Abstract
Background Transient receptor potential channel 5 (TRPC5) is a non-selective cationic ion channel expressed in brain, kidney and other organs where its activation underlies podocyte injury in chronic kidney diseases. Specifically, it has been suggested that a podocyte TRPC5 plasma membrane relocation and channel activation following injury results from activation of Rac-1, propagating podocyte dysfunction and proteinuria. However, previous TRPC5 transgenic mouse studies had questioned a pathogenic role for TRPC5 in podocytes. This investigation was designed to specifically evaluate podocyte Rac-1 activation in the context of functional TRPC5 or a TRPC5 pore mutant to assess effects on proteinuria.
Materials and Methods We employed single cell patch-clamp studies of cultured podocytes and studied proteinuria in transgenic mouse models to characterize the effects of TRPC5 following podocyte Rac-1 activation.
Results Inhibition of TRPC5 by small molecules reportedly ameliorated proteinuria in murine models of proteinuric kidney diseases. In order to directly examine TRPC5 function following Rac-1-induced podocyte injury, we analyzed TRPC5 inhibition in podocyte specific Rac-1 (active) transgenic mice. In addition, we generated a double-transgenic mouse constitutively overexpressing either TRPC5 (TRPC5WT) or a TRPC5 dominant-negative pore mutant (TRPC5DN) in concert with podocyte specific and inducible activation of active Rac-1 (Rac-1Dtg). In electrophysiological experiments, active TRPC5 was detected in primary podocytes overexpressing TRPC5 but not in podocytes with endogenous TRPC5 expression, nor with Rac-1 overexpressing podocytes. TRPC5 inhibition did not change proteinuria in mice with active podocyte Rac-1, nor did an increase or loss of TRPC5 activity affected podocyte injury in Rac-1Dtg animals. Administration of TRPC5 inhibitors, ML204 and AC1903, did not alleviate podocyte Rac-1 induced proteinuria.
Conclusion TRPC5 inhibition did not modify podocyte Rac-1 induced proteinuria in mice.
Significance Statement TRPC5 is a calcium conducting ion channel involved in a plethora of biological functions in the brain, kidney and other organs. In proteinuric kidney diseases, others proposed a model that links activation of small GTPase Rac-1 in podocytes to activation of TRPC5 channels propagating cellular injury and eventually leading to progressive kidney disease. To test this hypothesis, we have developed a novel transgenic mouse model that employs podocyte Rac-1 activation in the presence or absence of a functional TRPC5 channel. Our data shows that transgenic mice with activated Rac-1 in podocytes did not enhance endogenous TRPC5 expression or its activity. Furthermore, TRPC5 blockade or activation did not modify Rac-1 induced proteinuria in mice.
Competing Interest Statement
S.S. and J.R. are co-founders and shareholders of Walden Biosciences, a biotechnology company that develops novel kidney-protective therapies.
