit increased proportions of CD25⁺ FoxP3⁺ regulatory T cells and CD25⁺ FoxP3⁻ T cells (of CD45⁺ CD3⁺ CD4⁺ helper T cells) as compared to healthy controls.HLA tissue-typing of these lupus patients revealed a prominent subgroup (~ 30%) of patients possessing the HLA DRB1*1503 allele. The investigation of this subgroup demonstrated regulatory T cell composition similar to that of the total lupus group and to that of the non-HLA DRB1*1503 subgroup.Genetic analysis for molecular gene expression levels of various lupus-associated genes by real-time PCR demonstrated a unique profile as compared to healthy controls. Increased gene expression of FoxP3 together with decreased gene expression levels of GATA3, TNFAIP3, and TNFSF4 suggest that variations in gene products compared to healthy controls may be playing a role in the immune cell dysregulation and disproportionate CD25⁺ FoxP3⁺ regulatory T cells.     

CD4+CD45RA−FoxP3high activated regulatory T cells are functionally impaired and related to residual insulin‐secreting capacity in patients with type 1 diabetes

Accumulating lines of evidence have suggested that regulatory T cells (T_regs) play a central role in T cell-mediated immune response and the development of type 1A and fulminant type 1 diabetes. CD4⁺forkhead box protein 3 (FoxP3)⁺ T cells are composed of three phenotypically and functionally distinct subpopulations; CD45RA⁺FoxP3^low resting T_regs (r-T_regs), CD45RA⁻FoxP3^high activated T_regs (a-T_regs) and CD45RA⁻FoxP3^low non-suppressive T cells (non-T_regs). We aimed to clarify the frequency of these three subpopulations in CD4⁺FoxP3⁺ T cells and the function of a-T_regs with reference to subtypes of type 1 diabetes. We examined 20 patients with type 1A diabetes, 15 patients with fulminant type 1 diabetes, 20 patients with type 2 diabetes and 30 healthy control subjects. A flow cytometric analysis in the peripheral blood was performed for the frequency analysis. The suppressive function of a-T_regs was assessed by their ability to suppress the proliferation of responder cells in a 1/2:1 co-culture. A flow cytometric analysis in the peripheral blood demonstrated that the frequency of a-T_regs was significantly higher in type 1A diabetes, but not in fulminant type 1 diabetes, than the controls. Further, the proportion of a-T_regs among CD4⁺FoxP3⁺ T cells was significantly higher in patients with type 1A diabetes with detectable C-peptide but not in patients with type 1A diabetes without it and with fulminant type 1 diabetes. A proliferation suppression assay showed that a-T_regs were functionally impaired both in fulminant type 1 diabetes and in type 1A diabetes. In conclusion, a-T_regs were functionally impaired, related to residual insulin-secreting capacity and may be associated with the development of type 1 diabetes.     

Expansion of regulatory T cells from umbilical cord blood and adult peripheral blood CD4+CD25+ T cells

CD4⁺CD25⁺ regulatory T cells (Treg), if properly expanded from umbilical cord blood (UCB), may provide a promising immunotherapeutic tool. Our previous data demonstrated that UCB CD4⁺CD25⁺ T cells with 4-day stimulation have comparable phenotypes and suppressive function to that of adult peripheral blood (APB) CD4⁺CD25⁺ T cells. We further examined whether 2-week culture would achieve higher expansion levels of Tregs. UCB CD4⁺CD25⁺ T cells and their APB counterparts were stimulated with anti-CD3/anti-CD28 in the presence of IL-2 or IL-15 for 2 weeks. The cell proliferation and forkhead box P3 (FoxP3) expression were examined. The function of the expanded cells was then investigated by suppressive assay. IL-21 was applied to study whether it counteracts the function of UCB and APB CD4⁺CD25⁺ T cells. The results indicate that UCB CD4⁺CD25⁺ T cells expanded much better than their APB counterparts. IL-2 was superior to expand UCB and APB Tregs for 2 weeks than IL-15. FoxP3 expression which peaked on Day 10–14 was comparable. Most importantly, expanded UCB Tregs showed greater suppressive function in allogeneic mixed lymphocyte reaction. The addition of IL-21, however, counteracted the suppressive function of expanded UCB and APB Tregs. The results support using UCB as a source of Treg cells.     

T regulatory cells and Th1/Th2 cytokines in peripheral blood from tuberculosis patients

About 10% of people infected with Mycobacterium tuberculosis develop active tuberculosis (TB), and Th1 effector cells and Th1 cytokines play key roles in controlling M. tuberculosis infection. Here, we hypothesise that this susceptibility to M. tuberculosis infection is linked to increased T regulatory (Treg) cells and Th2 cytokines in TB patients. To test this, we recruited 101 participants (71 TB patients, 12 non-TB pulmonary diseases and 18 healthy subjects) and investigated Treg cells and Th1/Th2 cytokines in peripheral blood. CD4⁺CD25⁺ T cells and CD4⁺CD25⁺FoxP3⁺ T cells significantly increased and IL-5 dramatically decreased in TB patients relative to healthy subjects. CD8⁺CD28⁻ T cells, IFN-γ, TNF-α, IL-10 and IL-4 significantly increased in patients with culture and sputum smear-positive pulmonary TB (PTB(+)) compared with healthy subjects. CD4⁺CD25⁺FoxP3⁺ and CD8⁺CD28⁻ T cells significantly decreased in PTB(+) after one month of chemotherapy. CD4⁺, CD4⁺CD25⁺ and CD8⁺CD28⁺ T cells significantly increased in extra-pulmonary TB patients after one month of chemotherapy. These findings suggest that M. tuberculosis infection induces circulating CD4⁺CD25⁺FoxP3⁺ and CD8⁺CD28⁻ T cell expansion, which may be related to the progression of M. tuberculosis infection, and that the balance between effector immune responses and suppression immune responses is essential to control M. tuberculosis infection.