Abstract
Exogenous cyclic adenosine 3′,5′-monophosphate (cAMP) inhibits the growth of plasma cell tumor (MPC-11) in culture and the inhibition is entirely prevented by the addition of insulin to the culture medium (1). Opposite effects of cyclic AMP and insulin have been described in a number of nonneoplastic cells (2-6), a finding which prompted the search for biochemical effects altered in opposite directions by these two agents.
Materials and methods. Cyclic AMP, N6-O2-dibutyryl adenosine 3′,5′-cyclic monophosphate (DBcAMP) and 5-adenosine monophosphate (AMP) were purchased from Sigma Chemical Company, adenosine from Schwarz/Mann, crystalline zinc porcine insulin (containing 0.002% glucagon) was a gift from Lilly Research Laboratory. Denatured insulin was prepared as described by Lotten and Sneyd (7). 14C-thymidine was purchased from Schwarz/Mann, 3H-leucine and 14C-inulin were obtained from New England Nuclear. The purity of all nucleotides and nucleosides was checked by thinlayer chromatography in several systems.
Incubation conditions. MPC-11 cells, which have been in culture for many months, were used in these experiments. The culture and growth characteristics have been described previously (1). Cells were washed with Dulbecco's modified Eagle's medium without serum, and resuspended in regular growth medium at a density of 2 × 106 cells/ml. Incubations were carried out in 25 ml siliconized Erlenmeyer flasks in 5% CO2, 95% air at 37° in a Psycro Therm incubator shaker (80 rpm) for 60 min. Each incubation flask contained 5.0 ml of suspension and 0.5 ml Hanks' Balanced Salt Solution (HBSS) or other indicated additions. Incubations were carried out for 60 min prior to addition of tracers (0.2 μCi/ml 14C-thymidine, 0.5 μCi/ ml 3H-leucine, or 0.1 μCi/ml 14C-inulin and continued for 30 min thereafter.
After incubation the cells were centrifuged, washed three times with HBSS and resuspended in 5.0 ml HBSS.
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