Foxp3 In Patients Research Articles

The Effect of a Casein and Gluten-Free Diet on the Epigenetic Characteristics of FoxP3 in Patients With Hashimoto's Thyroiditis.

Background Hashimoto's thyroiditis (HT) is an autoimmune thyroid disease characterized by inflammation and dysfunction of the thyroid gland, resulting in hypothyroidism, it results in impaired thyroid hormone generation and mimics hypothyroidism. The disease involves complex interactions among genetic, environmental, and epigenetic factors, particularly affecting the regulation of T regulatory (Treg) cells, including CD4 +foxp3+ T cells. Treg cells, defined as CD4 + T cells, rely on the expression of the foxp3 transcription factor, which is crucial for their development and differentiation. Disruptions in this regulation can lead to immune dysregulation and potential proinflammatory responses. The study focuses on investigating the impact of dietary patterns on the epigenetic changes in the foxp3 gene, a key player in the development of HT. The primary aim was to evaluate how eliminating gluten and casein proteins from dietary regimens may influence the methylation levels of the foxp3 gene, considering the potential link between these dietary components and the triggering of autoimmune diseases. Methods An epigenetic analysis of thefoxp3 gene in HT patients who were strictly following a dietary plan compared with the control group. For the epigenetic study, a methylation analysis experiment was conducted. Results Our findings revealed a notable reduction in foxp3 gene methylation levels among HT patients who adhered to a diet excluding casein and gluten. The control maintained normal dietary guidelines and showed no significant alterations in methylation levels. Discussion The laboratory values showed a decrease in methylation levels of the foxp3 gene, with statistical significance indicated as *p<0.005, **p<0.001, ***p<0.0001, suggesting a potential enhancement in its expression which could have profound implications for immune system regulation. Disruptions in the foxp3 pathway are crucial in the development of autoimmune disorders, where altered activity hinders the regulation of T cell (Treg) development, ultimately contributing to conditions like HT disease. These findings imply that nutritional interventions, especially for individuals with HT, could potentially be a strategy for mitigating autoimmunity through epigenetic mechanisms.

The expression level of FOXP3 and CD25 in Iraqi patients with post burn injuries

Introduction and Aim: Regulatory T cells (Treg) are characterized by CD25+ expression and Foxp3+ transcription factors which is a characteristic marker for these cells. This study aimed to investigate the expression of Foxp3 in patients with post-burn injuries in Baghdad, Iraq. Materials and Methods: The study samples included 38 persons divided into two groups: the first group included patients with Total Body surface area (TBSA) of burn less than 45% (TBSA 45 %), and patients with TBSA of burn more than 45 % (TBSA 45%). The control group included healthy individuals (n=10). The expression of FOXP3 and CD25 of Treg cells was measured by using flow cytometry and data obtained was statistically analyzed. Results: The result of the study showed a significant increase in Foxp3 and CD25 expression in Treg cells that were isolated from peripheral blood in patients with TBSA 45 % and TBSA 45% following day 1 and day 5 of the burn injury in contrast to the control group, and the expression level of Foxp3 was (35.674, 63.768 ,16.147) respectively after the first day of the burn, while the expression level of Foxp3 after the fifth day of the burn was (39.588, 47.275, 16.1470) respectively. Similarly, the levels of CD5 recorded for patients in TBSA 45%, TBSA45% and control group on day1 was 29.747, 43.447 and 16.793 respectively, while on day 5 it was 33.870.197 and 16.793, for the three groups, respectively. Conclusion: The increase of Foxp3 and CD25 expression in Treg cells isolated from burn patients has an important effect in enhancing the activity of Treg cells in inhibiting the immune response.

