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ObjectiveTo investigate the effect of different doses of Jiedu Tongluo Shengjin prescription （JTSP） on serum interleukin-6 （IL-6）， tumor necrosis factor-α （TNF-α） and forkhead box P3 （FoxP3） in submandibular gland of NOD/Ltj mice with Sjögren's syndrome， and to explore the mechanism of JTSP on immune regulation in NOD/Ltj mice. MethodThirty NOD/Ltj mice （eight weeks old） were randomly divided into model group， JTSP low-dose group， JTSP medium-dose group， JTSP high-dose group and hydroxychloroquine group， and were administrated with normal saline， JTSP 9， 18， and 36 g·kg-1， and hydroxychloroquine 60 mg·kg-1 daily， respectively from the age of 12 weeks. Six ICR mice were given an equal amount of normal saline by gavage as the control group. During the experiment， daily water consumption and saliva secretion of mice at the age of 9， 12， 16 weeks were recorded. After 4 weeks of administration， submandibular gland and spleen tissues were dissected to calculate corresponding indexes. The pathological morphology of submandibular gland was observed by hematoxylin-eosin （HE） staining. Meso Scale Discovery （MSD） and immunohistochemistry were employed to detect the serum levels of IL-6， TNF-α and IL-10， and the expression and distribution of FoxP3 in submandibular gland， respectively. The protein expression of FoxP3 in mouse submandibular gland was determined by Western blot， and the mRNA expressions of FoxP3 and TNF-α were determined by real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the control group， the model group presented increased daily water consumption， decreased saliva secretion， lowered submandibular gland index， elevated pathological score of submandibular gland， up-regulated serum IL-6 and TNF-α and mRNA expression of TNF-α while down-regulated serum IL-10 and protein and mRNA expressions of FoxP3 in submandibular gland （P<0.05）. Compared with the conditions in model group， daily water consumption in JTSP groups was reduced while saliva secretion was increased， especially in medium-dose and high-dose groups （P<0.05）， and there was an increase in the submandibular gland index of JTSP medium-dose group （P<0.05） while a decrease in the spleen index of JTSP high-dose group （P<0.05）. Additionally， JTSP groups had lower pathological score of submandibular gland than the model group （P<0.05）， especially high-dose group， as well as lower serum IL-6 and TNF-α and mRNA expression of TNF-α while higher serum IL-10 （P<0.05）. JTSP at medium and high doses up-regulated the protein and mRNA expressions of FoxP3 in submandibular gland （P<0.05）. ConclusionJTSP may inhibit the secretion of inflammatory cytokines by regulating the stability of FoxP3+ regulatory T （Treg） cells， thus alleviating the systemic immune inflammation in Sjögren's syndrome.
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OBJECTIVES@#This study aimed to clarify the effects of Foxp3 silencing on the expression of inflammatory cytokines in human periodontal ligament cells (hPDLFs) in an inflammatory environment and on cell proliferation and invasiveness, as well as to explore the role of Foxp3 gene in the development of periodontitis.@*METHODS@#An small interfering RNA (siRNA) construct specific for Foxp3 was transfected into hPDLFs. Foxp3 silencing efficiency was verified by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, and the siRNA with the optimum silencing effect of Foxp3 gene was screened. Using lipopolysaccharide to simulate an inflammatory environment in vitro, CCK-8 detected the effect of silencing Foxp3 on hPDLFs proliferation under inflammatory conditions. Wound-healing experiments and transwell assays were conducted to detect the effect of silencing Foxp3 on hPDLF migration under inflammatory conditions. The expression of the inflammatory cytokines interleukin (IL)-6 and IL-8 was detected by RT-PCR and Western blotting under inflammatory conditions.@*RESULTS@#After siRNA transfection, RT-PCR and Western blotting analyses showed that the expression of Foxp3 mRNA in the Foxp3-si3 group decreased significantly (t=21.03, P<0.000 1), and the protein expression of Foxp3 also decreased significantly (t=12.8, P<0.001). In the inflammatory environment, Foxp3 gene silencing had no significant effect on hPDLFs proliferation (P>0.05), and Foxp3 gene silencing promoted hPDLFs migration (P<0.05). Moreover, the expression of IL-6 and IL-8 increased (P<0.05).@*CONCLUSIONS@#In an inflammatory environment, Foxp3 gene silencing promoted hPDLFs migration but had no significant effect on hPDLFs proliferation. The expression of inflammatory factors expressed in hPDLFs increased after Foxp3 gene silencing, indicating that Foxp3 gene inhibited inflammation in periodontitis.
