Inhibition of Na superset+/H superset+ exchange decreases albumin-induced NF-kappaB activation in renal proximal tubular cell lines (OK and LLC-PK1 cells).
Filtered proteins play a role in the pathogenesis of renal interstitial inflammation and fibrosis. At least part of these effects are mediated by the nuclear factor kappaB (NF-kappaB). Receptor-mediated endocytosis of proteins like albumin in renal proximal tubular cells is in part dependent on Na superset+/H superset+ exchanger (NHE) isoform 3. We tested the hypothesis that pharmacological inhibition of NHE-3 reduces albumin-induced NF-kappaB activation - and therefore albumin-induced renal interstitial inflammation and fibrosis - using established proximal tubular cell lines (OK and LLC-PK1). 5-(N-ethyl-N-isopropyl)-amiloride (EIPA) or HOE694 were used to inhibit NHE. Albumin endocytosis was determined by fluorometric analysis of FITC-BSA uptake. Electromobility gel shift assays were performed in order to determine the NF-kappaB-specific DNA-binding activity. EIPA reduced albumin uptake in OK and LLC-PK1 cells and HOE694 decreased albumin uptake in LLC-PK1 cells, with IC subset50 values corresponding to NHE-3 inhibition. Furthermore, albumin-induced increases in NF-kappaB DNA-binding activity were partially inhibited by EIPA in OK and LLC-PK1 cells. HOE694 at a concentration of 100 micromol/l similarly decreased albumin-induced NF-kappaB DNA-binding activity. Cytosolic acidification by propionic acid did not prevent BSA-induced activation of NF-kappaB. Inhibition of BSA endocytosis by chlorpromazine decreased NF-kappaB activation. NHE-dependent albumin endocytosis induces an increase in NF-kappaB-specific protein activity in renal proximal tubular cells in culture, which is decreased by EIPA and HOE694. Thus, inhibition of albumin uptake might be a therapeutical strategy to prevent albumin-induced NF-kappaB activation and albumin-associated inflammatory or fibrotic renal pathomechanisms in vivo.
Drumm
K
K
Institute of Physiology, University of Wuerzburg, Roentgenring 9, D-97070 Wuerzburg, Germany. karina.drumm@gmx.de
Gassner
B
B
Silbernagl
S
S
Gekle
M
M
D000584:Amiloride / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002746:Chlorpromazine / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D006146:Guanidines / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D018374:LLC-PK1 Cells; D016328:NF-kappa B / Q000378:metabolism*; D009893:Opossums; D012710:Serum Albumin, Bovine / Q000378:metabolism* / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors*; D013450:Sulfones / Q000494:pharmacology; D013552:Swine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D006146:Guanidines; D016328:NF-kappa B; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; C074107:3-methylsulfonyl-4-piperidinobenzoyl guanidine; D012710:Serum Albumin, Bovine; D000584:Amiloride; D004247:DNA; D013755:Tetradecanoylphorbol Acetate; D002746:Chlorpromazine; C039614:ethylisopropylamiloride