Mutations from patients with IPEX ported to mice reveal different patterns of FoxP3 and Treg dysfunction

Mutations of the transcription factor FoxP3 in patients with "IPEX" (immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome) disrupt regulatory Tcells (Treg), causing an array of multiorgan autoimmunity. To understand the functional impact of mutations across FoxP3 domains, without genetic and environmental confounders, six human FOXP3 missense mutations are engineered into mice. Two classes of mutations emerge from combined immunologic and genomic analyses. A mutation in the DNA-binding domain shows the same lymphoproliferation and multiorgan infiltration as complete FoxP3 knockouts but delayed by months. Tregs expressing this mutant FoxP3 are destabilized by normal Tregs in heterozygous females compared with hemizygous males. Mutations in other domains affect chromatin opening differently, involving different cofactors and provoking more specific autoimmune pathology (dermatitis, colitis, diabetes), unmasked by immunological challenges or incrossing NOD autoimmune-susceptibility alleles. This work establishes that IPEX disease heterogeneity results from the actual mutations, combined with genetic and environmental perturbations, explaining then the intra-familial variation in IPEX.

Targeting FoxP3 gene to check out the impairment of tolerance in breast cancer patients

ObjectiveFoxP3 gene is a potential gene in the development of Breast Cancer (BC). Mutated expression of this gene shows some correspondence with tumorigenic activity for this reason it is important to understand. The principal goal of this study was the mutational analysis of the FoxP3 gene in BC patients. This research also provides the basic functions and genetic details on the FoxP3 gene in the Lahore population, especially how the FoxP3 gene acts as a risk factor in BC development. The study design was Cross-sectional. MethodsGenomic DNA was extracted from peripheral blood samples of BC patients. Amplified products were electrophoretically separated and target PCR products were sequenced with a Sanger sequencing analyzer. The main purpose is to determine the order of nucleotides in the FoxP3 gene. Further mutations were analyzed by the chromatograms using Chromas, NCBI nucleotide BLAST X, and the Mutation Surveyor. After that, different protein-coding sequences were studied. ResultsThe substitution occurs at location 324 where T is replaced by A and in another sample G is replaced by A at position 328 while in the third sample InDel mutation was observed at position 28. ConclusionIn the current study, intronic variants typically in the FoxP3 gene exon 2 region were confirmed by computational analysis using a gene database. We investigated nucleotide mutations in FoxP3 in patients with malignant breast tumors, along with their association with various characteristics such as marriage status, age of menopause, miscarriage and lactation history, and family history. A better understanding of the FoxP3 gene in the etiology and progression of breast cancer can reveal many explorable problems, including the clinical implications of the FoxP3 gene. However, these intronic variations are of massive importance and demonstrated that this enhancer can establish a physical interaction with a promoter site binding nuclear factor-κB components necessary for T cell regulation.

The crosstalk between H. pylori virulence factors and the PD1:PD-L1 immune checkpoint inhibitors in progression to gastric cancer