Subject(s)
Humans , Cell Proliferation/genetics , Cells, Cultured , Cytokines/metabolism , Fibroblasts/metabolism , Forkhead Transcription Factors/metabolism , Gene Silencing , Interleukin-6/metabolism , Interleukin-8/metabolism , Periodontal Ligament/metabolism , Periodontitis/metabolism , RNA, Small Interfering/metabolism , Transcription Factors/metabolismABSTRACT
@#[摘 要] 目的：探讨PD-1/PD-L1通路及相关免疫细胞在宫颈鳞癌（cervical squamous cell cancer，CSCC）发生、发展中的变化特点及其临床意义。方法：收集2018年12月至2020年9月在福州市第一医院接受手术的CSCC患者和健康体检人员的癌组织/宫颈组织和外周血样本，分为健康对照组、宫颈上皮内癌变（cervical intraepithelial neoplasia，CIN）Ⅱ级组、CIN Ⅲ级组和CSCC组，代表CSCC发生、发展进程各阶段，每组50例。ELISA法检测各组人员的外周血血浆中PD-1、PD-L1、叉头状转录因子P3（FOXP3）的表达水平，FCM法检测各组人员外周血PD-1+CD4+CD25+CD127-/low细胞的数量，应用多色荧光组织染色法检测肿瘤浸润性淋巴细胞（TIL）在CSCC组织中的分布特点。结果：随着模拟的CSCC发生和发展，外周血中PD-1、PD-L1和FOXP3 的表达呈上升趋势，术后则呈下降趋势。在CSCC患者抗凝全血中，CD4+、CD4+CD25+CD127-/low以及PD-1+CD4+CD25+CD127-/low细胞占淋巴细胞的比例增加。在CSCC组织中可见大量CD4+、CD8+和FOXP3+细胞浸润，其中CD4+和FOXP3+细胞主要围绕肿瘤细胞聚集区分布、CD8+和PD-L1+细胞则呈广泛弥漫性分布。结论：PD-1、PD-L1、FOXP3和适应性调节性T细胞是促进CSCC发生发展的重要因素，其可作为ESCC免疫治疗的靶点和临床预后的潜在标志物。
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Objective:To investigate the relationship between CD8 +FoxP3 +CD25 + T cell subsets and the therapeutic effect of programmed death receptor 1 (PD-1) inhibitor pembrolizumab in treatment of uterine cervical cancer. Methods:The data of 105 patients with uterine cervical cancer who received pemblizumab therapy based on chemotherapy in the First Hospital of Qinhuangdao from January 2018 to January 2020 were retrospectively analyzed. Flow cytometry was used to detect the ratio of CD8 +FoxP3 +CD25 + T cell in peripheral blood of patients. The efficacy and safety were analyzed. According to the efficacy, all patients were divided into remission group (complete remission + partial remission) and non-remission group (stable disease + progressive disease). The clinical characteristics and CD8 +FoxP3 +CD25 + T cell ratio of the two groups were compared. Multivariate logistic regression model was used to analyze the influencing factors for the efficacy. The efficacy of CD8 +FoxP3 +CD25 + T cell ratio predicting the therapeutic effect of patients was analyzed by using receiver operating characteristic (ROC) curve. Results:The objective remission rate of all patients was 17.14% (18/105), and the incidence of adverse reaction was 39.05% (41/105). The proportion of patients with a family history of cervical cancer in the remission group was lower than that than in the non-remission group [5.56% (1/18) vs. 34.48% (30/87)], and the difference was statistically significant ( χ2=6.00, P=0.014). The proportion of CD8 +FoxP3 +CD25 + T cell of 105 patients before and after treatment was (0.83±0.21)% and (0.77±0.10)%, respectively; the proportion of CD8 +FoxP3 +CD25 + T cell before and after treatment in the remission group was (0.55±0.26)%, (0.31±0.12)%, respectively; the proportion of CD8 +FoxP3 +CD25 + T cell before and after treatment in the non-remission group was (0.89±0.30)%, (0.87±0.28)%, respectively. The proportion of CD8 +FoxP3 +CD25 + T cell after treatment in the remission group was lower than that before treatment ( P < 0.05); there was no statistically significant difference in the proportion of CD8 +FoxP3 +CD25 + T cell before and after treatment in the non-remission group ( P>0.05). The proportion of CD8 +FoxP3 +CD25 + T cell before and after treatment in the non-remission group was higher than that in the remission group (all P<0.001). The proportion of CD8 +FoxP3 +CD25 + T cell higher than the mean value of both groups before treatment and the proportion of CD8 +FoxP3 +CD25 + T cell higher than the mean value of both groups after treatment were independent risk factor of disease remission ( OR=2.542, 95% CI 1.649-3.918, P<0.001; OR=2.936, 95% CI 2.154-4.002, P<0.001). ROC curve analysis showed that the area under the curve of CD8 +FoxP3 +CD25 + T cell ratio predicting the disease remission before treatment was 0.720, and its best cut-off value was 0.77%, the senfitivity was 77.78%, the specificity was 70.11%. Conclusions:Early detection of CD8 +FoxP3 +CD25 + T cell ratio helps to predict the effect of PD-1 inhibitor pembrolizumab therapy for uterine cervical cancer.
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Human T-cell lymphotropic virus type 1 (HTLV-1) is associated with the development of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). It has been reported that the HTLV-1 proteins (specifically TAX and HBZ) can modulate FOXp3, resulting in an immune imbalance that can favor the progression of HAM/TSP. This review aims to summarize the literature in order to clarify the relationship between the expression of HTLV-1 mRNAs and/or viral proteins (TAX and HBZ) with the expression of mRNA and/or protein FOXp3 and their correlation with HAM/TSP development. This systematic review was conducted according to the recommendations of the Preferred Reporting Items for Systematic Reviews and Meta-Analysis. The search strategy was performed on the Medical Literature Analysis and Retrieval System Online and Latin American and Caribbean Literature in Health Sciences Platform using subject descriptors. After screening, six articles were included in this review. The studies suggested that TAX and HBZ have a directly proportional correlation with FOXp3 in individuals with HAM/TSP, which also presented an increased expression of FOXp3 compared to asymptomatic controls and/or healthy donors. This systematic review indicates that TAX and HBZ can interact with FOXp3 and that interaction may influence HAM/TSP development.
