Upregulation of mitochondrial Nox4 mediates TGF-β-induced apoptosis in cultured mouse podocytes

Area of Science:

• Nephrology
• Molecular Biology
• Cell Biology

Background:

• Podocyte injury is a key factor in chronic kidney diseases, often marked by proteinuria.
• Elevated transforming growth factor-beta (TGF-β) in renal tissue correlates with podocyte damage, leading to apoptosis and detachment.

Purpose of the Study:

• To elucidate the proapoptotic mechanism of TGF-β in immortalized mouse podocytes.
• To investigate the role of NADPH oxidase 4 (Nox4) and reactive oxygen species (ROS) in TGF-β-induced podocyte apoptosis.

Main Methods:

• Treatment of mouse podocytes with TGF-β1.
• Assessment of apoptosis, caspase-3 activation, ROS levels, and mitochondrial membrane potential.
• Upregulation and localization studies of NADPH oxidase 4 (Nox4).
• Inhibition studies using antioxidants, Nox inhibitors, TGF-β receptor blockers, and small interfering RNA (siRNA) for Nox4, Smad2, and Smad3.

Main Results:

• TGF-β1 induced podocyte apoptosis via caspase-3 activation, linked to increased ROS from Nox4 upregulation.
• Nox4, localized in mitochondria, was upregulated by TGF-β1, leading to mitochondrial membrane depolarization.
• Antioxidants, Nox inhibitors, and Nox4 knockdown mitigated TGF-β1-induced apoptosis and ROS production.
• Smad2/3 knockdown prevented TGF-β1-induced Nox4 increase, ROS generation, mitochondrial dysfunction, and apoptosis.

Conclusions:

• TGF-β1 upregulates mitochondrial Nox4 through the TGF-β receptor-Smad2/3 pathway, causing ROS production and mitochondrial dysfunction.
• This mechanism contributes to podocyte apoptosis and may play a role in proteinuric glomerular diseases like diabetic nephropathy.