BackgroundThe progression to gastric cancer has been linked to chronic infection with Helicobacter pylori (H. pylori). Immune checkpoint inhibitors (programmed cell death -1, PD-1; programmed cell death –ligand 1, PD-L1) have a role in cancer immune escape. The relationship between H. pylori virulence factors with PD-1, PD-L1 T helper 1 (Th1), T helper 17 (Th17), and regulatory T cell (Treg) response genes, has not been thoroughly investigated in the development of gastric cancer. Therefore, we evaluated how H. pylori virulence factors influence the expression levels of immune-related genes in the development of gastric immunopathology. MethodsA total of 92 gastric tissues of normal controls and patients with gastritis, gastric ulcer, and gastric cancer were examined for the expression of immune-checkpoint inhibitor genes (PD-1 PD-L1), Th1 (interferon- γ, IFN-γ), Th17 (interleukin- 17, IL-17, Retinoic-acid-receptor- related orphan nuclear receptor gamma t, RORγ-t), and Treg (Forkhead box P3, FOXP3) response genes with quantitative real-time PCR (qRT-PCR). Furthermore, correlation of H. pylori virulence factors’ (cytotoxin-associated gene A, cagA; vacuolating cytotoxin gene A, vacA (s1,s2,m1,m2); blood group antigen-binding adhesin gene A, babA, duodenal ulcer promoting gene A, dupA; the putative neuraminyllactose-binding hemagglutinin homolog, hpaA; neutrophil-activating protein A napA; outer inflammatory protein A, oipA; urease A, ureA; and urease B, ureB) genotypes with a degree of inflammation and density of H. pylori were investigated. Next, the relationship between H. pylori virulence factors and immune-checkpoint inhibitor genes, and T-cell response genes was evaluated. Eventually, a decision tree model was developed to determine the clinical outcome of patients using expression data. ResultsThe intensity of PD-1 and PD-L1 mRNA expression was increased significantly in gastric tissue of patients with gastric ulcer (PD-1: 2.3 fold, p=0.01; PD-L1: 2.1 fold, p=0.004), and gastric cancer (PD-1: 2 fold, p= 0.04; PD-L1: 1.8 fold, p=0.05) compared with control subjects. Also, PD-1: PD-L1 expression was significantly higher in patients with gastritis, who were infected with a marked density of H. pylori compared with its mildly infected counterparts. Furthermore, a novel negative correlation was found between PD-1 (r= -0.43) and PD-L1 (r= -0.42) with FOXP3 in patients with gastritis. CagA-positive H. pylori strain's negative association with PD-L1 expression (r=-0.34) was detected in patients with gastritis. Interestingly, PD-1 mRNA expression correlated positively with vacA s2/m2, in gastritis (r=0.43) and ulcer (r=0.43) patients. Furthermore, PD-1: PDL1 expression negatively correlated with vacA m1/m2 (r=-0.43 for PD-1; r=-0.38 for PD-L1) in gastritis patients. Moreover, an inverse correlation of PDL1 was present with vacA m1 (r=0.52) and vacA s1/m1 (r=0.46) versus vacA m2 (r=-0.44) and vacA m1 (r=0.52) and vacA s1/m2 (r=-0.14) in ulcer patients, respectively. Also, a correlation of vacA m2 (r=-0.47) and vacA s1/s2 (r= 0.45) with PD-1 was detected in ulcer patients. In addition, a novel negative correlation between FOXP3 mRNA levels and napA was shown in patients with gastritis and ulcer (r=-0.59). Finally, a computer-based model that was developed showed that knowing the expression levels of PD-L1, RORγ-t, and vacA s1/m2 would be useful to detect the clinical outcome of a patient. ConclusionOur results suggested that PD-1:PD-L1 immune checkpoint inhibitors were increased in gastric pre-cancerous lesions that progress to gastric cancer. Herein, we report the relationship between H. pylori virulence factors and expression of host immune checkpoint inhibitors for diagnostic prediction of gastric malignancies using computer-based models.

Effect of intraoperative blood transfusion on Treg and FOXP3 in patients with digestive tract malignancies and different ABO blood types

BackgroundBlood transfusion can cause immunosuppression and lead to worse outcomes in patients with digestive tract malignancies; however, the specific mechanism behind this is not completely understood. One theory is that increased numbers of regulatory CD3+CD4+CD25+FOXP3+ T cells (Tregs) and forkhead box protein-3 mRNA (FOXP3) expression in the blood after transfusion contribute to these outcomes. The effect of blood transfusion on immune function in patients with different ABO blood types is variable. This study investigates the effect of intraoperative blood transfusion on the number of Tregs and the expression of FOXP3 in the blood of patients with different ABO blood types and digestive tract malignancies.MethodsPatients with digestive tract malignancies who underwent radical resection and received intraoperative blood transfusion were divided into four groups according to their blood types:blood group A, blood group B, blood group O and blood group AB (n = 20 for each group). Blood was collected from all patients before surgery, immediately after transfusion, 1 day after transfusion, and 5 days after transfusion. The number of Tregs was measured by flow cytometry. The expression of FOXP3 was detected by real time reverse transcription polymerase chain reaction (RT-PCR).ResultsThere was no significant difference in the number of Tregs or expression of FOXP3 mRNA among patients with different blood types before surgery. However, the number of Tregs and the expression of FOXP3 increased after blood transfusion in all blood type groups. This increase was especially evident and statistically significant on the first day after blood transfusion when compared with measures obtained before the surgery. Measures returned to the preoperative level five days after surgery. There were significant differences in the increase of Tregs and expression of FOXP3 among patients with different blood types. The greatest increase was seen in patients with blood group B and the least in blood group A.ConclusionsIntraoperative blood transfusion can lead to an increase in blood Tregs and FOXP3 expression in patients with digestive tract malignancies. Increases were greatest on the first day after surgery and differed among patients with different blood types. Increases were greatest in blood type B and least in blood type A.