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Objective:To investigate the effects of Notch1 signaling on histone acetylation of Foxp3 gene and its roles in regulating regulatory T (Treg) cells in children with acute B-cell precursor lymphoblastic leukemia (BCP-ALL).Methods:Blood samples were collected form 38 children with BCP-ALL before treatment and 15 age-matched healthy children (control group). Flow cytometry was performed to detect the proportion of peripheral blood CD4 + CD25 hiFoxp3 + Treg cells and the expression of Foxp3, cytotoxic lymphocyte antigen 4 (CTLA4), glucocorticoid-induced tumor necrosis factor receptor (GITR), CD39 and Notch1 at protein level. Histone 4 acetylation (H4Ac) at Foxp3 gene promoter and the binding abilities of Foxp3 gene promoter to NICD1 and p300 in CD4 + T cells were measured by chromatin immunoprecipitation. Quantitative real-time PCR was performed to detect the expression of Foxp3, presenilin 1 (PSEN1), mastermind-like transcriptional coactivator 1 (MAML1), SKI-interacting protein (SKIP), F-box and WD40 domain protein 7 (FBXW7), glycogen synthase kinase-3 beta (GSK3β) and IKAROS at mRNA level in CD4 + T cells. The concentrations of TGF-β and IL-10 in plasma were evaluated by ELISA. Results:(1) The proportion of peripheral blood CD4 + CD25 hiFoxp3 + Treg cells, the expression of differentiation- and function-associated molecules (Foxp3, CTLA4, GITR and CD39) and the concentrations of TGF-β and IL-10 in plasma were higher in the BCP-ALL group than in the control group ( P<0.05). (2) In children with acute BCP-ALL, H4Ac at Foxp3 promoter and the binding abilities of Foxp3 gene promoter to NICD1 and p300 were significantly increased as compared with those in control group( P<0.05). The binding abilities of Foxp3 gene promoter to NICD1 and p300 were positively correlated with the expression of Foxp3 at mRNA level ( r=0.58 and 0.46, both P<0.05). After competitive inhibition, the three aforementioned indexes in the acute BCP-ALL group were significantly lower than those in untreated group ( P<0.05); the binding ability of Foxp3 gene promoter to NICD1 in the control group was also significantly lower than that in untreated control group ( P<0.05), but no statistical differences in the other two indexes were found between the control groups with or without treatment ( P>0.05). ⑶ Compared with the control group, the expression of Notch1, PSEN1, MAML1 and SKIP in CD4 + T cells were elevated significantly ( P<0.05), while the transcription level of negative regulatory factor FBXW7 was decreased remarkably in children with acute BCP-ALL ( P<0.05). No statistical differences in the expression of GSK3β or IKAROS were found between the two groups ( P>0.05). Conclusions:Overactivation of Notch1 signaling caused by low expression of FBXW7 might be the key factor resulting in histone 4 hyperacetylation at foxp3 gene promoter and Treg cell dysfunction in children with acute BCP-ALL.
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A large number of cancer patients relapse after chemotherapeutic treatment. The immune system is capable of identifying and destroying cancer cells, so recent studies have highlighted the growing importance of using combinatorial chemotherapy and immunotherapy. However, many patients have innate or acquired resistance to immunotherapies. Long-term follow-up in a pooled meta-analysis exhibited long-term survival in approximately 20% of patients treated with immune checkpoint inhibitors or the adoptive transfer of chimeric T cells. It has been reported that high levels of immunoregulatory cells in cancer patients contribute to immunotherapy resistance via immunosuppression. Among the most important regulatory cell subtypes are the CD4+ T-regulatory cells (Tregs), identified by their expression of the well-characterized, lineage-specific transcription factor FOXP3. In addition to CD4+ Tregs, other regulatory cells present in the tumor microenvironment, namely CD8+ Tregs and IL10-producing B-regulatory cells (Bregs) that also modulate the immune response in solid and lymphoid tumors. These cells together have detrimental effects on tumor immune surveillance and anti-tumor immunity. Therefore, targeting these regulatory lymphocytes will be crucial in improving treatment outcomes for immunotherapy.
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T-Lymphocytes, Regulatory , Immunotherapy , Neoplasms , Immunosuppression TherapyABSTRACT
INTRODUCCIÓN: El síndrome IPEX (inmunodesregulación, poliendocrinopatía y enteropatía autoinmune ligada a X) causado por mutaciones en el gen FOXP3, se caracteriza por diarrea prolongada, alteraciones endocrinológicas y dermatitis. El tratamiento consiste en la administración de medicamentos inmunosupresores, siendo el trasplante de médula ósea la única cura potencial. OBJETIVO: Describir una nueva mutación del gen FOXP3, así como los hallazgos y evolución de un paciente con síndrome IPEX. CASO CLÍNICO: Lactante menor masculino que debutó al mes de vida con diarrea cró nica, falla intestinal e infecciones recurrentes. Exámenes de laboratorio y biopsia intestinal sugerentes de enteropatía autoinmune. Durante el seguimiento, el paciente presentó refractariedad al manejo inmunosupresor con esteroides, ciclosporina y tacrolimus, falleciendo a los 7 meses de edad por complicaciones vasculares. Antecedente familiar por línea materna de múltiples muertes en hombres menores de 1 año. Ante la sospecha de síndrome IPEX se realizó exoma en trío que reportó una mutación probablemente patogénica en el gen FOXP3. CONCLUSIÓN: Se documentó una nueva mutación del gen FOXP3 en paciente con síndrome IPEX. A pesar de la baja prevalencia de esta enfermedad, es importante el reconocimiento de síntomas no específicos pero sugerentes del diagnóstico.
