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Nox-4 deletion reduces oxidative stress and injury by PKC-α-associated mechanisms in diabetic nephropathy.

Thallas-Bonke V, Jha JC, Gray SP, Barit D, Haller H, Schmidt HH, Coughlan MT, Cooper ME, Forbes JM, Jandeleit-Dahm KA - Physiol Rep (2014)

Bottom Line: Nox4 deletion attenuated diabetes-associated increases in albuminuria, glomerulosclerosis, and extracellular matrix accumulation.Lack of Nox4 resulted in a decrease in diabetes-induced renal cortical ROS derived from the mitochondria and the cytosol, urinary isoprostanes, and PKC activity.Downregulation of the PKC pathway was observed in tandem with reduced expression of vascular endothelial growth factor (VEGF), transforming growth factor (TGF)-β1 and restoration of the podocyte slit pore protein nephrin.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Complications Division, Baker IDI Heart & Diabetes Institute, JDRF Danielle Alberti Memorial Centre for Diabetic Complications, Melbourne, Victoria, Australia Department of Medicine, Austin and Northern Clinical Schools, University of Melbourne, Melbourne, Victoria, Australia.

No MeSH data available.


Related in: MedlinePlus

Oxidative stress markers at 20 weeks from renal cortex, A) Ex vivo NADH‐dependent mitochondrial tissue superoxide production, B) Ex vivo NADPH‐dependent cytosolic tissue superoxide production, C) Superoxide production from isolated mitochondria, D) Urinary 8‐isoprostane excretion, E) Computer‐aided analysis of immunohistochemical staining of renal cortical nitrotyrosine expressed as percent area, *P <0.05 versus WT‐C; †P <0.05 versus WT‐D; ‡P <0.001 versus WT‐C; §P <0.05 versus KO‐C, #P <0.01 versus WT‐D, n =8–10/group.
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fig06: Oxidative stress markers at 20 weeks from renal cortex, A) Ex vivo NADH‐dependent mitochondrial tissue superoxide production, B) Ex vivo NADPH‐dependent cytosolic tissue superoxide production, C) Superoxide production from isolated mitochondria, D) Urinary 8‐isoprostane excretion, E) Computer‐aided analysis of immunohistochemical staining of renal cortical nitrotyrosine expressed as percent area, *P <0.05 versus WT‐C; †P <0.05 versus WT‐D; ‡P <0.001 versus WT‐C; §P <0.05 versus KO‐C, #P <0.01 versus WT‐D, n =8–10/group.

Mentions: It has been suggested that mitochondria are a major source of ROS, which may play a role in the pathogenesis of DN (Forbes et al. 2008). Furthermore, Nox4 has recently been reported to localize within mitochondria (Block et al. 2009) but if Nox4 is a major source of mitochondrial ROS generation in diabetes remains unknown. Therefore, we measured superoxide production in live renal cortical tissue slices and in isolated mitochondria in Nox4 replete and KO mice in the presence and absence of diabetes. There was excess of superoxide production from both renal cortical mitochondrial (NADH‐driven) and cytosolic (NADPH‐driven) sources in diabetes (Fig. 6A and B), with more superoxide derived from the cytosolic compartment. Deletion in Nox4 resulted in normalization of superoxide production, suggesting that NADPH oxidase directly impact on mitochondrial superoxide production within the renal cortex. This was verified in isolated mitochondria (Fig. 6C). However, assessment of “whole body” superoxide production, by measurement of a stable marker of lipid peroxidation, 15‐isoprostane F2t showed residual oxidative stress status subsequent to Nox4 deletion (Fig. 6D), suggesting that there remain sources of oxidative stress that are independent of NADPH oxidase. Immunohistochemical evaluation of another oxidative stress marker, nitrotyrosine in glomeruli demonstrated a diabetes‐induced increase in the wild‐type mice, which was reduced in the diabetic KO mice (Fig. 6E).