Value of human mitochondrial transcription termination factor 3 and forkhead box protein 3 in predicting the prognosis of non-small cell lung cancer

Objective To explore the prognostic value of human mitochondrial transcription termination factor 3 (hMTERF3) and forkhead box protein 3 (Foxp3) in non-small cell lung cancer (NSCLC). Methods The clinical data of 88 patients with NSCLC who were admitted to the Third Medical Center of PLA General Hospital from March 2017 to March 2018 were retrospectively analyzed. All patients were diagnosed by pathological puncture. The patients were followed-up by telephone for 12 months, and according to the prognosis, the patients were divided into good prognosis group and poor prognosis group. The pathological tissues were taken from all patients, and the expressions of hMTERF3 and Foxp3 proteins were detected by immunohistochemistry. The expressions of hMTERF3 and Foxp3 in the good prognosis group and the poor prognosis group were compared. Logistic regression model was used to analyze the risk factors of poor prognosis in patients with NSCLC. Results Of 88 patients, 61 patients (69.3%) had good prognosis and 27 patients (30.7%) had poor prognosis. The positive expression rate of hMTERF3 in the good prognosis group was 57.4% (35/61), which was significantly lower than that in the poor prognosis group (81.5%, 22/27) (χ 2= 4.766, P= 0.029). The positive expression rate of Foxp3 in the good prognosis group was 55.7% (34/61), which was significantly lower than that in the poor prognosis group (85.2%, 23/27) (χ 2= 7.113, P= 0.008). The proportions of patients with medium and high differentiation or stage Ⅰ- Ⅱ in the good prognosis group were 82.0% (50/61) and 68.8% (42/61), respectively, which were significantly higher than those in the poor prognosis group [48.15% (13/27) and 25.93% (7/27)] (both P < 0.05). Logistic regression analysis showed that the poor differentiation, stage Ⅲ-Ⅳ, hMTERF3-positive and Foxp3-positive were the risk factors for poor prognosis in NSCLC patients (all P < 0.05). Conclusions The positive expression rates of hMTERF3 and Foxp3 in patients with good prognosis are lower. The hMTERF3-positive and Foxp3-positive are risk factors for poor prognosis in NSCLC patients. Key words: Carcinoma, non-small-cell lung; Mitochondrial proteins; Forkhead transcription factors; Human mitochondrial transcription termination factor 3; Forkhead box protein 3; Prognosis

Four SNPs and Systemic Level of FOXP3 in Smokers and Patients with Chronic Obstructive Pulmonary Disease

ABSTRACTForkhead box P3 (FOXP3) is the essential transcription factor for the function of regulatory T-cell (Treg). However, the gene mutation of FOXP3 in patients with chronic obstructive pulmonary disease (COPD) at different stages has not been reported. We aim to investigate four single nucleotide polymorphisms (SNPs) and the mRNA expression of FOXP3 in smokers with normal lung function and smokers with COPD at different stages. FOXP3 mRNA expression and SNPs in FOXP3 were assessed in nonsmokers with normal lung function (N), smokers with normal lung function (S), smokers with COPD in the Global Initiative for Chronic Obstructive Lung Disease (GOLD) 1 or 2 grade (COPD 1–2), and smokers with COPD in GOLD 3 or 4 grade (COPD 3–4). In peripheral blood sample, FOXP3 mRNA was assessed using real-time quantitative PCR and SNPs were analyzed by TaqMan PCR. FOXP3 mRNA level in peripheral blood sample was decreased when COPD was aggravated. The frequency of FOXP3 rs5902434 genotype del/del and allele del are lower in COPD 1–2 and COPD 3–4 than that in N or S. The rs5902434 genotype del/del and allele del were, respectively, associated with decreased risk of COPD and lung function decline. The rs5902434 genotypic distribution was correlated with FOXP3 mRNA level. In conclusion, both FOXP3 rs5902434 genotypes and alleles were differently distributed in COPD patients and smokers with normal lung function. The distribution of del/del genotype was associated with systemic expression of FOXP3 mRNA. More research is needed to explore the role of FOXP3 gene polymorphism in immunoinflammation of COPD.

