Modulation of metabolism in HepG2 cells upon treatment with cyclosporin A and Nva2-cyclosporin.
HepG2 cells were cultured in the presence of different concentrations of cyclosporin A (CsA) or Nva2-cyclosporin (Nva2-Cs) for up to 20 days. At a low concentration (2 micrograms/ml) of CsA or Nva2-Cs, the [3H]thymidine incorporation into DNA and the rate of incorporation of [3H]leucine into total protein decreased by 20-25%. Concentrations of 10 micrograms/ml resulted in a 70% reduction of the [3H]thymidine incorporation in comparison with controls. Low concentrations of CsA resulted in mitochondria in the condensed state together with autophagosomes, large vacuoles, and elevated numbers of coated vesicles, as shown by electron microscopy. Low concentrations of Nva2-Cs resulted in swollen mitochondria, increased autophagocytosis, and increased numbers of intermediate filaments and microtubules. Higher doses of these substances (5 micrograms/ml) caused disarrangement of mitochondrial cristae, vesiculation of the endoplasmic reticulum, an elevated number of free polysomes, and accelerated autophagocytosis. Labeling of phospholipids and triglycerides with [3H]glycerol and of cholesterol and dolichol with [3H]acetate was decreased after exposure of HepG2 cells to CsA, or, in particular, Nva2-Cs. Phospholipids secreted from the cells into the medium exhibited an increased level of labeling, but the specific radioactivity of the neutral lipids in the medium was significantly decreased. Treatment of HepG2 cells with either CsA or Nva2-Cs doubled the mitochondrial cytochrome oxidase and carnitine acetyl-transferase, as well as microsomal NADPH-cytochrome c reductase activities. Such treatment also increased the cyanide-insensitive beta-oxidation of fatty acids in peroxisomes, as well as cytoplasmic DT-diaphorase and glutathione transferase activities. Prolonged treatment of the cells with CsA did not result in any cumulative effect. HepG2 cells appear to be suitable for studying the effects of cyclosporins on cellular structure and metabolism and in this system the two drugs studied here exhibited similar effects.
Bäckman
L
L
Clinical Research Center, Huddinge Hospital, Karolinska Institute, Stockholm, Sweden.
Appelkvist
E L
EL
Sundberg
A
A
Teclebrhan
H
H
Brunk
U
U
D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000648:ultrastructure; D002332:Carnitine O-Acetyltransferase / Q000378:metabolism; D002374:Catalase / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D016572:Cyclosporine; D003524:Cyclosporins / Q000494:pharmacology*; D008058:Dihydrolipoamide Dehydrogenase / Q000378:metabolism; D004286:Dolichols / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000378:metabolism; D044943:Fatty Acid Desaturases / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D050356:Lipid Metabolism; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology; D008854:Microscopy, Electron; D009245:NADH Dehydrogenase / Q000378:metabolism; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured
D003524:Cyclosporins; D004286:Dolichols; D011506:Proteins; D010171:Palmitoyl Coenzyme A; C045571:cyclosporin G; D016572:Cyclosporine; D002784:Cholesterol; D002374:Catalase; D044943:Fatty Acid Desaturases; D009245:NADH Dehydrogenase; D008058:Dihydrolipoamide Dehydrogenase; D003576:Electron Transport Complex IV; D002332:Carnitine O-Acetyltransferase; D005982:Glutathione Transferase