Increased expression of plasminogen activator inhibitor-1 in hypoxic renal injury and its pathological significance in progression of advanced renal disease

Chronic hypoxia has been newly proposed as a common mechanism of progressive renal fibrosis where PAI-1 plays important roles in the accumulation of extracellular matrix (ECM) through inhibition of plasmin-dependent ECM degradation. Hypoxia is also presumably associated with macrophage recruitment and angiogenesis that form fibrotic lesions. In the present study, we examined the effects of hypoxia and TNF-alpha on PAI-1, MCP-1 and VEGF expression in cultured human proximal renal tubular cells (HPTECs). We also investigated the presence of PAI-1 in renal tubular cells by immunostaining renal biopsy samples and measuring urinary PAI-1 levels in different kidney diseases. cDNA array analysis identified PAI-1 as a major gene highly induced by hypoxia in HPTECs. Hypoxia, TNF-alpha and their combination induced a 2.8-fold, a 1.8-fold, and a 4.6-fold increase in PAI-1 protein secretion, and produced a 3.6-fold, a 3.3-fold, and a 12.1-fold increase at the PAI-1 mRNA level, respectively. Similar results were confirmed by luciferase assay. Immunoblot analysis and immunocytochemistry revealed that hypoxia-inducible factor-1alpha (HIF-1alpha) was markedly accumulated in nuclei after 16-hours of hypoxia. Hypoxia reduced basal and TNF-alpha-stimulated MCP-1 expression, while it induced VEGF expression in HPTECs. In crescentic glomerulonephritis (CrGN) or diabetic nephropathy (DN) with severe proteinuria, clusters of proximal tubules and a part of the fibrotic interstitium were specifically stained for PAI-1, while no stainings were found in minor glomerular abnormality or minimal change nephrotic syndrome. Urinary PAI-1 levels were significantly higher in CrGN and DN than in healthy controls. In DN, urinary TNF-alpha levels were significantly correlated with urinary PAI-1 levels. PAI-1 induced by hypoxia and inflammation may contribute to further progression of advanced kidney disease, CrGN or DN. Hypoxia together with inflammation may also be involved in promotion of renal fibrosis in part by modulating MCP-1 and VEGF expression.