FOXP3 variants are independently associated with transforming growth factor B1 plasma levels in female patients with inflammatory bowel disease

Abstract Objective: The aim of this study was to evaluate the association of -924 G>A (rs2232365) and -3279 C>A (rs3761548) FOXP3 variants with IBD susceptibility, clinical and endoscopic activity, and IL-10 and TGF-β1 plasma levels. Method: The study included 110 IBD female patients, 60 with Ulcerative Colitis (UC) and 50 with Crohn's Disease (CD), and 154 female controls. FOXP3 variants were determined with Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP). Plasma levels of IL-10 and TGF-β1 were determined using immuno-fluorimetric assay. Results: AA genotype of rs2232365 and rs3761548 was associated with CD (OR = 3.147, 95% CI 1.015-9.758, p = 0.047) and UC (OR = 3.221, 95% CI 1.050-9.876, p = 0.041) susceptibility, respectively. However, were not associated with TGF-β1 and IL-10 levels, and endoscopic/clinical activity disease. GAGA haplotype was associated with IBD (OR = 4.003, 95% CI 1.100-14.56, p = 0.035) and UC susceptibility (OR = 6.107, 95% CI 1.609-23.18, p = 0.008). In addition, IBD patients with the GAGA haplotype had lower TGF-β1 levels (p = 0.041). Moreover, G/C haplotype (dominant model) had a protective effect of 60% in CD susceptibility and lower Endoscopic Severity Index. Conclusions: These results suggest that FOXP3 variants could exert a role in the Treg, which could be one of the factors involved in the susceptibility and pathogenesis of IBD.

Value of human mitochondrial transcription termination factor 3 and forkhead box protein 3 in predicting the prognosis of non-small cell lung cancer / 肿瘤研究与临床

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.

Methylation of forkhead box protein 3 gene promoter in CD4+ T cells in patients with chronic hepatitis B / 中华实验和临床病毒学杂志

Objective@#To investigate the methylation status and expression of FOXP3 in CD4+ T cells of patients with chronic hepatitis B (CHB).@*Methods@#Peripheral blood mononuclear cells (PBMCs) from 59 CHB patients and 22 healthy controls (HC) were collected. The percentage of CD4+ CD25+ Foxp3+ Tregs in CD4+ T cells was estimated by flow cytometry. FOXP3 expression was measured by quantitative real time-polymerase chain reaction (RT-qPCR) and Western blotting. The methylation status of FOXP3 was determined by methylation-specific polymerase chain reaction.@*Results@#The percentage of CD4+ CD25+ pFoxp3+ Tregs in CD4+ T cells, FOXP3 mRNA and protein expression levels were significantly higher in patients with CHB than HCs (P<0.05). Meanwhile, the methylation frequency of FOXP3 was significantly lower in CHB patients than HCs (P<0.05). FOXP3 mRNA levels and the percentage of CD4+ CD25+ Foxp3+ Tregs were significantly lower (P<0.05) in patients with gene methylation than those without.@*Conclusions@#Aberrant demethylation of FOXP3 gene existed in CD4+ T cells of CHB, which contributed to an elevation in FOXP3 expression and percentage of CD4+ CD25+ Foxp3+ Tregs. It might provide a new target for prevention and treatment of CHB.

Effects of SMYD3 and MLL5 on histone methylation of Foxp3 gene in children with Kawasaki disease / 中华风湿病学杂志

