CAJ | 학술논문

目的探讨系统性红斑狼疮(SLE)患者T淋巴细胞CD70mRNA和蛋白的表达及CD70基因启动子DNA甲基化的状态.方法分离15例活动期SLE患者、15例非活动期SLE患者和15例健康对照外周血CD4+与CD8+细胞,用实时定量逆转录PCR(RT-PCR)方法检测CD4+和CD8+细胞CD70 mRNA转录水平,流式细胞仪检测CD4+CD70+细胞和CD8+CD70+细胞百分率,亚硫酸氢钠基因测序法检测CD4+细胞和CD8+细胞CD70基因启动子区域甲基化水平.组间比较采用单因素方差分析,组间两两比较采用SNK-q检验.结果 ①活动期、非活动期SLE患者CD4+细胞CD70 mRNA转录水平分别为0.82±0.12和0.73±0.11,明显高于健康对照组(0.45±0.09),F=53.017,P＜ 0.01,活动期SLE患者CD4+胞的CD70 mRNA转录水平显著高于非活动期SLE患者(P＜0.05).活动期、非活动期SLE患者外周血CD4+CD70+细胞百分率分别为80.30％±11.04％和66.80％±3.98％,明显高于健康对照组(12.48％±3.45％),F=311.517,P＜ 0.01,活动期SLE患者CD4+CD70+细胞百分率显著高于非活动期SLE患者(P值＜0.05).SLE患者外周血CD70+CD4+细胞百分率与SLE疾病活动度呈显著正相关(r=0.792,P=0.000).活动期、非活动期SLE患者组的CD4+细胞CD70基因启动子序列-600～-300 bp区域平均甲基化水平分别为0.32±0.05和0.36±0.05,明显低于健康对照组(0.62±0.05),F=152.64,P＜0.01,活动期平均甲基化水平明显低于非活动期SLE患者组(P＜0.05).结论 CD4+细胞CD70基因启动子区域处于低甲基化状态,这种低甲基化状态可能是CD70过度表达的直接原因.
목적 탐토계통성홍반랑창(SLE)환자T림파세포CD70mRNA화단백적표체급CD70기인계동자DNA갑기화적상태.방법 분리15례활동기SLE환자、15례비활동기SLE환자화15례건강대조외주혈CD4+여CD8+세포,용실시정량역전록PCR(RT-PCR)방법검측CD4+화CD8+세포CD70 mRNA전록수평,류식세포의검측CD4+CD70+세포화CD8+CD70+세포백분솔,아류산경납기인측서법검측CD4+세포화CD8+세포CD70기인계동자구역갑기화수평.조간비교채용단인소방차분석,조간량량비교채용SNK-q검험.결과 ①활동기、비활동기SLE환자CD4+세포CD70 mRNA전록수평분별위0.82±0.12화0.73±0.11,명현고우건강대조조(0.45±0.09),F=53.017,P＜ 0.01,활동기SLE환자CD4+포적CD70 mRNA전록수평현저고우비활동기SLE환자(P＜0.05).활동기、비활동기SLE환자외주혈CD4+CD70+세포백분솔분별위80.30％±11.04％화66.80％±3.98％,명현고우건강대조조(12.48％±3.45％),F=311.517,P＜ 0.01,활동기SLE환자CD4+CD70+세포백분솔현저고우비활동기SLE환자(P치＜0.05).SLE환자외주혈CD70+CD4+세포백분솔여SLE질병활동도정현저정상관(r=0.792,P=0.000).활동기、비활동기SLE환자조적CD4+세포CD70기인계동자서렬-600～-300 bp구역평균갑기화수평분별위0.32±0.05화0.36±0.05,명현저우건강대조조(0.62±0.05),F=152.64,P＜0.01,활동기평균갑기화수평명현저우비활동기SLE환자조(P＜0.05).결론 CD4+세포CD70기인계동자구역처우저갑기화상태,저충저갑기화상태가능시CD70과도표체적직접원인.
Objective To detect the expressions of CD70 mRNA and protein and to determine the methylation status of CD70 gene promoter in T cells from patients with systemic lupus erythematosus (SLE).Methods Peripheral blood CD4+ and CD8+ T cells were isolated from 15 patients with active SLE,15 patients with inactive SLE and 15 healthy control subjects.Real-time quantitative reverse transcription-PCR was carried out to quantify the mRNA expression of CD70,flow cytometry to determine the frequency of CD4+CD70+ and CD8+ CD70+ T cells,and bisulfite sequencing to evaluate the methylation status of CD70 gene promoter in CD4+ and CD8+ T cells.Differences in these parameters among these groups were analyzed by one-factor analysis of variance and SNK-q test.Results Compared with the healthy controls,the patients with active SLE and inactive SLE showed a significant increase in CD70 mRNA expression in CD4+ T cells (0.82 ± 0.12 and 0.73 ± 0.11 vs.0.45 ±0.09,F =53.017,P ＜ 0.01) and in the frequency of CD70+CD4+ T cells (80.30％ ± 11.04％ and 66.80％ ± 3.98％ vs.12.48％ ± 3.45％,F =311.517,P ＜ 0.01).Also,the expression of CD70 mRNA in CD4+ T cells and the frequency of CD70+CD4+ T cells were significantly higher in patients with active SLE than in patients with inactive SLE (both P ＜ 0.05).There was a positive correlation between the frequency of peripheral CD70+CD4+ T cells and disease activity in SLE in these patients (r =0.792,P ＜ 0.01).The average methylation index of the region between-600 bp and-300 bp of CD70 gene promoter in CD4+ T cells was 0.32 ± 0.05 and 0.36 ± 0.05 respectively in the patients with active and inactive SLE,significantly lower than that in the healthy controls (0.62 ± 0.05,F =152.64,P ＜ 0.01),and the patients with active SLE showed a significantly lower methylation index than those with inactive SLE (P ＜ 0.05).Conclusions The CD70 gene promoter in CD4+ T cells is significantly hypomethylated in patients with SLE,which may directly lead to the overexpression of CD70.