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目的 探讨重症肌无力(MG)患者细胞毒性T淋巴细胞相关抗原 -4 -(CTLA -4 )的表达状况及由CTLA -4基因启动区多态性导致的不同遗传易感性机制。方法 ELISA法测定MG患者血清中sCTLA 4的水平;限制性片段长度多态性分析用于检测启动区 1772、16 6 1位点的多态性;转录因子NF- 1和c -EBPβ结合位点通过染色质免疫沉淀实验(CHIP)得以验证。结果 MG患者血清sCTLA 4的表达水平与等位基因的突变相关,携带T→C -1 772 突变基因的患者可表达高水平的sCTLA -4。TC -1 772 基因型的频率在MG患者尤其是胸腺瘤亚组明显高于对照组,而G -1 6 6 1 等位基因和GG -1 6 6 1 基因型的频率在MG患者则显著降低。当 1772位点的等位基因是T而非C时,存在转录因子NF- 1结合位点;同样,当 16 6 1位点的等位基因是G而非A时,存在转录因子c -EBPβ结合位点,刀豆蛋白A(ConA)、植物血凝素(PHA)能促进NF -1和c- EBPβ的这种位点特异性转录活性。结论 MG患者CTLA 4表达异常,启动区C T 1 772 和A G 1 6 6 1 多态性可导致无效转录,T→C 1 772 的突变能影响基因的剪接,干扰蛋白的表达和功能,阻止了负性调节信号的传递而致发病。
Objective To investigate the expression of cytotoxic T lymphocyte antigen-4 (CTLA-4) in patients with myasthenia gravis (MG) and the mechanism of different genetic susceptibility to CTLA-4 promoter polymorphism. Methods Serum levels of sCTLA 4 in MG patients were measured by ELISA. Restriction fragment length polymorphism (RFLP) was used to detect the polymorphism of 1772,16 6 1 in promoter region. The transcription factor NF-1 and c -EBPβ binding sites Confirmed by Chromatin Immunoprecipitation Assay (CHIP). Results The serum level of sCTLA 4 in MG patients was correlated with allelic mutation. Patients with T → C -1 772 mutation could express high level of sCTLA -4. The frequency of TC-1 772 genotype was significantly higher in patients with MG, especially in thymoma subgroups, while the frequencies of G-1 6 6 1 allele and GG-1 6 6 1 genotype were significantly lower in MG patients . Similarly, when the allele at site 1772 is T but not C, there is a transcription factor NF-1 binding site; similarly, when the allele at site 1661 is G but not A, the transcription factor c -EBP [beta] The binding sites, ConA, PHA, promote this site-specific transcriptional activity of NF-I and c-EBP [beta]. Conclusions The expression of CTLA 4 in MG patients is abnormal. The polymorphisms of CT 1 772 and AG 1 6 6 1 in the promoter region can lead to inefficient transcription. The mutation of T → C 1 772 can affect the splicing and interfering protein expression and function of genes and prevent the negative Sexually Transmitted Signal Causes Incidence.