Foxp3 Cells Research Articles

The Contribution of the Thymus to Autoimmune Phenotypes in GvHD and Colitis

The thymus plays a crucial role in preventing autoimmune reactions in a variety of species. Through the expression of the autoimmune regulator (Aire), a protein found in medullary thymic epithelial cells (mTECs), the thymus is able to express tissue-restricted peripheral self-antigens (TRAs). Through their interaction with these TRAs, developing thymocytes that target host antigens can be tagged for clonal deletion. Graft versus host disease (GvHD), an autoimmune reaction occurring in approximately 40% of allogeneic stem cell transplantation patients, is an immune mediated reaction in which donor T cells recognize the host as antigenically foreign, causing donor T cells to expand and attack host tissues. GvHD is often associated with significant morbidity and mortality, and because the thymus is a target tissue of T-cell alloimmunity and can be severely damaged in many autoimmune diseases, we focused on significant morphological and cellular changes within the thymus as possible contributors to the continuing proliferation and survival of alloimmune T-cells in GvHD.Previous data collected from a mouse model of GvHD showed a reduction in expression of Aire and a reduced diversity of TRAs in the thymus. We also found that the thymus is significantly smaller in size when compared to wild-type mice, which correlates to overall thymic involution as both the cortical and medullary TEC levels were reduced. This reduction of Aire expression also correlated with a reduction in thymically-derived FoxP3+NrpI+regulatory T-cells, which were significantly reduced in both spleen and blood of GvHD mice when compared to bone marrow transplant controls (blood p < 0.0005, spleen p < 0.0001). SEMA4A, a transmembrane protein known to interact with Nrp1 to promote regulatory T-cell function and survival, was also significantly changed in GvHD compared to healthy controls (p = 0.014). In addition, cytokine data collected from a GvHD mouse model also showed increases in several chemokines associated with thymic atrophy and aging, indicating that thymii found in GvHD mice are subject to significant damage due to autoimmune reactions.Interestingly, there is evidence of thymic damage in mouse models of colitis as well. Ulcerative colitis is a form of inflammatory bowel disease caused by T-cell infiltration into the colon, resembling some of the pathophysiology of G.I. GvHD. Thymii from DSS colitis mice showed a similarly significant reduction of Aire and FoxP3 expression, suggesting a new, more prominent role for thymic damage in colitis severity (p < 0.005). This damage was observed to occur in a generalized way, as FACS analysis of thymic epithelial cell types showed that both medullary and cortical epithelial cells in diseased thymus were reduced, not just mTECs. The reduction in expression of Aire protein is due directly to RNA reduction, with a mean 8 fold reduction of Aire RNA expression in colitis mice compared to healthy mice (p = 0.0154).While SEMA4A was seen to be increased in GvHD, there was a significant reduction in expression in colitis thymii (p=0.0138). Reductions in SEMA4A have previously been shown to cause impaired immune cell differentiation and function, followed by impaired homeostasis and function of FoxP3+T-cells, indicating that this reduction in SEMA4A may be the cause of reduced FoxP3 expression in the colitis thymus. It was also observed that NrpI expressing FoxP3 cells in the spleen and blood were unchanged in colitis mice, indicating that reduction in FoxP3 expressing cells in colitis are not entirely due to thymic damage. This is supported by cytokine data, in which no inflammatory or aging related cytokines were changed in colitis thymii.The significant thymic involution and reduced expression of the Aire protein in the thymus is a likely contributor to overall increases of autoimmune T-cells in both colitis and GvHD. Though colitis and GvHD clearly use different pathways to affect the thymus, thymic damage plays a significant role in both diseases. The reduced expression of the FoxP3 protein also indicates a role for loss of Aire in lowered immune tolerance, contributing to the overall autoimmune phenotype in both diseases. DisclosuresNo relevant conflicts of interest to declare.

Erratum to: Effects of aging on human leukocytes (part II): immunophenotyping of adaptive immune B and T cell subsets.

