组蛋白去甲基化酶JMJD3促进弥漫性大B细胞淋巴瘤细胞的干性

Histone demethylase JMJD3 promotes the stemness of diffuse large B cell lymphoma cells

目的:探讨组蛋白去甲基化酶JMJD3(jumonji domain-containing protein 3)对弥漫性大B细胞淋巴瘤(diffuse large B-cell lymphoma,DLBCL)细胞干性的影响。方法:利用TCGA数据库中DLBCL患者的临床资料,分析JMJD3的表达水平与DLBCL患者总生存率的关系。应用脂质体转染方法分别将对照质粒(pCMV)和JMJD3表达质粒(pCMV-JMJD3)转染到ABC和GCB亚型DLBCL细胞中,用RT-PCR和qPCR检测转染细胞中JMJD3、ALDH1、OCT4和SOX2 mRNA表达水平,用流式细胞术检测细胞中ALDH1酶活性,用Western blotting检测细胞中OCT4和SOX2蛋白的表达水平。通过基因集富集分析法(gene set enrichment analysis,GSEA)分析高表达JMJD3的DLBCL患者的基因集富集情况。结果:患者预后分析结果显示,高表达水平的JMJD3与DLBCL患者的不良预后有关(P<0.05),但多因素分析结果显示JMJD3的表达水平不是DLBCL患者预后的独立影响因素(均P>0.05)。pCMV-JMJD3转染后细胞中JMJD3表达显著增加,同时导致DLBCL细胞中ALDH1 mRNA水平和酶活性以及OCT4和SOX2 mRNA和蛋白的表达水平均明显升高(P<0.05或P<0.01)。GSEA分析结果显示,高表达JMJD3的DLBCL患者样本富集于Wnt/β-catenin信号通路基因集(P<0.05)。结论:JMJD3具有促进DLBCL细胞干性的作用,其可能是DLBCL患者潜在的治疗靶点。

Objective: To investigate the effect of histone demethylase JMJD3(jumonji domain-containing protein 3) on the stemness of diffuse large B-cell lymphoma(DLBCL) cells. Methods: The relationship between the expression of JMJD3 and the overall survival of DLBCL patients was analyzed using the clinical data of DLBCL patients in TCGA database. The control plasmid(pCMV) and JMJD3 expression plasmid(pCMV-JMJD3) were transfected into DLBCL cells of ABC and GCB subtype via lipofectamine transfection.Then, the mRNA levels of JMJD3, ALDH1, OCT4 and SOX2 were detected by RT-PCR and qPCR;the activity of ALDH1 enzyme was detected by Flow cytometry;the protein expressions of OCT4 and SOX2 were detected by Western blotting. Gene enrichment in DLBCL patients with high JMJD3 expression was analyzed by gene set enrichment analysis(GSEA). Results: The result of prognosis analysis showed that high expression of JMJD3 was related with poor prognosis in DLBCL patients(P<0.05);however, multivariate analysis showed that the expression of JMJD3 was not the independent factor affecting the prognosis of DLBCL patients(all P>0.05).The expression of JMJD3 was remarkably increased in DLBCL cells transfected with pCMV-JMJD3, which led to significantly increased m RNA level and enzyme activity of ALDH1 as well as up-regulated mRNA and protein expressions of OCT4 and SOX2(P<0.05 or P<0.01). GSEA analysis showed that enrichment of Wnt/β-catenin signaling pathway related gene set was observed in DLBCL patients with high JMJD3 expression(P<0.05). Conclusion: JMJD3 promotes the stemness of DLBCL cells, which may be a potential therapeutic target for DLBCL patients.