INTRODUCTION: The IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked) syn drome is caused by the mutations of the FOXP3 gene, characterized by persistent diarrhea, endo crine disorders, and dermatitis. The treatment is the administration of immunosuppressive drugs, where hematopoietic stem cell transplantation is the only potential cure. OBJECTIVE: To describe a new FOXP3 gene mutation, as well as the findings and evolution of a patient with IPEX syndrome. CLINICAL CASE: Male infant presenting at one month of age with chronic diarrhea, intestinal failure, and recurrent infections. Lab tests and intestinal biopsy suggested autoimmune enteropathy. During follow-up, the patient presented resistance to immunosuppressive treatment with corticosteroids, cyclosporine, and tacrolimus, dying at 7 months of age due to vascular complications. He had a ma ternal family history of multiple deaths of men under 1 year of age. IPEX syndrome was suspected therefore a trio whole-exome sequencing was performed that showed a probably pathogenic FOXP3 gene mutation. CONCLUSION: A new FOXP3 gene mutation is reported in a patient with IPEX syndro me. Despite the low prevalence of this disease, it is important to recognize non-specific but suggestive symptoms for its diagnosis.
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Humans , Male , Infant , Genetic Diseases, X-Linked/diagnosis , Diabetes Mellitus, Type 1/congenital , Diarrhea/diagnosis , Forkhead Transcription Factors/genetics , Immune System Diseases/congenital , Pedigree , Genetic Markers , Chronic Disease , Fatal Outcome , Genetic Diseases, X-Linked/genetics , Diabetes Mellitus, Type 1/diagnosis , Diabetes Mellitus, Type 1/genetics , Diarrhea/genetics , Immune System Diseases/diagnosis , Immune System Diseases/genetics , MutationABSTRACT
Objective:To investigate the clinical efficacy of Loulu Shengma Tang combined with azithromycin in the treatment of pediatric mycoplasma pneumoniae pneumonia（MPP） with obstruction of lung by pathogenic heat， and its effect on inflammatory factors， treg and Foxp3 mRNA. Method:Totally 274 children with MPP were divided into observation group （137 cases） and control group （137 cases）. Observation group was treated with Loulu Shengma Tang combined with azithromycin dry suspension， while control group was treated with azithromycin dry suspension alone. The changes of traditional Chinese medicine（TCM） syndrome score of pathogenic-heat obstruction in the lung， serum inflammatory cytokines ［interleukin-6 （IL-6），interleukin-10 （IL-10），tumor necrosis factor-α （TNF-α），C-reactive protein （CRP）］， CD4+CD25+Treg， CD4+Foxp3+Treg and Foxp3 mRNA expressions were observed after treatment. The clinical efficacy and the incidence of adverse reactions were compared between two groups. Result:The total effective rate of observation group was 94.16%（129/137） after treatment， which was significantly higher than 77.37% （106/137）of observation group （P<0.05）. The disappearance times of cough， lung rale， fever and lung shadow in observation group were shorter than that in control group （P<0.05）. After treatment， TCM syndrome score of pathogenic-heat obstruction in lung was significantly higher than those in control group （P<0.05）， serum IL-6， IL-10， TNF-α and CRP levels in observation group were significantly lower than those in control group （P<0.05， P<0.01）， while CD4+CD25+Treg， CD4+Foxp3+Treg and Foxp3 mRNA expressions were significantly higher than those in control group （P<0.05）. The incidence of adverse reactions in observation group was 7/137 （5.11%）， which was significantly lower than 16/137 （11.68%） in control group. Conclusion:The clinical efficacy of Loulu Shengma Tang combined with azithromycin dry suspension in the treatment of pediatric MPP and its effect on serum inflammatory factors （IL-6， IL-10， TNF-α， CRP）， regulatory T cells and Foxp3 mRNA expressions were better than those of azithromycin dry suspension alone. The incidence of adverse reactions of Loulu Shengma Tang was lower than that of azithromycin dry suspension alone.
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Objective::To study the effect of warming and heat-clearing method (Wenyang Jiedu Huayu decoction) on the expressions of Forkhead box P3 (FoxP3), Retinoic acid-related orphan receptor gamma t (ROR-γt) in colon tissue of mice with acute-on-chronic liver failure (ACLF), in order to explore the possible regulatory mechanism on intestinal endotoxemia (IETM) in liver failure mice. Method::The 130 SD rats were randomly divided into normal group (10 rats) and model group (120 rats). The ACLF mice model was established through the subcutaneous injection with bovine serum albumin and the intraperitoneal injection with D-galactosamine(D-Gal) and lipopolysaccharide (LPS). The model mice were randomly divided into model group, heat-clearing group (Yinchenhao decoction, 6.68 g·kg-1), warming group (Yinchen Zhufu decoction, 7.09 g·kg-1) and warming and heat-clearing group (Wenyang Jiedu Huayu decoction, 19.53 g·kg-1). The normal group and the model group were given distilled water by gastric lavage, while the other groups were given equal volume of corresponding Chinese herbal medicines for a week. The value of each index at 1, 12 and 24 h was measured. The ratio of Treg/Th17 cell in peripheral blood were detected and calculated by flow cytometry. Real-time fluorescence quantitative PCR (Real-time PCR) was used to detect the expressions of FoxP3 and ROR-γt in colon tissues of mice at different time points. In situ hybridization and immunohistochemistry were used to observe the expressions of FoxP3 and ROR-γt genes and proteins. Result::Compared with normal group, the ratio of Treg/Th17 in the model group decreased significantly at each time point (P<0.01). Compared with the model group, the Treg/Th17 ratio increased only in the warming and heat-clearing method group (P<0.05). Compared with normal group, the expression of ROR-γt in the model group was significantly higher (P<0.01), and the expression of ROR-γt in the model group was higher than FoxP3.Compared with the model group, the expressions of FoxP3 and ROR-γt mRNA in the heat-clearing group and the warming group decreased at each time point (P<0.05), and the expressions of FoxP3 and ROR-γt in the warming and heat-clearing method group decreased significantly (P<0.01). The expressions of FoxP3 and ROR-γt mRNA in warming and heat-clearing group decreased compared with those in the warming group and heat-clearing group (P<0.05). Conclusion::The mechanism of the warming and heat-clearing method on IETM in liver failure may be related to the regulation of FoxP3 and ROR-γt expressions.
