Calcium-dependent dual oxidase 2 is a novel source of reactive oxygen species implicated in glomerular mesangial cell fibrotic response to angiotensin II and glucose

Abstract

The vasoactive peptide angiotensin II (Ang II) and hyperglycemia contribute to the initiation and the progression of glomerular fibrosis via activation of glomerular mesangial cells (MCs) and subsequent extracellular matrix expansion. We have previously shown that oxidative stress is critical for MC fibrotic response to Ang II or high concentrations of glucose (HG). Here, we demonstrate that Dual oxidase 2 (Duox2), a member of the Nox/Duox family of NADPH oxidases, is present in MCs and that its protein expression, along with its associated maturation factor, DuoxA2, are upregulated by Ang II and HG. Small interfering RNA (siRNA)-mediated downregulation of Duox2 significantly reduces Ang II-induced increase in reactive oxygen (ROS) generation and prevents the stimulatory effect of Ang II on MC fibrotic injury (as assessed by measuring a-smooth muscle actin and fibronectin expression). To demonstrate that Duox2 activation is Ca2+-dependent, we show that the extracellular Ca2+ chelator BAPTA prevented Ang II-induced ROS generation and the stimulation of MC fibrotic injury by Ang II. Moreover, treatment of MCs with ionomycin resulted in increased ROS production and enhanced MC fibrotic injury. These effects were abrogated by siRNA targeting Duox2. Fura-2 fluorescence was utilized as well to show calcium mobilization in response to Ang II and this effect was abrogated with siDuox2 transfection. These data indicate that Ang II-stimulated Duox2 activation and ROS generation subsequently lead to MC fibrotic injury. In summary, we have identified a novel role for Duox2 and its cognate maturation factor, DuoxA2, as a major source of ROS in response to Ang II and HG and established the significance of Duox2 in Ang II-mediated MC activation and fibrotic injury. Therapeutic targeting of this pathway may prevent or reverse pathophysiologic manifestations of renal fibrotic diseases. Research reported in this poster was supported by the University of the Incarnate Word Offce of Research and Graduate Studies and the National Institute of General Medical Sciences of the National Institutes of Health under Award Number SC2GM136569. The content is solely the responsibility of the authors and does not necessarily represent the offcial views of the National Institutes of Health. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.