Mechanism of LncRNA MEG3 regulating podocyte injury in diabetic nephropathy
Objective To investigate the mechanism of long non-coding RNA maternally expressed gene 3(LncRNA MEG3)on podocyte injury in diabetic kidney disease(DKD).Methods Mouse podocyte clone-5(MPC5)cells were used to construct the glucose-induced podocyte injury model.The MPC5 cells were treated with high glucose,high glucose+short hairpin RNA-negative control(sh-NC)group,high glucose+sh-LncRNA MEG3 group,high glucose+sh-LncRNA MEG3+3-methyladenine(3-MA)group,high glucose+sh-LncRNA MEG3+sh-tuberous sclerosis complex 1(TSC1)group and control group.Cell viability and apoptosis rate were detected by MTT assay and flow cytometry.The relative expression level of LncRNA MEG3 was detected by real-time PCR.Western blot was used to detect autophagy-related proteins recombinant Beclin 1(Beclin-1),TSC1,selective autophagy joint protein(p62)and microtubule-associated protein light chain 3(LC3).Fluorescence in situ hybridization was used to detect the expression and localization of LncRNA MEG3 in cells.Results The MPC5 activity of the high glucose group was lower than that of the control group,the MPC5 activity of the high glucose+sh-NC group was lower than that of the high glucose+sh-LncRNA MEG3 group,the MPC5 activity of high glucose+sh-LncRNA MEG3+sh-TSC1 group and high glucose+sh-LncRNA MEG3+3-MA group was lower than that of high glucose+sh-LncRNA MEG3 group(all P<0.01).The apoptosis rate of MPC5 in the high glucose group was higher than that in the control group,the apoptosis rate of MPC5 in the high glucose+sh-NC group was higher than that in the high glucose+sh-LncRNA MEG3 group,the apoptosis rate of MPC5 in high glucose+sh-LncRNA MEG3+sh-TSC1 group and high glucose+sh-LncRNA MEG3+3-MA group was higher than that in high glucose+sh-LncRNA MEG3 group(all P<0.01).The relative expression level of LncRNA MEG3 in the high glucose group was higher than that in the control group,the relative expression level of LncRNA MEG3 in the high glucose+sh-NC group was higher than that in the high glucose+sh-LncRNA MEG3 group(all P<0.01).The level of LC3,Beclin-1 and TSC1 in the high glucose group were lower than those in the control group,the level of p62 were higher than those in the control group.The level of LC3,Beclin-1 and TSC1 in the high glucose+sh-NC group were higher than those in the high glucose+sh-NC group,the level of p62 were lower than those in the high glucose+sh-NC group.The level of TSC1 in high glucose+sh-LncRNA MEG3+sh-TSC1 group was lower than that in high glucose+sh-LncRNA MEG3 group.The level of LC3 and Beclin-1 in high glucose+sh-LncRNA MEG3+3-MA group and high glucose+sh-LncRNA MEG3+sh-TSC1 group were lower than those in high glucose+sh-LncRNA MEG3 group,the level of p62 was higher than that in high glucose+sh-LncRNA MEG3 group(all P<0.01).The level of TSCI in high glucose+sh-LncRNA MEG3 was higher than that in high glucose+sh-NC group(P<0.01).LncRNA MEG3 was expressed in both nucleus and cytoplasm.Conclusion The study indicates that LncRNA MEG3 alleviating podocyte damage in DKD may be mediated by MPC5 autophagy through targeting TSC1.