Immunosenescence results from a continuous deterioration of immune responses resulting in a decreased response to vaccines. A well-described age-related alteration of the immune system is the decrease of de novo generation of T and B cells. In addition, the accumulation of memory cells and loss of diversity in antigen specificities resulting from a lifetime of exposure to pathogens has also been described. However, the effect of aging on subsets of γδTCR(+) T cells and Tregs has been poorly described, and the efficacy of the recall response to common persistent infections in the elderly remains obscure. Here, we investigated alterations in the subpopulations of the B and T cells among 24 healthy young (aged 19-30) and 26 healthy elderly (aged 53-67) individuals. The analysis was performed by flow cytometry using freshly collected peripheral blood. γδTCR(+) T cells were overall decreased, while CD4(+)CD8(-) cells among γδTCR(+) T cells were increased in the elderly. Helios(+)Foxp3(+) and Helios(-)Foxp3(+) Treg cells were unaffected with age. Recent thymic emigrants, based on CD31 expression, were decreased among the Helios(+)Foxp3(+), but not the Helios(-)Foxp3(+) cell populations. We observed a decrease in Adenovirus-specific CD4(+) and CD8(+) T cells and an increase in CMV-specific CD4(+) T cells in the elderly. Similarly, INFγ(+)TNFα(+) double-positive cells were decreased among activated T cells after Adenovirus stimulation but increased after CMV stimulation. The data presented here indicate that γδTCR(+) T cells might stabilize B cells, and functional senescence might dominate at higher ages than those studied here.

Foxp3 and Toll-like receptor signaling balance Treg cell anabolic metabolism for suppression.

CD4+ effector T cells (Teff cells) and regulatory T cells (Treg cells) undergo metabolic reprogramming to support proliferation and immunological function. Although signaling via the lipid kinase PI(3)K (phosphatidylinositol-3-OH kinase), the serine-threonine kinase Akt and the metabolic checkpoint kinase complex mTORC1 induces both expression of the glucose transporter Glut1 and aerobic glycolysis for Teff cell proliferation and inflammatory function, the mechanisms that regulate Treg cell metabolism and function remain unclear. We found that Toll-like receptor (TLR) signals that promote Treg cell proliferation increased PI(3)K-Akt-mTORC1 signaling, glycolysis and expression of Glut1. However, TLR-induced mTORC1 signaling also impaired Treg cell suppressive capacity. Conversely, the transcription factor Foxp3 opposed PI(3)K-Akt-mTORC1 signaling to diminish glycolysis and anabolic metabolism while increasing oxidative and catabolic metabolism. Notably, Glut1 expression was sufficient to increase the number of Treg cells, but it reduced their suppressive capacity and Foxp3 expression. Thus, inflammatory signals and Foxp3 balance mTORC1 signaling and glucose metabolism to control the proliferation and suppressive function of Treg cells.

Overexpression of CD39 in hepatocellular carcinoma is an independent indicator of poor outcome after radical resection.

Nucleoside triphosphate diphosphohydrolase-1 (ENTPD1/CD39) is the rate-limiting enzyme in a cascade leading to the generation of immunosuppressive adenosine and plays an important role in tumor progression. This study aimed to evaluate the expression of CD39 and CD39Foxp3 regulatory T cells (Tregs) and to determine their prognostic role in patients with hepatocellular carcinoma (HCC) after radical resection.Immunohistochemistry (IHC) and double IHC were used to analyze CD39 expression or the expression of CD39 and Foxp3 in a cohort of 324 HCC patients who underwent curative resection. The quantification of CD39 expression levels was determined using a computerized image analysis system and was evaluated by mean optical density (MOD), which corresponded to the positive staining intensity of CD39. The number of positive Foxp3 cells and both CD39 and Foxp3 positive cells in each 1-mm-diameter cylinder were counted under high-power magnification (×400). The "minimum P value" approach was used to obtain the optimal cutoff value for the best separation between groups of patients in relation to time to recurrence (TTR) or overall survival (OS). The expression of CD39 in HCC cell lines with stepwise metastatic potential and in human umbilical vein endothelial cells was determined by reverse transcription-polymerase chain reaction, Western blotting, and immunofluorescence. The SPSS 17.0 statistical package was used for statistics.CD39 was principally expressed on vascular endothelial cells, macrophagocytes, Tregs, and tumor cells in HCC. Compared with paired peritumoral tissues, tumoral tissues had a significantly higher expression level of CD39 (P < 0.0001). Overexpression of tumoral CD39 was related to increased tumor recurrence and shortened overall survival. Furthermore, the expression level of peritumoral CD39 showed a prognostic role in TTR and OS. Double IHC showed that tumoral tissues had significantly higher Foxp3Tregs and CD39Foxp3Tregs count per 1 mm core (14.1659 vs 4.9877, P = 0.001; 11.5254 vs 3.3930, P < 0.001) and a higher CD39Foxp3/Foxp3 ratio compared with paired peritumoral tissues. CD39Foxp3Tregs were a better prognosticator than CD39Tregs for TTR.Overexpression of CD39 protein in HCC was an independent predictor of poor outcome after radical resection. The CD39Foxp3Tregs count added prognostic power to Foxp3Tregs, providing a potential target for tumor immunotherapy.

