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INHIBITION OF RAT GLOMERULAR MESANGIAL CELL SODIUM/HYDROGEN EXCHANGE BY HYDROGEN PEROXIDE
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.tb01941.x
Subject(s) - amiloride , chemistry , hydrogen peroxide , dids , sodium–hydrogen antiporter , sodium , intracellular ph , extracellular , biophysics , biochemistry , biology , membrane , organic chemistry
SUMMARY 1. pH i regulation In glomerular mesangial cells (GMC) includes both Na + /H + and Cl‐/HCO 3 ‐ exchange. As a fall in pH i may protect against H 2 O 2 ‐mediated GMC damage during ischaemia‐reperfusion, the involvement of these mechanisms in the GMC pH i response to H 2 O 2 was assessed using confluent GMC grown in RPMI medium with 20% fetal calf serum (10–15 passages). 2. Cells were loaded with BCECF‐AM and pH i evaluated using standard fluorometric‐ratio techniques. In HEPES buffer, GMC exposure to H 2 O 2 dose‐dependently (25μmol/L‐1 mmol/L) decreased pH i over 10 rnin from 7.3 ± 0.1 to 6.7 ± 0.1 (at 100 μmol/L) partly due to rapid non‐competitive inhibition of amiloride‐sensltive Na + /H + exchange. 3. BCECF fluorescence in free solution was unchanged by H 2 O 2 whereas the pH i decrease was abolished by nigericin/K + clamp. Buffer capacity by direct titration was not changed by H 2 O 2 and averaged 100 ± 9 nmol/2.6±10 6 cells/pH unit. Similarly, zero‐Na + /high‐K + buffer, used to minimize Passive H + entry, did not prevent the fall in pH i while GMC H + ‐formation/extrusion, assessed by the rate of extracellular acidification in low‐capacity buffer (0.05 mmol/L), was rapidly inhibited. 4. In contrast, following only a brief 3 min exposure to 1 mmol/L H 2 O 2 , HCO 3 ‐/CO 2 buffer potentiated the inhibition of Na + / H + exchange from 50 to 80% of control and reduced the acidification from pH i 6.6 ± 0.1 to 7.15 ± 00.05. This effect was reversed (to pH i 6.8 ± 0.07) by pretreatment with 200 μmol/L DIDS, an inhibitor of Cl‐/HCO 3 ‐ exchange. 5. Thus, the decrease in GMC pH i in Fespohsë to H 2 O 2 In HEPES, partly mediated by inhibition of Na + /H + exchange and a possible redistribution of intraceliular H + , is antagonized in HCO 3 ‐/CO 2 through a DIDS‐sensitive Cl‐/HCO 3 ‐ exchange mechanism. This may act to negate potentially protective effects of low pH i and potentiate oxidative damage to membrane lipids, enzymes and intracellular organelles on reperfusion.