Role of Nox4 and p67phox in ROS production in response to increased tubular flow in the mTAL of Dahl salt‐sensitive (SS) rats

Abstract

Reactive Oxygen Species (ROS) in the renal outer medulla play an important role in the development of salt‐sensitive hypertension. NADPH oxidases (Nox) are the major source of ROS production in the kidney. We have examined the contribution of Nox4 and p67phox, related to the most abundant isoforms of NADPH‐oxidase Nox4 and Nox2 in the kidney, to the ROS production in response to sodium delivery and tubular flow in medullary thick ascending limb (mTAL) isolated from Dahl salt‐sensitive (SS) rats. We have shown in our previous studies that SSNox4−/− and SSp67phox−/− mutant rats exhibited a significant reduction of salt‐induced hypertension compared to SS rats after 21 day of 4.0% NaCl and a large reduction of albuminuria, tubular casts, and glomerular injury. In this study we have shown that increase in mTAL luminal flow stimulates cellular hydrogen peroxide (H2O2) and superoxide (O2·−), as well as mitochondrial H2O2 production in SS rats indicated by changes in fluorescence of sensitive dyes for detection of ROS (PF6‐AM, DHE, Mito‐PY1). In the present study we have determined that deletion of Nox4 (SSNox4−/−) resulted in significant reduction of intracellular H2O2 compared to SS in response to increased luminal flow in mTAL isolated from rats fed a 4% NaCl diet for 3 days. Deletion of the p67phox subunit in SS rat (SSp67phox−/−) also decreased H2O2 production compared to SS but to a lesser extent than that observed in mTAL of Nox4 rats (SSNox4−/−). There were no differences between cellular O2·− production in mTAL of SSNox4−/− rats compared to SS. However there was a significant decrease in cellular O2·− production in mTAL of mutant SSp67phox−/− rats. We conclude from this study that Nox4 is a major player in the production of cellular H2O2 in response to increased mTAL tubular flow of the SS rat whereas the p67phox subunit of Nox2 is involved in cellular O2·− production. Neither Nox4 nor the p67phox subunit of Nox2 seems to contribute to the mitochondrial H2O2 production in response to increased mTAL luminal flow.Support or Funding InformationHL 122662HL 082798