CLEC2B基因对淋巴细胞细胞因子的调节及对黑素细胞的影响

发布时间：2018-04-28 14:41

本文选题：白瘢风 + siRNA　；参考：《天津医科大学》2012年硕士论文

【摘要】：目的通过对新发现的白癜风相关基因CLEC2B进行功能学研究以揭示该基因可能的相关致病靶点。建立淋巴细胞瞬时转染体系,沉默淋巴细胞中CLEC2B基因表达,观察其对淋巴细胞增殖及其细胞因子IL-4mRNA、sIL-2RmRNA、 IL-6mRNA表达的调节作用,观察CLEC2B基因沉默后淋巴细胞上清液对黑素细胞增殖活性的影响从而揭示CLEC2B基因参与白癜风发病的免疫学机制。方法本实验选用Jurkat淋巴细胞及B16黑素瘤细胞为研究对象,应用RNA干扰技术下调淋巴细胞CLEC2BmRNA的表达,实时荧光定量PCR法分析CLEC2B基因沉默后淋巴细胞细胞因子IL-4mRNA、sIL-2RmRNA、IL-6mRNA表达情况,MTT法检测CLEC2B基因沉默后淋巴细胞增殖情况和淋巴细胞上清液对黑素细胞增殖的影响。结果 1. DMRIE-C脂质体与CLEC2BsiRNA按合理配比(2μl:80nM)可进行对淋巴细胞的瞬时转染并最终实现CLEC2B基因沉默。 2. CLEC2B基因沉默后淋巴细胞增殖变化无显著统计学差异；淋巴细胞细胞因子表达变化：IL-4mRNA表达上调,sIL-2RmRNA表达下降,IL-6mRNA变化无统计学差异。 3. CLEC2B基因沉默后淋巴细胞上清液对黑素细胞增殖活性抑制减少,有利于黑素细胞存活。结论淋巴细胞中存在CLEC2B基因表达,且该基因沉默后可直接调节淋巴细胞多种细胞因子IL-4、sIL-2RmRNA的转录表达,但对淋巴细胞本身的增殖可能无明显影响。CLEC2B基因也可通过淋巴细胞上清液影响黑素细胞增殖活性,提示CLEC2B基因在淋巴细胞中的表达水平对调节其功能有重要作用并可通过淋巴细胞改变黑素细胞的生存环境。揭示了CLEC2B基因可通过调节细胞因子水平及间接影响黑素增殖参与白癜风发病的免疫学机制,为CLEC2B信号传导途径研究提供了新的实验依据。
[Abstract]:Purpose A novel vitiligo associated gene (CLEC2B) was studied by functional analysis to reveal its possible pathogenicity targets. Lymphocyte transient transfection system was established to silence the expression of CLEC2B gene in lymphocytes and to observe its regulatory effect on lymphocyte proliferation and cytokine IL-4mRNAsIL-2RmRNAs, IL-6mRNA expression. To observe the effect of lymphocyte supernatant of CLEC2B gene silencing on the proliferation of melanocytes, so as to reveal the immunological mechanism of CLEC2B gene involved in the pathogenesis of vitiligo. Method In this study, Jurkat lymphocytes and B16 melanoma cells were used to down-regulate the expression of CLEC2BmRNA in lymphocytes by RNA interference technique. The expression of IL-4mRNAsIL-2RmRNA-6 mRNA after CLEC2B gene silencing was analyzed by real-time fluorescence quantitative PCR. The lymphocyte proliferation after CLEC2B gene silencing and the effect of lymphocyte supernatant on the proliferation of melanocytes were detected by MTT assay. Result 1. DMRIE-C liposomes and CLEC2BsiRNA can transfect lymphocytes at a reasonable ratio of 2 渭 l: 80nM and finally achieve CLEC2B gene silencing. 2. There was no significant difference in lymphocyte proliferation after CLEC2B gene silencing, while there was no significant difference in the expression of cytokines and the up-regulation of IL-4 mRNA expression and the decrease of IL-2R mRNA expression. There was no significant difference in the changes of IL-6 mRNA expression. 3. After CLEC2B gene silencing, the lymphocyte supernatant inhibited the proliferation of melanocytes, which was beneficial to the survival of melanocytes. Conclusion There is CLEC2B gene expression in lymphocytes, and the silencing of this gene can directly regulate the transcription expression of IL-4 / sIL-2R mRNA in lymphocytes. However, the proliferation of melanocytes may not be affected by the proliferation of lymphocytes. CLEC2B gene can also affect the proliferation activity of melanocytes through the supernatant of lymphocytes. It is suggested that the expression of CLEC2B gene in lymphocytes plays an important role in regulating its function and can change the living environment of melanocytes through lymphocytes. It is revealed that CLEC2B gene can play an important role in the pathogenesis of vitiligo by regulating cytokine level and indirectly affecting melanin proliferation, which provides a new experimental basis for the study of CLEC2B signal transduction pathway.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R758.41

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