NFAT Inhibitor

Human mast cells express functional A(2A) and A(2B) adenosine receptors. However, only stimulation of A(2B), not A(2A), leads to secretion of interleukin (IL)-4, an important step in adenosine receptor-mediated induction of IgE synthesis by B-cells. In this study, we investigate intracellular pathways that link stimulation of A(2B) receptors to IL-4 up-regulation in HMC-1 mast cells. Both A(2A) and A(2B) receptors couple to G(s) proteins and stimulate adenylate cyclase, but only A(2B) stimulates phospholipase Cbeta through coupling to G(q) proteins leading to activation of protein kinase C and calcium mobilization. Inhibition of phospholipase Cbeta completely blocked A(2B) receptor-dependent IL-4 secretion. The protein kinase C inhibitor 2-{8-[(dimethylamino)-methyl]-6,7,8,9-tetrahydropyrido[1,2-a]indol-3-yl}-3-(1-methyl-1H-indol-3-yl)maleimide (Ro-32-0432) had no effect on A(2B) receptor-mediated IL-4 secretion but inhibited phorbol 12-myristate 13-acetate-stimulated IL-4 secretion. In contrast, chelation of intracellular Ca(2+) inhibited both A(2B) receptor- and ionomycin-dependent IL-4 secretion. This Ca(2+)-sensitive pathway probably includes calcineurin and nuclear factor of activated T cells, because A(2B) receptor-dependent IL-4 secretion was blocked with cyclosporin A or 11R-VIVIT peptide.

A heightened sympathetic tone accelerates the development of lethal arrhythmias after myocardial infarction (MI) and the progression of heart failure (HF). Cardiomyocytes control their local neural milieu by expression of nerve growth factor (NGF), which triggers sympathetic neural growth (sympathetic nerve sprouting: SNS). The molecular mechanisms that regulate NGF expression are largely unknown. During HF or MI the myocytes are exposed to increased mechanical load and adrenergic stimulation. Both stimuli induce myocyte hypertrophy. The angiotensin-II-calcineurin-NFAT (nuclear factor of activated t-cells) pathway is a well characterized signaling cascade in the pathogenesis of myocyte hypertrophy. The present study aims to investigate the molecular mechanisms by which mechanical stretch and/or alpha-1-adrenergic stimulation affect NGF expression in neonatal rat ventricular myocytes. Both stimuli resulted in a down-regulation of NGF gene and protein expression. Angiotensin-II type 1 receptor blockade with losartan blunted the stretch-induced NGF down-regulation. Specific calcineurin inhibition with cyclosporine A and FK506 or NFAT inhibition with 11R-VIVIT reversed the stretch or alpha-1-adrenergic induced decrease of NGF.

Bone formation, which is inhibited by particulate wear debris, is a pathological factor that contributes to periprosthetic osteolysis. Although the nuclear factor of activated T cells c1 (NFATc1) is known to be involved in osteoblast differentiation, and its effect on osteoblasts in response to wear particles remains unclear. In this study, we investigated the role of NFATc1 in the regulation of osteoblastic differentiation of rat calvaria (RC) cells (a cell-culture model comprising many osteoprogenitors) that were challenged with titanium (Ti) particles. The results showed that the Ti particles inhibited osteoblastic differentiation and mineralization of RC cells. NFATc1 plays a critical role in the Ti-particle inhibition process of the osteoblastic differentiation in RC cells. Inactivation of NFATc1 by the 11R-VIVIT peptide potently enhanced osteoblast differentiation and mineralization inhibition by the Ti particles. The 11R-VIVIT peptide does not have a toxic effect on the RC cells. On the basis of these data, we conclude that inactivation of NFATc1 by the 11R-VIVIT peptide may provide a promising therapeutic target for the treatment of periprosthetic osteolysis by increasing bone formation.