Objective: Cancer immunotherapy has made remarkable advances in recent years, but its effectiveness in treating gastric cancer is often limited by the complexity of the tumor microenvironment and the lack of effective...Objective: Cancer immunotherapy has made remarkable advances in recent years, but its effectiveness in treating gastric cancer is often limited by the complexity of the tumor microenvironment and the lack of effective biomarkers. This study aimed to identify effective biomarkers for immunotherapy treatment by characterizing the tumor microenvironment.Methods: We retrieved the RNA-seq data from gastric cancer patients treated with the programmed death 1(PD-1) blockade pembrolizumab. Differentially expressed genes associated with clinical outcomes were identified and further analyzed using gene ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway analysis. Gene signature scores were calculated by single sample Gene Set Enrichment Analysis(ssGSEA). The infiltration levels of immune cells were quantified using the xCell website. Cell type enrichment analysis was performed to compare treatment response and non-response groups, and regression analysis was used to investigate the relationship between interferon gamma(IFNγ) immune response and immune cell infiltration. Biomarkers were identified using least absolute shrinkage and selection operator(LASSO) analysis.Results: Compared to normal tissues, cytokine activity and interleukin-6 production were highly activated in gastric tumors. Responders to pembrolizumab showed significantly up-regulated expression of IFNγ responserelated genes. Cell type enrichment analysis revealed that Th1 cells were significantly enriched in the tumor microenvironment of responders. Regression analysis indicated that Th1 cells induced IFNγ response more efficiently than other cell types. Using signatures of Th1 cells, stromal cells and IFNγ response, a set of eight genes were identified that effectively predicted the efficacy of immunotherapy treatment and patient prognosis.Conclusions: Th1 cells promote therapeutic efficacy of PD-1 blockade by promoting IFNγ immune response in gastric cancer. The identified biomarkers have the potential to improve the effectiveness of immunotherapy treatment for gastric cancer patients.展开更多
· AIM: To determine the effects of rapamycin on experimental autoimmune uveoretinitis(EAU) and investigate of role of rapamycin on T cell subsets in the disease.·METHODS: EAU was induced in rats using peptid...· AIM: To determine the effects of rapamycin on experimental autoimmune uveoretinitis(EAU) and investigate of role of rapamycin on T cell subsets in the disease.·METHODS: EAU was induced in rats using peptides1169 to 1191 of the interphotoreceptor binding protein(IRBP). Rapamycin(0.2 mg/kg/d) was administrated by intraperitoneal injection for a consecutive 7d after immunization. Th1/Th2/Th17 cytokines, TGF-β1, and IL-6produced by lymphocyteswere measured by ELISA, while Th17 cells and CD4 +CD25 + regulatory T cells(Tregs)from rat spleen were detected by flow cytometry.·RESULTS: Intraperitoneal treatment immediately after immunization dramatically ameliorated the clinical course of EAU. Clinical responses were associated with reduced retinal inflammatory cell infiltration and tissue destruction. Rapamycin induced suppression of Th1/Th2/Th17 cytokines, including IFN-γ, IL-2, IL-17, IL-4, and IL-10 release from T lymphocytes of EAU rats, in vitro.Rapamycin also significantly increased TGF-β1production but had no effect on IL-6 productionof T lymphocytes from EAU rats in vitro. Furthermore,rapamycin decreased the ratio of Th17 cells/CD4 +T cells and upregulated Tregs in EAU, as detected by flow cytometry.·CONCLUSION: Rapamycin effectively interferes with T cell mediated autoimmune uveitis by inhibiting antigen-specific T cell functions and enhancing Tregs in EAU.Rapamycin is a promising new alternative as an adjunct corticosteroid-sparing agent for treating uveitis.展开更多
