pd-150606 and Reperfusion-Injury

Calpain activation has been implicated in the development of ischemia-reperfusion (I-R) injury. Here we investigate the effects of two inhibitors of calpain activity, PD150606 and E-64, on the renal dysfunction and injury caused by I-R of rat kidneys in vivo. Male Wistar rats were administered PD150606 or E-64 (3mg/kg i.p.) or vehicle (10%, v/v, DMSO) 30min prior to I-R. Rats were subjected to bilateral renal ischemia (45min) followed by reperfusion (6h). Serum and urinary biochemical indicators of renal dysfunction and injury were measured; serum creatinine (for glomerular dysfunction), fractional excretion of Na(+) (FE(Na), for tubular dysfunction) and urinary N-acetyl-beta-d-glucosaminidase (NAG, for tubular injury). Additionally, kidney tissues were used for histological analysis of renal injury, immunohistochemical analysis of intercellular adhesion molecule-1 (ICAM-1) expression and nitrotyrosine formation. Renal myeloperoxidase (MPO) activity (for polymorphonuclear leukocyte infiltration) and malondialdehyde (MDA) levels (for tissue lipid peroxidation) were determined. Both PD150606 and E-64 significantly reduced the increases in serum creatinine, FE(Na) and NAG caused by renal I-R, indicating attenuation of renal dysfunction and injury and reduced histological evidence of renal damage caused by I-R. Both PD150606 and E-64 markedly reduced the evidence of oxidative stress (ICAM-1 expression, MPO activity, MDA levels) and nitrosative stress (nitrotyrosine formation) in rat kidneys subjected to I-R. These findings provide the first evidence that calpain inhibitors can reduce the renal dysfunction and injury caused by I-R of the kidney and may be useful in enhancing the tolerance of the kidney against renal injury associated with aortovascular surgery or renal transplantation.

Topics: Acrylates; Animals; Calpain; Cathepsins; Cysteine Proteinase Inhibitors; Kidney; Leucine; Male; Rats; Rats, Wistar; Renal Circulation; Reperfusion Injury

The goals of this study were to determine 1) the expression of calpain isoforms in rabbit renal proximal tubules (RPT); 2) calpain autolysis and translocation, and calpastatin levels during RPT injury; and 3) the effect of a calpain inhibitor (PD-150606) on calpain levels, mitochondrial function, and ion transport during RPT injury. RT-PCR, immunoblot analysis, and FITC-casein zymography demonstrated the presence of only mu- and m-calpains in rabbit RPT. The mitochondrial inhibitor antimycin A decreased RPT mu- and m-calpain and calpastatin levels in conjunction with cell death and increased plasma membrane permeability. No increases in either mu- or m-calpain were observed in the membrane nor were increases observed in autolytic forms of either mu- or m-calpain in antimycin A-exposed RPT. PD-150606 blocked antimycin A-induced cell death, preserved calpain levels in antimycin A-exposed RPT, and promoted the recovery of mitochondrial function and active Na+ transport in RPT after hypoxia and reoxygenation. The present study suggests that calpains mediate RPT injury without undergoing autolysis or translocation, and ultimately they leak from cells subsequent to RPT injury/death. Furthermore, PD-150606 allows functional recovery after injury.

Topics: Acrylates; Animals; Anti-Bacterial Agents; Antimycin A; Autolysis; Biological Transport, Active; Calcium-Binding Proteins; Calpain; Caseins; Cell Death; Cell Membrane; Cysteine Proteinase Inhibitors; Cytosol; Female; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Gene Expression Regulation, Enzymologic; Immunoblotting; Isoenzymes; Kidney Tubules, Proximal; Mitochondria; Rabbits; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Sodium