过表达IL-17RLM的人脐带间充质干细胞对TNBS诱导的结肠炎小鼠脾脏淋巴细胞的免疫调节作用

目的:研究过表达白细胞介素17受体样分子的人脐带间充质干细胞(IL-17RLM-hUCMSCs)对三硝基苯磺酸(TNBS)诱导的结肠炎小鼠脾脏淋巴细胞的免疫调节作用,为炎症性肠病的干细胞治疗提供优化的种子细胞。方法:体外分离培养hUCMSCs,利用慢病毒载体向干细胞内转入IL-17RLM基因,构建IL-17RLM-hUCMSCs。采用TNBS诱导小鼠实验性结肠炎模型,无菌取炎症小鼠脾脏制备淋巴细胞悬液,在刀豆蛋白A(Con A)刺激下将淋巴细胞分别与不同浓度的IL-17RLM-hUCMSCs及hUCMSCs共培养,72 h后以淋巴细胞+Con A为阳性对照,CCK8法及CFSE标记法检测淋巴细胞的增殖情况;同时用流式细胞术检测T淋巴细胞亚群(Th1、Th2、Th17及Treg)比例的改变。结果:hUCMSCs及IL-17RLM-hUCMSCs均对ConA刺激下的淋巴细胞增殖有抑制作用(P0.05);当MSCs/淋巴细胞为1∶1~1∶10时,MSCs对淋巴细胞增殖的抑制作用呈现浓度依赖性。在有效浓度范围内,IL-17RLM-hUCMSCs较h UCMSCs抑制作用更强(P0.05)。hUCMSCs及IL-17RLM-h UCMSCs均能够下调结肠炎小鼠脾脏淋巴细胞的Th1和Th17细胞亚群比例,上调Treg细胞亚群比例,但IL-17RLM-h UCMSCs对Th17细胞亚群的抑制作用更显著(P0.05)。结论:IL-17RLM-hUCMSCs呈浓度依赖性地抑制TNBS诱导的结肠炎小鼠脾脏淋巴细胞增殖,且该作用优于h UCMSCs。同时,IL-17RLM-hUCMSCs可调节T细胞亚群的免疫平衡,且抑制Th17细胞亚群作用强于hUCMSCs。

Immunomodulatory effect of human umbilical cord mesenchymal stem cells with interleukin-17 receptor-like molecule overexpression on spleen lymphocytes from mice with trinitrobenzene sulfonic acid-induced colitis

AIM: To investigate the immunomodulatory effect of human umbilical cord mesenchymal stem cells with interleukin-17 receptor-like molecule overexpression (IL-17RLM-hUCMSCs) on the spleen lymphocytes from the mice with trinitrobenzene sulfonic acid (TNBS)-induced colitis for providing optimal seed cells to treat inflammatory bowel disease. METHODS: Mesenchymal stem cells were isolated from human umbilical cord. The IL-17RLM gene was transferred into mesenchymal stem cells by lentivirus vector to establish IL-17RLM-hUCMSCs. The experimental colitis mice were induced by TNBS enema, and the spleen lymphocyte suspension was isolated. The lymphocytes were co-cultured with different concentrations of IL-17RLM-hUCMSCs and hUCMSCs under the stimulation of concanavalin A (ConA) for 72 h. The proliferation of lymphocytes was detected by the methods of CCK8 assay and CFSE labeling with lymphocytes+ConA as positive control. The changes of lymphocyte subsets (Th1, Th2, Th17 and Treg) were examined by flow cytometry.RESULTS: Both hUCMSCs and IL-17RLM-hUCMSCs inhibited T-cell proliferation in vitro co-culture system (P < 0.05). When the ratios of MSCs to lymphocytes ranged from 1:1 to 1:10, the inhibitory rates were in a dose-dependent manner. IL-17RLM-hUCMSCs showed higher inhibitory rate than hUCMSCs within the effective concentration range (P < 0.05). Both hUCMSCs and IL-17RLM-hUCMSCs reduced the proportions of Th1 and Th17 cell subsets and increased Treg cell population of spleen lymphocytes from TNBS-induced colitis mice, and IL-17RLM-hUCMSCs showed a stronger inhibitory effect on Th17 cell subset (P < 0.05). CONCLUSION: IL-17RLM-hUCMSCs remarkably inhibit the proliferation of spleen lymphocytes from TNBS-induced colitis mice in a dose-dependent manner. Meanwhile, they regulate immune balance of T cells and have stronger inhibitory effect on Th17 subset.