Nox-4 deletion reduces oxidative stress and injury by PKC-α-associated mechanisms in diabetic nephropathy.

Thallas-Bonke V, Jha JC, Gray SP, Barit D, Haller H, Schmidt HH, Coughlan MT, Cooper ME, Forbes JM, Jandeleit-Dahm KA - Physiol Rep (2014)

Oxidative stress markers at 20 weeks from renal cortex, A) Ex vivo NADH‐dependent mitochondrial tissue superoxide production, B) Ex vivo NADPH‐dependent cytosolic tissue superoxide production, C) Superoxide production from isolated mitochondria, D) Urinary 8‐isoprostane excretion, E) Computer‐aided analysis of immunohistochemical staining of renal cortical nitrotyrosine expressed as percent area, *P <0.05 versus WT‐C; †P <0.05 versus WT‐D; ‡P <0.001 versus WT‐C; §P <0.05 versus KO‐C, #P <0.01 versus WT‐D, n =8–10/group.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4255803&req=5

fig06: Oxidative stress markers at 20 weeks from renal cortex, A) Ex vivo NADH‐dependent mitochondrial tissue superoxide production, B) Ex vivo NADPH‐dependent cytosolic tissue superoxide production, C) Superoxide production from isolated mitochondria, D) Urinary 8‐isoprostane excretion, E) Computer‐aided analysis of immunohistochemical staining of renal cortical nitrotyrosine expressed as percent area, *P <0.05 versus WT‐C; †P <0.05 versus WT‐D; ‡P <0.001 versus WT‐C; §P <0.05 versus KO‐C, #P <0.01 versus WT‐D, n =8–10/group.
Mentions: It has been suggested that mitochondria are a major source of ROS, which may play a role in the pathogenesis of DN (Forbes et al. 2008). Furthermore, Nox4 has recently been reported to localize within mitochondria (Block et al. 2009) but if Nox4 is a major source of mitochondrial ROS generation in diabetes remains unknown. Therefore, we measured superoxide production in live renal cortical tissue slices and in isolated mitochondria in Nox4 replete and KO mice in the presence and absence of diabetes. There was excess of superoxide production from both renal cortical mitochondrial (NADH‐driven) and cytosolic (NADPH‐driven) sources in diabetes (Fig. 6A and B), with more superoxide derived from the cytosolic compartment. Deletion in Nox4 resulted in normalization of superoxide production, suggesting that NADPH oxidase directly impact on mitochondrial superoxide production within the renal cortex. This was verified in isolated mitochondria (Fig. 6C). However, assessment of “whole body” superoxide production, by measurement of a stable marker of lipid peroxidation, 15‐isoprostane F2t showed residual oxidative stress status subsequent to Nox4 deletion (Fig. 6D), suggesting that there remain sources of oxidative stress that are independent of NADPH oxidase. Immunohistochemical evaluation of another oxidative stress marker, nitrotyrosine in glomeruli demonstrated a diabetes‐induced increase in the wild‐type mice, which was reduced in the diabetic KO mice (Fig. 6E).

Bottom Line: Nox4 deletion attenuated diabetes-associated increases in albuminuria, glomerulosclerosis, and extracellular matrix accumulation.Lack of Nox4 resulted in a decrease in diabetes-induced renal cortical ROS derived from the mitochondria and the cytosol, urinary isoprostanes, and PKC activity.Downregulation of the PKC pathway was observed in tandem with reduced expression of vascular endothelial growth factor (VEGF), transforming growth factor (TGF)-β1 and restoration of the podocyte slit pore protein nephrin.

View Article: PubMed Central - PubMed

Affiliation: Diabetes Complications Division, Baker IDI Heart & Diabetes Institute, JDRF Danielle Alberti Memorial Centre for Diabetic Complications, Melbourne, Victoria, Australia Department of Medicine, Austin and Northern Clinical Schools, University of Melbourne, Melbourne, Victoria, Australia.

No MeSH data available.


Related in: MedlinePlus