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Abstract
The transport activity for cystine in cultured human fibroblasts decreased after incubation of the cells under a low oxygen concentration. After the incubation for 48 h under 3% oxygen, the Vmax of the transport was decreased to less than one-third of that of the control cells, with little change in Km. The similar transport activity was observed in the cells cultured under 3% oxygen for 10-40 days with several times of passages. When these low oxygen-cultured cells were incubated under room air, the activity was enhanced with a lag of about 4 h and was almost completely restored within 24 h. This restoration required protein synthesis. The cystine transport activity increased by 50% after exposure of the cells to hyperoxia (40% oxygen). From these results it is concluded that the transport activity for cystine is induced by oxygen. In contrast, little change in the transport activities for alanine and leucine occurred in the cells exposed to the corresponding hypoxia or hyperoxia. Since the cystine transported into the cells is reduced to cysteine and the cysteine readily exits to the culture medium where it autoxidizes to cystine, a cystine-cysteine cycle across the plasma membrane has been postulated. Since the autoxidation of cysteine in the culture medium was markedly slowed down under the low oxygen concentration, the change in the cystine transport activity in response to the oxygen concentration was regarded as pertinent. Induction of the cystine transport activity may constitute a protective mechanism against the oxidative stress, to which the culture cells are exposed, by providing the cells with cysteine which is mainly incorporated into glutathione.
Highlights
The transport activity for cystine in culthuuremdan amino acid transport systems, primarily by System ASC in fibroblasts decreased afterincubation of the cells un- humanfibroblasts (4)
After the incubation transport systemsof mammalian cells generallymediate both for 48 h under 3% oxygen, the Vmaxof the transport the inward and outward transport (5)
Thesimilar transport activity was observed in the cells cultured under 3% oxygen for 10-40 days with several timeosf passages. When these low oxygen-cultured cells were incubated under room air, the activity wasenhanced with a lag of about 4 h and was almost completely this bidirectional function of the transport systems, cysteine constantly exitsfrom the cell because it autoxidizes to cystine in the extracellular fluid
Summary
Thesimilar transport activity was observed in the cells cultured under 3% oxygen for 10-40 days with several timeosf passages. Cells plated at 1X 106/35-mmdiameter dish tration, the change in the cystine transport activity oinr at 2.5 X 105/60-mmdiameter dish were incubated for 2 days (unless response to theoxygen concentration was regardedas otherwise stated)before the experiment. The transport of amino acids is a cell membrane function dish were rinsed three times in warmed phosphate-buffered saline sensitive to environmental changes.We have described in Cultured human fibroblasatsNa’-independent, anionic amino glucose, and 10 mM phosphate, pH 7.4) They were incubated in acid transport system highly specific for cystine and glutamate.
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