Toll-like receptor (TLR4) shedding and depletion: acute proximal tubular cell responses to hypoxic and toxic injury.
Acute renal failure (ARF) induces tubular hyperresponsiveness to TLR4 ligands, culminating in exaggerated renal cytokine/chemokine production. However, the fate of TLR4 protein during acute tubular injury remains unknown. The study sought new insights into this issue. Male CD-1 mice were subjected to 1) unilateral ischemia-reperfusion (I/R), 2) cisplatin (CP) nephrotoxicity, or 3) glycerol-induced myohemoglobinuric ARF. Renal cortical TLR4 protein (Western blotting, immunohistochemistry) and TLR4 mRNA levels (RT-PCR) were determined thereafter (90 min-4 days). Urinary TLR4 excretion post-I/R or CP injection was also assessed. To gain proximal tubule-specific results, TLR4 protein and mRNA were quantified in posthypoxic or oxidant (Fe)-challenged isolated mouse tubules. Finally, TLR4 mRNA was determined in antimycin A-injured cultured proximal tubular (HK-2) cells. Acute in vivo renal injury reduced proximal tubule TLR4 content. These changes corresponded with the appearance of TLR4 fragment(s) in urine and a persistent increase in renal cortical TLR4 mRNA. Isolated proximal tubules responded to injury with rapid TLR4 reductions, dramatic extracellular TLR4 release, and increases in TLR4 mRNA. Glycine blocked these processes, implying membrane pore formation was involved. HK-2 cell injury increased TLR4 mRNA, but not protein levels, suggesting intact transcriptional, but not translational, pathways. Diverse forms of acute tubular injury rapidly reduce proximal tubular TLR4 content. Plasma membrane TLR4 release through glycine-suppressible pores, possibly coupled with a translation block, appears to be involved. Rapid postinjury urinary TLR4 excretion suggests its potential utility as a "biomarker" of impending ARF.
Zager
Richard A
RA
Department of Medicine, University of Washington, and the Fred Hutchinson Cancer Research Center, Seattle, Washington 98109-1024, USA. dzager@fhcrc.org
Johnson
Ali C M
AC
Lund
Steven
S
Randolph-Habecker
Julie
J
D000818:Animals; D000970:Antineoplastic Agents; D015153:Blotting, Western; D002460:Cell Line; D002945:Cisplatin; D005990:Glycerol; D006456:Hemoglobinuria / Q000139:chemically induced / Q000473:pathology; D006801:Humans; D000860:Hypoxia / Q000473:pathology*; D007150:Immunohistochemistry; D007672:Kidney Cortex / Q000378:metabolism / Q000473:pathology; D007676:Kidney Failure, Chronic / Q000139:chemically induced* / Q000473:pathology*; D007687:Kidney Tubules, Proximal / Q000166:cytology* / Q000378:metabolism* / Q000473:pathology; D008297:Male; D051379:Mice; D012333:RNA, Messenger / Q000096:biosynthesis; D051197:Toll-Like Receptor 4 / Q000187:drug effects / Q000378:metabolism*
D000970:Antineoplastic Agents; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D005990:Glycerol; D002945:Cisplatin