Background: Insulitis is defined by the presence of immune cells infiltrating in the pancreatic islets that might progress into the complete β-cell loss. The immunomodulatory properties of bone marrow-derived mesench...Background: Insulitis is defined by the presence of immune cells infiltrating in the pancreatic islets that might progress into the complete β-cell loss. The immunomodulatory properties of bone marrow-derived mesenchymal stem cells(BM-MSCs) have attracted much attention. This study aimed to evaluate the possible immunomodulatory effects of rat BM-MSCs and MSCs-derived insulin-producing cells(IPCs) in a mouse model of pancreatic insulitis. Methods: Insulitis was induced in BALB/c mice using five consecuti ve doses of streptozotocin. MSCs or IPCs were directly injected into the pancreas of mice and their effects on the expression of Th subsetsrelated genes were evaluated. Results: Both BM-MSCs and IPCs significantly reduced the expression of pancreatic Th1-related IFN-γ( P < 0.001 and P < 0.05, respectively) and T-bet genes(both P < 0.001). Moreover, the expression of IL-10 gene was significantly increased in IPC-treated compared to BM-MSC-or PBS-treated mice( P < 0.001 both comparisons). Conclusions: BM-MSCs and IPCs could successfully suppress pathologic Th1 immune responses in the mouse model of insulitis. However, the marked increase in IL-10 gene expression by IPCs compared to BM-MSCs suggests that their simultaneous use at the initial phase of autoimmune diabetes might be a better option to reduce inflammation but these results need to be verified by further experiments.展开更多
The essential effect of vitamin A on immune function occurs through various mechanisms including direct effect on ThloTh2 balance modulation. However, it is unclear whether or not vitamin A can regulate Thl-Th2 balanc...The essential effect of vitamin A on immune function occurs through various mechanisms including direct effect on ThloTh2 balance modulation. However, it is unclear whether or not vitamin A can regulate Thl-Th2 balance under a strong Thl-polarizing condition. Therefore, the purpose of our study was to examine the effect of vitamin A metabolite allotrans retinoic acid （ATRA） on ThloTh2 differentiation in CD4~ T cells under GATA-3 deficiency, which can induce Thl-polarizing condition. In the present study, GATA-3 deficiency T cells were induced by siRNA and checked by real-time quantitative PCR and western blot. GATA-3 deficiency CD4＋ T cells and normal CD4＋ T were treated for 48 h with or without ATRA.展开更多
Heat shock proteins (HSPs) are reported to act as effective adjuvants to elicit anti-tumor and anti-infection immunity. Here, we report that Hsp70-like protein 1 (Hsp70L1), a novel HSP derived from human dendritic cel...Heat shock proteins (HSPs) are reported to act as effective adjuvants to elicit anti-tumor and anti-infection immunity. Here, we report that Hsp70-like protein 1 (Hsp70L1), a novel HSP derived from human dendritic cells (DCs), has potent adjuvant effects that polarize responses toward Th1. With a calculated molecular weight of 54.8 kDa, Hsp70L1 is smaller in size than Hsp70 but resembles it both structurally and functionally. Hsp70L1 shares common receptors on DCs with Hsp70 and can interact with DCs, promoting DC maturation and stimulating secretion of the proinflammatory cytokines interleukin 12p70 (IL-12p70), IL-1beta, tumor necrosis factor-alpha (TNF-alpha), and the chemokines IP-10, macrophage inflammatory protein-1alpha (MIP-1alpha), MIP-1beta, and normal T cell expressed and secreted (RANTES). The induction of interferon-gamma-inducible protein 10 (IP-10) secretion by Hsp70L1 is not shared