circVRK1靶向miR-4428调控急性淋巴细胞白血病KOCL44细胞增殖及凋亡的分子机制研究  

Molecular Mechanism of circVRK1 Regulating the Proliferation and Apoptosis of Acute Lymphoblastic Leukemia KOCL44 Cells by Targeting miR-4428

作　　者：张欢[1] 吴斌[1] 王月娇 ZHANG Huan;WU Bin;WANG Yuejiao(Department of Hematology,Shengjing Hospital Affiliated to China Medical University,Shenyang 110004,China;Department of Rheumatology and Immunology,Shengjing Hospital Affiliated to China Medical University,Shenyang 110004,China)

机构地区：[1]中国医科大学附属盛京医院血液内科,沈阳110004 [2]中国医科大学附属盛京医院风湿免疫科,沈阳110004

出　　处：《四川大学学报（医学版）》2024年第4期872-877,共6页Journal of Sichuan University(Medical Sciences)

基　　金：国家自然科学基金(No.82101898);辽宁省民生科技计划联合计划项目(No.2021JH2/10300092)资助。

摘　　要：目的 分析circVRK1和miR-4428在急性淋巴细胞白血病(acute lymphoblastic leukemia, ALL)细胞增殖和凋亡中的关系。方法 体外培养ALL细胞KOCL44,实验分组为:pcDNA、pcDNA-circVRK1、anti-miR-NC、anti-miR-4428、si-NC、si-circVRK1、pcDNA-circVRK1+miR-NC和pcDNA-circVRK1+miR-4428组。qRT-PCR检测细胞circVRK1和miR-4428的表达水平;CCK-8法、流式细胞术分别检测细胞增殖及凋亡;双荧光素酶报告实验检测circVRK1与miR-4428的靶向关系[实验分为circVRK1野生型报告质粒(WT-circVRK1)+miR-NC、WT-circVRK1+miR-4428、circVRK1突变报告质粒(MUT-circVRK1)+miR-NC和MUT-circVRK1+miR-4428组];Western blot检测Ki-67、cleaved caspase-3、cleaved caspase-9蛋白表达量。结果 相较于pcDNA组,pcDNA-circVRK1组的circVRK1表达上调(P<0.05);与转染pcDNA或者转染anti-miRNC相比,转染pcDNA-circVRK1或anti-miR-4428后,KOCL44细胞活力和Ki-67蛋白表达降低(P<0.05),凋亡率和cleaved caspase-3、cleaved caspase-9蛋白水平增加(P<0.05);circVRK1可负向调控miR-4428的表达,但该作用仅在转染WTcircVRK1的组别中体现;与pcDNA组相比,pcDNA-circVRK1组miR-4428表达降低(P<0.05);与si-NC组相比,si-circVRK1组miR-4428表达增加(P<0.05);与共转染pcDNA-circVRK1+miR-NC相比,共转染pcDNA-circVRK1+miR-4428后细胞活力升高(P<0.05),Ki-67蛋白表达增加(P<0.05),凋亡率和cleaved caspase-3、cleaved caspase-9蛋白水平降低(P<0.05)。结论 circVRK1过表达可通过下调miR-4428表达而减弱ALL细胞增殖能力及诱导细胞凋亡。Objective To elucidate the role of circVRK1 and its interaction with miR-4428 in regulating proliferation and apoptosis in acute lymphoblastic leukemia(ALL)cells.Methods KOCL44 ALL cells were cultured in vitro,and experimental groups included pcDNA,pcDNA-circVRK1,anti-miR-NC,anti-miR-4428,si-NC,si-circVRK1,pcDNA-circVRK1+miR-NC,and pcDNA-circVRK1+miR-4428.The expression levels of circVRK1 and miR-4428 were detected using qRT-PCR.CCK-8 assays and flow cytometry were used to assess cell proliferation and apoptosis,respectively.The dual luciferase reporter assays were employed to investigate the interaction between circVRK1 and miR-4428,with groups categorized as WT-circVRK1+miR-NC,WT-circVRK1+miR-4428,MUT-circVRK1+miR-NC,and MUT-circVRK1+miR-4428.Western blotting was utilized to detect the expression levels of Ki-67,cleaved caspase-3,and cleaved caspase-9 proteins.Results Compared to the pcDNA group,circVRK1 expression was up-regulated in the pcDNA-circVRK1 group(P<0.05).Compared to transfection with pcDNA or anti-miR-NC,transfection with pcDNA-circVRK1 or anti-miR-4428 led to decreased cell viability and Ki-67 protein levels in KOCL44 cells(P<0.05),and increased apoptosis rates and levels of cleaved caspase-3 and cleaved caspase-9(P<0.05).circVRK1 was found to negatively regulate miR-4428 expression,with this effect observed only in the WT-circVRK1 group.miR-4428 levels were lower in the pcDNA-circVRK1 group compared to the pcDNA group(P<0.05)and higher in the si-circVRK1 group compared to the si-NC group(P<0.05).Co-transfection with pcDNA-circVRK1+miR-4428 resulted in increased cell viability(P<0.05)and Ki-67 expression(P<0.05),and decreased apoptosis rates and levels of cleaved caspase-3 and cleaved caspase-9(P<0.05)compared to co-transfection with pcDNA-circVRK1+miR-NC.Conclusion Overexpression of circVRK1 reduces the proliferation ability of acute ALL cells and induces cell apoptosis by downregulating miR-4428 expression.

关 键 词：急性淋巴细胞白血病 circVRK1 miR-4428 细胞增殖 细胞凋亡 

分 类 号：R733.71[医药卫生—肿瘤]