lncRNA HCG18靶向miR-34b-5p/FOXP1轴促进骨肉瘤细胞增殖、迁移和侵袭的机制研究

刘文斌; 李艳兵; 周焱涛

doi:10.13898/j.cnki.issn.1000-2200.2023.06.004

lncRNA HCG18靶向miR-34b-5p/FOXP1轴促进骨肉瘤细胞增殖、迁移和侵袭的机制研究

Study on the mechanism of lncRNA HCG18 promoting the proliferation, migration and invasion of osteosarcoma cells by targeting miR-34b-5p/FOXP1 axis

摘要

摘要:
目的探讨长链非编码RNA(lncRNA)人类白细胞抗原复合体18(HCG18)对骨肉瘤细胞增殖、迁移和侵袭的影响及可能机制。
方法收集47例骨肉瘤组织及对应瘤旁组织，RT-qPCR法检测组织中HCG18和miR-34b-5p表达，Pearson相关分析评价骨肉瘤组织中HCG18和miR-34b-5p表达的相关性。体外培养骨肉瘤U2OS细胞，分别转染si-HCG18、miR-34b-5p mimics、共转染si-HCG18与anti-miR-34b-5p或miR-34b-5p mimics与pcDNA-FOXP1后，采用CCK-8法和克隆形成实验检测细胞增殖，划痕实验和Transwell分别检测细胞迁移和侵袭，蛋白印迹法检测细胞中FOXP1蛋白表达。双荧光素酶报告基因实验验证HCG18和miR-34b-5p及miR-34b-5p和FOXP1调控关系。
结果骨肉瘤组织中HCG18表达量高于瘤旁组织(P < 0.01)，而miR-34b-5p表达量低于瘤旁组织(P < 0.01)。Pearson相关分析结果显示，骨肉瘤组织中HCG18与miR-34b-5p的表达呈负相关关系(r=-0.812, P < 0.05)。下调HCG18或上调miR-34b-5p后，U2OS细胞吸光度值和克隆形成数降低(P < 0.05)，划痕愈合率和细胞侵袭数降低(P < 0.05)。HCG18靶向负调控miR-34b-5p，而FOXP1是miR-34b-5p靶基因。下调miR-34b-5p逆转下调HCG18对骨肉瘤U2OS细胞增殖、迁移和侵袭的抑制作用。上调FOXP1可逆转上调miR-34b-5p对骨肉瘤U2OS细胞增殖、迁移和侵袭的抑制作用。
结论HCG18在骨肉瘤组织中表达升高，其可能通过靶向miR-34b-5p/FOXP1轴促进骨肉瘤细胞增殖、迁移和侵袭。

Abstract:
ObjectiveTo investigate the effect and possible mechanism of long non-coding RNA (lncRNA) human leukocyte antigen complex 18 (HCG18) on the proliferation, migration and invasion of osteosarcoma cells.
MethodsForty-seven cases of osteosarcoma tissues and corresponding adjacent tissues were collected. RT-qPCR was used to detect the expression of HCG18 and miR-34b-5p in the tissues. Pearson correlation analysis was used to evaluate the correlation between the expression of HCG18 and miR-34b-5p in osteosarcoma tissue. Osteosarcoma U2OS cells were cultured in vitro and transfected with si-HCG18, miR-34b-5p mimics, co-transfected with si-HCG18 and anti-miR-34b-5p or miR-34b-5p mimics and pcDNA-FOXP1. Then CCK-8 method and colony formation test were used to detect cell proliferation, scratch test and Transwell were used to detect cell migration and invasion, and Western blotting was used to detect the expression of FOXP1 protein in cells. The dual luciferase reporter gene experiment was applied to verify the regulatory relationship between HCG18 and miR-34b-5p, and between miR-34b-5p and FOXP1.
ResultsCompared with adjacent tissues, the expression of HCG18 in osteosarcoma tissue was increased (P < 0.01), and the expression of miR-34b-5p was decreased (P < 0.01). Pearson correlation analysis showed that the expression HCG18 and miR-34b-5p in osteosarcoma tissue was negatively correlated (r=-0.812, P < 0.05). After down-regulating HCG18 or up-regulating miR-34b-5p, the OD value of U2OS cells and number of clone formation decreased (P < 0.05), and scratch healing rate and number of invaded cells decreased (P < 0.05). HCG18 targeted and negatively regulated miR-34b-5p, while FOXP1 was the target gene of miR-34b-5p. Down-regulating miR-34b-5p reversed the inhibitory effect of down-regulating HCG18 on the proliferation, migration and invasion of osteosarcoma U2OS cells. Up-regulating FOXP1 reversed the inhibitory effect of up-regulating miR-34b-5p on the proliferation, migration and invasion of osteosarcoma U2OS cells.
ConclusionsThe expression of HCG18 is elevated in osteosarcoma tissues, which may promote the proliferation, migration and invasion of osteosarcoma cells by targeting the miR-34b-5p/FOXP1 axis.

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