Acid mucopolysaccharides in fibroblast cultures. 4. 35S-sulfate incorporation in dependence on pH-value cell density and lactate.

UNLABELLED

Cultures of embryonic rat fibroblasts were incubated with 35S-sulfate at pH 6.6 and 7.4 (Eagle medium, Hepes buffer) for 48 hours. The MPS were isolated and fractionated. Determination of hexuronic acid was done according to BITTER and MUIR, measuring of sulfate incorporation by liquid scintillation counting.

RESULTS

1. HS, Ch-6-S and DS showed different synthesis rates. They rised at increasing cell density and were lower at pH 6.6 than at 7.4. 2. At pH 6.6 the sulfate incorporation rates (de novo-synthesis) of HS, Ch-6-S and DS are lowered. The percent increase of the MPS fractions at pH 6.6 and their specific activities are explained by individual differences in the unlabelled MPS pools as well as by differences in the rates of synthesis and catabolism. The cell density also has an influence on that. Increase of the MPS at pH 6.6 is by mainly due to an inhibition of catabolism. 3. Contrary to CH-6-S and DS heparan sulfate is catabolized more slowly at high cell densities. 4. The volume of the unlabelled MPS pool is modified by lactate at pH 6.6, e.g. it effectuates enhancement of the DS pool at low cell densities. De novo-synthesis and specific activity are scarcely influenced. CH-6-S and DS catabolism are considerably inhibited, their total amount is increased. The relative synthesis rate of DS is enhanced by lactate. 5. The possible importance of the results for inflammatory processes and wound healing is mentioned.