Cystinotic and normal fibroblasts: differential protection in cystine-free medium by dithiothreitol.
We studied survival of normal (n=4) and cystinotic (n=3) fibroblast strains in selective (cystine-free) medium, with or without dithiothreitol (DTT)-thiol or disulfide. Cells were plated at low density first in maintenance medium (containing cystine), then washed and exposed to selective medium for 24 hr or longer. Cultures were trypsinized at specified times and the cells counted. Selective medium caused progressive detachment (greater than 90% by 72 hr) of control and cystinotic cells: DTT-disulfide (4 mM) and DTT-thiol (0.5 mM) both protected cystinotic cells (p less than 0.02) but not control cells in selective medium. DTT-thiol (0.5 mM) and DTT-disulfide (4mM) both depleted the intracellular non-protein cystine pool of cystinotic cells to less than 25 percent of pretreatment levels. Cystinotic cells preincubated with cysteamine (1 mM X 4 hr) to deplete the intracellular cystine pool were not protected by DTT-disulfide than with DTT-thiol. These findings imply that mobilization of intracellular cystine protects cystinotic cells in selective medium; cystinotic fibroblasts can utilize DTT-disulfide as well as DTT-thiol to mobilize cystine.
D003470:Culture Media; D003553:Cystine / Q000378:metabolism*; D003554:Cystinosis / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D004229:Dithiothreitol / Q000494:pharmacology* / Q000633:toxicity; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans
D003470:Culture Media; D003553:Cystine; D004229:Dithiothreitol