The effects of actinomycin D on the primary and secondary immune responses induced in vitro Public Deposited

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  • In order to assess the importance of RNA synthesis in the induction and antibody-producing phases of immune responses, actinomycin D was used to inhibit the primary and secondary in vitro immune responses of mouse spleen cells against heterologous erythrocytes. The presence of actinomycin D (0.1 or 0.125 μg /ml) throughout the entire four day culture period inhibited the development of plaque-forming cells (PFCs) in both primary and secondary responses by more than 99%. The same drug concentration, if added after 1, 2, or 3 days of culture, inhibited the number of PFCs on the fourth day by 81-99%. Utilizing cultures of spleen cells from mice already undergoing a secondary immune response, no stimulation of the PFC number upon a 24 hour treatment with actinomycin D (1.25 μg /ml) was found, unlike the observation of Harris (J. Experimental Medicine 127:675, 1968), with a similar system of cultured rabbit spleen cells. The treatment of these mouse spleen suspensions for 24 hours with actinomycin D (1.25 μg /ml) usually brought about a 90-95% decrease in the number of PFCs. For both the primary and secondary in vitro immune responses the sensitivity to actinomycin D treatment throughout the culture period was determined, A drug concentration of 10⁻³ μg/ml caused no inhibition in either type of response, but rather caused some stimulation of PFC numbers by the end of the fourth day of culture, The secondary response was inhibited by 43% and 98% by actinomycin D concentrations of 3x10⁻³ and 10⁻² μg /ml respectively, The primary response was inhibited by 95% and 99% by actinomycin D concentrations of 3x10⁻³ and 10⁻² μg /ml respectively, This sensitivity of the entire response is much greater than that reported by Uyeki and Llacer (Biochemical Pharmacology 18:948, 1969) for actinomycin D additions at a later stage of the response, The spleen cell suspension was separated into a macrophagerich fraction and a lymphocyte-rich fraction. Treatment of the macrophage population for one hour with actinomycin D (0.1 μg /ml), either during or before the addition of antigen, inhibited by 95% the immune response which resulted following removal of the drug and addition of the lymphocyte-rich population to reconstitute the original cell suspension. The results are discussed with respect to the possible events occurring during the response which would account for the observed sensitivity to actinomycin D. A suggestion is made that the extremely high sensitivity of the early phases of the immune response may be due to the need for synthesis of some very large RNA molecule.
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