Δ9-tetrahydrocannabinol (Δ9-THC), the main psychoactive congener in marijuana, modulates a variety of immunological responses, of which humoral immune responses against T cell antigens are particularly sensitive to suppression by Δ9-THC. Among different types of contact-mediated B cell activation, the CD40-CD40L interaction between B cells and activated CD4+ T cells plays an important role in all stages involved in plasma cell differentiation. Given that both B cells and activated CD4+ T cells are crucial for T cell-dependent humoral immune response, it is important to assess the effects of ∆9-THC on both the indirect suppression of accessory T-cell function and/or the direct suppression of B-cell function. The ultimate goal of this project was to identify potential molecular targets responsible for Δ9-THC-impairment of signal transduction cascades and gene expression during lymphocyte activation by testing the following hypothesis: Δ9-THC attenuates the human T cell-dependent IgM antibody response by suppression of CD40L upregulation in activated CD4+ T cells and impairment of the CD40-mediated B cell activation. Therefore, the objectives of the present studies were three-fold; of which the first two objectives focused on T cells, whereas the third objective focused on B cells. The first objective investigated the effects of ∆9-THC and the involvement of cannabinoid receptor (CB) 1, CB2, and glucocorticoid receptor (GR) on CD40L expression in mouse splenic T cells. The induction of CD40L on activated mouse splenic CD4+ T cells was differently modulated by ∆9-THC, depending on the mode of T cell activation, (i.e., antibodies directed to CD3 and CD28; anti-CD3/CD28 or phorbol ester plus calcium ionophore; PMA/Io). ∆9-THC significantly impaired the upregulation of surface CD40L on CD4+ T cells induced by anti-CD3/CD28, but not by PMA/Io. Further, suppression of anti-CD3/CD28-induced CD40L expression in mouse splenic CD4+ T cells by ∆9-THC likely occurred at the transcriptional level, independently of CB1, CB2 or GR. The second objective was to investigate the mechanisms by which ∆9-THC suppressed CD40L expression by activated primary human CD4+ T cells. Attenuation of anti-CD3/CD28-induced CD40L expression in activated primary human CD4+ T cells by ∆9-THC was mediated, at least in part, through suppression of NFAT and NFκB-DNA binding activity. The inhibitory effect of Δ9-THC on the activation of NFAT and NFκB in primary human CD4+ T cells also involved impairment of Ca2+ elevation without perturbation of proximal T cell receptor signaling events (e.g., tyrosine phosphorylation of ZAP70, Akt, PLCγ1/2, and GSK3β). The third objective was to assess the direct effect of ∆9-THC on the effector function of primary human B cells using a polyclonal in vitro activation model to generate primary immunoglobulin M (IgM) antibody secreting cells. Δ9-THC significantly decreased the number of IgM secreting cells, which correlated with the impairment of plasma cell differentiation as evidenced by suppression of immunoglobulin joining chain (IgJ) mRNA expression. Δ9-THC specifically decreased surface expression of CD80, but not other B cell activation markers (CD69, CD86, and ICAM1). Despite the modest effect on B cell activation, Δ9-THC profoundly attenuated the proliferation of activated B cells, preventing the progression of B cells toward plasmacytic differentiation. In addition, pretreatment with ∆9-THC was accompanied by a robust decrease of STAT3 phosphorylation, whereas the phosphorylation of p65 NFκB subunit was not affected in activated B cells. Collectively, this dissertation research demonstrated that ∆9-THC exhibits stimulation- and/or cell type-specific selectivity of NFκB inhibition, and identified several aspects of the multi-faceted mechanism by which ∆9-THC suppresses T cell-dependent humoral immunity in humans. A better understanding of the mechanism of cannabinoid-mediated immune function and can be used to address a risk-to-benefit ratio for therapeutic marijuana use.
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