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Background: Systemic lupus erythromatosus (SLE) is an autoimmune disease with 20–65% of patients developing lupus nephritis (LN). Studies have reported 10% of LN patients will end up with end stage renal disease and their mortality rate is higher compared to patients without LN. Abnormality of regulatory T cells (Tregs) level is thought to be a potential factor for this LN development. The aim of study was to evaluate the percentage of Tregs in LN patients.Methods: This was a comparative cross sectional study involving LN patients and age and gender matched controls with a 2:1 ratio. The patients were grouped into active and inactive LN based on their lupus activity index; complement levels, ANA, dsDNA antibodies, ESR, SLE Disease Activity Index (SLEDAI2K) score and also urine PCI (uPCI>0.05 for active group). Disease history, demographic data, routine blood test, peripheral blood for differentials count were taken and recorded. Peripheral blood mononuclear cells were stained with CD4, CD25 and Foxp3 antibodies and percentage of Tregs was analysed using BD fluorescence-activated cell sorting (FACS) cytometer. We compared demographic and laboratory parameters between healthy controls and LN patients as well as active and inactive LN patients.Results: A total of 34 LN patients (32 females, 2 males) were recruited. Their mean age and disease duration were 37.97±11.14 years and 110.95±65.07 months respectively. Thirteen matched controls with mean age 35.23±7.89 years were enrolled. There was no demographic difference between 2 groups of LN patients. Tregs were significantly lower in active LN compared to inactive LN and healthy control (0.44±0.37% vs. 1.89±0.46% vs. 3.12±0.56% of the CD4+, P<0.001). C3 and C4 complement fragments were significantly reduced in patients with active disease (C3; 50.92±28.43 vs. 76.31±25.63, P=0.011) and (C4; 11.17±8.41 vs. 16.70±6.50 P=0.044). Proteinuria was significantly higher while serum albumin levels were significantly lower in active patients compared to inactive patients and healthy control (urine PCI; 0.25(0.15-0.3) vs. 0.03(0.01-0.05) vs. 0.01, P<0.001) and (albumin; 29.89±6.87 vs. 36.87±3.58 vs. 40.62±1.89mmol/L, P<0.001). We found positive inversely correlation between Tregs with SLEDAI2K (r = -0.572, P=0.011) and proteinuria (r = -0.451, P=0.007).Conclusions: Tregs, C3 and C4 complements, and albumin were significantly lower while proteinuria was significantly higher in active LN. There was positive inversely correlation between the percentage of Tregs with SLEDAI2K score and proteinuria.
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Objective To investigate the effect of Toxoplasma gondii excretory-secretory antigens (ESA) on CD4+ CD25+ Foxp3+ T (Treg) cells in mice carrying Lewis lung carcinoma, and examine the inhibitory effect of T. gondii ESA on tumor growth. Methods C57BL/6 mice were randomly assigned into the PBS group (n = 14) and the Lewis group (n = 34). Mice in the Lewis group were subcutaneously injected with 2 × 105 Lewis lung carcinoma cells in the right axilla, while animals in the PBS group were injected with the same volume of sterile PBS. On day 7 post-injection (D7), mice in the PBS group were further divided into the PBS2 group and the PBS2 + ESA group, of 7 mice in each group, and mice in the Lewis group were further divided into the Lewis2 group and the Lewis2 + ESA group, of 17 mice in each group. Then, mice in the PBS2 + ESA group and the Lewis2 + ESA group were intraperitoneally injected with 100 μL of ESA. The mouse spleen coefficient was calculated in each group 7 days post-injection with ESA, and the changes of Treg cell counts and the long-term tumor growth were measured in tumor-bearing mice. Results The spleen coefficient was significantly greater in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (0.66% ± 0.09% vs. 0.30% ± 0.02%, P < 0.05) and Lewis2 groups (0.69% ± 0.07% vs. 0.33% ± 0.03%, P < 0.05) 7 days post-treatment with ESA, respectively, and the percentage of splenic Treg cells in splenocytes was significantly lower in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (1.28% ± 0.14% vs. 2.06% ± 0.07%, P < 0.05) and Lewis2 groups (1.58% ± 0.14% vs. 2.44% ± 0.23%, P < 0.05), respectively. T. gondii ESA treatment caused a delay in tumor growth, and the tumor size was significantly smaller in the Lewis2 + ESA group than in the Lewis2 group (P < 0.05). Conclusion T. gondii ESA may reduce the proportion of splenic Treg cells in splenocytes and inhibit tumor growth in mice carrying Lewis lung carcinoma.
