Cyclic Nucleotide Changes in Murine Lymphocytes Following Thymosin Incubation In Vitro Paul H. Naylor, Gary B. Thurman, and Allan L. Goldstein Abstract: Thymosin fraction 5 and a more purified acidic fraction of thymosin at a 100-fold lower concen- tration elevated cGMP levels, but not cAMPlevels,in murine thymocytes. When both thymus and spleen lympbocytes were fractionated via a BSA gradient procedure, both cAMP and cGMP basal values varied depending on the density of the subpopulations. Thymosin Fr5 did not elevate cAMP in any thymus subpopulafions of lymphocytes obtained from the BSA density gradients. The cGMP elevation due to thymosin Fr5 in thymocytes was maximal in the most buoyant thymocytes, and no elevation of cGMP was detected in nude mouse spleen lymphocytes. These results suggest that the cGMP elevation may be an early event in the thymosin-mediated differentiation of a more mature subpopulation of thymocytes. They also suggest that utilization of subpopulations may be necessary for the complete determination of the effects of agents on cyclic nucleotide values of lymphocytes. Key Words: Thymosin; T-cell differentiation;Thymic hormones; Cyclic nucleotides INTRODUCTION The importance of the endocrine thymus in the control of immunity via secretion of a family of polypeptide hormones called thymosin has been established over the past 15 years (Goldstein et al., 1966; Trainin, 1974; White and Goldstein, 1975; Bach J-F, 1976). Recently it has been shown that thymosin is active in increasing T-cell numbers and responses in cancer patients and increasing cell-mediated immunity and resistance to infection in children with thymic- dependent primary immunodeficiency diseases (Goldstein et al., 1976). Although the effects of thymosin on various cell populations both in vivo and in vitro have been repeatedly demonstrated, the mechanism by which thymosin activates T-cell populations is not known. In many cases the effects of thymosin are very rapid; e.g., in vitro thymosin-induced induction of surface T-cell markers (I-2 hr) such as Thy-1 or Ly (Kumuro and Boyse, 1973), or expression of azathioprine or anti-0 sensitivity as assessed by the mouse rosette assay (5 rain) (Bach J-F et al., 1971). In addition, in vivo injections of thymosin have been shown to increase the response of murine spleen cells to the T-ceU mitogen, Con A, within 2 hr (Thurman and Goldstein, 1977). Received August 23, 1978; revised and accepted October 23, 1978. From the Department of ExperimentalTherapeutics, Grace Cancer Drug Center, RoswellPark Memorial Institute, Buffalo, New York (P.H.N.); and the Department of Biochemistry, George Washington University Medical Center, Washington, D.C. (G.B.T. and A.L.G.). Portions of this report have been previously presented as an abstract at the American Society for Cell Biology meeting in San Diego, November 1977 (Cell Biol 75: 100a). Address reprint requests to: Dr. Paul H. Naylor, Department of ExperimentalTherapeutics, RoswellPark Memorial Institute, 666 Elm St., Buffalo, NY 14263 USA. © Elsevier North Holland, Inc., 1979 lmmunopharmacology 1, 89-101 (1979) 0162-3109/79/01008913502.25 89