系统性红斑狼疮患者血浆对间充质干细胞免疫抑制作用的影响

发布时间：2019-06-02 01:48

【摘要】：目的:探究系统性红斑狼疮(Systemic Lupus Erythematosus,SLE)患者血浆对健康者骨髓来源的间充质干细胞(Mesenchymal Stem Cells,MSCs)免疫抑制作用的影响。方法:实验采用添加SLE患者富集血浆或胎牛血清的细胞培养体系(胎牛血清作为对照组),在体外共同培养SLE患者来源的B淋巴细胞与健康者骨髓来源的MSCs,检测MSCs对B淋巴细胞的增殖能力和分化成熟的影响。采用细胞增殖试验检测分析B淋巴细胞的增殖能力情况;使用流式细胞仪检测B淋巴细胞的表面标志物,统计分析B淋巴细胞的分化成熟情况。实验数据统计分析:采用SPSS 12.0软件进行学生t检验统计学分析,P0.05视为两组差异有统计学意义。结果:1、MSCs的分离、扩增培养和诱导分化:实验成功培养获得健康骨髓捐献者来源的MSCs;(1)MSCs经过流式细胞仪检测结果显示,高表达CD44、CD73、CD90、和CD105,不表达CD34、CD45、CD14和CD106;(2)通过添加特殊诱导剂诱导培养MSCs后,成功分化为成脂细胞和成骨细胞。2、正常MSCs可以抑制SLE患者来源B淋巴细胞的增殖(脂多糖为增殖刺激分子)。3、SLE患者富集血浆能够抵制正常MSCs对SLE患者来源B淋巴细胞的增殖抑制作用(脂多糖为增殖刺激分子)。4、正常MSCs可以抑制SLE患者来源B淋巴细胞的成熟:在胎牛血清培养系统中,加入MSC干预后,SLE患者来源B淋巴细胞表面CD27与CD38的表达量下调,CD19的表达水平无明显影响。5、SLE患者富集血浆能够抵制正常MSCs对SLE患者来源B淋巴细胞成熟的抑制作用:在SLE患者富集血浆培养系统中,加入MSCs干预后,SLE患者来源B淋巴细胞表面CD27与CD38的表达量明显上调,CD19的表达水平受到抑制作用。结论:正常MSCs可以抑制脂多糖刺激的SLE患者B淋巴细胞的增殖与成熟,调节B淋巴细胞亚群所占比例。而SLE患者富集血浆却能够抵制正常MSCs对B淋巴细胞的免疫抑制作用,这将负向干扰正常MSCs移植对SLE的治疗效果。
[Abstract]:Objective: to investigate the effect of plasma on bone marrow derived mesenchymal stem cells (Mesenchymal Stem Cells,MSCs) immunosuppressive effect in patients with systemic lupus erythematosus (Systemic Lupus Erythematosus,SLE). Methods: the cell culture system supplemented with SLE patients to enrich plasma or fetal bovine serum (fetal bovine serum as control group) was used to co-culture B lymphocytes from SLE patients and MSCs, from healthy bone marrow in vitro. The effects of MSCs on the proliferation and differentiation and maturation of B lymphocytes were detected. Cell proliferation test was used to detect the proliferation ability of B lymphocytes, flow cytometry was used to detect the surface markers of B lymphocytes, and the differentiation and maturation of B lymphocytes were statistically analyzed. Statistical analysis of experimental data: SPSS 12.0 software was used to analyze the t test of students, and the difference between the two groups was statistically significant (P 0.05). Results: 1. Isolation, amplification, culture and differentiation of MSCs: MSCs; from healthy bone marrow donors was successfully cultured. The main results were as follows: (1) the results of flow cytometry showed that high expression of CD44,CD73,CD90, and CD105, did not express CD34,CD45,CD14 and CD106;. (2) after MSCs was induced by adding special inducer, it was successfully differentiated into adipocytes and osteoblasts. 2, normal MSCs could inhibit the proliferation of B lymphocytes from SLE patients (lipopolysaccharide was a proliferation stimulator). The enrichment of plasma in SLE patients can resist the inhibitory effect of normal MSCs on the proliferation of B lymphocytes from SLE patients (lipopolysaccharide is a proliferation stimulator). Normal MSCs could inhibit the maturation of B lymphocytes from SLE patients: in fetal bovine serum culture system, the expression of CD27 and CD38 on the surface of B lymphocytes from SLE patients was down-regulated after adding MSC intervention, but the expression level of CD19 was not significantly affected. The enrichment of plasma in SLE patients can resist the inhibitory effect of normal MSCs on the maturation of B lymphocytes from SLE patients: in the enrichment plasma culture system of SLE patients, MSCs intervention was added. The expression of CD27 and CD38 on the surface of B lymphocytes from SLE patients was significantly up-regulated, and the expression level of CD19 was inhibited. Conclusion: normal MSCs can inhibit the proliferation and maturation of B lymphocytes and regulate the proportion of B lymphocytes subsets in SLE patients stimulated by lipopolysaccharide. However, the enrichment of plasma in patients with SLE can resist the immunosuppressive effect of normal MSCs on B lymphocytes, which will negatively interfere with the therapeutic effect of normal MSCs transplantation on SLE.
【学位授予单位】：贵州大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.241

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