by Hsp70, and other functional differences include more potent stimulation of DC IL-12p70, CC-chemokine, and CCR7 and CXCR4 expression by Hsp70L1. Immunization of mice with the hybrid peptide Hsp70L1-ovalbumin(OVA)(257-264) induces an OVA(257-264)-specific Th1 response and cytotoxic T lymphocyte (CTL) that results in significant inhibition of E.G7-OVA tumor growth. The ability of Hsp70L1 to activate DCs indicates its potential as a novel adjuvant for use with peptide immunizations; the Hsp70L1 antigen peptide hybrid may serve as a more effective vaccine for the control of cancer and infectious diseases.展开更多
To investigate the effects of overall alkali of a traditional Chinese medicine “Tongbiling” (brucine and strychnine alkaloids in main) on the cytokines expression in Th1 and Th2 cells in the synovial fluid of patien...To investigate the effects of overall alkali of a traditional Chinese medicine “Tongbiling” (brucine and strychnine alkaloids in main) on the cytokines expression in Th1 and Th2 cells in the synovial fluid of patients with rheumatism arthritis and their signal pathway, the mononuclear cells in the synovial fluid (SFMC) of patients were isolated by Ficoll-Hypaque gradient centrifugation, and the CD3 + CD69 + and CD3 + HLA-DR antigen were analyzed by flow cytometry in comparison with those of the peripheral blood. The rest of cells were cultured after resuspension with RPMI 1640 culture medium. Phorbol 12,13-dibutyrate (PDB) and ionomycin were added successively into the culture with various concentration of overall alkali Tongbiling (TBL). After 4 h of cultivation, the expression of IFN-γ and IL-4 in CD3 + cells were analyzed by flow cytometry. The influence of overall alkali TBL (100?mg/L) on the intracellular calcium was investigated after Fluo-3/AM labeling and stimulation with PDB and ionomycin at 1, 2, 4 and 10?min, and the influence of TBL on the expression of CD3 + CD69 + cells were determined with stimulation of PDB for 24?h in the whole blood lymphocytes culture. It was found that the percentage of T cells bearing CD69 was significantly up-regulated (77%), while that of T cells bearing HLA-DR was 44% in the synovial mononucleated cells. After PDB and ionomycin stimulation, the expression of IFN-γ in CD3 + cells were up-regulated, but there was no change on the expression of IL-4 in CD3 + cells, indicating that ratio of Th1/Th2 was significantly increased and Th cells differentiate to Th1 cells in mainly. Four concentrations of overall alkaloid of TBL (200?mg/L, 100?mg/L, 50?mg/L, 25?mg/L) could down-regulated the expression of IFN-γ in CD3 + cells and the Th1/Th2 ratio obviously, but all the concentrations of the overall alkaloids had no effect on the expression of IL-4 in CD3 + cells. 100?mg/L concentration of the overall alkaloid did not down-regulate the intracellular calcium level. Each concentration of the overall alkaloid could down-regulated the expression CD69 obviously on the PDB-activated mouse T cells. It concluded from the above observations that the overall alkaloid of TBL could relieve the inflammatory and immune damages by suppressing the expression of Th1 type cytokines and Th1 cell differentiation, regulating the imbalance of Th1/Th2 cells and inhibiting the early activation of the T lymphocytes bearing CD69. There was no remarkable influence on the intracellular calcium signaling transduction pathway. The inhibitory effected on T cells to express IFN-γ might be due to the suppression of PKC-MAPK signaling pathway. From the standpoint of traditional Chinese medicine, this might be due to the regulation of “Yin” and “Yang” imbalance of joints to modify the pathological status in rheumatoid arthritis. This study provided an experimental basis for the application of overall alkaloids of TBL in the treatment of rheumatoid arthritis.展开更多
