长链非编码RNA C5orf66-AS1通过靶向miR-637对胃癌细胞增殖、凋亡的影响

胡广军; 宗殿亮; 孙清森; 赵俊卿; 张新如; 谷斌

doi:10.13898/j.cnki.issn.1000-2200.2023.07.006

长链非编码RNA C5orf66-AS1通过靶向miR-637对胃癌细胞增殖、凋亡的影响

Effects of long non-coding RNA C5orf66-AS1 on the proliferation and apoptosis of gastric cancer cells by targeting miR-637

摘要

摘要:
目的探讨长链非编码RNA C5orf66-AS1通过靶向miR-637对胃癌(GC)细胞增殖、凋亡的影响，为GC临床治疗提供依据。
方法收集32例GC病人的癌组织及其癌旁组织并体外培养人胃腺癌细胞系MKN28、MKN451、BGC823、MGC803、AGS及正常胃上皮细胞系GES-1，qRT-PCR实验检测其C5orf66-AS1和miR-637表达。将对数生长期的AGS细胞利用脂质体转染试剂进行转染并分为对照组、GV144组、GV144-C5orf66-AS1组、miR-NC组、miR-637 mimics组、GV144-C5orf66-AS1+miR-NC组、GV144-C5orf66-AS1+miR-637 mimics组。CCK-8法检测细胞增殖能力；流式细胞术分析细胞凋亡情况；Western blotting实验测定细胞周期蛋白CyclinD1、B淋巴细胞瘤-2基因(Bcl-2)及其同源蛋白Bax相对表达水平；RNA免疫共沉淀实验验证C5orf66-AS1和miR-637的作用关系。
结果与癌旁组织或正常胃上皮细胞系GES-1相比，C5orf66-AS1和miR-637在GC组织和细胞系MKN28、MKN451、BGC823、MGC803、AGS中的表达水平均降低(P < 0.05)，选择C5orf66-AS1表达水平最低的AGS细胞进行转染实验。转染GV144-C5orf66-AS1后，AGS细胞中C5orf66-AS1、miR-637表达水平、凋亡率及Bax蛋白表达升高，细胞活性(48、72、96 h)及CyclinD1、Bcl-2蛋白降低(P < 0.05)。C5orf66-AS1和miR-637存在相互作用关系。miR-637过表达可使C5orf66-AS1过表达对AGS细胞增殖的抑制作用和凋亡的促进作用更明显(P < 0.05)。
结论C5orf66-AS1和miR-637在GC组织和细胞中低表达，过表达C5orf66-AS1和上调miR-637，可协同抑制GC细胞的增殖，促进凋亡。

Abstract:
ObjectiveTo investigate the effects of long non-coding RNA C5orf66-AS1 on the proliferation and apoptosis of gastric cancer (GC) cells by targeting miR-637, so as to provide a basis for the clinical treatment of GC.
MethodsThe cancer tissues and adjacent tissues of 32 GC patients were collected.Human gastric adenocarcinoma cell lines MKN28, MKN451, BGC823, MGC803, AGS and normal gastric epithelial cell line GES-1 were cultured in vitro.The expression of C5orf66-AS1 and miR-637 was detected by qRT-PCR.AGS cells in logarithmic growth phase were transfected with liposome transfection reagent and divided into control group, GV144 group, GV144-C5orf66-AS1 group, miR-NC group, miR-637 mimics group, GV144-C5orf66-AS1 + miR-NC group, and GV144-C5orf66-AS1 + miR-637 mimics group.CCK-8 method was used to detect cell proliferation ability, flow cytometry was used to analyze the apoptosis, Western blotting was used to determine the expression levels of CyclinD1, B-cell lymphoma-2 gene (Bcl-2) and its homologous protein Bax of cells.RNA immunoprecipitation experiment was used to verify the relationship between C5orf66-AS1 and miR-637.
ResultsCompared with adjacent tissues or normal gastric epithelial cell line GES-1, the expression levels of C5orf66-AS1 and miR-637 in GC tissues and cell lines MKN28, MKN451, BGC823, MGC803, AGS were reduced (P < 0.05).The AGS cells with the lowest expression level of C5orf66-AS1 were selected for transfection experiments.After transfection of GV144-C5orf66-AS1, the C5orf66-AS1, miR-637 expression levels, apoptosis rate and Bax protein expression in AGS cells were significantly increased, and cell viability (48 h, 72 h, 96 h), CyclinD1 and Bcl-2 proteins were reduced (P < 0.05).There was an interaction between C5orf66-AS1 and miR-637.Moreover, the overexpression of miR-637 could make the overexpression of C5orf66-AS1 more effective in inhibiting AGS cell proliferation and promoting apoptosis (P < 0.05).
ConclusionsC5orf66-AS1 and miR-637 are lowly expressed in GC tissues and cells.Overexpression of C5orf66-AS1 and up-regulation of miR-637 can synergistically inhibit the proliferation of GC cells and promote apoptosis.

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