Objective To establish the model of autoimmune dry eye and analyze the level of cytokines and the pathological changes of autoimmune dry eye rabbits treated with human umbilical cord mesenchymal stem cells (MSCs).Methods Twentyfour female New Zealand rabbits were randomly divided into three groups:Normal control group,dry eye group and MSCs treatment group,8 cases in each group.After treated with MSCs for six weeks,the pathological changes were observed,and the expression of Th1 and Th17 cell differentiation related cytokines mRNA in lacrimal gland tissue was detected by real-time quantitative PCR.The percentage of regulatory T cells (CD4 +Foxp3 + cells) in the lacrimal gland and spleen tissues of the dry eye model group and MSCs treatment group was measured by flow cytometry.Results After treatment with MSCs for 6 weeks,HE staining from each rabbit lacrimal gland showed that,the normal control group showed no or only a few lymphocytes;The lacrimal gland cells were atrophied in the dry eye group,the distribution of lymphocyte scattered around the gland and small blood vessels;Compared with dry eye group,the lacrimal gland lymphocyte infiltration reduced in MSCs treatment group,and the cell morphology were better.Compared with dry eye group,lymphocyte infiltration and aggregation in the conjunctival conjunctiva in MSCs treatment group were significantly reduced,the epithelial structure was intact,and the degeneration and atrophy cells were rare.The expression of inflammatory factors including interferon-γ(IFN-γ),Interleukin-17 (IL-17) and transcription factor T-bet mRNA were decreased in MSCs treatment group,there were significant difference compared with dry eye group (all P ＜ 0.05);Thl7 cell associated cytokines IL-17 expression levels were decreased,but no significant difference was found between two groups (P ＞ 0.05);The expression of transcription factor RORrt mRNA in dry eye group was significantly decreased,the difference was statistically significant (P ＜ 0.05).The expression of inflammatory factors TNF-α in the lacrimal gland tissue of MSCs treatment group was significantly lower than that in the dry eye model group,while the expression of the anti-inflammatory factor TGF-β was significantly higher than that in the dry eye model group,and the differences were statistically significant (all P＜0.05).The regulatory T cells (CD4 + Foxp3 + cells) in the lacrimal gland of dry eye model group accounted for 10％ of lymphocytes was,while the treatment group accounted for 27.8％,MSCs group was significantly higher than dry eye group,the difference was statistically significant (P ＜ 0.05).Conclusion MSCs can reduce the histopathological changes of the immune dry eye,and they may have immunoregulatory effects on autoimmune dry eye.The mechanisms of immunomodulatory effects may be related to the balance of inflammatory cytokines and anti-inflammatory cytokines.%目的 建立兔自身免疫性干眼模型,观察间充质干细胞(mesenchymal stem cells,MSCs)治疗后兔自身免疫性干眼的组织病理学变化及泪腺组织中细胞因子表达的变化.方法 取健康成年雌性新西兰白兔24只24眼,采用随机数学表法将兔分为正常对照组、干眼模型组和MSCs治疗组,每组各8只8眼.MSCs治疗6周后,观察兔眼组织病理学改变,实时荧光定量PCR检测泪腺组织中Th1、Th17细胞分化相关细胞因子mRNA相对表达量.流式细胞仪检测干眼模型组和MSCs治疗组兔泪腺组织和脾脏组织中调节性T细胞(CD4+ Foxp3+细胞)占淋巴细胞的比例.结果 MSCs治疗6周后,取各组兔泪腺并行HE染色显示:正常对照组未见或仅见少量淋巴细胞浸润;干眼模型组泪腺内部分腺体细胞出现萎缩,腺管周围和小血管周围可见散在分布的淋巴细胞汇集区;与干眼模型组相比,MSCs治疗组泪腺内淋巴细胞浸润减轻,腺泡细胞形态较好.与干眼模型组比较,MSCs治疗组结膜组织中淋巴细胞浸润聚集显著减轻,上皮结构完整,变性和萎缩细胞较少见.MSCs治疗组泪腺组织中Th1特征性细胞因子IFN-γ以及转录因子T-bet mRNA表达水平较干眼模型组明显下降,差异均有统计学意义(均为P ＜0.05);Th17细胞分化相关细胞因子IL-17表达水平有下降趋势,但两组差异无统计学意义(P＞0.05);其转录因子RORa mRNA的表达水平较干眼模型组显著降低,差异有统计学意义(P＜0.05).MSCs治疗组泪腺组织中炎症因子TNF-α表达较干眼模型组显著降低,而抑炎因子TGF-β的表达较干眼模型组显著升高,差异均有统计学意义(均为P＜0.05).泪腺组织中干眼模型组调节性T细胞(CD4+ Foxp3+细胞)占淋巴细胞的比例为10.0％,而MSCs治疗组中调节性T细胞占淋巴细胞的比例为27.8％,MSCs治疗组调节性T细胞较干眼模型组显著升高,差异有统计学意义(P＜0.05).结论 MSCs可以减轻自身免疫性干眼的组织病理学改变,对自身免疫性干眼具有一定的免疫调节及治疗作用,其免疫调控作用可能与调节抑炎因子和促炎因子平衡有关.
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