As one of the most serious types of psoriasis, pathogenesis of erythrodermic psoriasis（EP） is unclear so far. In this study, we aimed to detect the levels of Th1/Th2 cytokine-associated transcription factors and T-l...As one of the most serious types of psoriasis, pathogenesis of erythrodermic psoriasis（EP） is unclear so far. In this study, we aimed to detect the levels of Th1/Th2 cytokine-associated transcription factors and T-lymphocyte clone in peripheral blood mononuclear cells（PBMCs） derived from EP patients, and gene expression level of T-bet/GATA-3 in skin lesion. The potential role of Th1/Th2 reaction pattern played in the pathogenesis of EP was also discussed. Serum levels of IFN-γ, IL-2, IL-4 and IL-10 were quantified by ELISA among 16 EP patients, 20 psoriasis vulgaris（PV） patients and 15 healthy controls. The expression levels of T-bet/GATA-3 in the skin lesion and PBMCs were examined by real-time qPCR. The ratio of Th1/Th2 was measured by flow cytometry. The levels of IFN-γ, IL-2, IL-4 and IL-10 were higher in EP patients than in the healthy controls. The levels of IL-4 and IL-10 were 69.44±11.45 and 12.62±4.57 pg/mL, respectively, in EP patients, significantly higher than those in PV patients and healthy controls（P〈0.05）. Flow cytometry revealed the levels of both Th1 and Th2 in PBMCs from EP patients were higher than those in healthy controls, and the Th1/Th2 ratio was dramatically lower than in PV patients（P〈0.01）. The ratios of IFN-γ/IL-4 and T-bet/GATA-3 in EP patients were both less than 1.0, suggesting a reversal when compared with the other two groups. Our study indicated that the EP patients exerted a Th1/Th2 bidirectional response pattern, and the balance of Th cell subsets inclines to Th2, which might be one of the important mechanisms of EP pathogenesis.展开更多
目的探讨支气管哮喘患者外周程序性死亡受体1(programmend cell death protein l,PD-1)、干扰素-γ(intorferon-γ,IFN-γ)、白细胞介素-4(interleukin-4,IL-4)与辅助性T细胞(helper-T cell,Th细胞)及T细胞亚群水平表达的临床意义。方...目的探讨支气管哮喘患者外周程序性死亡受体1(programmend cell death protein l,PD-1)、干扰素-γ(intorferon-γ,IFN-γ)、白细胞介素-4(interleukin-4,IL-4)与辅助性T细胞(helper-T cell,Th细胞)及T细胞亚群水平表达的临床意义。方法选取2019年9月~2020年9月就诊于西安国际医学中心医院的80例哮喘患者为研究组,另以同期在该院体检的80例健康成人为对照组,采用肺功能仪检测两组研究对象的肺功能指标;ELISA法检测两组研究对象血清PD-1,IFN-γ和IL-4水平;单克隆抗体间接免疫荧光法检测两组研究对象外周血Th细胞和T淋巴细胞亚群占比,比较两组以上指标差异并分析其临床意义。结果与对照组相比,研究组肺功能指标FEV1和FEV1/预计值百分比(fevl%pred)、血清PD-1和IFN-γ水平、外周血Th1细胞占比、Th1/Th2比值、CD8+水平以及CD4^(+)/CD8+比值均显著降低,血清IL-4表达水平和外周血Th2细胞及CD4^(+)水平占比均显著升高,差异均有统计学意义(t=3.221~19.847,均P<0.01),而两组CD3+水平无显著差异(t=1.132,P=0.259)。结论支气管哮喘发作伴随着患者血清PD-1水平降低以及IL-4水平增高,从而抑制了Th1细胞产生IFN-γ,并使得Th1/Th2和CD4^(+)/CD8+平衡状态破坏,各细胞及细胞因子相互制约、相互调节,构成复杂的调节网络,对哮喘的诊断治疗有重要意义。展开更多
目的:探讨慢性乙型肝炎(乙肝)患者外周血CD4^(+)Th细胞内赖氨酸特异性组蛋白去甲基化酶1(histone lysine specific demethylase 1,LSD1)表达水平与Th1/Th2平衡的关系,初步探讨LSD1介导的组蛋白修饰对慢性乙肝患者CD4^(+)Th细胞分化的作...目的:探讨慢性乙型肝炎(乙肝)患者外周血CD4^(+)Th细胞内赖氨酸特异性组蛋白去甲基化酶1(histone lysine specific demethylase 1,LSD1)表达水平与Th1/Th2平衡的关系,初步探讨LSD1介导的组蛋白修饰对慢性乙肝患者CD4^(+)Th细胞分化的作用。方法:选择2017年6月至12月在南通大学附属医院感染科住院的慢性乙肝患者65例,分为丙氨酸转氨酶(ALT)高水平[>4×ULN(正常上限)]组与低水平(≤4×ULN)组,HBV-DNA高载量(≥10^(6)拷贝/mL)组与低载量(<106拷贝/mL)组。以同期在该院体检的30例健康人为对照。采用密度梯度离心法分离外周血单个核细胞,免疫磁珠法分选CD4^(+)Th细胞;提取CD4^(+)Th细胞总蛋白,以Western印迹法检测LSD1表达水平。采用ELISA法检测血清中干扰素γ(IFN-γ)、白细胞介素4(IL-4)含量;用微粒子化学发光法检测血清乙型肝炎表面抗原(HBsAg)含量;用实时定量PCR法检测血清HBV-DNA载量。结果:慢性乙肝组患者外周血CD4^(+)Th细胞中LSD1表达水平高于健康人(0.52±0.21 vs 0.28±0.09,t=-7.49,P<0.001)。慢性乙肝患者中,CD4^(+)Th细胞中LSD1表达量在ALT高水平组(n=38)低于低水平组(n=27,0.39±0.18 vs 0.64±0.16;t=-5.79,P<0.001),在HBV-DNA高载量组(n=32)高于HBV-DNA低载量组(n=33,0.69±0.08 vs 0.35±0.16;t=10.80,P<0.001)。与健康人相比,慢性乙肝组血清IFN-γ含量降低、IL-4含量增高、IFN-γ/IL-4比值减小(P<0.05)。慢性乙肝患者外周血CD4^(+)Th细胞中LSD1表达量与其血清ALT水平(r=-0.590)、IFN-γ水平(r=-0.379)及IFN-γ/IL-4(-0.285)负相关(P<0.01),与HBV-DNA载量正相关(r=0.880,P<0.001),与血清IL-4水平无相关性(r=0.169,P=0.102)。结论:LSD1高表达可能是慢性乙肝患者体内Th1/Th2失衡、Th1反应水平下降的原因之一,并导致机体清除HBV能力减弱或抑制HBV复制能力减弱。展开更多