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Objective To investigate the effect of Toxoplasma gondii excretory-secretory antigens (ESA) on CD4+ CD25+ Foxp3+ T (Treg) cells in mice carrying Lewis lung carcinoma, and examine the inhibitory effect of T. gondii ESA on tumor growth. Methods C57BL/6 mice were randomly assigned into the PBS group (n = 14) and the Lewis group (n = 34). Mice in the Lewis group were subcutaneously injected with 2 × 105 Lewis lung carcinoma cells in the right axilla, while animals in the PBS group were injected with the same volume of sterile PBS. On day 7 post-injection (D7), mice in the PBS group were further divided into the PBS2 group and the PBS2 + ESA group, of 7 mice in each group, and mice in the Lewis group were further divided into the Lewis2 group and the Lewis2 + ESA group, of 17 mice in each group. Then, mice in the PBS2 + ESA group and the Lewis2 + ESA group were intraperitoneally injected with 100 μL of ESA. The mouse spleen coefficient was calculated in each group 7 days post-injection with ESA, and the changes of Treg cell counts and the long-term tumor growth were measured in tumor-bearing mice. Results The spleen coefficient was significantly greater in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (0.66% ± 0.09% vs. 0.30% ± 0.02%, P < 0.05) and Lewis2 groups (0.69% ± 0.07% vs. 0.33% ± 0.03%, P < 0.05) 7 days post-treatment with ESA, respectively, and the percentage of splenic Treg cells in splenocytes was significantly lower in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (1.28% ± 0.14% vs. 2.06% ± 0.07%, P < 0.05) and Lewis2 groups (1.58% ± 0.14% vs. 2.44% ± 0.23%, P < 0.05), respectively. T. gondii ESA treatment caused a delay in tumor growth, and the tumor size was significantly smaller in the Lewis2 + ESA group than in the Lewis2 group (P < 0.05). Conclusion T. gondii ESA may reduce the proportion of splenic Treg cells in splenocytes and inhibit tumor growth in mice carrying Lewis lung carcinoma.
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Introdução: Nas últimas décadas, progressos significativos foram feitos no manejo do carcinoma de células renais no cenário de doença avançada ou metastática. O uso de imunoterápicos estão aumentando tendo em vista que as células inatas e adaptativas do sistema imune desempenham um papel importante na regulação do crescimento do câncer. Objetivos: Os autores avaliaram as expressões imuno-histoquímicas de FOXP3, CD4 e CD8 e seus impactos prognósticos na sobrevida global e as compararam com fatores clínicos e patológicos. Metodologia: As expressões imuno-histoquímicas de FOXP3, CD4 e CD8 foram analisadas em 103 casos de carcinomas renais de células claras dispostos em tissue microarray. Para análise de seu impacto na sobrevida e associação com variáveis clínicas, dados epidemiológicos e clínicos foram coletados. Resultados: Em uma análise univariada, a razão de linfócitos CD4+/CD8+ apresentou impacto na sobrevida câncer específica (SCE) (p=0,045). Conclusão: A relação de linfócitos CD4+/CD8+ esteve associada a piores taxas de SCE. Nenhuma das variáveis clínicas e anatomopatológicas estudadas mostrou relação significativa com linfócitos positivos para FOXP3, CD4 e CD8 (AU)
Introduction: Significant progress has been made in the management of renal cell carcinoma during the last decades in the advanced or metastatic scenario. The use of immunotherapy is increasing as innate and adaptive cells of the immune system play an important role in regulating cancer growth. Objectives: The authors evaluated the immunohistochemical expressions of FOXP3, CD4 and CD8 and their prognostic impacts on overall survival and compared them with clinical and pathological factors. Methodology: Epidemiological and clinical data were collected. Immunohistochemical expressions were analyzed in 103 cases arranged in tissue microarray. Results: In a univariate analysis, the CD4+/CD8+ lymphocyte ratio impacted cancer-specific survival (ECS) (p = 0.045). Conclusion: The ratio of CD4+/CD8+ lymphocytes was associated with worse ECS rates. None of the clinical and anatomopathological variables studied showed a significant relationship with FOXP3, CD4 and CD8 positive lymphocytes (AU)
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Carcinoma, Renal Cell , Adenocarcinoma, Clear Cell , Inflammation , Kidney NeoplasmsABSTRACT
Objective To investigate the immune tolerance of Qi-Boosting Toxin-Resolving Granules (QBTRF) to Treg in the micro- environment of nasopharyngeal carcinoma (NPC) transplanted tumor, and reveal the possible anti-tumor mechanism of traditional Chinese medicine. Methods Using tumor cells CNE2 to build grafted tumor models, the anti-tumor effect of the QBTRF and cisplatin was observed, and the protein expression levels of Foxp3, ROR-γt and the content of IFN-γ, TGF-β and IL-17 in serum were detected. Results It was found that the content of TGF-β and IL-17 in serum of the tumor granule group and the combination therapy group were significantly lower than the model group, the IFN-γ content was obviously higher than that of the model group and cisplatin group, the apoptotic rate was significantly higher than that of the model group, the protein expression of Foxp3 was significantly increased, and the expression of ROR-γt was obviously decreased, in which the combination group was obviously better than that of single-use groups (P < 0.01). Conclusion QBTRF can effectively regulate the balance of Treg/Th17 in mice, restore the function of Treg, enhance the immune function of T cells during chemotherapy in mice, and exert anticancer effect. Combined with cisplatin, it can reduce the toxicity of chemotherapy.
