The effect of the CD28 activation pathway on the immunosuppressive action of CsA was assessed. Human peripheral blood lymphocytes were stimulated with anti-CD3, bryostatin (Bryo) a novel activator of protein kinase C (PKC) and anti-CD28 singly or in combination, to which graded doses of CsA were added to determine relative sensitivity. Proliferation, IL-2 production, and IL-2 receptor expression were assessed and the IC50 determined. Lymphocytes stimulated with Bryo exhibited a marginal proliferative response but expressed the IL-2 receptor despite the presence of CsA. Addition of anti-CD3 or anti-CD28 to Bryo-stimulated lymphocytes promoted a vigorous proliferative response. CsA effectively inhibited the proliferative response and IL-2 production induced with anti-CD3 and Bryo but did not inhibit the response of cells stimulated with anti-CD28 and Bryo. However, II-2 receptor expression in both sets of cultures were comparable due to the induction of IL-2 receptor by Bryo and was not inhibited by CsA. Costimulation of lymphocytes with anti-CD3 plus anti- CD28 resulted in a 2-3-fold enhancement of proliferation compared with lymphocytes stimulated with anti- CD3 alone. Addition of CsA to lymphocytes stimulated with anti-CD3 resulted in the dose-dependent suppression of the proliferative response and IL-2 production (ICSo= 10-25 nM) but less so for IL-2 receptor expression (IC50=100-150 nM). In comparison, the proliferative response and IL-2 production elicited by anti-CD3 + anti-CD28 was more resistant to the effects of CsA (IC50=100-200 nM). However, IL-2 receptor expression exhibited comparable sensitivity to CsA (IC50= 100-200 nM) in the presence of anti-CD28. Combination drug:drug studies revealed that CsA and the protein kinase C inhibitor H-7 were additive for both anti-CD3 and anti-CD3 plus anti-CD28 response. On the other hand, the cGMP-dependent protein kinase inhibitor H- 8 was synergistic with CsA in inhibiting the response of lymphocytes to anti-CD3 plus anti-CD28 but only additive for responses to anti-CD3. Taken together, these data suggest that CsA inhibits T cell activation at two distinct levels, leading to inhibition of IL-2 production and inhibition of IL-2 receptor expression. Activation of the CD28 pathway partially overcomes the inhibitory activity of CsA on IL-2 production and may be mediated by indirect activation of a cGMP-dependent protein kinase. The failure of the CD28 activation pathway to completely overcome the inhibitory effects of CsA when used in conjunction with anti-CD3 stimulation (compared with complete CsA resistance of IL-2 production when used with activators of protein kinase C) suggests that another pathway involved in T cell activation leading to IL-2 receptor expression is inhibited by CsA.
|Original language||English (US)|
|Number of pages||9|
|State||Published - Jun 1991|
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