Characterization of oxygen-tolerant Chinese hamster ovary cells. II. Energy metabolism and antioxidant status.
Further characteristics of an oxygen-tolerant variant of Chinese hamster ovary cells (CHO-99) capable of stable proliferation at 99% O2/1% CO2, and O2 level that is lethal to the parental line (CHO-20), are described. Previous work has revealed that CHO-99 cells have 2- to 4-fold increased activities of superoxide dismutases, catalase and glutathione peroxidase, and substantially increased relative volumes of mitochondria and peroxisomes. To document possible additional mechanisms of O2 tolerance we compared CHO-20 cells growing at 20% O2 (normoxia) and CHO-99 cells at 99% O2 (normobaric hyperoxia). We show the following: (1) the estimated total (oxidative and glycolytic) ATP production in CHO-99 cells was 36% decreased. ATP production through oxidative phosphorylation was 52% lower in CHO-99 cells, while the relative contribution from glycolysis was increased from 6% to 30%. The ATP content was 29% lower in CHO-99 cells, the adenylate energy charge being also significantly decreased, indicating that energy production through oxidative phosphorylation is compromised in CHO-99 cells. Cyanide-resistant respiration was 4-fold higher in CHO-99 cells, probably reflecting, at least partly, the increased peroxisomal activity in these cells. (2) The level of reduced glutathione was several fold increased in CHO-99 cells, oxidized glutathione being unaltered; (NADPH + NADP+) levels were elevated 2.7-fold, while the ratio of NADPH to NADP+ was increased almost two-fold. These changes were associated with a 50% increased metabolism of glucose through the hexose monophosphate pathway. (3) No evidence was obtained for an increased steady-state level of endogenous lipid peroxidation in CHO-99 cells, in spite of a 50% increased content of polyunsaturated fatty acids in the phospholipid fraction.
van der Valk
P
P
Institute of Human Genetics, Free University, Amsterdam, The Netherlands.
Gille
J J
JJ
van der Plas
L H
LH
Jongkind
J F
JF
Verkerk
A
A
Konings
A W
AW
Joenje
H
H
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D003486:Cyanides / Q000494:pharmacology; D004351:Drug Resistance; D004734:Energy Metabolism / Q000187:drug effects*; D005227:Fatty Acids / Q000032:analysis; D005260:Female; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D007424:Intracellular Fluid / Q000378:metabolism; D008054:Lipid Peroxides / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010053:Ovary / Q000166:cytology*; D010101:Oxygen Consumption / Q000187:drug effects*; D010427:Pentose Phosphate Pathway / Q000187:drug effects
D000975:Antioxidants; D003486:Cyanides; D005227:Fatty Acids; D008054:Lipid Peroxides; D009249:NADP; D005954:Glucosephosphate Dehydrogenase; D005978:Glutathione