Objective To investigate the effects of SMYD3 and MLL5 on histone methylation of Transcription factor forkhead box protein 3 (Foxp3) gene and its roles in the immunological pathogenesis of Kawasaki disease (KD). Methods Forty-two children with KD and 26 age-matched healthy children were consented to participate in this study. Co-Immunoprec-ipitation and real-time polymerase chain reaction (PCR) was performed to determine Foxp3-associated histone methylation levels of H3K4me3 and H3K27me3, and binding levels of SMYD3 and MLL5 with Foxp3 gene in CD4+T cells. The proportion of CD4+CD25high Foxp3+cells (Treg) and protein levels of Foxp3, cytotoxic T lymphocyte associated antigen-4 (CTLA4), TGF-βRⅡand pSmad3 were analyzed by flow cytometry. Quantitative real-time PCR was used to evaluate levels of Foxp3, interleukin (IL)-10, GITR, TGF-βRⅠand RARαmRNA in CD4+T cells. Plasma concentrations of TGF-βand retinol acid (RA) were measured by enzyme-linked Immunosorbent assay. Independent-samples t-test was used as the statistical method in this study. Results ① The proportion of Treg, expression levels of Foxp3 and molecules associated with suppressive function of Treg cells(IL-10, GITR and CTLA4), and histone methylation levels of H3K4me3 associating with promoter, conserved non-coding DNA sequence (CNS) 1 and CNS2 of Foxp3 gene decreased remarkably during acute KD [Promoter:(5.4±1.8)%vs (9.1±2.2)%;CNS1:(2.6±0.9)% vs (3.8±1.1)%; CNS2: (2.4±0.8)% vs (4.2±1.0)%; t=5.50, 6.02, 9.56, 7.92, 7.97, 4.76, 7.73, 5.01, 8.66; P0.05). ② Binding levels of SMYD3 and MLL5 with Foxp3 gene in CD4+T cells were down-regulated significantly during acute KD (t=6.63, 6.15; P<0.05), and restored to some extent after IVIG treatment (t=5.36, 4.56; P<0.05). Positive correlations between binding levels of SMYD3 and MLL5 and expression level of Foxp3 mRNA were detected in patients with acute KD (r=0.62、0.45, P<0.05). Furthermore, Binding levels of SMYD3 and MLL5 with Foxp3 gene in KD-CAL+group were lower than those in KD-CAL- group (t=4.11, 4.31; P<0.05). ③ Compared with healthy controls, plasma concentration of TGF-β and RA, and expressions of TGF-βRⅡ, TGF-βRⅠ, pSmad3 and RARα were down-regulated during acute KD (t=11.54, 12.81, 7.43, 16.10, 8.25, 12.06; P<0.05), and elevated remarkably after IVIG treatment (t=8.40, 6.24, 5.94, 11.78, 6.27, 8.30; P<0.05). Simultaneously, all the items aforementioned in KD-CAL+ group were found to be lower than those in KD-CAL-group (t=3.58, 3.30, 3.82, 5.27, 4.71, 3.78; P<0.05). Conclusion Hypomethylation of H3K4me3 associated with Foxp3 gene caused by insufficient binding levels of SMYD3/MLL5 may be involved with immune dysfunction in Kawasaki disease.

Study of NO and FOXP3 in patients with pulmonary tuberculosis / 国际检验医学杂志

Objective To Study the expressions and significance of nitric oxide (NO) and forkhead box protein 3 (FOXP3) in patients with pulmonary tuberculosis .Methods Serum NO levels were measured by Griess colorimetric reaction .Flow cytometry was used to determine the number of CD4+CD25+ T cells in peripheral blood .The expression of FOXP3 mRNA was measured by real‐time polymerase chain reaction .Results Serum NO level in patients was (15 .71 ± 1 .26)μmol/L ,higher than the (5 .45 ± 0 .98)μmol/L of healthy controls (P<0 .05) .CD4+ CD25+ T cells comprised (4 .57 ± 0 .85)% of CD4+ T cells in patients ,higher than the (1 .83 ± 0 .49)% in healthy controls (P<0 .05) .CD4+ CD25+ T cells in the peripheral blood of patients with pulmonary tuberculosis highly expressed FOXP3 .Conclusion Patients with pulmonary tuberculosis could be with an increased level of NO and FOXP3 ,which might have important role in the pathogenesis of pulmonary tuberculosis .

Expression of TIM-3, Human beta-defensin-2, and FOXP3 and Correlation with Disease Activity in Pediatric Crohn's Disease with Infliximab Therapy

BACKGROUND/AIMS: This study investigated the expression of T cell immunoglobulin- and mucin-domain-containing molecule 3 (TIM-3), human beta-defensin (HBD)-2, forkhead box protein 3 (FOXP3), and the frequency of CD4+ CD25+ FOXP3+ regulatory T cells (Tregs) in children with Crohn's disease (CD) during infliximab therapy. METHODS: We enrolled 20 CD patients who received infliximab treatment for 1 year. Peripheral blood and colonic mucosal specimens were collected from all CD patients and from healthy control individuals. RESULTS: A significant difference in TIM-3 mRNA expression was evident in peripheral blood mononuclear cells and colonic mucosa between CD patients before infliximab therapy and the healthy controls (p<0.001 and p=0.005, respectively). A significant difference in HBD-2 mRNA expression was found in colonic mucosa between CD patients before infliximab therapy and the healthy controls (p=0.013). In the active phase of CD, at baseline, the median percentage of T cells that were CD25+ FOXP3+ was 1.5% (range, 0.32% to 3.49%), which increased after inflixmab treatment for 1 year to 2.2% (range, 0.54% to 5.02%) (p=0.008). CONCLUSIONS: Our study suggests that both the adaptive and innate immune systems are closely linked to each other in CD pathogenesis. And the results of our study indicate that it could be a useful therapeutic tool, where restoration of TIM-3, HBD-2 and the function of Tregs may repair the dysfunctional immunoregulation in CD.

