Hydrogen sulfide (H2S) is produced in various species and at low concentrations it is reported to limit apoptosis and to activate mitochondrial K + ATP channels. The study was planned to investigate how H2S donor can protect myoblasts against oxidative stress. H9C2 myoblasts were kept in DMEM culture enriched with fetal bovine serum (10%) and penicillin/streptomycin (1%). In Group I oxidative stress only was induced with hydrogen peroxide (H2O2, 400 μM). After 2 h, the concentration of lactate dehydrogenase (LHD) was measured with enzyme-spectrophotometric tests. In Group II the each culture of myoblasts was pre-treated for 24 h with a selected concentration (0.1, 1, 10, 100 mM and 1 mM) of H2S. After the washout of H2S, cells were challenged with 400 μM of H2O2. In Group III the effect of H2S in activating p44/p/42 MAPK was studied. Cells did not undergo oxidative stress, but simply received the dose of H2S (10 mM) which was seen to be the most effective dose in reducing LDH in Group II. After the treatment, p44/p/42 MAPK was tested with Western blot using a specific antibody. Hydrogen sulfide shows a dose-dependent protective activity from 0.1 to 10 mM. In fact, in Group II, at 10 mM of H2S, oxidative stress reduced LDH release to 50% with respect to the value observed in Group I. When H2S was given at doses higher than 10 mM, the protective effect disappeared. Western blot analysis showed that 10 mM of H2S induced a marked increase of the concentration of phosphorylated p-44/p/42 MAPK. Our study has demonstrated that a pre-treatment with an appropriate dose of a H2S donor protects H9C2 myoblasts against oxidative stress via a signaling cascade that include p44/p/42 MAPK.
PROTECTIVE EFFECT OF HYDROGEN SULFIDE AGAINST OXIDATIVE STRESS OF MYOBLASTS
MANCARDI, Daniele;PENNA, Claudia;TULLIO, FRANCESCA;RAIMONDO, Stefania;RASTALDO, Raffaella;LOSANO, Giovanni;PAGLIARO, Pasquale
2008-01-01
Abstract
Hydrogen sulfide (H2S) is produced in various species and at low concentrations it is reported to limit apoptosis and to activate mitochondrial K + ATP channels. The study was planned to investigate how H2S donor can protect myoblasts against oxidative stress. H9C2 myoblasts were kept in DMEM culture enriched with fetal bovine serum (10%) and penicillin/streptomycin (1%). In Group I oxidative stress only was induced with hydrogen peroxide (H2O2, 400 μM). After 2 h, the concentration of lactate dehydrogenase (LHD) was measured with enzyme-spectrophotometric tests. In Group II the each culture of myoblasts was pre-treated for 24 h with a selected concentration (0.1, 1, 10, 100 mM and 1 mM) of H2S. After the washout of H2S, cells were challenged with 400 μM of H2O2. In Group III the effect of H2S in activating p44/p/42 MAPK was studied. Cells did not undergo oxidative stress, but simply received the dose of H2S (10 mM) which was seen to be the most effective dose in reducing LDH in Group II. After the treatment, p44/p/42 MAPK was tested with Western blot using a specific antibody. Hydrogen sulfide shows a dose-dependent protective activity from 0.1 to 10 mM. In fact, in Group II, at 10 mM of H2S, oxidative stress reduced LDH release to 50% with respect to the value observed in Group I. When H2S was given at doses higher than 10 mM, the protective effect disappeared. Western blot analysis showed that 10 mM of H2S induced a marked increase of the concentration of phosphorylated p-44/p/42 MAPK. Our study has demonstrated that a pre-treatment with an appropriate dose of a H2S donor protects H9C2 myoblasts against oxidative stress via a signaling cascade that include p44/p/42 MAPK.
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