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目的:建立荧光定量PCR法检测CD3ε链表达水平的方法,了解多发性骨髓瘤(MM)T细胞信号传导分子CD3ε基因的表达特点。方法:采用SYBR GreenⅠ荧光定量PCR和相对定量分析法检测22例MM患者及22例正常人外周血单个核细胞CD3ε基因的表达情况,以β2微球蛋白基因(β2M)作为内参,22例健康人作为对照,采用相对定量公式:2-△Ct×100%,计算MM患者CD3ε基因相对mRNA表达量。结果:MM患者和正常人外周血单个核细胞均表达CD3ε基因,具体表达量呈现很大的个体差异性。MM患者CD3ε基因mRNA相对表达量最高为39.78%,最低为0.48%,正常人CD3ε基因mRNA相对表达量最高为5.15%,最低为0.01%。MM患者CD3ε基因表达量明显高于正常人(P<0.05)。结论:成功建立SYBR GreenⅠ荧光定量PCR法检测CD3ε链表达水平的方法,率先报道了多发性骨髓瘤中CD3ε基因高表达的特点,为了解多发性骨髓瘤T细胞免疫学特点提供新的资料。
OBJECTIVE: To establish a method for detecting the expression of CD3ε chain by fluorescence quantitative PCR and to investigate the expression characteristics of CD3ε gene in multiple myeloma (MM) T cells. Methods: The expression of CD3ε gene in peripheral blood mononuclear cells from 22 patients with MM and 22 normal controls was detected by SYBR Green Ⅰ real-time quantitative PCR and relative quantitative analysis. The β2 microglobulin gene (β2M) As a control, the relative quantification formula: 2-Δ Ct × 100% was used to calculate the relative mRNA level of CD3ε gene in MM patients. Results: CD3ε gene was expressed in peripheral blood mononuclear cells of MM patients and normal individuals, and the specific expression of CD3ε gene showed a great individual difference. The highest relative expression of CD3ε mRNA in MM patients was 39.78%, the lowest was 0.48%. The highest relative expression of CD3ε mRNA in normal subjects was 5.15% and the lowest was 0.01%. The expression of CD3ε gene in MM patients was significantly higher than that in normal controls (P <0.05). Conclusion: The SYBR Green Ⅰ fluorescent quantitative PCR method was successfully established to detect the expression of CD3 epsilon chain. The first report of the high expression of CD3 epsilon gene in multiple myeloma was provided, and new information was provided to understand the characteristics of T cell immunology in multiple myeloma.