寻常性银屑病患者外周血T细胞受体β可变区优势表达分析

目的 分析寻常性银屑病患者外周血中T细胞受体β可变区(TRBV)的优势表达状况,探讨其在银屑病发病中的作用.方法 以人功能性TRBV家族设计33个上游引物,共同的T细胞受体β恒定区(TRBC)基因设计下游引物,在T细胞受体α恒定区(TRAC)设计引物作为内参,分析寻常性银屑病患者、正常人对照外周血样本各10例,检测各PCR反应管荧光强度,以大于正常人外周血T细胞相应TRBV基因相对表达量(-x+3s)筛选TRBV优势表达基因家族.结果 TRAC扩增产物阈值循环数(Ct)值集中于21～24,银屑病组TRBV2扩增产物Ct与TRAC产物Ct差值平均数银屑病患者为2.98,TRBV5-7为3.24,TRBV6-6/6-9为2.52,TRBV12为2.04,TRBV24为3.56,TRBV29为4.12,其表达与正常人比较均明显增高(P值均＜0.05),其中TRBV6-6/6-9、TRBV12、TRBV29在银屑病患者呈优势表达.结论 银屑病患者外周血TRBV基因家族存在优势表达,可能在银屑病T细胞异常免疫反应中起重要作用.

Abstract：

Objective To assess the preferential expressions of peripheral blood T cell receptor beta chain variable region (TRBV) subfamilies in patients with psoriasis vulgaris(PV), and to estimate their role in the pathogenesis of psoriasis. Methods Thirty-three upstream primers were designed to target the human functional TRBV genes, downstream primers to target the common T cell receptor beta constant (TRBC) gene,with T cell receptor alpha constant (TRAC) gene as the internal reference. Total RNA was extracted from the peripheral blood T cells of 10 health human controls and 10 patients with PV, and transcribed into cDNA.Then, TRBV genes were amplified by real-time fluorescence quantitative PCR (RFQ-PCR) and the fluorescence intensity of each samples was detected. The expression levels of TRBV genes in the control group were used to calculate the cut-off values (mean expression levels of TRBV subfamilies in the 10 normal controls + 3 standard deviations). When the expression level of a TRBV subfamily from patients with PV was equal to or higher than the cut-off value, it was considered as the preferentially expressed TRBV subfamily. Results The threshold cycle (Ct) value varied from 21 to 24 for TRAC gene. The difference in the Ct value between TRBV subfamily genes and TRAC gene in patients with PV was 2.98 for TRBV2 gene, 3.24 for TRBV5-7 gene, 2.52 for TRBV6-6/6-9 gene, 2.04 for TRBV 12 gene, 3.56 for TRBV 24 gene, and 4.12 for TRBV 29 gene, and the expression levels of these subfamily genes were significantly higher than those in the normal controls (all P ＜ 0.05). According to the above standard, TRBV6-6/6-9, TRBV12 and TRBV29 were considered to be preferentially expressed subfamilies. Conclusions There is a preferential expression of TRBV gene subfamilies in peripheral blood of patients with psoriasis vulgaris, which may play a vital role in the abnormal T cell-mediated immune responses in psoriasis.

Preferential expressions of peripheral blood T cell receptor beta chain variable region subfamilies in patients with psoriasis vulgaris

Objective To assess the preferential expressions of peripheral blood T cell receptor beta chain variable region (TRBV) subfamilies in patients with psoriasis vulgaris(PV), and to estimate their role in the pathogenesis of psoriasis. Methods Thirty-three upstream primers were designed to target the human functional TRBV genes, downstream primers to target the common T cell receptor beta constant (TRBC) gene,with T cell receptor alpha constant (TRAC) gene as the internal reference. Total RNA was extracted from the peripheral blood T cells of 10 health human controls and 10 patients with PV, and transcribed into cDNA.Then, TRBV genes were amplified by real-time fluorescence quantitative PCR (RFQ-PCR) and the fluorescence intensity of each samples was detected. The expression levels of TRBV genes in the control group were used to calculate the cut-off values (mean expression levels of TRBV subfamilies in the 10 normal controls + 3 standard deviations). When the expression level of a TRBV subfamily from patients with PV was equal to or higher than the cut-off value, it was considered as the preferentially expressed TRBV subfamily. Results The threshold cycle (Ct) value varied from 21 to 24 for TRAC gene. The difference in the Ct value between TRBV subfamily genes and TRAC gene in patients with PV was 2.98 for TRBV2 gene, 3.24 for TRBV5-7 gene, 2.52 for TRBV6-6/6-9 gene, 2.04 for TRBV 12 gene, 3.56 for TRBV 24 gene, and 4.12 for TRBV 29 gene, and the expression levels of these subfamily genes were significantly higher than those in the normal controls (all P ＜ 0.05). According to the above standard, TRBV6-6/6-9, TRBV12 and TRBV29 were considered to be preferentially expressed subfamilies. Conclusions There is a preferential expression of TRBV gene subfamilies in peripheral blood of patients with psoriasis vulgaris, which may play a vital role in the abnormal T cell-mediated immune responses in psoriasis.