Regulatory T-cells and IL17A+ cells infiltrate oral lichen planus lesions

Oral lichen planus (OLP) is a complex immunological disorder, mediated in part by the release of cytokines from activated T-cells. Of late, two closely related T-helper (Th) cell subsets; regulatory T-cells (Tregs; FoxP3(+)) and Th17 cells (IL17(+)) have been described in various chronic inflammatory diseases. The aim of this study was to determine the expression of FoxP3 and IL17 in OLP using immunohistochemistry (IHC) and quantitative real-time reverse transcriptase polymerase chain reaction (qPCR). For IHC, formalin fixed, paraffin embedded archival specimens, an OLP group (n=10) and a non-specific inflammatory (NSI) control group (n=9) were used. In addition, 12 fresh tissue samples were used to determine gene expression of FoxP3 and IL17. Significantly more FoxP3(+) cells were present in OLP than in NSI. IL17(+) cells were significantly more frequent in the control tissues than in OLP. The gene expression experiments revealed a significantly higher expression of FoxP3 in OLP when compared to the controls. IL17 gene expression was not different between the groups. Double labelling immunofluorescence indicated co-localisation of IL17 with tryptase(+) mast cells. These findings suggest FoxP3(+) Tregs have a more prominent role in the pathogenesis of OLP when compared to IL17(+)cells.

Rapid and Efficient Generation of Regulatory T Cells to Commensal Antigens in the Periphery

Commensal bacteria shape the colonic regulatory T (Treg) cell population required for intestinal tolerance. However, little is known about this process. Here, we use the transfer of naive commensal-reactive transgenic Tcells expressing colonic Treg Tcell receptors (TCRs) to study peripheral Treg (pTreg) cell development in normal hosts. We found that Tcells were activated primarily in the distal mesenteric lymph node. Treg cell induction was rapid, generating >40% Foxp3(+) cells 1week after transfer. Contrary to prior reports, Foxp3(+) cells underwent the most cell divisions, demonstrating that pTreg cell generation can be the dominant outcome from naive Tcell activation. Moreover, Notch2-dependent, but not Batf3-dependent, dendritic cells were involved in Treg cell selection. Finally, neither deletion of the conserved nucleotide sequence 1 (CNS1) region in Foxp3 nor blockade of TGF-β (transforming growth factor-β)-receptor signaling completely abrogated Foxp3 induction. Thus, these data show that pTreg cell selection to commensal bacteria is rapid, is robust, and may be specified by TGF-β-independent signals.

Assessment of the Effects of Rituximab Monotherapy on Different Subsets of Circulating T-Regulatory Cells and Clinical Disease Severity in Severe Pemphigus Vulgaris

Background: Robust evidence for the efficacy of rituximab monotherapy in pemphigus is lacking. The effects of rituximab on T-regulatory cells (Tregs) in pemphigus have not been studied. Objective: The primary objective was to assess the efficacy of rituximab monotherapy in severe pemphigus vulgaris. The secondary objectives were to assess whether counts of different subsets of Tregs in the peripheral blood correlate with baseline clinical severity and whether clinical response in severe pemphigus is associated with an alteration in the Treg count. Methods: Eighteen eligible subjects with severe pemphigus vulgaris were recruited and were treated with 1 g of intravenous rituximab on days 0 and 15. Efficacy was assessed in terms of disease control, time to disease control, complete remission off therapy, and relapse. Flow cytometric analysis of CD4+CD25+FoxP3, IL-10-secreting Tr1, and TGF-β secreting Th3 regulatory cells was performed. Clinical evaluation and flow cytometric analysis of Tregs was performed periodically until follow-up at 26 weeks. Results: Rituximab monotherapy was able to induce complete remission in all but 5 (68.75%) patients and was well tolerated. No direct relationship between clinical severity and CD4+CD25+FoxP3 cell counts was found. There were inverse correlations between serially measured values of the cutaneous and mucosal Autoimmune Bullous Skin Disorder Intensity Score (ABSIS) and Th3 cell count. Conclusion: Rituximab is a safe and effective monotherapy option for severe pemphigus. As the immunological findings were somewhat different from those observed in other autoimmune conditions treated with rituximab, further studies are required to substantiate the findings of our study in pemphigus patients.

