摘要
目的探讨长链非编码RNA母系表达基因3(LncRNA MEG3)在不同浓度葡萄糖下表达情况及对血管内皮生长因子(VEGF)、转化生长因子β1(TGF-β1)表达的影响。方法按照不同浓度d-葡萄糖培养ARPE-19细胞,随机分为5 mmol·L^(-1)d-葡萄糖组、15 mmol·L^(-1)d-葡萄糖组、30 mmol·L^(-1)d-葡萄糖组。实时荧光定量PCR(RT-qPCR)检测各组ARPE-19细胞LncRNA MEG3、VEGF mRNA、TGF-β1 mRNA相对表达水平。ARPE-19细胞转染PCDNA-MEG3及PCDNA-NC质粒,建立LncRNA MEG3过表达ARPE-19细胞及转染空质粒的ARPE-19细胞。以含30 mmol·L^(-1)d-葡萄糖的DMEM分别培养以下各组ARPE-19细胞:LncRNA MEG3过表达ARPE-19细胞(O组);转染空质粒的ARPE-19细胞(NC组);加入PBS的正常ARPE-19细胞(G组);同时以含5 mmol·L^(-1)d-葡萄糖的DMEM培养正常ARPE-19细胞(C组);以上各组细胞培养24 h后,采用RT-qPCR法及Western blot法检测各组ARPE-19细胞VEGF mRNA和VEGF蛋白、TGF-β1 mRNA和TGF-β1蛋白的相对表达水平。结果与5 mmol·L^(-1)d-葡萄糖组比较,15 mmol·L^(-1)d-葡萄糖组和30 mmol·L^(-1)d-葡萄糖组ARPE-19细胞LncRNA MEG3相对表达水平降低,VEGF mRNA、TGF-β1 mRNA相对表达水平升高,差异均有统计学意义(均为P<0.05)。与15 mmol·L^(-1)d-葡萄糖组比较,30 mmol·L^(-1)d-葡萄糖组ARPE-19细胞LncRNA MEG3相对表达水平降低,VEGF mRNA、TGF-β1 mRNA相对表达水平升高,差异均有统计学意义(均为P<0.05)。与G组和NC组比较,O组ARPE-19细胞VEGF mRNA和VEGF蛋白、TGF-β1 mRNA和TGF-β1蛋白相对表达水平均降低,差异均有统计学意义(均为P<0.05)。结论高糖下调ARPE-19细胞LncRNA MEG3的表达,上调VEGF和TGF-β1的表达。LncRNA MEG3过表达降低高糖诱导的VEGF和TGF-β1表达的升高。LncRNA MEG3可能通过抑制VEGF和TGF-β1的表达调节糖尿病视网膜病变的发展。
Objective To investigate the expression of long non-coding RNA maternally expressed gene 3(LncRNA MEG3)under different glucose concentrations and its effect on the expression of vascular endothelial growth factor(VEGF)and transforming growth factorβ1(TGF-β1).Methods ARPE-19 cells cultured in media with different d-glucose concentrations were randomly divided into 5 mmol·L^(-1) d-glucose group,15 mmol·L^(-1) d-glucose group,and 30 mmol·L^(-1) d-glucose group.The relative expression levels of LncRNA MEG3,VEGF mRNA,and TGF-β1 mRNA in each group were detected by real-time quantitative polymerase chain reaction(RT-qPCR).ARPE-19 cells were transfected with PCDNA-MEG3 and PCDNA-NC plasmids to establish LncRNA MEG3 overexpressed ARPE-19 cells and empty plasmid transfected ARPE-19 cells.LncRNA MEG3 overexpressed ARPE-19 cells cultured in DMEM containing 30 mmol·L^(-1) d-glucose were set as group O.Empty plasmid transfected ARPE-19 cells cultured in DMEM containing 30 mmol·L^(-1) d-glucose were set as group NC.Normal ARPE-19 cells added with PBS cultured in DMEM containing 30 mmol·L^(-1) d-glucose were set as group G.Normal ARPE-19 cells cultured in DMEM containing 5 mmol·L^(-1) d-glucose were set as group C.After the cells were incubated for 24 h,the relative expression levels of VEGF mRNA and VEGF protein,TGF-β1 mRNA and TGF-β1 protein in each group were detected by RT-qPCR and Western blot.Results Compared with the 5 mmol·L^(-1) d-glucose group,the relative expression level of LncRNA MEG3 in the 15 mmol·L^(-1) d-glucose and 30 mmol·L^(-1) d-glucose groups was decreased,while the relative expression levels of VEGF mRNA and TGF-β1 mRNA were increased(all P<0.05).Compared with the 15 mmol·L^(-1) d-glucose group,the relative expression level of LncRNA MEG3 in the 30 mmol·L^(-1) d-glucose group was decreased,while the relative expression levels of VEGF mRNA and TGF-β1 mRNA were increased(all P<0.05).Compared with group G and group NC,the relative expression levels of VEGF mRNA and VEGF protein,TGF-β1 mRNA and TGF-β1 protein in group O were decreased(all P<0.05).Conclusion High glucose down-regulates the expression of LncRNA MEG3 and up-regulates the expression of VEGF and TGF-β1 in ARPE-19 cells.The overexpression of LncRNA MEG3 reduces the elevated expression of VEGF and TGF-β1 induced by high glucose.LncRNA MEG3 may suppress the development of diabetic retinopathy by inhibiting the expression of VEGF and TGF-β1.
作者
张达宁
冷瀛
秦海翔
李相军
白丹
石盛玲
王敬怡
卢奕安
张富程
陈艳
ZHANG Daning;LENG Ying;QIN Haixiang;LI Xiangjun;BAI Dan;SHI Shengling;WANG Jingyi;LU Yian;ZHANG Fucheng;CHEN Yan(Department of Ophthalmology,Affiliated Hospital of Beihua University,Jilin 132011,Jilin Province,China)
出处
《眼科新进展》
CAS
北大核心
2022年第7期524-528,共5页
Recent Advances in Ophthalmology
基金
吉林省卫生健康技术创新项目(编号:2019J048)。
作者简介
张达宁(ORCID:0000-0003-3317-2370),女,1974年4月出生,吉林人,硕士,副主任医师。研究方向:眼底病。E-mail:qinhyapple@126.com。