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OBJECTIVE: To observe the effect of acupoint injection on expression of fork head/winged helix protein 3 (Foxp3), retinoic acid-related orphan receptor γt (RORγt) in nasal mucosa and serum interleukin-17 (IL-17) level in allergic rhinitis (AR) rats, so as to explore its mechanism underlying improvement of AR in terms of balancing Th17/Treg. METHODS: Thirty-two SD rats (half male and half female) were randomized into normal control, AR model, acupoint injection and non-acupoint injection groups (n＝8 in each group). The AR model was established by ovalbumin sensitization. In the acupoint injection group, "Yintang" (EX-HN3) and bilateral "Yingxiang"(LI20) were selected for injection of mixture solution of dexamethasone (DEX) and transfer factor and lidocaine (0.1 mL/acupoint), once every 3 days for a total of 4 times. The non-acupoints, located at the mid-point between the "Houhai" (GV1) and "Huantiao"(GB30) on the bilateral hips and the sites 5 cm inferior to the axillary were injected with the same dose of mixture solution as that in the acupoint injection. The AR severity was assessed by cumulative quantification scoring methods (including the numbers of nose-catching and sneezes, and the amount of nasal secretions in 30 min). The expressions of Foxp3 and RORγt in the nasal mucosa were detected by immunohistochemistry. The serum IL-17 content was detected by enzyme linked immunosorbent assay (ELISA)．. RESULTS: The AR symptom score and serum IL-17 content were significantly higher in the AR model group than those in the normal control group (P0.05). The percentage of the Foxp3 positive cells and the ratio of Foxp3/RORγt were negatively correlated with the AR symptom score(P<0.05), the expression of RORγt and the content of IL-17 were positively correlated with the symptom score (P<0.05)．. CONCLUSION: Acupoint injection is able to improve symptoms of RA rats, which may be related with its function in up-regulating the level of nasal mucosal Foxp3 and suppressing the levels of nasal RORγt and serum IL-17 to correct the imbalance of Th17/Treg.
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@#Objective To investigate the effect of artesunate inhibiting the expression of FOXP3 on proliferation, apoptosis and multidrug resistance of adriamycin (ADR) -resistant K562/ADR cells in chronic myeloid leukemia (CML), and to explore its mechanism. Methods The expressions of FOXP3 mRNA in K562 and K562/ADR cells were detected by real-time PCR. The expressions of FOXP3 proteins in K562 and K562/ADR cells were detected by Western blot assay. The K562/ADR cells were treated with different concentrations of artesunate (2.5, 5.0, 7.5, 10.0 and 12.5 mg/L) for 24 h. The toxicities of different concentrations of artesunate to K562/ADR cells were detected by CCK-8 assay, and the non-cytotoxic concentrations were screened. The expressions of FOXP3 mRNA and proteins in K562/ADR cells treated by non-cytotoxic concentration of artesunate were detected by RT-PCR and Western blot assay. The changes of toxicities of ADR in K562/ ADR cells were detected by CCK-8 assay. The average fluorescence intensities of ADR were detected by FCM assay. Results The expressions of FOXP3 were higher in K562/ADR cells than those in K562 cells. The mRNA and proteins expressions of FOXP3 were significantly lower in 2.5 mg/L group, 5 mg/L group and 7.5 mg/L group than those in the control group. The toxicities and concentrations of ADR were increased in K562/ADR cells treated by artesunate (both P＜0.05). Conclusion FOXP3 gene is highly expressed in adriamycin-resistant K562/ADR cells in CML. Artesunate can increase the concentrations of ADR and reverse multidrug resistance in K562/ADR cells by inhibiting the expression of FOXP3 in a dose-dependent manner.
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Objective The aim of this study was to investigate the effect of complete lung cancer resection on the balance of Th17/Treg cells in the peripheral blood of lung cancer patients. Methods Flow cytometry was used to detect the percentage of Th17 and Treg cells in the peripheral blood of 21 patients with lung cancer before and after operation and 21 healthy controls. RT-PCR was used to detect the expression of fork-head/winged helix transcription factor( Foxp3) and retinoic acid-related orphan receptor γt ( RORγt)in PBMC. The plasma levels of interleukin(IL) -17 and transforming growth factor(TGF) -β1 were detected by enzyme-linked immunosorbent assay(ELISA). Results After surgery,the percentage of Th17 cells in the peripheral blood decreased and the percentage of Treg cells increased in patients when compared to the pre -operation ( P <0. 01). After surgery,the expression of RORγt was decreased and the expression of Foxp3 was increased in the CD4 +T cells of patients in comparison with the pre-opera-tion(P<0. 01). After surgery,the expression of IL-17 was decreased and the expression of TGF-β1 was increased in the plasma of patients in comparison with the pre -operation( P <0. 01). In addition,the percentage of Th17 and Treg cells,the expression of RORγt and Foxp3,and the expression of IL-17 and TGF-β in the peripheral blood were increased in preoperative lung cancer pa-tients when compared to healthy controls. Conclusion There is a Th17/Treg imbalance in the peripheral blood of lung cancer pa-tients after complete resection of lung cancer.
