Primary focal and segmental glomerulosclerosis (FSGS) is a common cause of end-stage renal disease. Many of these patients are refractory to current treatments and primary FSGS often recurs after kidney transplantation. There is now a consensus that many cases of primary and recurrent FSGS are driven by circulating permeability factors that induce leakiness in the glomerular filtration barrier and ultrastructurl changes in podocytes. Some rare non-recurring genetic forms of FSGS are associated with gain-of-function mutations in Ca2+-permeable TRPC6 channels, resulting in chronic Ca2+ overload. Our central hypothesis is that increased podocyte TRPC6 activity drives pathology in FSGS, and this occurs in response to circulating permeability factors in primary FSGS, and possibly in response to hyperfiltration in secondary FSGS. If so, inhibition of TRPC6 could be a broadly useful therapeutic approach for FSGS of several etiologies. We also propose that primary FSGS can be caused by multiple circulating permeability factors with distinct but synergistic modes of action that converge on podocyte TRPC6. This research will focus on two circulating factors, TNF and suPAR, that have been implicated in some clinical studies and case reports. Our preliminary studies indicate that sera from several recurrent FSGS patients cause profound changes in podocyte TRPC6 channels, including markedly increased TRPC6 activation by mechanical stimuli, and increased expression on the podocyte cell surface. These sera also cause loss of podocin, which can explain increased mechanical activation of TRPC6. Some of these sera also evoke increases in ?3-integrin. The activity in sera from two recurrent FSGS patients taken during relapse and remission correlates with the patient's clinical status. Similar changes in TRPC6 gating and podocin were seen in glomeruli isolated from chronic PAN-treated rats, a model of secondary FSGS, and in podocytes treated with suPAR. TNF also increases TRPC6 activity in podocytes, but probably through mechanisms distinct and additive with those of suPAR. Here we will characterize modulation of endogenous podocyte TRPC6 channels by a larger sample of recurrent FSGS sera and we will determine if TNF and/or suPAR are required for their effects on podocytes. We present data showing that both of these factors contribute to activity in one of the recurrent FSGS serum samples. We will also examine transduction pathways whereby FSGS sera, suPAR, and TNF modulate TRPC6 channels. We will also examine changes in podocyte TRPC6 gating and maximal currents in in vivo mouse models of FSGS, using novel electrophysiological methods that we developed for our studies of podocin and chronic PAN nephrosis. Finally, we will also determine if TRPC6 knockout protects glomerular function in mice with albuminuria evoked by LPS, and in a transgenic mouse with albuminuria due to elevated circulating low molecular weight suPAR. The results could provide a strong rationale for drug discovery programs based on inhibition of mechanical and/or chemical activation of TRPC6 channels.
Primary FSGS is a leading cause of kidney failure that often recurs after the patient has received a kidney transplant. A role for TRPC6 channels was suggested from studies of patients with rare genetic forms of FSGS. This research will examine how substances in the blood of patients with primary FSGS increases activity of TRPC6 channels, will examine animal models of primary and secondary FSGS, and will determine if inhibition of TRPC6 might be a valid therapeutic approach for treatment of these patients.
