背景:我们先前的研究表明,Hcy通过促进肠道T辅助细胞17(Th17)的分化而加剧了DSS诱导的结肠炎,但确切的机制仍不明。因此,本研究旨在阐明Hcy促进Th17细胞分化的信号通路.
方法:BALb/c小鼠随机分为6组。用3%DSS诱导小鼠结肠炎模型,而高同型半胱氨酸血症模型是用1.7%的蛋氨酸诱导的。使用酶联免疫吸附测定(ELISA)测量Hcy和前列腺素E2(PGE2)的浓度。胞浆磷脂酶A2(cPLA2)的蛋白表达,磷酸化-CPLA2(p-CPLA2),环氧合酶2(COX2),环磷酸腺苷(cAMP),信号转导和转录激活因子3(STAT3),磷酸化STAT3(p-STAT3),白细胞介素-17A(IL-17A),和类维生素A相关的孤儿核受体γt(RORγt)使用蛋白质印迹分析进行评估。
结果:与DSS+HHcy组相比,COX抑制剂的加入并没有显著改变p-PLA2/PLA2的蛋白表达,但导致血清PGE2浓度显著降低,cAMP,和p-STAT3/STAT3蛋白表达。STAT3抑制剂添加上游的p-PLA2/PLA2,COX2和cAMP的蛋白表达没有显着变化。然而,PGE2浓度和p-STAT3/STAT3蛋白表达显著降低。添加COX抑制剂和STAT3抑制剂后,CD4+T细胞中IL-17A和RORγt的蛋白表达以及IL-17A和IL-23R的水平显著降低。
结论:HHcy通过PGE2/STAT3信号通路促进Th17细胞的分化和增殖,加重DSS诱导的结肠炎。
BACKGROUND: Our previous study indicated that Hcy exacerbated DSS-induced colitis by facilitating the differentiation of intestinal T helper cell 17 (Th17), but the precise mechanism remains unidentified. Therefore, our current research aims to elucidate the signaling pathway through which Hcy promotes the differentiation of Th17 cells.
METHODS: BALb/c mice were randomly assigned into six groups. The model of mice colitis was induced using 3% DSS, while the model of Hyperhomocysteinemia was induced using 1.7% methionine. The concentrations of Hcy and prostaglandin E2 (
PGE2) were measured using enzyme-linked immunosorbent assay (ELISA). The protein expressions of cytosolic phospholipase A2 (cPLA2), phosphorylated-cPLA2 (p-cPLA2), cyclooxygenase 2 (COX2), cyclic adenosine monophosphate (cAMP), signal transducer and activator of transcription 3 (STAT3), phosphorylated-STAT3 (p-STAT3), interleukin-17A (IL-17A), and retinoid-related orphan nuclear receptor-γt (RORγt) were assessed using western blot analysis.
RESULTS: Compared to the DSS + HHcy group, the addition of the COX inhibitor did not significantly alter the protein expression of p-PLA2/PLA2, but led to significant decreases in serum
PGE2 concentration, cAMP, and p-STAT3/STAT3 protein expression. The protein expressions of p-PLA2/PLA2, COX2, and cAMP upstream of STAT3 inhibitor addition did not exhibit significant changes. However,
PGE2 concentration and p-STAT3/STAT3 protein expression were notably reduced. After the COX inhibitor and STAT3 inhibitor added, the protein expression of IL-17A and RORγt and the levels of IL-17A and IL-23R in CD4+ T cells were significantly reduced.
CONCLUSIONS: HHcy aggravated DSS-induced colitis by promoting the differentiation and proliferation of Th17 cells through the
PGE2 / STAT3 signaling pathway.