基金financially supported by the National Natural Science Foundation of China (No. 82030079, 82341005, 81972656 and 82173035)the National Science and Technology Major Project of China (No. 2022YFC3400 901)Sino-Russian Math Center in PKU。
文摘Objective: Cancer immunotherapy has made remarkable advances in recent years, but its effectiveness in treating gastric cancer is often limited by the complexity of the tumor microenvironment and the lack of effective biomarkers. This study aimed to identify effective biomarkers for immunotherapy treatment by characterizing the tumor microenvironment.Methods: We retrieved the RNA-seq data from gastric cancer patients treated with the programmed death 1(PD-1) blockade pembrolizumab. Differentially expressed genes associated with clinical outcomes were identified and further analyzed using gene ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway analysis. Gene signature scores were calculated by single sample Gene Set Enrichment Analysis(ssGSEA). The infiltration levels of immune cells were quantified using the xCell website. Cell type enrichment analysis was performed to compare treatment response and non-response groups, and regression analysis was used to investigate the relationship between interferon gamma(IFNγ) immune response and immune cell infiltration. Biomarkers were identified using least absolute shrinkage and selection operator(LASSO) analysis.Results: Compared to normal tissues, cytokine activity and interleukin-6 production were highly activated in gastric tumors. Responders to pembrolizumab showed significantly up-regulated expression of IFNγ responserelated genes. Cell type enrichment analysis revealed that Th1 cells were significantly enriched in the tumor microenvironment of responders. Regression analysis indicated that Th1 cells induced IFNγ response more efficiently than other cell types. Using signatures of Th1 cells, stromal cells and IFNγ response, a set of eight genes were identified that effectively predicted the efficacy of immunotherapy treatment and patient prognosis.Conclusions: Th1 cells promote therapeutic efficacy of PD-1 blockade by promoting IFNγ immune response in gastric cancer. The identified biomarkers have the potential to improve the effectiveness of immunotherapy treatment for gastric cancer patients.
基金Supported by National Natural Science Foundation of China(No.81371005)
文摘· AIM: To determine the effects of rapamycin on experimental autoimmune uveoretinitis(EAU) and investigate of role of rapamycin on T cell subsets in the disease.·METHODS: EAU was induced in rats using peptides1169 to 1191 of the interphotoreceptor binding protein(IRBP). Rapamycin(0.2 mg/kg/d) was administrated by intraperitoneal injection for a consecutive 7d after immunization. Th1/Th2/Th17 cytokines, TGF-β1, and IL-6produced by lymphocyteswere measured by ELISA, while Th17 cells and CD4 +CD25 + regulatory T cells(Tregs)from rat spleen were detected by flow cytometry.·RESULTS: Intraperitoneal treatment immediately after immunization dramatically ameliorated the clinical course of EAU. Clinical responses were associated with reduced retinal inflammatory cell infiltration and tissue destruction. Rapamycin induced suppression of Th1/Th2/Th17 cytokines, including IFN-γ, IL-2, IL-17, IL-4, and IL-10 release from T lymphocytes of EAU rats, in vitro.Rapamycin also significantly increased TGF-β1production but had no effect on IL-6 productionof T lymphocytes from EAU rats in vitro. Furthermore,rapamycin decreased the ratio of Th17 cells/CD4 +T cells and upregulated Tregs in EAU, as detected by flow cytometry.·CONCLUSION: Rapamycin effectively interferes with T cell mediated autoimmune uveitis by inhibiting antigen-specific T cell functions and enhancing Tregs in EAU.Rapamycin is a promising new alternative as an adjunct corticosteroid-sparing agent for treating uveitis.