Role of FOXP3 in tumor immunity / 第二军医大学学报

FOXP3, a member of the forkhead/winged-helix family, exerts its immunosupression effect by regulating the development, differentiation and maturation of CD4+ CD25+ regulatory T cells (Treg). Treg plays an important role in immune escape mechanism in oncology research. This article reviews the expression of FOXP3 in various types of tumors and its role in tumor immunology and immunotherapy.

Effect of ionizing radiation on expressions of regulatory T cells and related molecules in mice spleen / 中华放射医学与防护杂志

Objective To observe the changes in expressions of spleen regulatory T cells (Tregs)and the related factor forkhead box protein-3 (Foxp3) after irradiation with different doses of X-ray in mice at different times,and to elaborate the effects of X-rays on regulatory T cells and Foxp3.Methods 112male ICR mice were randomly divided into 2 groups and irradiated by X-rays at the doses of 0.075 and 2 Gy,respectively.The mice were killed at 0,4,8,16,24,48,and 72 h post-irradiation and the spleens removed.Flow cytometry was used to detect the percentage of CD4 + CD25 + Treg and protein expression of (Foxp3),and RT-PCR was used to exmiamine the mRNA expression of Fox3.Results Compared with those before irradiation,the CD4 + CD25 + Treg positive rates began to increase and peaked at 8 h post-irradiation with 0.075 Gy at 8,16,24,72 h(t = 8.73,10.55,4.21,4.65 ,P ＜ 0.05) and 2 Gy at 8,16,48,72 h(t = 4.65,4.28,3.71,2.88,P ＜ 0.05),and then slightly decreased,but still remained at high levels.The mRNA protein levels of Fox3 did not change significantly after exposure to the dose of 0.075 Gy,but began to significantly increase at 8 h after exposure to the dose of 2 Gy.However,the Foxp3 protein level began to increase 4 h post-irradiation,peaked at 16 h,and then slightly decreased,but still ramained at high levels (t =2.59,3.37,3.70,3.20,P＜0.05).Conclusions The changes in expressions of Tregs and Foxp3 after high- and low-dose X-ray irradiation may be used to explain the differences in immune effects induced by ionizing radiation at different doses.

Expression of regulatory T cells and helper T cells in human IgA nephropathy and its significance / 中华肾脏病杂志

Objective To investigate the effects of CD4+CD25high regulatory T cells(Treg)and the imbalance of helper T lymphocyte subsets(Th1/Th2)on the immunological mechanism of IgA nephropathy(IgAN)patients. Methods The percentage of Treg and helper T cells subpopulation (Th1/Th2)in the peripheral blood of IgAN patients and healthy controls was examined by flow cytometry.The FOXP3 expression was detected through intracellular staining.The correlation of Treg or Th1/Th2 with clinical parameters of IgAN was analyzed by Spearman or Pearson rank correlation test. Results The percentages of Treg and Th2 cells were significantly higher in peripheral blood of IgAN patients compared to that of healthy controls[Treg (2.14±0.82)%vs[1.59±0.53)%,Th2(2.57±0.72)%vs(1.81±1.10)%,all P＜0.05].Th1/Th2 ratio was significantly reduced in IgAN patients(5.75±1.89 vs 12.73±9.79,P＜0.05).The percentage of circulating Treg cells was positively correlated with serunl IgA concentration(r=0.397,P＜0.05),and was negatively correlated with eGFR(r=-0.376,P＜0.05).The percentage of circulating Th2 cells was positively correlated with serum IgA(r=0.468,P＜0.05). Conclusions There is a disorder of T lymphocyte population in the peripheral blood of IgAN patients.The increased Treg and Th2 cells may play an important role in the pathogenesis of IgAN.

Role of FOXP3 in tumor immunity / 第二军医大学学报

FOXP3,a member of the forkhead/winged-helix family,exerts its immunosupression effect by regulating the development,differentiation and maturation of CD4+ CD25+ regulatory T cells(Treg).Treg plays an important role in immune escape mechanism in oncology research.This article reviews the expression of FOXP3 in various types of tumors and its role in tumor immunology and immunotherapy.