Lin Mei1,2, Hong Yan1,2, Song Wang2, Chenjun Guo2, Xiaoliang Zheng2, Bo Yan3, Jing Zhao3, Angang Yang3. 1. Department of Ophthalmology, Affiliated Guangren Hospital School of Medicine, Xi'an Jiaotong University, Xi'an No. 4 Hospital, Shaanxi Eye Hospital, Xi'an, Shaanxi Province, China. 2. Department of Ophthalmology, Tangdu Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, China. 3. Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an, Shaanxi Province, China.
Abstract
Purpose: To investigate the role of miRNAs in regulating oxidative damage during cataract formation. Methods: Microarray analysis and gene expression profiling assay were used to separately evaluate the miRNAs and mRNAs profiles in normal human lens epithelium cell line HLE-B3 treated by H2O2. The expression level of miR-630 was detected by RT-qPCR and the gene expression profiles were performed with gene ontology analysis using Bio Informatical database. The targets of miR-630 were predicted using miRecords and the results were used for screening targets of miR-630 combined with the GO analysis above. The mRNA levels of ALCAM, PCDH7, COL12A2, and EDIL3 in HLE-B3 cells after oxidative stimulation or miR-630 mimics transfection were measured by RT-qPCR, and the expression of ALCAM regulated by miR-630 was confirmed by Western blot and dual-luciferase reporter gene assay. The level of cell migration was measured by transwell assay and scratching test after transfection of miR-630 mimics and ALCAM siRNAs. Results: The microarray analysis demonstrated that miR-630 was significantly increased in HLE-B3 cells after oxidative stimulation. ALCAM, PCDH7, COL12A2, and EDIL3 were screened to be the possible targets of miR-630 by miRecords combined with GO analysis, but the results of RT-qPCR, Western blot and dual-luciferase reporter gene assay showed that only the expression of ALCAM was repressed by miR-630 transfection. Cell migration was inhibited through transfection of miR-630 mimics or ALCAM siRNAs and the upregulation of miR-630 partly reduced the cell migration increased by oxidative stimulation. Conclusion: miR-630 is one of the miRNAs increased by oxidative stimulation in human lens epithelium cells. Its upregulation may inhibit cell migration by targeting on ALCAM, which is important for HLECs to resist behavioral changes induced by oxidative damage and may delay the progression of cataract.
Purpose: To investigate the role of miRNAs in regulating oxidative damage during cataract formation. Methods: Microarray analysis and gene expression profiling assay were used to separately evaluate the miRNAs and mRNAs profiles in normal human lens epithelium cell line HLE-B3 treated by H2O2. The expression level of miR-630 was detected by RT-qPCR and the gene expression profiles were performed with gene ontology analysis using Bio Informatical database. The targets of miR-630 were predicted using miRecords and the results were used for screening targets of miR-630 combined with the GO analysis above. The mRNA levels of ALCAM, PCDH7, COL12A2, and EDIL3 in HLE-B3 cells after oxidative stimulation or miR-630 mimics transfection were measured by RT-qPCR, and the expression of ALCAM regulated by miR-630 was confirmed by Western blot and dual-luciferase reporter gene assay. The level of cell migration was measured by transwell assay and scratching test after transfection of miR-630 mimics and ALCAM siRNAs. Results: The microarray analysis demonstrated that miR-630 was significantly increased in HLE-B3 cells after oxidative stimulation. ALCAM, PCDH7, COL12A2, and EDIL3 were screened to be the possible targets of miR-630 by miRecords combined with GO analysis, but the results of RT-qPCR, Western blot and dual-luciferase reporter gene assay showed that only the expression of ALCAM was repressed by miR-630 transfection. Cell migration was inhibited through transfection of miR-630 mimics or ALCAM siRNAs and the upregulation of miR-630 partly reduced the cell migration increased by oxidative stimulation. Conclusion:miR-630 is one of the miRNAs increased by oxidative stimulation in human lens epithelium cells. Its upregulation may inhibit cell migration by targeting on ALCAM, which is important for HLECs to resist behavioral changes induced by oxidative damage and may delay the progression of cataract.