MTX周期联合CTX经JAK-STAT3通路对CIA小鼠Th17细胞分化的影响
发布时间：2018-08-13 08:44
【摘要】：目的:明确MTX周期联合CTX是否通过JAK-STAT3通路抑制CIA小鼠脾脏Na?ve T细胞向Th17细胞分化。方法:本实验以DBA1小鼠为研究对象,包括体内实验和体外实验两部分。体内试验:选取雄性DBA1小鼠56只,随机分为正常对照组10只和CIA组46只。建立CIA模型后根据拟用药不同随机分为4组:CIA组、MTX组(1.5 mg/kg/3d)、CTX组(30 mg/kg/10d)和MTX+CTX组(联合组,MTX:1.5 mg/kg/3d,CTX:30mg/kg/10d)。加强免疫3周后开始给药,治疗过程中连续监测各组小鼠左踝关节肿胀度及关节炎指数d(arthritis index,AI)。给药11周后处死小鼠,取双膝、左踝关节包埋、切片,进行HE染色,观察病理改变。分选脾细胞中的Na?ve T细胞,经细胞因子IL-6、TGF-β1、IL-1β、IL-23刺激于37℃5%CO2细胞培养箱中培养96小时,用流式细胞术检测培养后Th17细胞所占比例,用RT-PCR检测分化后Th17细胞中STAT3的表达水平。体外实验:选取雄性的DBA1小鼠20只建立CIA模型,7周时处死,分选脾淋巴细胞中的Na?ve T细胞,分为空白对照组、抗CD3CD28激活组(激活组)、刺激Th17细胞定向分化组(Th17分化组)、CTX组、低浓度MTX组(MTX低组)、中浓度MTX组(MTX中组)、高浓度MTX组(MTX高组)、MTX低+CTX组(低联合组),在37℃5%CO2细胞培养箱中培养96小时,用流式细胞术检测培养后Th17细胞所占比例,用RT-PCR检测分化后Th17细胞中STAT3的表达水平。结果:1.体内实验(1)治疗前,各治疗组小鼠关节肿胀度、AI与CIA组相比,差异无统计学意义;治疗4周后,各治疗组小鼠关节肿胀度、AI较CIA组改善明显,差异有统计学意义,各治疗组间差异无统计学意义;治疗11周后,联合组小鼠关节肿胀度、AI较CIA组、CTX组改善明显,差异有统计学意义,联合组较MTX组改善明显,但差异无统计学意义;(2)治疗11周后,各治疗组Th17细胞所占百分较CIA组明显降低,差异有统计学意义,联合组Th17百分率较单用药组降低明显,但差异无统计学意义;(3)治疗11周后,各治疗组Th17细胞中STAT3 m RNA的表达均低于CIA组,差异有统计学意义,联合组Th17细胞中STAT3 m RNA的表达低于单用药组,但差异无统计学意义。2.体外实验(1)体外培养96h后,Th17分化组Th17细胞所占百分比与空白组、MTX中组、MTX高组、低联合组相比,差异有统计学意义;低联合组与单用药组相比,培养后Th17细胞所占百分比降低,差异无统计学意义;(2)体外培养96h后,Th17分化组Th17细胞中STAT3 m RNA的表达与空白组、MTX中组、MTX高组、低联合组相比,差异有统计学意义;低联合组与单用药组相比,培养后Th17中STAT3 m RNA的表达降低,差异无统计学意义。结论:MTX周期联合CTX可能通过JAK-STAT3通路影响CIA小鼠Th17细胞的分化。
[Abstract]:Aim: to determine whether MTX cycle combined with CTX inhibits the differentiation of Na?ve T cells into Th17 cells in the spleen of CIA mice via JAK-STAT3 pathway. Methods: DBA1 mice were studied in vivo and in vitro. In vivo test: 56 male DBA1 mice were randomly divided into normal control group (n = 10) and CIA group (n = 46). The CIA model was established and randomly divided into 4 groups according to the drug use. They were divided into 4 groups: 1. 5 mg/kg/3d group (1. 5 mg/kg/3d), CTX group (30 mg/kg/10d) and MTX CTX group (CTX: 1. 5 mg / kg / 3 d ~ (-1) CTX: 30 mg / kg / 10 d). Three weeks after immunization, the swelling degree of left ankle joint and arthritis index (d (arthritis index AI) were continuously monitored. After 11 weeks of administration, mice were killed, bilateral knees were taken, left ankle was embedded, sections were sliced, HE staining was performed and pathological changes were observed. Na?ve T cells were isolated from splenocytes and stimulated by IL-6 TGF- 尾 1 and IL-1 尾 -IL-23 in 37 鈩，
本文编号：2180459
[Abstract]:Aim: to determine whether MTX cycle combined with CTX inhibits the differentiation of Na?ve T cells into Th17 cells in the spleen of CIA mice via JAK-STAT3 pathway. Methods: DBA1 mice were studied in vivo and in vitro. In vivo test: 56 male DBA1 mice were randomly divided into normal control group (n = 10) and CIA group (n = 46). The CIA model was established and randomly divided into 4 groups according to the drug use. They were divided into 4 groups: 1. 5 mg/kg/3d group (1. 5 mg/kg/3d), CTX group (30 mg/kg/10d) and MTX CTX group (CTX: 1. 5 mg / kg / 3 d ~ (-1) CTX: 30 mg / kg / 10 d). Three weeks after immunization, the swelling degree of left ankle joint and arthritis index (d (arthritis index AI) were continuously monitored. After 11 weeks of administration, mice were killed, bilateral knees were taken, left ankle was embedded, sections were sliced, HE staining was performed and pathological changes were observed. Na?ve T cells were isolated from splenocytes and stimulated by IL-6 TGF- 尾 1 and IL-1 尾 -IL-23 in 37 鈩，
本文编号：2180459
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