| Objective: Micro RNA is a class of single-stranded non-coding RNAs with a length of 20-25 nucleotides,which are regulated intracellularly by classical pathways that bind directly to the non-coding region of target gene m RNA 3',as well as by non-classical pathways such as precursor RNAs that target and regulate other non-coding RNAs.Some studies have shown that micro RNA-125 b is closely related to apoptosis,while apoptosis of lens epithelial cells is a common cytological basis for the formation of non-congenital cataract.At the same time,this study used bioinformatics software to predict that Bcl-2 antagonist/killer 1(BAK1)may have direct binding sites to micro RNA-125 b.BAK1 is a member of the pro-apoptotic family bcl-2,a protein that locates on the outer membrane of mitochondria and is a necessary component for the transmission of apoptotic signals through the mitochondrial pathway.therefore,this study intends to observe the expression of mi R-125 b in uv-induced murine cataract model and human lens epithelial cell apoptosis model,and its regulatory effect on predicting target gene BAK1 and human lens epithelial cell apoptosis.Methods: The human lens epithelial cell line(SRA01/04)with logarithmic long-term growth was selected and transfected with mi R-125 b mimic(mimic),mimic negative control(mimic control),mi R-125 b inhibitor,inhibitor negative control(inhibitor control)by Lipofectamine TM RNAi MAX.The expression of mi R-125 b in human lens epithelial cells was up-regulated and down regulated respectively.After 48 hours,the apoptosis model of human lens epithelial cells was constructed by ultraviolet UVB(irradiation intensity 360?W / cm2)for 25 minutes.Ten 8-week-old SPF C57 BL / 6mice were selected.After mydriatic treatment,ultraviolet UVB(irradiation intensity 360?W / cm2)was used to directly irradiate the left eye for 5 minutes every time,once a day for 7 days.The right eye was not treated.The lens capsule was collected after the mice were killed by cervical dislocation.In the cataract apoptosis model of human lens epithelial cells,human lens epithelial cells of different transfection groups and mouse lens capsule tissues,real time q PCR was used to verify the transfection efficiency,mi R-125 b and its predicted target gene BAK1 m RNA expression were detected,Western blot was used to detect the expression of mi R-125 b and BAK1 m RNA The expression of BAK1 protein was detected by blot,and the apoptosis rate of human lens epithelial cells was detected by TUNEL.The experimental data were analyzed by Graph Pad prism 8.Results: Compared with normal human lens epithelial cells,the expression of mi R-125 b was significantly decreased,BAK1 was significantly increased and apoptosis was increased(p < 0.05);Compared with the mi R-125 b group,the expression of mi R-125 b in the mi R-125 b group was significantly higher,the expression of BAK1 was significantly lower and the apoptosis was decreased(p < 0.05);compared with the mi R-125 b group,the expression of mi R-125 b in the mi R-125 b group was significantly lower and the expression of BAK1 was significantly higher(p < 0.05);Compared with the control group,the expression of miR-125 b,BAK1 and apoptosis of lens capsule in UVB group were significantly decreased,significantly increased(p < 0.05).Conclusion: The expression of micro RNA125 b is low in the UV induced cataract model of mice and the apoptosis model of human lens epithelial cells,which may target to negatively regulate the expression of BAK1,and then inhibit the apoptosis of human lens epithelial cells,and play a regulatory role in the pathogenesis of cataract,which may provide a new idea for non-surgical targeted treatment of cataract. |
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