Nonspecific activation of murine lymphocytes. I. Proliferation and polyclonal activation induced by 2-mercaptoethanol and alpha-thioglycerol

Abstract

The effect of 2-mercaptoethanol (2-ME) and alpha-thioglycerol (alpha TG) on proliferation and polyclonal activation of lymphocytes was studied in cultures of spleen cells from C3H mice. Inclusion in serum-free or serum-containing medium of the optimal concentration (5 x 10(-5) M) of either 2-ME or alpha TG resulted in highly significant uptake and incorporation of tritiated thymidine ([3H]TdR) into DNA and in morphological blast transformation. These phenomena were dose-dependent, with both lower and higher doses causing less marked effects. The kinetic peak of these responses was found to occur at day 3 of culture. Improved cellular viability could not explain these results, because by day 3 there was no significant difference in viability between cells cultured in the presence or absence of 2-ME. 2-ME evoked a proliferative response in cultures of congenitally athymic (nu/nu) spleen cells that exhibited a similar but lower dose-response profile compared with that of heterozygous (nu/+) littermates. Cultures of bone marrow-derived (B) lymphocytes, generated by treatment of spleen cells with rabbit antithymocyte serum and complement, incorporated [3H]TdR to a degree at least equal to that of normal spleen cell cultures. Thymus-dependent (T) cells did not support significant 2-ME, alpha TG, or Concanavalin A responses in the absence of serum. However, when cultured in 5% fetal calf serum, definite T-cell responses occurred, though always of a lower magnitude than B-cell responses in this system. When the enriched B-cell and T-cell preparations were co-cultured, a synergistic response was noted. Macrophage dependency of the 2-ME and alpha TG effect was shown to be minimal. It is likely that the greater effectiveness of alpha TG relative to 2-ME is due to differences in the chemical structure of these two thiol compounds. The advantages of utilizing 2-ME and alpha TG as probes in the study of lymphocyte activation are evaluated and their possible mechanisms of action are discussed.