基金This study was supported by a grant from Shiraz University of Medical Sciences(No.94-7616).
文摘Background: Insulitis is defined by the presence of immune cells infiltrating in the pancreatic islets that might progress into the complete β-cell loss. The immunomodulatory properties of bone marrow-derived mesenchymal stem cells(BM-MSCs) have attracted much attention. This study aimed to evaluate the possible immunomodulatory effects of rat BM-MSCs and MSCs-derived insulin-producing cells(IPCs) in a mouse model of pancreatic insulitis. Methods: Insulitis was induced in BALB/c mice using five consecuti ve doses of streptozotocin. MSCs or IPCs were directly injected into the pancreas of mice and their effects on the expression of Th subsetsrelated genes were evaluated. Results: Both BM-MSCs and IPCs significantly reduced the expression of pancreatic Th1-related IFN-γ( P < 0.001 and P < 0.05, respectively) and T-bet genes(both P < 0.001). Moreover, the expression of IL-10 gene was significantly increased in IPC-treated compared to BM-MSC-or PBS-treated mice( P < 0.001 both comparisons). Conclusions: BM-MSCs and IPCs could successfully suppress pathologic Th1 immune responses in the mouse model of insulitis. However, the marked increase in IL-10 gene expression by IPCs compared to BM-MSCs suggests that their simultaneous use at the initial phase of autoimmune diabetes might be a better option to reduce inflammation but these results need to be verified by further experiments.
基金supported by the National Natural Science Foundation of China(No.30671761)
文摘The essential effect of vitamin A on immune function occurs through various mechanisms including direct effect on ThloTh2 balance modulation. However, it is unclear whether or not vitamin A can regulate Thl-Th2 balance under a strong Thl-polarizing condition. Therefore, the purpose of our study was to examine the effect of vitamin A metabolite allotrans retinoic acid （ATRA） on ThloTh2 differentiation in CD4~ T cells under GATA-3 deficiency, which can induce Thl-polarizing condition. In the present study, GATA-3 deficiency T cells were induced by siRNA and checked by real-time quantitative PCR and western blot. GATA-3 deficiency CD4＋ T cells and normal CD4＋ T were treated for 48 h with or without ATRA.
文摘Heat shock proteins (HSPs) are reported to act as effective adjuvants to elicit anti-tumor and anti-infection immunity. Here, we report that Hsp70-like protein 1 (Hsp70L1), a novel HSP derived from human dendritic cells (DCs), has potent adjuvant effects that polarize responses toward Th1. With a calculated molecular weight of 54.8 kDa, Hsp70L1 is smaller in size than Hsp70 but resembles it both structurally and functionally. Hsp70L1 shares common receptors on DCs with Hsp70 and can interact with DCs, promoting DC maturation and stimulating secretion of the proinflammatory cytokines interleukin 12p70 (IL-12p70), IL-1beta, tumor necrosis factor-alpha (TNF-alpha), and the chemokines IP-10, macrophage inflammatory protein-1alpha (MIP-1alpha), MIP-1beta, and normal T cell expressed and secreted (RANTES). The induction of interferon-gamma-inducible protein 10 (IP-10) secretion by Hsp70L1 is not shared by Hsp70, and other functional differences include more potent stimulation of DC IL-12p70, CC-chemokine, and CCR7 and CXCR4 expression by Hsp70L1. Immunization of mice with the hybrid peptide Hsp70L1-ovalbumin(OVA)(257-264) induces an OVA(257-264)-specific Th1 response and cytotoxic T lymphocyte (CTL) that results in significant inhibition of E.G7-OVA tumor growth. The ability of Hsp70L1 to activate DCs indicates its potential as a novel adjuvant for use with peptide immunizations; the Hsp70L1 antigen peptide hybrid may serve as a more effective vaccine for the control of cancer and infectious diseases.
