Abstract
The final control of renal water reabsorption occurs in the collecting duct (CD) and relies on regulated expression of aquaporin-2 (AQP2) in principal CD cells. AQP2 transcription is primarily induced by type 2 vasopressin receptor (V2R)-cAMP-protein kinase A (PKA) signaling but also by other factors, including TonEBP and NF-κB. NAPDH oxidase 4 (NOX4) represents a major source of reactive oxygen species (ROS) in the kidney. Because NOX-derived ROS may alter PKA, TonEBP and NF-κB activity, we examined the effects of NOX4 depletion on AQP2 expression. Depleted NOX4 expression by siRNA (siNOX4) in mpkCCDcl4 cells attenuated increased AQP2 mRNA expression by arginine vasopressin (AVP) but not by hypertonicity, which induces both TonEBP and NF-κB activity. AVP-induced AQP2 expression was similarly decreased by the flavoprotein inhibitor diphenyleneiodonium. siNOX4 altered neither TonEBP nor NF-κB activity but attenuated AVP-inducible cellular cAMP concentration, PKA activity and CREB phosphorylation as well as AQP2 mRNA expression induced by forskolin, a potent activator of adenylate cyclase. The repressive effect of siNOX4 on AVP-induced AQP2 mRNA expression was abolished by the non-selective phosphodiesterase (PDE) inhibitor 3-isobutyl-1-methylxanthine (IBMX) and was significantly decreased by selective PDE antagonists cilostamide and rolipram, but not vinpocetine, which respectively target PDE3, PDE4 and PDE1. Thus, by inhibiting PDE3 and PDE4 activity NOX4-derived ROS may contribute to V2R-cAMP-PKA signaling and enhance AQP2 transcription.
Highlights
Despite variations of water intake and loss, whole body water homeostasis is maintained within a narrow range by the continuous adjustment of water reabsorption by the kidney collecting duct (CD) [1]
NAPDH oxidase 4 (NOX4) deficiency decreases AQP2 expression We have previously demonstrated by Amplex Red assay that basal levels of H2O2 produced in cultured CD principal cells is decreased two-fold by siRNA targeting NOX4 [34]
We investigated the possibility that the absence of NOX4 might alter intracellular signaling that controls AQP2 expression
Summary
Despite variations of water intake and loss, whole body water homeostasis is maintained within a narrow range by the continuous adjustment of water reabsorption by the kidney collecting duct (CD) [1]. This process critically relies on the kidney’s ability to modulate both the corticomedullary osmotic gradient and aquaporin-2 (AQP2) water channel abundance that respectively provide the driving force and the permeability for water reabsorption [2,3]. The antidiuretic hormone arginine vasopressin plays a key role by increasing both AQP2 transcription as well as AQP2 expression at the apical cell surface [7].
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