AIM： To investigate the effects and mechanism of miR-211 in mediating the antioxidant function of lens epithelial cells affected by age-related cataracts. METHODS： Real-time quantitative polymerase chain reaction ...AIM： To investigate the effects and mechanism of miR-211 in mediating the antioxidant function of lens epithelial cells affected by age-related cataracts. METHODS： Real-time quantitative polymerase chain reaction （RT-qPCR） was used to detect miR-211 expression in the anterior lens capsules of healthy people, the anterior lens capsules of patients with age-related cataracts, and human epithelial cell line （SRA01/04） cells exposed to oxidative stress. A 2＇, 7＇-dichloro-fluorescein diacetate （DCFH-DA） probe was used to measure the levels of endogenous reactive oxygen species （ROS） in human lens epithelial cells （hLECs） exposed to 400 pmol/L H2O2 for lh. SRA01/04 cells were transfected with either miR-211 mimics, mimic controls, miR-211 inhibitors or inhibitor controls. After 72h, these cells were exposed to 400 IJmollL H2O2 for lh, then p53 and Bax mRNA expression were measured using RT-qPCR. p53 and Bax protein expression were also measured by Western blotting analysis. Finally, cell viability was assessed using an MTS assay. RESULTS： Compared to the control group, expression of miR-211 in the anterior lens capsules of age-related cataract patients and in SRA01/04 cells exposed to oxidative stress was significantly increased （P〈0.001）. Levels of endogenous ROS were significantly elevated in hLECs exposed to oxidative stress （P〈0.001）. Compared to the mimic control group, the hLECs in the miR-211 mimic group expressed significantly higher levels of p53 and Bax mRNA and protein while cell viability was significantly reduced （P〈0.001）. Conversely, p53 and Bax mRNA and protein expression were significantly reduced in the miR-211 inhibitor group as compared to the control group, while the cells in this group had much higher levels of call viability （P〈0.001）. CONCLUSION： miR-211 is upregulatsd in the anterior lens capsules of age-related cataract patients, miR-211 decreased the antioxidative stress capacity of lens epithelial cells by upregulating p53 and Bax, while inhibiting cell proliferation and repair. This finding suggests that miR-211 may play a key role in the development of age-related cataracts.展开更多
AIM： To detect the expression of miR-211 in age-related cataract tissue, explore the effects of miR-211 on lens epithelial cell proliferation and apoptosis, and identify its target gene.METHODS： This study used real...AIM： To detect the expression of miR-211 in age-related cataract tissue, explore the effects of miR-211 on lens epithelial cell proliferation and apoptosis, and identify its target gene.METHODS： This study used real-time quantitative polymerase chain reaction（RT-q PCR） to measure the expression of miR-211 and its predicted target gene [silent matingtype information regulation 2 homolog 1（SIRT1）] in 46 anterior lens capsules collected from age-related cataract patients. Human lens epithelial cell line（SRA01/04） cells were transfected with either miR-211 mimics, mimic controls, miR-211 inhibitors or inhibitor controls, 72 h after transfection, miR NA and protein expression of SIRT1 were measured using RT-qP CR and Western blotting; then cells were exposed to 200 μmol/L H2O2 for 1h, whereupon cell viability was measured by MTS assay, caspase-3 assay was performed. Dual luciferase reporter assay was performed to verify the relationship between miR-211 of SIRT1.RESULTS： Compared to the control group, expression of miR-211 was significantly increased（P〈0.001）, the miR NA and protein expression of SIRT1 were significantly decreased（P〈0.001） in the anterior lens capsules of patients with age-related cataracts. Relative to the control group, SIRT1 miR NA and protein levels in the miR-211 mimic group were significantly reduced, cell proliferation activity significantly decreased, and caspase-3 activity was significantly increased（P〈0.001）. In the miR-211 inhibitor group, SIRT1 miRNA and protein expression were significantly increased, cell proliferation activity significantly increased, and caspase-3 activity was significantly decreased（P〈0.001）. A dual luciferase reporter assay confirmed that SIRT1 is a direct target of miR-211.CONCLUSION： miR-211 is highly expressed in the anterior lens capsules of patients with age-related cataracts. By negatively regulating the expression of SIRT1, miR-211 promotes lens epithelial cell apoptosis and inhibits lens epithelial cell proliferation.展开更多
