title: Hydrogen peroxide increases the intracellular calcium activity in rat mesangial cells in primary culture.
abstract: Oxygen radicals are known to be mediators of renal injury under several pathophysiological conditions. We have examined the effect of hydrogen peroxide (H2O2) on intracellular calcium activity ([Ca2+]i) in mesangial cells in primary culture. Mesangial cells were loaded with 1 mumol/liter fura-2, and kept in a Ringer-like solution. Fura-2 fluorescence was measured in an inverted microscope at 37 degrees C. Angiotensin II (0.1 nmol/liter) and ATP (0.1 mumol/liter) induced a rapid transient increase of [Ca2+]i, which was followed by a sustained plateau (N = 37 and N = 24). In contrast, the addition of H2O2 (0.01 to 10 mmol/liter, N = 157) caused a time- and concentration-dependent slow increase of [Ca2+]i, which reached a stable [Ca2+]i plateau after 3 to 10 minutes (ED50: 100 mumol/liter). After the removal of H2O2 [Ca2+]i decreased partially and reached a stable value approximately 90% above the resting [Ca2+]i value. Addition of 100 mumol/liter H2O2 to an extracellular Ca(2+)-free solution resulted either in no rise of [Ca2+]i in some experiments (N = 7), or [Ca2+]i oscillations in others (N = 10). In the presence of H2O2 (> 25 mumol/liter), the angiotensin II or ATP mediated increases in [Ca2+]i were almost completely inhibited (N = 15 and N = 10). The cations Ni2+ and La3+ and the Ca(2+)-antagonist verapamil (10 mumol/liter) did not inhibit the H2O2 mediated increase of -Ca2+-i (N = 6 to 9). Flufenamate (100 mumol/liter), an inhibitor of non-selective cation channels inhibited the H2O2 induced increase of [Ca2+]i by 63 +/- 11% (N = 7). Preincubation of the cells with a disulphide reducing agent (dithiothreitol, 500 mumol/liter, N = 5) or an iron-chelator (deferoxamine, 100 mumol/liter, N = 5) attenuated the H2O2 mediated effect by 95 +/- 15% and 74 +/- 6%, respectively. The H2O2 mediated [Ca2+]i increase was completely inhibited when mesangial cells were preincubated with 1 mumol/liter U-83836E, an inhibitor of lipid peroxidation (N = 7), and inhibited by 84 +/- 6% when the cells were pretreated with 1 mmol/liter pyruvate (N = 5). The data indicate that H2O2: (i) increases [Ca2+]i in mesangial cells by a mechanism distinct from angiotensin II or ATP and (ii) that it inhibits the [Ca2+]i response to both agonists.
authors.affiliation: Department of Nephrology, Albert-Ludwig Universität, Freiburg, Germany.
authors.forename: T N
authors.initials: TN
authors.lastname: Meyer
authors.forename: J
authors.initials: J
authors.lastname: Gloy
authors.forename: M J
authors.initials: MJ
authors.lastname: Hug
authors.forename: R
authors.initials: R
authors.lastname: Greger
authors.forename: P
authors.initials: P
authors.lastname: Schollmeyer
authors.forename: H
authors.initials: H
authors.lastname: Pavenstädt
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D000931:Antidotes / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002374:Catalase / Q000494:pharmacology; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D002839:Chromans / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D003676:Deferoxamine / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D005439:Flufenamic Acid / Q000494:pharmacology; D005920:Glomerular Mesangium / Q000166:cytology* / Q000201:enzymology; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007811:Lanthanum / Q000494:pharmacology; D009532:Nickel / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D019289:Pyruvic Acid / Q000494:pharmacology; D051381:Rats; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D000931:Antidotes; D000975:Antioxidants; D002839:Chromans; D010879:Piperazines; D013439:Sulfhydryl Reagents; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; D000804:Angiotensin II; C066382:U 78517F; D005439:Flufenamic Acid; D007811:Lanthanum; D009532:Nickel; D019289:Pyruvic Acid; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D014700:Verapamil; D002374:Catalase; D003676:Deferoxamine; D002118:Calcium; D004229:Dithiothreitol