文摘To investigate the effects of overall alkali of a traditional Chinese medicine “Tongbiling” (brucine and strychnine alkaloids in main) on the cytokines expression in Th1 and Th2 cells in the synovial fluid of patients with rheumatism arthritis and their signal pathway, the mononuclear cells in the synovial fluid (SFMC) of patients were isolated by Ficoll-Hypaque gradient centrifugation, and the CD3 + CD69 + and CD3 + HLA-DR antigen were analyzed by flow cytometry in comparison with those of the peripheral blood. The rest of cells were cultured after resuspension with RPMI 1640 culture medium. Phorbol 12,13-dibutyrate (PDB) and ionomycin were added successively into the culture with various concentration of overall alkali Tongbiling (TBL). After 4 h of cultivation, the expression of IFN-γ and IL-4 in CD3 + cells were analyzed by flow cytometry. The influence of overall alkali TBL (100?mg/L) on the intracellular calcium was investigated after Fluo-3/AM labeling and stimulation with PDB and ionomycin at 1, 2, 4 and 10?min, and the influence of TBL on the expression of CD3 + CD69 + cells were determined with stimulation of PDB for 24?h in the whole blood lymphocytes culture. It was found that the percentage of T cells bearing CD69 was significantly up-regulated (77%), while that of T cells bearing HLA-DR was 44% in the synovial mononucleated cells. After PDB and ionomycin stimulation, the expression of IFN-γ in CD3 + cells were up-regulated, but there was no change on the expression of IL-4 in CD3 + cells, indicating that ratio of Th1/Th2 was significantly increased and Th cells differentiate to Th1 cells in mainly. Four concentrations of overall alkaloid of TBL (200?mg/L, 100?mg/L, 50?mg/L, 25?mg/L) could down-regulated the expression of IFN-γ in CD3 + cells and the Th1/Th2 ratio obviously, but all the concentrations of the overall alkaloids had no effect on the expression of IL-4 in CD3 + cells. 100?mg/L concentration of the overall alkaloid did not down-regulate the intracellular calcium level. Each concentration of the overall alkaloid could down-regulated the expression CD69 obviously on the PDB-activated mouse T cells. It concluded from the above observations that the overall alkaloid of TBL could relieve the inflammatory and immune damages by suppressing the expression of Th1 type cytokines and Th1 cell differentiation, regulating the imbalance of Th1/Th2 cells and inhibiting the early activation of the T lymphocytes bearing CD69. There was no remarkable influence on the intracellular calcium signaling transduction pathway. The inhibitory effected on T cells to express IFN-γ might be due to the suppression of PKC-MAPK signaling pathway. From the standpoint of traditional Chinese medicine, this might be due to the regulation of “Yin” and “Yang” imbalance of joints to modify the pathological status in rheumatoid arthritis. This study provided an experimental basis for the application of overall alkaloids of TBL in the treatment of rheumatoid arthritis.
基金supported by grants from the National Natural Science Foundation of China(Nos.81171495 and 81271765)
文摘As one of the most serious types of psoriasis, pathogenesis of erythrodermic psoriasis（EP） is unclear so far. In this study, we aimed to detect the levels of Th1/Th2 cytokine-associated transcription factors and T-lymphocyte clone in peripheral blood mononuclear cells（PBMCs） derived from EP patients, and gene expression level of T-bet/GATA-3 in skin lesion. The potential role of Th1/Th2 reaction pattern played in the pathogenesis of EP was also discussed. Serum levels of IFN-γ, IL-2, IL-4 and IL-10 were quantified by ELISA among 16 EP patients, 20 psoriasis vulgaris（PV） patients and 15 healthy controls. The expression levels of T-bet/GATA-3 in the skin lesion and PBMCs were examined by real-time qPCR. The ratio of Th1/Th2 was measured by flow cytometry. The levels of IFN-γ, IL-2, IL-4 and IL-10 were higher in EP patients than in the healthy controls. The levels of IL-4 and IL-10 were 69.44±11.45 and 12.62±4.57 pg/mL, respectively, in EP patients, significantly higher than those in PV patients and healthy controls（P〈0.05）. Flow cytometry revealed the levels of both Th1 and Th2 in PBMCs from EP patients were higher than those in healthy controls, and the Th1/Th2 ratio was dramatically lower than in PV patients（P〈0.01）. The ratios of IFN-γ/IL-4 and T-bet/GATA-3 in EP patients were both less than 1.0, suggesting a reversal when compared with the other two groups. Our study indicated that the EP patients exerted a Th1/Th2 bidirectional response pattern, and the balance of Th cell subsets inclines to Th2, which might be one of the important mechanisms of EP pathogenesis.
