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目的:探讨细胞凋亡清除障碍及瓣状内切核酸酶1(Fen1)基因变异在狼疮性肾炎(LN)发病机制中的作用。方法:首先收集30例LN患者肾穿刺组织,用TUNEL法检测细胞凋亡,探讨LN病理分型、系统性红斑狼疮(SLE)活动度评分2000(SLEDAI2000)与肾脏组织细胞凋亡指数(AI)的关系;其次,收集50例LN患者,25例健康者,20例胃癌患者的外周血标本,采用全血基因组DNA柱式试剂盒提取DNA,筑巢PCR扩增Fen1基因61563142～61563342目的片段,扩增后对目的基因进行测序,并将测序结果与基因数据库中人Fen1基因进行比较,计算突变位点频率,比较LN患者、胃癌患者、健康者Fen1基因突变情况。结果:(1)所有患者肾组织凋亡检测均为阳性,凋亡细胞在肾小球及肾小管均呈散在分布,SLEDAI与其AI呈正相关(r=0.39,P<0.05)。根据ISN/RPS2003制定的病理分型标准,30例LN患者中,病理类型以Ⅳ型最多(40.00%),其次为Ⅱ型(26.67%),Ⅲ型16.67%,Ⅴ型13.33%,Ⅵ型3.33%,无Ⅰ型病变患者。(2)三组Fen1基因61563142～61563342目的片段未发现有统计学意义的碱基变异。结论:LN患者存在细胞凋亡清除障碍,凋亡细胞的数量越多,SLE病情越活跃;LN患者Fen1基因61563142～61563342目的片段中未发现碱基变异,尚需要扩大样本量,或进行该基因的全基因组关联研究进一步证实Fen1基因在LN中的作用。
OBJECTIVE: To investigate the role of apoptosis clearance and variation of flap-like endonuclease-1 (Fen1) gene in the pathogenesis of lupus nephritis (LN). Methods: Thirty renal biopsy specimens from patients with LN were collected and TUNEL method was used to detect apoptosis. The pathological types of LN, SLE activity 2000 (SLEDAI2000) and apoptosis index (AI) Secondly, peripheral blood samples of 50 patients with LN, 25 healthy individuals and 20 patients with gastric cancer were collected. DNA was extracted by whole blood genomic DNA column kit and nested for PCR amplification of Fen6 gene 61563142 ~ 61563342, After amplification, the target gene was sequenced, and the sequencing results were compared with that of human Fen1 gene in the gene database to calculate the mutation site frequency. The mutations of Fen1 gene in LN patients, gastric cancer patients and healthy subjects were compared. Results: (1) Apoptosis was detected in all the renal tissues. Apoptotic cells were scattered in the glomerulus and tubules. SLEDAI was positively correlated with AI (r = 0.39, P <0.05). According to the criteria of pathological classification established by ISN / RPS2003, the pathological type of type Ⅳ was the most (40.00%), followed by type Ⅱ (26.67%), type Ⅲ 16.67%, type V 13.33% and type Ⅲ 3.33 %, No type I disease patients. (2) There was no statistically significant nucleotide variation among the three groups of 61563142 ~ 61563342 Fen1 genes. CONCLUSION: There is a disorder of cell apoptosis in LN patients. The more the number of apoptotic cells, the more active the SLE patients. In LN patients, there is no base variation in Fengyang gene 61563142 ~ 61563342, but the sample size needs to be expanded or the gene Genome-wide association study further confirmed Fen1 gene in LN.