MECHANISM OF CIGARETTE SMOKE-INDUCED IMMUNO SUPRESSION

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Cigarette smoking is associated with elevated risks of upper respiratory tract infections, cancers of several organs, and cardiovascular disease. To determine whether this increased susceptibility to infections correlated with cigarette smoke-induced changes in the immune system, we studied the effects of cigarette smoke in rodents. Our results indicated that chronic exposure of rats/mice to cigarette smoke from R1 cigarettes (high tar, high nicotine, University of Kentucky Reference Cigarette) affects primarily the B lymphocyte function. Moreover, the B cell responses, most susceptible to cigarette smoke, are those which are activated via the antigen receptors (cross-linking of surface immunoglobulins). The goals of this proposal are to: (a) understand the underlying mechanism(s) for the immunosuppressive effects of cigarette smoke, and (b) to identify specific components of cigarette smoke condensate that are responsible for the observed immunosuppression. Experiments proposed herein are directed at evaluating the effects of cigarette smoke on the known stages of B cell proliferation and differentiation. B cells from smoke-exposed and sham control animals will be examined for their ability to activate, receptor crosslinking-dependent, metabolism of phosphatidylinositol, including activation of tyrosine phosphokinase (western blot analysis using mAb to phosphotyrosines), phospholipase C (metabolism of inositol phosphates), increase in intracellular calcium content (changes in indo-1 fluorescence), and the activation of protein kinase C (32P-labeling). Flow cytometry, using acridine orange staining will be employed to determine cell cycle phases. Activation of protooncogenes known to correlate with lymphocyte activation, like c-fos and c-myc, will be assayed by northern blotting using 32P- labeled cDNA probes. In addition, B cells will be examined for their ability to produce immunoglobulin (assayed by ELISA) in response to antigenic/mitogenic activation, and in the presence or absence of lymphokines such as IL4, IL5, IL6, and tau-IFN. Established protocols will be employed to obtain fractions and subfractions of cigarette smoke condensate. These fractions will be incorporated into constant-release inert pellets (pellets containing specific fractions will be made by Innovative Research of America, Toledo, OH), implanted intradermally into animals, and effects on the immune system monitored at various times after implantation. The effective fractions will be further evaluated for their ability to influence lymphocyte functions in vitro. Results from these experiments are likely to provide ideas abut the manner in which cigarette smoke affects the immune system. The applicability of these results to human smokers has been strengthened by the recent results from Weissman et al. (see Appendix 1) in which cigarette smoke affects the human immune system in a manner similar to those observed, in our laboratory, in the rat.