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Objective To determine the proportion of CD4+ CD25+ regulatory T (Treg) cells,mRNA expression of the forkhead box protein 3 (Foxp3) gene,and DNA methylation status of the Foxp3 promoter in peripheral CD4+ T cells from patients with Henoch-Sch(o)nlein purpura.Methods Totally,20 inpatients with Henoch-Sch(o)nlein purpura and 20 healthy controls were enrolled from Department of Dermatology,the Second Xiangya Hospital of Central South University between 2015 and 2016,and there were no significant differences in the gender and age between the two groups (both P ＞ 0.05).CD4+ T cells were isolated from the peripheral blood samples of these subjects.Real-time fluorescence-based quantitative PCR was performed to detect the mRNA expression of the Foxp3 gene,flow cytometry to determine the proportion of CD4 + CD25+ Treg cells,and sodium bisulfite sequencing PCR (BSP) to determine the DNA methylation status of the Foxp3 promoter.Statistical analysis was carried out with SPSS16.0 software by using two-sample t test for the comparison between the two groups,and linear correlation analysis for evaluating the correlations of the DNA methylation status of the Foxp3 promoter with clinical severity scores and the proportion of CD4+CD25+ Treg cells.Results Compared with the healthy control group,the Henoch-Sch(o)nlein purpura group showed significantly decreased mRNA expression of the Foxp3 gene in CD4+ T cells (0.380 ± 0.226 vs.1,t =9.503,P ＜ 0.01),proportion of CD4+CD25+ Treg cells (1.668％ ± 0.959％ vs.2.741％ ± 1.131％,t =2.552,P ＜ 0.05),but significantly increased DNA methylation status of the Foxp3 promoter (0.712 ± 0.164 vs.0.453 ± 0.147,t =3.610,P ＜ 0.01).In the Henoch-Sch(o)nlein purpura group,the DNA methylation status of the Foxp3 promoter was negatively correlated with the percentage of CD4+CD25+ Treg cells (r =-0.490,P ＜ 0.05),but positively correlated with the clinical severity scores (r =0.486,P ＜ 0.05).The DNA methylation level of the Foxp3 promoter was significantly higher in the patients with renal impairment than in those without renal impairment (P ＜0.05).Conclusion The patients with Henoch-Sch(o)nlein purpura showed increased DNA methylation status of the Foxp3 promoter in CD4+ T cells,decreased mRNA expression of the Foxp3 gene and proportion of CD4+CD25+ Treg cells,which may be related to the occurrence of Henoch-Sch(o)nlein purpura,and affect disease development and prognosis.
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Objective To investigate the expression of transcription factor forkhead/winged helix transcription factor 3 (Foxp3),immune factor transforming growth factor-beta 1 (TGF-β1),and T-lymphocyte activation related factor interleukin-2 (IL-2) in peripheral blood of patients with coal-burning arsenic poisoning,and to analyze the effects of arsenic exposure on immune function.Methods A case-control study was conducted to investigate 149 cases [94 males and 55 females,(50.69 ± 6.14) years old] of arsenic poisoning in Yuzhang coalburning arsenic poisoning area,southwestern Guizhou Province,and the cases were diagnosed based on the "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015) and confirmed by clinical review.According to skin damage,the patients were divided into mild arsenic poisoning group (39 cases),moderate arsenic poisoning group (54 cases) and severe arsenic poisoning group (56 cases);and 41 cases [12 males and 29 females,(45.76 ± 7.88) years old] of non-arsenic exposed residents from 12 km of Yuzhang coal-burning area were selected as control group.Morning urine and peripheral blood samples were collected with informed consent.Urine arsenic content was measured by inductively coupled plasma mass spectrometry (ICP-MS).Urine arsenic was corrected by creatinine (Cr).Detection of regulatory T cell (Treg)-specific transcription factor Foxp3 gene expression in human peripheral blood was done by real-time fluorescence quantitative PCR,and the levels of Treg-related immune factor TGF-β1 and IL-2 in serum were detected by enzyme linked immunosorbent assay (ELISA).Results The urinary arsenic contents [median (quartile):29.13 (19.75-54.50),31.81 (17.52-53.31),30.51 (18.35-45.76) μg/g Cr] in each arsenic poisoning group were higher than that in the control group [21.62 (17.65-28.44) μg/g Cr,P ＜ 0.05].The expression levels of Foxp3 mRNA in peripheral blood of each arsenic poisoning group [median (quartile):0.58 (0.17-1.27),0.32 (0.17-0.61),0.33 (0.13-0.62)] were significantly lower than that in the control group [0.87 (0.64-1.50),P ＜ 0.05];compared with mild arsenic poisoning group,the expression of Foxp3 mRNA in peripheral blood of moderate and severe arsenic poisoning groups decreased (P ＜ 0.05).The contents of serum TGF-β1 [(13.14 ± 5.19),(12.85 ± 5.51),(12.78 ± 4.95) μg/L] in each arsenic poisoning group were significantly higher than that in the control group [(3.90 ± 1.53) μg/L,P ＜ 0.05].The levels of IL-2 in serum of each arsenic poisoning group [(9.85 ± 5.38),(11.64 ± 6.40),(12.27 ± 6.19) ng/L] were lower than that in the control group [(34.30 ± 4.84) ng/L,P ＜ 0.05];the serum level of IL-2 in severe arsenic poisoning group was higher than that in mild arsenic poisoning group (P ＜ 0.05).Conclusions Arsenic exposure can cause abnormal changes of Treg-specific transcription factor Foxp3 and related immune factors TGF-β1 and IL-2 in peripheral blood of patients.It is suggested that Treg dysfunction may be related to arsenic poisoning.