circ_NEK6/miR-503-5p对多发性骨髓瘤细胞增殖、凋亡及迁移的影响

Effect of circ_NEK6/miR-503-5p on the Proliferation,Apoptosis and Migration of Multiple Myeloma Cells

目的探讨circ_NEK6/miR-503-5p分子轴对多发性骨髓瘤细胞增殖、凋亡及迁移的影响及其可能的作用机制。方法选取2020年2月至2021年7月遵义医科大学第三附属医院收治的30例多发性骨髓瘤患者为研究组,同时选取在该院体检的55例健康志愿者为对照组;qRT-PCR法检测血清中circ_NEK6、miR-503-5p的表达量;体外培养人多发性骨髓瘤U266细胞,分为si-circ_NEK6组、miR-503-5p组和si-circ_NEK6+anti-miR-503-5p组以及相应的对照组(si-NC组、miR-NC组、si-circ_NEK6+anti-miR-NC组);细胞增殖、凋亡及迁移用MTT法、克隆形成实验、流式细胞术与Transwell实验测定;双荧光素酶报告基因实验验证circ_NEK6与miR-503-5p之间的靶向关系;Western blot检测Bax、Bcl-2蛋白表达量。结果与对照组比较,研究组患者血清中circ_NEK6高表达(P<0.05),miR-503-5p低表达(P<0.05);si-circ_NEK6组U266细胞miR-503-5p的表达、细胞增殖抑制率、凋亡率和Bax水平增加(均P<0.05),而克隆数、迁移数和Bcl-2水平降低(均P<0.05);circ_NEK6可靶向结合miR-503-5p;在miR-503-5p组U266细胞,细胞增殖抑制率、凋亡率和Bax水平上升(均P<0.05),而克隆数、迁移数和Bcl-2水平降低(均P<0.05);此外,si-circ_NEK6+anti-miR-503-5p组细胞增殖抑制率、凋亡率和Bax水平降低(均P<0.05),而克隆数、迁移数和Bcl-2水平升高(P<0.05)。结论敲低circ_NEK6可以通过靶向miR-503-5p降低多发性骨髓瘤细胞生长和迁移能力。

Objective To explore the effect of circ_NEK6/miR-503-5p axis on the proliferation,apoptosis and migration of multiple myeloma(MM)cells and its possible mechanism.Methods Thirty patients with MM admitted in our hospital from February 2020to July 2021were selected as the study group,and 55healthy volunteers who received physical examination in our hospital were selected as the control group.The qRT-PCR method was used to detect the expression of circ_NEK6and miR-503-5p in serum.U266cells were cultured for in vitro assay,and cells were randomly divided into the following groups:si-circ_NEK6group,miR-503-5p group,si-circ_NEK6+anti-miR-503-5p group and negative control group(si-NC group,miR-NC group,and si-circ_NEK6+anti-miR-NC group).MTT assay,colony formation assay,flow cytometry assay and Transwell assay were used to detect cell proliferation,apoptosis and migration.Dual-luciferase reporter assay was used to detect the targeting relationship between circ_NEK6and miR-503-5p.Western blot was used to detect the expression of Bax and Bcl-2protein.Results Compared with the control group,the expression of circ_NEK6was increased(P<0.05),while the expression of miR-503-5p was decreased(P<0.05)in the serum of MM patients.The miR-503-5p expression,cell proliferation inhibition rate,apoptosis rate and Bax level were increased(all P<0.05),while cell colony number,cell migration number and Bcl-2level were decreased(all P<0.05)in the si-circ_NEK6group.The circ_NEK6could target miR-503-5p.Cell proliferation inhibition rate,apoptosis rate and Bax level were increased(all P<0.05),while cell colony number,cell migration number and Bcl-2level were decreased(all P<0.05)in the miR-503-5p group.Besides,cell proliferation inhibition rate,apoptosis rate and Bax level were decreased(all P<0.05),while cell colony number,cell migration number and Bcl-2level were increased(all P<0.05)in the si-circ_NEK6+anti-miR-503-5p group.Conclusion Knockdown of circ_NEK6could inhibit the growth and migration of MM cells by targeting miR-503-5p.