T-cell proliferation involving the CD28 pathway is associated with cyclosporine-resistant interleukin 2 gene expression.

Abstract

CD28 is a homodimeric glycoprotein expressed on the surface of a major subset of human T cells that has recently been identified as a member of the immunoglobulin supergene family. The binding of monoclonal antibodies to the CD28 antigen on purified T cells does not result in proliferation; however, previous studies have shown that the combination of CD28 stimulation and protein kinase C activation by phorbol myristate acetate (PMA) results in T-cell proliferation that is independent of both accessory cells and activation of the T-cell receptor-CD3 complex. In the present study, effects of stimulation by anti-CD28 on cell cycle progression and on the interleukin 2 (IL-2) and IL-2 receptor system have been investigated on primary cultures of purified peripheral-blood CD28+ T cells. There was no measurable effect on cell size or on DNA synthesis after stimulation of resting (GO) cells by CD28 alone. After 3 h of activation of T cells by PMA alone, a slight (8%) increase in cell volume occurred that did not progress to DNA synthesis. In contrast, T-cell stimulation by CD28 in combination with PMA resulted in a progressive increase in cell volume in --100% of cells at 12 to 14 h after stimulation. Northern blot (RNA blot) analysis revealed that CD28 stimulation alone failed to cause expression of the alpha chain of the IL-2 receptor or of IL-2 mRNA, and in accord with previous studies, stimulation by PMA alone resulted in the accumulation of IL-2 receptor transcripts but no detectable IL-2 mRNA. In contrast, T-cell stimulation by the combination of CD28 and PMA resulted in the appearance of IL-2 transcripts and enhanced expression of IL-2 receptor mRNA. Functional studies revealed that the proliferation induced by CD28 and PMA stimulation was entirely resistant to cyclosporine, in contrast to T-cell activation induced by the CD3-T-cell receptor complex. Cyclosporine was found not to affect the accumulation of IL-2 mRNA after CD28 plus PMA stimulation, although there was no detectable IL-2 mRNA after stimulation by CD3 in the presence of the drug. Furthermore, stimulation by CD28 in combination with immobilized CD3 antibodies caused a striking enhancement of IL-2 mRNA expression that was, in part, resistant to the effects of cyclosporine. These studies indicate that the CD28 molecule synergizes with protein kinase C activation to induce IL-2 gene expression and demonstrate that stimulation by the CD28 pathway can cause vigorous T-cell proliferation even in the presence of cyclosporine and that cyclosporine does not prevent transcription of 16-2 mRNA, as has been suggested previously. Moreover, these findings suggest that a potential role for the CD28 molecule in vivo may be to augment IL-2 production after stimulation of the CD3-T-cell receptor molecular complex and thereby to amplify an antigen-specific immune response. Finally, these results provide further evidence that the CD28 molecule triggers T-cell proliferation in a manner that differs biochemically from CD3-T-cell receptor-induced proliferation.The binding ...

CD28 Utilizes Vav-1 to Enhance TCR-Proximal Signaling and NF-AT Activation

“…A recent report has suggested that Vav-1 has no apparent role in co-stimulation based on the evidence that Vav-1-deficient T cells activated by anti-CD28 Ab and PMA showed no defect in proliferation and IL-2 production (55). This type of stimulation which generates CsA-resistant lymphokine expression (56), is likely to be supraphysiological and to bypass relevant signaling steps elicited by cell surface receptors (57). Indeed, CD28 enhancement of a TCR-induced increase in IL-2 mRNA is potently, though not entirely, inhibited by CsA (56, 58) and we found that NF-AT activation induced by Vav-1 overexpression/CD28 engagement (Fig.…”

Late Signals from CD27 Prevent Fas-Dependent Apoptosis of Primary CD8+ T Cells

“…The number of known costimulatory molecules has increased drastically since the characterization of CD28 as a costimulator of T cells nearly 20 years ago (7). Many TNFR family members such as CD30 (8,9), OX40 (10), and 4-1BB (11,12) have also been shown to be involved in T cell costimulation.…”

Arterial and venular endothelial cell costimulation of cytokine secretion by human T cell clones

“…Costimulation signals delivered through CD28 has been reported to confer resistance to the immunosuppressant cyclosporin A [26]. The A17 and M2 T cell clones used in these experiments do not express CD28 and resting VIEC and AIEC do not express B7.…”

“…HUVEC [28], but not AIEC or VIEC (Fig. 8), confer upon costimulated T cells partial resistance to the immunosuppressant cyclosporin A. Cyclosporin A resistance is also observed after costimulation through CD28 [26], which activates the principle factor for IL-2 transcription, NFAT, through a separate pathway [41] involving phosphorylation of the jun subunit of AP-1 [42]. However, costimulation by AIEC, VIEC, or HUVEC does not involve the B7-CD28 pathway because these EC do not express B7 and costimulation is not inhibited by soluble CTLA4 (CTLA4Ig), which blocks signaling through B7 (Fig.…”

“…HUVEC are thought to costimulate IL-2 production by increasing transcription of the IL-2 gene, which correlates with an increase in the fraction of the fos subunit in the heterodimeric NFAT [43]. In contrast, antibodies to CD28 both activate transcription of the IL-2 gene and stabilize IL-2 mRNA [44], which may contribute to cyclosporin A resistance [26]. Cyclosporin A inhibition of VIEC and AIEC costimulation suggests that costimulated IL-2 production occurs at the level of increased transcription of the IL-2 gene.…”