Amplification of pyridine nucleotide pools in mitogen-stimulated human lymphocytes

Abstract

NAD + levels in resting human lymphocytes obtained from 20 donors were found to be 69.9 ± 21.7 pmols/10 6 cells. After 3 days of phytohemagglutinin (PHA) stimulation the NAD + levels rose to 452 ± 198 pmols/10 6 cells. NADH, NADP + and NADPH also increased in mitogen-stimulated lymphocytes, but the major portion of the increase in total pyridine nucleotide pools was accounted for by the increase in NAD +. When PHA-stimulated lymphocytes were incubated in nicotinamide-deficient growth medium, there was no significant increase in their total pyridine nucleotide pools; however, the ratios of oxidized to reduced pyridine nucleotides changed in a similar fashion to cells grown in medium containing nicotinamide. When lymphocytes in nicotinamide-deficient medium were stimulated with PHA they increased their levels of DNA synthesis and cell replication in a similar fashion to cells growing in nicotinamide-supplemented media. Human lymphocytes were able to synthesize pyridine nucleotides from nicotinamide or nicotinic acid; however, in the absence of a preformed pyridine ring they did not efficiently use tryptophan for the synthesis of NAD. Uptake of [carbonyl- 14C]nicotinamide and conversion to NAD was markedly increased in PHA-stimulated lymphocytes; these cells also showed a marked increase in activity of the enzyme adenosine-triphosphate-nicotinamide mononucleotide (ATP-NMN) adenylyl transferase.