文摘目的探讨支气管哮喘患者外周程序性死亡受体1(programmend cell death protein l,PD-1)、干扰素-γ(intorferon-γ,IFN-γ)、白细胞介素-4(interleukin-4,IL-4)与辅助性T细胞(helper-T cell,Th细胞)及T细胞亚群水平表达的临床意义。方法选取2019年9月~2020年9月就诊于西安国际医学中心医院的80例哮喘患者为研究组,另以同期在该院体检的80例健康成人为对照组,采用肺功能仪检测两组研究对象的肺功能指标;ELISA法检测两组研究对象血清PD-1,IFN-γ和IL-4水平;单克隆抗体间接免疫荧光法检测两组研究对象外周血Th细胞和T淋巴细胞亚群占比,比较两组以上指标差异并分析其临床意义。结果与对照组相比,研究组肺功能指标FEV1和FEV1/预计值百分比(fevl%pred)、血清PD-1和IFN-γ水平、外周血Th1细胞占比、Th1/Th2比值、CD8+水平以及CD4^(+)/CD8+比值均显著降低,血清IL-4表达水平和外周血Th2细胞及CD4^(+)水平占比均显著升高,差异均有统计学意义(t=3.221~19.847,均P<0.01),而两组CD3+水平无显著差异(t=1.132,P=0.259)。结论支气管哮喘发作伴随着患者血清PD-1水平降低以及IL-4水平增高,从而抑制了Th1细胞产生IFN-γ,并使得Th1/Th2和CD4^(+)/CD8+平衡状态破坏,各细胞及细胞因子相互制约、相互调节,构成复杂的调节网络,对哮喘的诊断治疗有重要意义。
文摘目的:探讨慢性乙型肝炎(乙肝)患者外周血CD4^(+)Th细胞内赖氨酸特异性组蛋白去甲基化酶1(histone lysine specific demethylase 1,LSD1)表达水平与Th1/Th2平衡的关系,初步探讨LSD1介导的组蛋白修饰对慢性乙肝患者CD4^(+)Th细胞分化的作用。方法:选择2017年6月至12月在南通大学附属医院感染科住院的慢性乙肝患者65例,分为丙氨酸转氨酶(ALT)高水平[>4×ULN(正常上限)]组与低水平(≤4×ULN)组,HBV-DNA高载量(≥10^(6)拷贝/mL)组与低载量(<106拷贝/mL)组。以同期在该院体检的30例健康人为对照。采用密度梯度离心法分离外周血单个核细胞,免疫磁珠法分选CD4^(+)Th细胞;提取CD4^(+)Th细胞总蛋白,以Western印迹法检测LSD1表达水平。采用ELISA法检测血清中干扰素γ(IFN-γ)、白细胞介素4(IL-4)含量;用微粒子化学发光法检测血清乙型肝炎表面抗原(HBsAg)含量;用实时定量PCR法检测血清HBV-DNA载量。结果:慢性乙肝组患者外周血CD4^(+)Th细胞中LSD1表达水平高于健康人(0.52±0.21 vs 0.28±0.09,t=-7.49,P<0.001)。慢性乙肝患者中,CD4^(+)Th细胞中LSD1表达量在ALT高水平组(n=38)低于低水平组(n=27,0.39±0.18 vs 0.64±0.16;t=-5.79,P<0.001),在HBV-DNA高载量组(n=32)高于HBV-DNA低载量组(n=33,0.69±0.08 vs 0.35±0.16;t=10.80,P<0.001)。与健康人相比,慢性乙肝组血清IFN-γ含量降低、IL-4含量增高、IFN-γ/IL-4比值减小(P<0.05)。慢性乙肝患者外周血CD4^(+)Th细胞中LSD1表达量与其血清ALT水平(r=-0.590)、IFN-γ水平(r=-0.379)及IFN-γ/IL-4(-0.285)负相关(P<0.01),与HBV-DNA载量正相关(r=0.880,P<0.001),与血清IL-4水平无相关性(r=0.169,P=0.102)。结论:LSD1高表达可能是慢性乙肝患者体内Th1/Th2失衡、Th1反应水平下降的原因之一,并导致机体清除HBV能力减弱或抑制HBV复制能力减弱。