Abnormalities in the T cell receptor Vδ repertoire and Foxp3 expression in refractory anemia with ringed sideroblasts.

Immune dysregulation has been implicated in myelodysplastic syndrome (MDS) pathogenesis. Refractory anemia with ringed sideroblasts (RARS) is a low-risk subtype of MDS. Our previous study has reported an abnormal γδ T cell receptor (TCR) repertoire in patients with intermediate or high-risk MDS. To characterize the status of T cell immunity in RARS, we investigated the distribution and clonality of the TCR Vδ repertoire and the expression of Foxp3 in patients with RARS. The number of expressed Vδ subfamily members in the RARS group (4.8±2.25) was significantly lower than that in the control group (7.6±0.52, p=0.0012). A significantly lower expression frequency for the Vδ4 (p=0.007), Vδ5 (p=0.0049) and Vδ7 subfamilies (p=0.0225) could be detected in the RARS group. The most frequent clonally expanded T cell subfamily member in the RARS group was Vδ7 (100%, 3/3). Foxp3 mRNA expression was significantly lower and higher than that in the controls in 60% and 40% RARS patients, respectively. In conclusion, marked restriction of the TCR Vδ subfamily expression pattern and great heterogeneity in Foxp3 expression were characteristics found in RARS. These results provide data regarding the immunodeficiency and immune reactive characteristics of patients with RARS, which may provide a basis for immunotherapy options.

THU0555 Immunopathological Roles of the Novel Anti-Inflammatory Cytokine Interleukin-35 in Patients with Systemic Lupus Erythematosus

BackgroundIL-35, a dimeric protein with two subunits, IL-12A (p35) and Epstein-Barr virus induced 3, is a novel IL-12 cytokine family regulatory T-cells (Treg)-specific immunosuppressive/anti-inflammatory cytokine. IL-35 is expressed by resting...

The expression and significance of CD4+CD25+CD127low/- regulatory T cells and Foxp3 in patients with portal hypertension and hypersplenism.

To study the expression of CD4+CD25+CD127low/- regulatory T cells and Foxp3 in patients with portal hypertension (PH) and hypersplenism, and explore the significance of Treg in immunomodulation of hypersplenism. Testing of CD4+CD25+CD127low/- regulatory T cells and CD3+CD4+CD8+ T cells in peripheral venous blood with flow cytometry in 20 patients with PH and hypersplenism; testing for Foxp3 in spleen tissue of 30 patients with PH and hypersplenism with immunohistochemistry. The percentage of CD4+CD25+CD127low/-/CD4+ in patients with PH and hypersplenism was 5.3%+-3.0%. It was obviously increased compared with normal control samples 2.5%+-0.9%, with significant difference (p=0.001). There was a negative correlation between CD4+CD25+CD127low/-/CD4+ rate and CD3+CD4+ (r=-0.630, p=0.003; r=-0.561, p=0.01). The spleen tissue Foxp3 expression (IOD=293.1+-180.0) in patients with PH and hypersplenism were markedly improved compared with the normal expressions of spleen-cutting groups (IOD=115.2+-84.1), the difference was significant (p<0.01). The percentage of CD4+CD25+CD127low/- Treg and Foxp3 in patients with PH and hypersplenism was significantly increased, suggesting that they may take part in the regulation of immune function and be related to the change of T lymphocyte subsets in patients with PH and hypersplenism, which may have an important clinical significance.

IPEX Syndrome, FOXP3 and Cancer.

In this review, we introduce the IPEX syndrome and its relationship with germline mutations of the FOXP3 gene. We then describe the multiple functional roles of FOXP3 in regulatory T cells and epithelial cells as well as in IPEX syndrome and tumor progression. Potential mechanisms of FOXP3 inactivation and transcriptional regulation are discussed with recent advances. Finally, we point out current issues and a potential FOXP3-mediated therapeutic strategy as well as the reactivation of FOXP3 in patients with IPEX syndrome and cancer.