Cell autonomous or systemic EGFR blockade alters the immune-environment in squamous cell carcinomas.

Targeting mutations and amplifications in the EGFR has been successful precision therapy for cancers of the lung, oral cavity and gastrointestinal track. However, a systemic immune reaction manifested by dose-limiting inflammation in the skin and gut has been a consistent adverse effect. To address the possibility that intra-tumoral immune changes contribute to the anti-cancer activity of EGFR inhibition, squamous cancers were produced by syngeneic orthografts of either EGFR null or wildtype mouse primary keratinocytes transduced with an oncogenic H-ras retrovirus. Flow cytometric, RNA and Bioplex immunoassay analyses of the tumor immune milieu were performed. Cancers forming from keratinocytes genetically depleted of EGFR were smaller than wildtype cancers and had fewer infiltrating FoxP3 Treg cells, lower Foxp3 RNA and a lower percentage of CD4 PD1 positive cells indicating a tumor cell autonomous regulation of its microenvironment. Hosts bearing wildtype cancers treated with gefitinib for 1 week showed a trend for smaller tumors. In this short term pharmacological model, there was also a trend to reduced FoxP3 cells and FoxP3 RNA in the tumors of treated mice as well as a substantial increase in the ratio of IL-1A/IL-1RA transcripts. These results suggest that relatively brief systemic inhibition of EGFR signaling alters the immune environment of the targeted cancer. Together these data imply that an EGFR dependent Treg function supports the growth of squamous cancers and is a target for the therapeutic activity of EGFR inhibition.

Foxp3 downregulation in NSCLC mediates epithelial-mesenchymal transition via NF-κB signaling.

Forkhead box P3 (Foxp3) is a member of forkhead box transcription factor family and it was identified as a tumor suppressor in various solid tumors. This study evaluated the expression of Foxp3 in non-small cell lung cancer(NSCLC) and investigated its role in epithelial‑mesenchymal transition(EMT) of cancer cells. qRT-PCR and western blot analysis were used for examining the expression of Foxp3 in NSCLC tissues and the non-tumor tissues. A tissue microarray was constructed and scored for evaluating the clinical significance of Foxp3 expression in NSCLC tissues. RNAi was employed for downregulating Foxp3 expression and cell proliferation was done with MTT assay. Transwell with or without basement membrane matrix was used for cell migration and invasion assay respectively. Foxp3 was found downregulated in NSCLC tissues compared with non-tumoral tissues; downregulation of Foxp3 predicted adverse tumor stage and overall survival; silencing of FOXP3 promoted the proliferation, migration and invasion ability of NSCLC cells and influenced the expression level of EMT-associated proteins. However, forced expression of Foxp3 could reverse this effect. Moreover, Foxp3 could interact with LMO2 and affect the expression level of TAL1, which was in accordance with the findings in T-cell acute lymphoblastic leukemia. By screening the signalling pathways, we observed an obvious upregulation of phosphorylated NF-κB in A549 and H520 cells after silencing of FOXP3. Our results suggest that Foxp3 suppressed NSCLC cell metastasis, at least partially, via NF-κB signaling.

Divergence of helper, cytotoxic, and regulatory T cells in the decidua from miscarriage

The aim of this prospective study was to evaluate phenotypic differences of helper T (Th), cytotoxic T (Tc), and regulatory T (Treg) cells in the deciduae of missed miscarriage with a normal chromosome karyotype of a fetus (MN) and missed miscarriage with an abnormal chromosome karyotype of a fetus (MA). The decidua of 19 MN and 28 MA was obtained. Additionally, the decidua of 15 induced abortion (IA) and the endometrium of 19 non-pregnant women (EM) were obtained. IFN-γ(+) , IL-17(+) , CD25(high) Foxp3(+) cells in CD4(+) (Th) cells, and IFN-γ(+) cells in CD8(+) (Tc) cells were evaluated by flow cytometry. The percentages of IFN-γ(+) Tc and CD4(+) CD25(high) Foxp3(+) (Treg) cells in MN were significantly increased as compared with MA and IA. The percentage of IFN-γ(+) Th in MN was increased as compared with IA. Activation of IFN-γ(+) Tc and Treg cells in the decidua might be associated with the pathophysiology underlying MN.

