Protection of LLC-PK1 cells against hydrogen peroxide-induced cell death by modulation of ceramide level.
Oxidative stress has been reported to elevate ceramide level during cell death. The purpose of the present study was to modulate cell death in relation to cellular glutathione (GSH) level and GST (glutathione S-transferase) expression by regulating the sphingolipid metabolism. LLC-PK1 cells were treated with H2O2 in the absence of serum to induce cell death. Subsequent to exposure to H2O2, LLC-PK1 cells were treated with desipramine, sphingomyelinase inhibitor, and N-acetylcysteine (NAC), GSH substrate. Based on comparative visual observation with H2O2-treated control cells, it was observed that 0.5 microM of desipramine and 25 mM of NAC exhibited about 90 and 95% of cytoprotection, respectively, against H2O2-induced cell death. Desipramine and NAC lowered the release of LDH activity by 36 and 3%, respectively, when compared to 71% in H2O2-exposed cells. Cellular glutathione level in 500 microM H2O2-treated cells was reduced to 890 pmol as compared to control level of 1198 pmol per mg protein. GST P1-1 expression was decreased in H2O2-treated cells compared to healthy normal cells. In conclusion, it has been inferred that H2O2-induced cell death is closely related to cellular GSH level and GST P1-1 expression in LLC-PK1 cells and occurs via ceramide elevation by sphingomyelinase activation.
Yoo
Jae-Myung
JM
College of Pharmacy, Chungbuk National University, Cheongju 361-763, Korea.
Lee
Youn-Sun
YS
Choi
Heon-Kyo
HK
Lee
Yong-Moon
YM
Hong
Jin-Tae
JT
Yun
Yeo-Pyo
YP
Oh
Seikwan
S
Yoo
Hwan-Soo
HS
D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D016923:Cell Death / Q000187:drug effects; D002518:Ceramides / Q000378:metabolism*; D003891:Desipramine / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase / Q000096:biosynthesis; D006861:Hydrogen Peroxide / Q000378:metabolism*; D007527:Isoenzymes / Q000096:biosynthesis; D018374:LLC-PK1 Cells; D018384:Oxidative Stress; D013107:Sphingolipids / Q000096:biosynthesis; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors; D013552:Swine
D002518:Ceramides; D007527:Isoenzymes; D013107:Sphingolipids; D006861:Hydrogen Peroxide; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase; D013108:Sphingomyelin Phosphodiesterase; D005978:Glutathione; D003891:Desipramine; D000111:Acetylcysteine