Molecular damage to rat liver mitochondrial H(+)-ATPase during cold preservation with UW solution.
Deterioration of rat liver mitochondrial function during cold preservation with UW solution was studied. Mitochondria were isolated from control liver (0-hr preservation), 24-hr preserved liver, and 48-hr preserved liver with UW solution at 4 degrees C. Respiration assay revealed that phosphorylation was damaged more rapidly than oxidation. Inside-out submitochondrial particles prepared from each sample by sonication in the presence of EDTA were subjected to ATPase assay. ATP hydrolyzing activity of H(+)-ATPase (ATP synthase), which plays a key role in phosphorylation in mitochondria, decreased markedly to 58% after 24-hr preservation. After 48-hr preservation, reduction to 40% of control was noted. When an intrinsic H(+)-ATPase inhibitor protein was removed from ESMP by Sephadex gel filtration, decrease of the ATPase activity was enhanced down to 49% and 29% of the control for 24-hr and 48-hr preserved liver, respectively. Molecular damage of the enzyme was confirmed by SDS-PAGE. Alpha subunit of the enzyme decreased time-dependently, and H(+)-ATPase molecules that lost alpha subunit seemed to lose their catalytic activity. Although the cause of the molecular damage of H(+)-ATPase is not clear yet, some mitochondrial protease(s) may be involved.
Tsunekawa
S
S
Second Department of Surgery, Faculty of Medicine, Kyoto University, Japan.
Tanaka
A
A
Ozawa
K
K
D000241:Adenosine; D000493:Allopurinol; D000818:Animals; D002725:Chloroform; D003080:Cold Temperature; D004492:Edetic Acid / Q000494:pharmacology; D005978:Glutathione; D007328:Insulin; D016031:Liver Transplantation / Q000502:physiology; D046911:Macromolecular Substances; D008297:Male; D008930:Mitochondria, Liver / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D009926:Organ Preservation / Q000379:methods*; D019188:Organ Preservation Solutions; D010085:Oxidative Phosphorylation; D006180:Proton-Translocating ATPases / Q000502:physiology*; D011887:Raffinose; D051381:Rats; D011919:Rats, Inbred Strains; D012996:Solutions
D007328:Insulin; D046911:Macromolecular Substances; D019188:Organ Preservation Solutions; D012996:Solutions; C055206:University of Wisconsin-lactobionate solution; D000493:Allopurinol; D002725:Chloroform; D004492:Edetic Acid; D006180:Proton-Translocating ATPases; D005978:Glutathione; D000241:Adenosine; D011887:Raffinose