Prevalence of FoxP3+ Cells in Canine Tumours and Lymph Nodes Correlates Positively with Glucose Transporter 1 Expression

Hypoxia and regulatory T cells (Tregs) in tumours are both known to be negative prognostic factors in cancer, and this study demonstrated a correlation between the two factors in canine neoplasia. Samples of 57 canine tumours and 29 canine lymph nodes categorized as tumour-draining, with or without metastasis, or reactive and not tumour-associated, were examined. Sequential sections were labelled by immunohistochemistry for glucose transporter 1 (Glut1) and FoxP3 as markers of hypoxia and Tregs, respectively. Up to 21 regions of interest (ROI) were selected in each section in a representative pattern and were assigned a semiquantitative score based on Glut1 labelling. The number of FoxP3(+) cells within each ROI was counted. A generalized estimating equation with negative binomial log link function was used to determine an association between Glut1 expression and FoxP3(+) cell count. Higher Glut1 immunoreactivity was correlated with significantly higher numbers of FoxP3(+) cells in the total tumour sample pool and total lymph node sample pool. Analysis of various subcategories of tumours and lymph nodes showed that this correlation was also present within samples characterized as malignant, haemopoietic mesenchymal tumours, non-haemopoietic mesenchymal tumours, epithelial tumours, lymphoma, lymph nodes containing metastases and reactive lymph nodes. These results indicate that hypoxia in canine tumours may result in an increased infiltration by Tregs.

Abstract 5006: Docetaxel-platinum-fluorouracil (TPF) induction chemotherapy increases PD-L1 positivity and CD8+ / FOXP3+ immune infiltrates in advanced head and neck squamous cell carcinoma (HNSCC)

Abstract Introduction: Checkpoint inhibitors targeting programmed cell death-1/-ligand1 (PD-1/-L1) have demonstrated promising clinical activity in locally advanced head and neck squamous cell carcinoma (LA-HNSCC), with overall response rates (ORR) of ≈20% in unselected patients (pts). Moreover, significant improvements have been observed when selecting pts on PD-L1 positivity, with ORR reaching 50-78%. It is therefore crucial to optimize therapeutic sequence between induction chemotherapy (Docetaxel, Platinum, Fluorouracile,TPF) and anti-PD-1/-L1 in order to maximize the number of pts that benefit from these drugs. Methods: All patients with LA-HNSCC treated at Gustave Roussy between 2006 and 2013 by induction TPF followed by surgery were considered. Only patients for who paired samples (pre and post-TPF) were available were kept for further analysis. PD-L1 expression was quantified by immunochemistry (IHC) (clone E1L3N, Cell Signaling) on a Ventana Discovery Ultra autostainer by a senior pathologist according to a validated protocol. PD-L1 IHC staining on tumor cells (TC) and tumor-infiltrating immune cells (IC) was quantified with a positivity threshold of ≥5%. The density of CD8+ and FOXP3+ lymphocytes was also assessed. Results: Out of 313 pts receiving induction TPF, 86 underwent surgery; paired samples were available for 21. Baseline PD-L1 expression was positive in only 2 and 5 samples for TC and IC respectively. An increased PD-L1 expression was observed post-TPF in 3 (14%) and 15 (71%; p = 0.0003, T-test) samples for TC and IC respectively. Tumor-infiltrating CD8+ and FOXP3+ populations also significantly increased post-TPF. Conclusion: TPF induction in LA-HNSCC increases PD-L1 positivity on tumor-infiltrating immune cells as well as CD8+ and FOXP3+ populations. This suggests that anti-PD-1/-L1 therapy might be more effective as a second-line than first-line strategy, but warrants clinical evaluation. IHC staining resultspre-TPFpost-TPFpaired samplesmedianCImedianCIT-testPD-L1 positive IC (%)2110-10100-300.0003***PD-L1 positive IC (%)2100-2010-200.08 NSCD8+ cell density (n/mm3)171194-77942653-11900.016*FoxP3 cell density (n/mm3)179126-1801774-4670.018* Citation Format: Charlotte Leduc, Julien Adam, Emilie Louvet, Marine Bernard, Elodie Maingot, Caroline Even, Ken A. Olaussen, Stéphane Temam, Jean-Charles Soria, Sophie Postel-Vinay. Docetaxel-platinum-fluorouracil (TPF) induction chemotherapy increases PD-L1 positivity and CD8+ / FOXP3+ immune infiltrates in advanced head and neck squamous cell carcinoma (HNSCC). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5006.