文摘AIM： To investigate the effects and mechanism of miR-211 in mediating the antioxidant function of lens epithelial cells affected by age-related cataracts. METHODS： Real-time quantitative polymerase chain reaction （RT-qPCR） was used to detect miR-211 expression in the anterior lens capsules of healthy people, the anterior lens capsules of patients with age-related cataracts, and human epithelial cell line （SRA01/04） cells exposed to oxidative stress. A 2＇, 7＇-dichloro-fluorescein diacetate （DCFH-DA） probe was used to measure the levels of endogenous reactive oxygen species （ROS） in human lens epithelial cells （hLECs） exposed to 400 pmol/L H2O2 for lh. SRA01/04 cells were transfected with either miR-211 mimics, mimic controls, miR-211 inhibitors or inhibitor controls. After 72h, these cells were exposed to 400 IJmollL H2O2 for lh, then p53 and Bax mRNA expression were measured using RT-qPCR. p53 and Bax protein expression were also measured by Western blotting analysis. Finally, cell viability was assessed using an MTS assay. RESULTS： Compared to the control group, expression of miR-211 in the anterior lens capsules of age-related cataract patients and in SRA01/04 cells exposed to oxidative stress was significantly increased （P〈0.001）. Levels of endogenous ROS were significantly elevated in hLECs exposed to oxidative stress （P〈0.001）. Compared to the mimic control group, the hLECs in the miR-211 mimic group expressed significantly higher levels of p53 and Bax mRNA and protein while cell viability was significantly reduced （P〈0.001）. Conversely, p53 and Bax mRNA and protein expression were significantly reduced in the miR-211 inhibitor group as compared to the control group, while the cells in this group had much higher levels of call viability （P〈0.001）. CONCLUSION： miR-211 is upregulatsd in the anterior lens capsules of age-related cataract patients, miR-211 decreased the antioxidative stress capacity of lens epithelial cells by upregulating p53 and Bax, while inhibiting cell proliferation and repair. This finding suggests that miR-211 may play a key role in the development of age-related cataracts.
基金Supported by the National Natural Science Foundation of China(No.81170836No.81570838)+1 种基金the Natural Science Foundation of Liaoning Province,China(No.2015020474)the Liaoning Provincial Hospital Program for Building Treatment Capacity in Key Clinical Departments(No.LNCCC-D15-2015)
文摘AIM： To detect the expression of miR-211 in age-related cataract tissue, explore the effects of miR-211 on lens epithelial cell proliferation and apoptosis, and identify its target gene.METHODS： This study used real-time quantitative polymerase chain reaction（RT-q PCR） to measure the expression of miR-211 and its predicted target gene [silent matingtype information regulation 2 homolog 1（SIRT1）] in 46 anterior lens capsules collected from age-related cataract patients. Human lens epithelial cell line（SRA01/04） cells were transfected with either miR-211 mimics, mimic controls, miR-211 inhibitors or inhibitor controls, 72 h after transfection, miR NA and protein expression of SIRT1 were measured using RT-qP CR and Western blotting; then cells were exposed to 200 μmol/L H2O2 for 1h, whereupon cell viability was measured by MTS assay, caspase-3 assay was performed. Dual luciferase reporter assay was performed to verify the relationship between miR-211 of SIRT1.RESULTS： Compared to the control group, expression of miR-211 was significantly increased（P〈0.001）, the miR NA and protein expression of SIRT1 were significantly decreased（P〈0.001） in the anterior lens capsules of patients with age-related cataracts. Relative to the control group, SIRT1 miR NA and protein levels in the miR-211 mimic group were significantly reduced, cell proliferation activity significantly decreased, and caspase-3 activity was significantly increased（P〈0.001）. In the miR-211 inhibitor group, SIRT1 miRNA and protein expression were significantly increased, cell proliferation activity significantly increased, and caspase-3 activity was significantly decreased（P〈0.001）. A dual luciferase reporter assay confirmed that SIRT1 is a direct target of miR-211.CONCLUSION： miR-211 is highly expressed in the anterior lens capsules of patients with age-related cataracts. By negatively regulating the expression of SIRT1, miR-211 promotes lens epithelial cell apoptosis and inhibits lens epithelial cell proliferation.
