Premium
ROLE OF INTRACELLULAR SIGNALLING PATHWAYS IN HYDROGEN PEROXIDE‐INDUCED INJURY TO RAT GLOMERULAR MESANGIAL CELLS
Author(s) -
Shaw Sidney,
Naegeli Patricia,
Etter JeanDaniel,
Weidmann Peter
Publication year - 1995
Publication title -
clinical and experimental pharmacology and physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.752
H-Index - 103
eISSN - 1440-1681
pISSN - 0305-1870
DOI - 10.1111/j.1440-1681.1995.tb02328.x
Subject(s) - chemistry , ruthenium red , phospholipase c , intracellular , pertussis toxin , thapsigargin , extracellular , biochemistry , calcium , pharmacology , biology , signal transduction , g protein , organic chemistry
SUMMARY 1. Brief exposure of cultured rat glomerular mesangial cells (GMC) to H202 in nominally bicarbonate‐free solution induced a rapid dose dependent, dantrolene‐inhibitable increase in intracellular free Ca 2+ from 65 ± 6 to 203 ± 14 nmol/L and a prolonged release of [ 14 C]‐arachidonic acid [ 14 C]‐AA which preceded the onset of cell membrane damage assessed by trypan‐blue uptake. 2. Ca 2+ responses were potentiated in HCO 3 − /CO 2 containing buffers and reached values of 1145 ± 100 nmol/L at 1 mmol/L H 2 O 2 . In HCO 3 − /CO 2 solutions, but not HEPES buffer, HzOz‐induced Ca 2+ increases were markedly attenuated by verapamil (100 μmol/L) or removal of extracellular calcium. 3. Enhanced release of [ 14 C]‐AA was partially attenuated by inhibitors of key intracellular signalling mechanisms including the phospholipase‐A 2 (PLA 2 ) inhibitor mepacrine (100 μmol/L), the NADPH oxidase inhibitor diphenyliodonium (10 μmol/L), the mitochondrial calcium‐cycling inhibitor ruthenium red (100 μmol/L) and the iron chelator dipyridyl (100 μmol/L). Release was unaffected by protein kinase C inhibition with H7 (100 μmol/L), inositol triphosphate antagonism with neomycin (1 mmol/L) or overnight treatment with the G‐protein antagonist pertussis toxin (5 μg/mL). 4. Several structurally diverse lipoxygenase inhibitors, including esculetin, baicalein and phenidone, over the dose range 1–100 μmol/L, also prevented [ 14 C]‐AA release and markedly protected against cell membrane damage. No drug directly scavenged H202 assessed by UV absorption. 5. These results indicate that H 2 O 2 activates in GMC a complex series of interrelated pathological mechanisms which in turn contribute to a prolongation of oxidative damage beyond the time of the initial exposure. These include an increase in intracellular calcium which, depending upon conditions, appears to be mediated by release from intracellular stores as well as Ca 2+ entry from the extracellular space. In turn there is a sustained release of arachidonic acid, which may partly depend on prolonged activation of PLA 2 but not phospholipase C. 6. Release of [ 14 C]‐AA could be attenuated by inhibitors of NADPH oxidase, mitochondrial calcium‐cycling, iron chelators and a structurally diverse range of lipoxygenase inhibitors in association with protection from H 2 Otrnediated cell membrane damage.