Trophoblast Major Histocompatibility Complex Class I Expression Is Associated with Immune-Mediated Rejection of Bovine Fetuses Produced by Cloning.

Trophoblast cells from bovine somatic cell nuclear transfer (SCNT) conceptuses express major histocompatibility complex class I (MHC-I) proteins early in gestation, and this may be one cause of the significant first-trimester embryonic mortality observed in these pregnancies. MHC-I homozygous-compatible (n = 9), homozygous-incompatible (n = 8), and heterozygous-incompatible (n = 5) SCNT pregnancies were established. The control group consisted of eight pregnancies produced by artificial insemination. Uterine and placental samples were collected on Day 35 ± 1 of pregnancy, and expression of MHC-I, leukocyte markers, and cytokines were examined by immunohistochemistry. Trophoblast cells from all SCNT pregnancies expressed MHC-I, while trophoblast cells from age-matched control pregnancies were negative for MHC-I expression. Expression of MHC-I antigens by trophoblast cells from SCNT pregnancies was associated with lymphocytic infiltration in the endometrium. Furthermore, MHC-I-incompatible conceptuses, particularly the heterozygous-incompatible ones, induced a more pronounced lymphocytic infiltration than MHC-I-compatible conceptuses. Cells expressing cluster of differentiation (CD) 3, gamma/deltaTCR, and MHC-II were increased in the endometrium of SCNT pregnancies compared to the control group. CD4+ lymphocytes were increased in MHC-I-incompatible pregnancies compared to MHC-I-compatible and control pregnancies. CD8+, FOXP3+, and natural killer cells were increased in MHC-I heterozygous-incompatible SCNT pregnancies compared to homozygous SCNT and control pregnancies.

Tolerogenic Vaccination with MOG/VitD Overcomes Aggravating Effect of C. albicans in Experimental Encephalomyelitis.

Multiple sclerosis (MS) is an immune-mediated demyelinating disorder of the central nervous system (CNS). We described that Candida albicans (Ca) aggravates experimental autoimmune encephalomyelitis (EAE) that is a model to study MS. We also observed that vaccination with a myelin peptide (MOG) in the presence of vitamin D (VitD) protected mice against EAE. In this work, we investigated whether Ca infection interferes with the efficacy of this vaccine. EAE was induced in C57BL/6 female mice previously vaccinated with MOG+VitD and then infected 3 days before encephalomyelitis induction. Vaccination was able to control EAE development in infected mice. These animals gained weight, and only a few progressed to very low clinical scores. Protection was confirmed by a lower inflammatory infiltration in the CNS and was also associated with a reduced production of encephalitogenic cytokines by spleen and CNS cell cultures. The elevated percentage of CD25(+) FoxP3(+) cells suggests that regulatory T cells are involved in the protection. Adoptive transfer of splenocytes from mice vaccinated with MOG+VitD supports the view that protection is mediated by immunoregulatory cells. Together, these experiments provide evidence demonstrating that EAE can be prevented by the inverse vaccination with MOG+VitD even in the presence of a disease-aggravating infectious agent.