Abstract 3651: Foxp3 expression correlates with survival in urothelial carcinoma of the bladder

Abstract Introduction and Objective The transcription factor Forkhead box p3 (Foxp3), originally described during the development of CD4+CD25+ T regulatory cells, has recently been implicated in divergent roles in human prostate, breast, pancreatic cancer, and melanoma. Here, we investigate the inducibility of Foxp3 in bladder cancer models and assess the prognostic role of Foxp3 in patients with bladder cancer. Methods Foxp3 expression in the human bladder cancer cell line SW780 was examined in in vitro cultures by flow cytometry and in in vivo subcutaneous implanted tumor models in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice by immunohistochemistry. Proliferation and invasion of SW780 cells in the absence of Foxp3 was performed by transfection of siRNA in vitro. Correlation of Foxp3 expression to urothelial carcinoma grade, stage, recurrence-free survival, and overall survival was examined in a human tumor microarray from a cohort of 280 patients by Cox proportional hazards regression and Kaplan-Meier analysis. Results Foxp3 expression correlates to higher cellular proliferation in vitro and in vivo. Knockdown of Foxp3 in SW780 cells resulted in decreased proliferation as well as invasion in vitro. Foxp3 expression increases with higher T stage, but not grade, as expression in adjacent normal bladder, Tis, Ta-1, T2-4, and lymph node metastasis was 1.2%, 3.8%, 15.2%, 26.5%, and 23.4% respectively, suggesting higher expression in muscle invasive and lymph node metastatic disease, but not carcinoma in situ. Using a cutoff of 10%, Foxp3 expression negatively correlates with survival following radical cystectomy with a median survival of 9.3 vs 2.8 years and a 10-year survival probability of 47% vs 27%. Conclusions This work implicates Foxp3 as novel transcription factor driving proliferation and tumor invasion in urothelial carcinoma. Expression of Foxp3 correlates with T stage and is a novel predictor of overall survival following radical cystectomy. Future studies will need to understand the mechanistic regulation of Foxp3 expression, as well examine Foxp3-mediated gene expression in urothelial carcinoma. Foxp3 expression may improve selection of patients to therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3651. doi:1538-7445.AM2012-3651

The Regulation of FOXP3 Expression by the Treatment of TGF-β and the Modification of DNA Methylation in Lung Cancer Cell Lines

Background: Transcription factor FOXP3 characterizes the thymically derived regulatory T cells. FOXP3 is expressed by cancer cell itself and FOXP3 expression was induced by TGF-β treatment in pancreatic cancer cell line. However, the expression of FOXP3 expression is not well known in patients with lung cancer. This study was conducted to investigate the expression of FOXP3 in patients with lung cancer and to investigate the regulation of FOXP3 expression by the treatment of TGF- β and DNA methyltransferase inhibitor in lung cancer cell lines. Methods: FOXP3 expression in the tissue of patients with resected non-small cell lung cancer (NSCLC) was evaluated by immunohistochemistry. The regulation of FOXP3 expression was investigated by Western blot and RT-PCR after lung cancer cell lines were stimulated with TGF-β1 and TGF-β2. The regulation of FOXP3 expression was also investigated by RT-PCR and flow cytometry after lung c ancer cell lines were treated with DNA methyltransferase inhibitor (5-AZA-dC). Results: FOXP3 expression was confirmed in 27% of patients with NSCLC. In NCI-H460 cell line, TGF-β2 decreased FOXP3 mRNA and protein expressions. In A549 cell line, both TGF-β1 and TGF-β2 decreased FOXP3 mRNA and protein expressions. 5-AZA-dC increased FOXP3 mRNA expression in NCI-H460 and A549 cell lines. Moreover, 5-AZA-dC increased intracellular FOXP3 protein expression in A549 cell lines. Conclusion: It was shown that FOXP3 is expressed by cancer cell itself in patients with NSCLC. Treatment of TGF- β2 and DNA methyltransferase inhibitor seems to be associated with the regulation of FOXP3 expression in lung cancer cell lines.

CD4+CD25+ Regulatory T Cells in Patients with Acute Lymphocytic Leukemia.

