Succinic Dehydrogenase and Cytochrome Oxidase Activities in Cell Cultures

Abstract

Succinic dehydrogenase and cytochrome oxidase have been assayed in permanent cell lines (HEP 1, HEP 2, and HLM), in short-term cultures of chick embryo heart cells, and in various tissues. Their activities in different cells are compared by relating them to deoxyribonucleic acid. They are very low in HEP 1, HEP 2, and HLM cells by comparison with the activities in any normal tissues examined. All the succinic dehydrogenase was shown to be located in the mitochondria of the permanent cell lines by staining with tetrazolium derivatives. Both enzymes were more active in tissues of 19-day chick embryos than in those of 11-or 14-day embryos. The increasing activities found during normal development were quickly curtailed or reversed when heart cells were grown as monolayer cultures.Digitonin-treated mitochondria produced preparations with much higher activities of cytochrome oxidase than untreated samples. Activities measured in this way were again very much lower in HEP 1, HEP 2, and HLM cells than in the normal tissues. From the derived ratio of cytochrome oxidase:succinic dehydrogenase, it was apparent that cytochrome oxidase is diminished to a greater extent than succinic dehydrogenase in both permanent cell lines and short-term cultures, by comparison with the corresponding activities in embryonic and adult tissues. The features common to the metabolism of proliferating cells in vitro and malignant cells are discussed.

Factors affecting the lactate dehydrogenase isoenzyme pattern of cultured kidney-cortex cells

“…Many cells grow satisfactorily irrespective of whether oxidative or glycolytic pathways are predominant (Leslie & Jarnell, 1960). The content of cytochrome oxidase and the formation of mitochondria in cultured cells depends on oxygen partial pressure (Leslie & Jarnell, 1960;Linnae, 1965;Hakami & Pious, 1967). The described decrease in H subunit/ M subunit ratio may reflect the adaptation of cultured cells to a diminished supply of oxygen.…”

Utilization of 14c-Labelled Glucose by Human Cells (Strain Hlm) in Tissue Culture

“…Previous investigations have shown that embryonic and malignant cells in tissue culture respond to the presence of insulin. In cultures of freshly explanted embryonic-chick-heart cells insulin stimulated cell proliferation, as measured by increments in DNA phosphorus, RNA phosphorus, lipid phosphorus and protein nitrogen (Leslie & Davidson, 1951; Leslie & Paul, 1954); it also stimulated glucose uptake, but decreased total a-oxo acid production by these cells (Leslie & Paul, 1954). The hormone elicited a different response, however, in some experiments on established strains.…”

“…and FeSO4,7H,O (0-092 mg./100 ml.). As insulin stimulates culture growth (Leslie & Paul, 1954;Leslie et al 1957), Boots Pure Drug Co. Ltd. crystalline insulin (1 unit/ml.) was included in the stock medium.…”

“…The cells were scraped off the glass surface into 3 ml. of ice-cold N-perchloric acid, and the suspension was transferred to a pointed-end roller tube (Leslie & Paul, 1954). After thorough mixing to ensure extraction of acid-soluble material, the cells were centrifuged down at 1800 g for 5 min.…”

“…Chemical analysis of the medium. The deproteinized medium was analysed for the uptake or production of the following components: glucose by the anthrone method of Trevelyan & Harrison (1952); lactic acid as described by Hullin & Noble (1953); and oa-oxo acids (mainly pyruvic acid) by the method of Friedemann & Haugen (1943) as modified by Leslie & Paul (1954).…”