CD4+CD25hiFoxp3+ Cells Exacerbate Bleomycin-Induced Pulmonary Fibrosis

Idiopathic pulmonary fibrosis is a fatal lung disease with a median survival of 2 to 5 years. A decade of studies has downplayed inflammation contributing to its pathogenesis. However, these studies preceded the discovery of regulatory T cells (Tregs) and all of their functions. On the basis of human studies demonstrating Tregs can decrease graft-versus-host disease and vasculitides, there is consideration of their use to treat idiopathic pulmonary fibrosis. We hypothesized that Treg therapy would attenuate the fibroplasia involved in a preclinical murine model of pulmonary fibrosis. IL-2 complex was used invivo to expand CD4(+)CD25(hi)Foxp3(+) cells in the lung during intratracheal bleomycin challenge; however, this unexpectedly led to an increase in lung fibrosis. More important, this increase in fibrosis was a lymphocyte-dependent process. We corroborated these results using a CD4(+)CD25(hi)Foxp3(+) cellular-based therapy. Mechanistically, we demonstrated that CD4(+)CD25(hi)Foxp3(+) cells undergo alterations during bleomycin challenge and the IL-2 complex had no effect on profibrotic (eg, transforming growth factor-β) or type 17 immune response cytokines; however, there was a marked down-regulation of the type 1 and augmentation of the type 2 immune response cytokines from the lungs. Collectively, our animal studies show that a specific lung injury can induce Treg alterations, which can augment pulmonary fibrosis.

Tumor-infiltrating lymphocytes in breast cancer predict the response to chemotherapy and survival outcome: A meta-analysis

Tumor-infiltrating lymphocytes (TILs) influence tumor prognosis and the chemotherapeutic response. Here, we quantified the clinical relevance of TILs, including the effect of TILs on lymphocyte subpopulations and assessed their consistency in breast cancer. We searched published literature from January 2000 to January 2016. The main parameters analyzed were pathological complete response (pCR) and survival outcome following chemotherapy in patients with breast cancer. Pooled odds ratio (OR) or relative risk (RR) values with 95% confidence intervals (CIs) were computed using random and fixed-effects models. Subgroup and heterogeneity analyses were also conducted. Twenty-three studies, which included 13,100 patients, met the inclusion criteria. The pooled results showed that TILs were associated with clinicopathological parameters of biologically aggressive phenotypes, such as high tumor grade or estrogen/progesterone receptor negativity, but they were not correlated with human epidermal growth factor receptor-2 expression. Moreover, a high TIL level was associated with a significantly improved pCR rate compared with a low TIL level (OR, 2.81; P < 0.001), particularly in the triple-negative breast cancer subtype (OR, 4.67; P < 0.001). An analysis of lymphocyte subpopulations showed that infiltration by CD8 lymphocytes, but not by CD4 lymphocytes and Foxp3 cells, was associated with a high pCR rate. Furthermore, a high TIL level was associated with significantly longer disease-free survival and overall survival. Our present meta-analysis indicates that an increased number of TILs predicted pCR to chemotherapy and improved survival. A high TIL level, characterized mainly by the infiltration of CD8 lymphocytes, is a strong predictive and prognostic factor.

Role of Interleukin - 1 alpha (IL-1α) and Regulatory T (Treg) Cells In The Pathogenesis of Inflammatory Bowel Disease

Background: Continuous effort has been paid to deeply understanding the pathophysiology of Inflammatory bowel diseases (IBD).It has been postulated that multiple factors such as genetic susceptibility, immunological and environmental factors are involved and generate a dysregulated response of the mucosal immune system toward intraluminal antigens. Accumulating evidence indicates that discrepancy between the number of T regulatory cells (Treg) in peripheral blood and in inflamed colonic mucosa has a central role and Interleukin (IL)-1 is a key mediator in the pathogenesis of IBD. Objective: The aim of this study was to determine the role of Interleukin1 alpha (IL-1α) and T regulatory (Treg) cells in the pathogenesis of IBD. Study Design: This study was conducted in internal medicine Department of ElHussein university hospital, AL-Azhar University. In this study, 50 subjects were selected and divided into two groups; Group A (n=30) were patients with IBD and Group B (n=20) were healthy control group. The two groups were subjected to the following; full history taking, clinical evaluation, Abdominal ultrasonography, CBC with differential count, Liver function tests, renal function tests, C-reactive protein , ESR and serum level of both Treg cells and IL-1α . IL-1α was detected in serum by ELISA and (Treg) (FoxP3) cells by flow cytometry techniques. Colonoscopy with multiple biopsies and histopathological assessment was performed to group A only. Results: Our results showed that the serum level of Treg cells was significantly decreased in Group A (3.16±2.26) in comparison to control group B (9.14±0.639) (P<0.001). moreover ,the serum level of Treg cells was significantly decreased in moderate to severe disease activity (1.755± 0.556) as compared with patients with mild disease activity (5.980±1.616) (P<0.001) . The present study also revealed that, the serum level of IL-1α was significantly increased in patients Group A (11.74±3.24) in comparison to control Group B (2.84±0.877) (P<0.001). Additionally, the level IL-1α was significantly higher in moderate to sever disease activity (13.750 ± 1.602) than patients with mild disease activity (7.720±1.289) (P<0.001). Conclusion: The level of Treg cells and Interleukin1 alpha have an important role in the pathogenesis and propagation of the inflammatory response in IBD and had a dynamic changes in both remission and active disease stages. These results suggested that Treg cells and Interleukin1 alpha can be used as a diagnostic markers and a potential therapeutic targets for IBD.