Human CD4+CD25+ regulatory T (Treg) cells suppress antigen-specific T cell immune response and contribute to peripheral tolerance. However, little is known about the role of Treg cells in the pathogenesis of acute lymphocytic leukemia (ALL). In this study, the proportion of CD4+CD25+ Treg cells in the peripheral blood of three groups of people [the patients with ALL who had never received any chemotherapy before, the patients with ALL who had achieved partial remission (PR) or complete remission (CR) after chemotherapy and the healthy donors] was evaluated by flow cytometry. The expression level of Foxp3 mRNA of each group was examined by real-time RT-PCR. In vitro, the peripheral blood mononuclear cells (PBMC) from healthy donors were cultured in the serum of ALL patients without chemotherapy; each group was divided into sub-groups according to various serum doses. After co-cultured for 72 hours, the cells of all the groups were harvested separately and further tested the number of CD4+CD25+ Treg cells and the mRNA expression of Foxp3. The results showed that both the percentage of CD4+CD25+ T cells and the mRNA expression level of Foxp3 in patients with chemotherapy were significantly higher than those of the healthy donors and patients without chemotherapy (P<0.01 and P<0.01). Although the population of CD4+CD25+ T cells in ALL patients without chemotherapy was almost the same to that in the healthy donors (P>0.05), the expression of Foxp3 mRNA in former was higher (P<0.05) .The serum derived from ALL patients without chemotherapy remarkably increased the proportion of CD4+CD25+ T cells and the expression level of Foxp3 mRNA in co-cultured system, both of which were statistically higher than those in controls (P<0.05 and P<0.01) and did not vary with the serum dose. These results suggest that the chemotherapy may exert an influence on the production of CD4+CD25+ Treg cells and CD4+CD25+ Treg cells may play an important role in the pathogenesis of ALL.

High Proportions of CD4+CD25hi T Cells Pretransplant Correlate with Worse Overall Survival after Stem Cell Transplant.

Regulatory cells may have a modulatory effect on alloreactive phenomena following hematopoietic stem cell transplantation (HSCT), although their role in this setting in patients remains controversial. We have analysed the effect of pre-conditioning peripheral blood levels of T-regs on the outcome of 89 adult patients undergoing HLA-identical (10/10) unrelated donor (UD)-HSCT. Allografts were T-cell depleted with Alemtuzumab, which has been described to spare T-regs from, and we found that higher proportion of CD4+CD25hi T-cells correlated with worse survival (p=0.002). This higher proportion also correlated with higher incidence of relapse (p=0.0274) and with higher incidence of chronic (c) GvHD (p=0.033) but not with the incidence of aGvHD.Regulatory T cells (T-regs) are a naturally occurring regulatory population of CD4+CD25hi T-cells known to express the transcription repressor FoxP3 and to produce anti-inflammatory cytokines such as TGFβ and may therefore affect patient survival. We analyse the profile of the CD4+CD25hiT cells in our cohort. In our healthy donor control group (n=30), CD4+CD25hi-expression associated with both, FoxP3 mRNA-levels (r=0.649; p=0.001) and TGFβ production (r=0.912; p<0.001), as previously described for T-regs. In preconditioning patient samples however, CD4+CD25hi-expression had a strong correlation with TGFβ regulatory cytokine production (r=0.863; p<0.001) as expected. However, FoxP3 mRNA-expression correlated neither with the CD4+CD25hi T-regs phenotype (r=0.280; p=0.040), nor with the production of TGFβ (r=0.229; p=0.156), and appeared not be an accurate marker for regulatory T-cell function in this patient setting. In addition, an inverse correlation with TNFα expression (r=−0.458 p<0.001) was found that may argue that the CD4+CD25hi cells represent a true regulatory population and not activated cells. This suggests either that activated cells that are not expressing FOXP3 might be included in the CD4+CD25hi population or that CD4+CD25loFOXP3+ cells have not acquire the regulatory function (i.e. they are not expressing TGFβ).In summary, CD4+CD25hi Regulatory T cells may have an impact suppressing allo-responses against the tumour, decreasing the overall survival by increasing the rates of relapse.Although, in mice CD4+CD25hi population have been clearly identified by the expression of the FOXP3, which encodes a transcription repressor, in humans this remains controversial, where FOXP3 expression is not exclusive of the CD4+CD25hi population.Relying on the expression of CD4+CD25hi and FOXP3 in patients is insufficient to determine regulatory T cell numbers and suggests that other parameters of regulatory function should be taken in to account.