| Podocyte injury is commonly considered as the vital trigger for albuminuria and the eventual progression of glomerular disease.Although the mechanisms of podocyte injury have been explored,effective means to alleviate podocyte injury are still lacking.Previous studies have shown that mitochondrial abnormalities are involved in a variety of podocyte injury models,including diabetic models and FSGS models,and studies have shown that abnormalities in mitochondria-related genes are involved in podocyte injury.In the first part of this study,primary podocytes,diabetic and adriamycin animal models were used to elucidate the relationship between mitochondrial abnormalities and podocyte injury in detail.The results showed that with the progression of diabetes in mice,urinary microalbumin increased gradually and podocyte injury worsened.The adriamycin also induced urinary albumin and podocyte injury in mice.We found that mitochondrial abnormalities occurred in the early stage of the diabetic model while no significant podocyte injury was observed,which also appeared in the adriamycin model,indicating that mitochondrial abnormalities may be one of the initiating factors in podocyte injury.SS31,a mitochondrial-targeted drug,protected mitochondria in the early stage of diabetes by reducing albuminuria in diabetic mice,which further indicated the importance of mitochondria in podocyte.UCP2 is located in the inner mitochondrial membrane and belongs to the uncoupling protein family members.UCP2 is widely expressed and can be regulated at the transcriptional and translational levels,which draws extensive attention to UCP2.However,the role of UCP2 in podocytes has not been reported.In this study,we focused on diabetic and adriamycin glomerular injury models,primary podocytes,podocyte Ucp2-/-mice,and DKD patients to elucidate the role of UCP2 in podocyte injury.We found that in glomeruli UCP2 was mainly distributed in podocytes.Glomerular UCP2 expression was elevated in response to diabetes and ADR stimulation.Inhibition of podocyte UCP2 expression in vitro resulted in increased apoptosis under injury.In vivo renal overexpression of UCP2 alleviated diabetesinduced podocyte injury.More importantly,podocyte Ucp2-/-mice developed agedependent proteinuria,with more severe proteinuria and podocyte injury in diabetic and ADR-induced renal injury models.Finally,we performed a correlation study between glomerular UCP2 expression and proteinuria in DKD patients and found that glomerular UCP2 expression was negatively correlated with proteinuria and positively correlated with e GFR in patients with DKD.We demonstrated the role of UCP2 in podocyte injury.Podocytes upregulate UCP2 expression in response to injury,and inhibition of UCP2 expression in vitro and in vivo aggravates podocyte injury.Our previous results showed that UCP2 is important for podocyte structure and function,we further clarified the role of UCP2 in podocyte mitochondria.We examined podocyte mitochondrial morphology in control and Ucp2-/-mice,and isolated primary podocytes to detect mitochondrial morphology and function.In parallel,we downregulated UCP2 with adenovirus Ucp2 AV-sh RNA in vitro to observe mitochondrial morphology and function.Our results showed that 3-month-old Ucp2-/-mice had abnormal mitochondrial morphology in podocytes compared with control mice.In vitro results showed that loss of UCP2 in podocytes resulted in mitochondria changing from linear to punctate,a decrease in the number of mitochondrial cristae,and abnormal expression of fusion fission related proteins.Meanwhile,mitochondria showed abnormalities in function,including altered membrane potential,reduced ATP production,and respiratory depression.Inhibition of podocyte UCP2 expression in vitro also showed the above mitochondrial morphological and functional abnormalities.In this part,we detailed the effects of UCP2 deletion on mitochondrial structure and function in vitro and in vivo,indicating that UCP2 is involved in the regulation of podocyte mitochondrial homeostasis.Mitochondria are highly dynamic organelles that continuously undergo fusion and fission to maintain homeostasis.Disruption of mitochondrial dynamics leads to mitochondrial dysfunction and causes human disease.Fusion of mitochondrial outer and inner membranes is mediated by two classes of dynamin-like proteins,including MFN(Mitofusin)and OPA1(optic atrophy1).Mitochondrial inner membrane fusion is more complex than the outer membrane.OPA1 is involved not only in the fusion of the inner mitochondrial membrane but also in the formation of the mitochondrial cristae.We examined glomerular OPA1 expression in diabetic model,and down-regulated OPA1 by transfection of small interfering RNA(si RNA).The results showed that glomerular OPA1 was down-regulated in a time-dependent manner in diabetic model.Inhibition of OPA1 in vitro leads to abnormal mitochondrial morphology and function.In addition,our previous results showed that UCP2 deficency caused reduced OPA1 expression in vitro and in vivo,we then overexpressed UCP2 under HG,Which could significantly inhibit the down-regulation of OPA1,suggesting that UCP2 acts by stabilizing OPA1.We overexpressed OPA1 in the presence of UCP2 depletion and found that it could significantly improve mitochondrial function and inhibit podocyte injury.Further studies showed that UCP2 interacts with OPA1 and loss of UCP2 leads to metalloprotease OMA1 activation,which degrades OPA1.In this part,we clarified the importance of OPA1 in podocytes.We found that UCP2 depletion leads to down-regulation of OPA1,and we further clarified that UCP2 interacts with OPA1 and maintains mitochondrial homeostasis by inhibiting OMA1 activation to stable OPA1. |