Neonatal respiratory syncytial virus infection has an effect on lung inflammation and the CD4(+) CD25(+) T cell subpopulation during ovalbumin sensitization in adult mice.

In BALB/c adult mice, respiratory syncytial virus (RSV) infection enhances the degree of lung inflammation before and/or after ovalbumin (OVA) respiratory sensitization. However, it is unclear whether RSV infection in newborn mice has an effect on the immune response to OVA respiratory sensitization in adult mice. The aim of this study was to determine if RSV neonatal infection alters T CD4(+) population and lung inflammation during OVA respiratory sensitization in adult mice. BALB/c mice were infected with RSV on the fourth day of life and challenged by OVA 4 weeks later. We found that in adult mice, RSV neonatal infection prior to OVA sensitization reduces the CD4(+) CD25(+) and CD4(+) CD25(+) forkhead protein 3 (FoxP3)(+) cell populations in the lungs and bronchoalveolar lavage. Furthermore, it also attenuates the inflammatory infiltrate and cytokine/chemokine expression levels in the mouse airways. In conclusion, the magnitude of the immune response to a non-viral respiratory perturbation in adult mice is not enhanced by a neonatal RSV infection.

Hepatic Infiltrates in Operational Tolerant Patients After Liver Transplantation Show Enrichment of Regulatory T Cells Before Proinflammatory Genes Are Downregulated.

Immunosuppression can be discontinued from selected and stable patients after liver transplantation resulting in spontaneous operational tolerance (SOT), although the underlying mechanisms remain elusive. Thus, we analyzed serial liver biopsy specimens from adult liver recipients enrolled in a prospective multicenter immunosuppression withdrawal trial that used immunophenotyping and transcriptional profiling. Liver specimens were collected before the initiation of weaning, at the time of rejection, or at 1 and 3 years after complete drug discontinuation. Unexpectedly, the tolerated grafts developed portal tract expansion with increased T cell infiltration after immunosuppression withdrawal. This was associated with transient and preferential accumulation of CD4(+) FOXP3(+) cells and a trend toward upregulation of immune activation and regulatory genes, without signs of rejection. At the same time, no markers of endothelial damage or activation were noted. Portal infiltrates persisted at 3 years but were characterized by decreased expression of genes associated with chronic immunological damage. Further, SOT was not associated with a progressive liver fibrosis up to 5 years. These data suggest that SOT involves several mechanisms: a long-lasting local immune cell persistence with a transient regulatory T cells accumulation followed by a downregulation of immune-activated genes over years. These results have important implications for designs and follow-up of weaning trials.

The Role of Lymphoid Neogenesis in Allografts.

De novo induction of organized lymphoid aggregates at nonlymphoid sites has been observed in many chronic inflammatory conditions where foreign antigens such as infectious agents, autoantigens or alloantigens, persist. The prevailing opinion in the field of transplantation is that lymphoid neogenesis within allografts is detrimental to the establishment of immune tolerance. These structures, commonly referred to as tertiary lymphoid organs (TLOs), are thought to contribute to graft rejection by generating and propagating local alloimmune responses. However, recent studies have shown that TLOs rich in regulatory Foxp3(+) cells are present in long-term accepting allografts. The notion that TLOs can contribute to the local downregulation of immune responses has been corroborated in other chronic inflammation models. These findings suggest that contrary to previous suggestions that the induction of TLOs in allografts is necessarily harmful, the induction of "tolerogenic" TLOs may prove advantageous. In this review, we discuss our current understanding of how TLOs are induced and how they regulate immune responses with a particular focus on alloimmunity.