Thymulin and zinc (Zn2+)-mediated inhibition of endotoxin-induced production of proinflammatory cytokines and NF-κB nuclear translocation and activation in the alveolar epithelium: Unraveling the molecular immunomodulatory, anti-inflammatory effect of thymulin/Zn2+ in vitro

The immunomodulatory potential of thymulin and zinc (Zn2+) in the perinatal alveolar epithelium is not well characterized. In an in vitro model of fetal alveolar type 11 epithelial cells (FATEII), we have investigated the exhibition of an anti-inflammatory activity of this peptide hormone. Thymulin selectively ameliorated, in a dose-dependent manner, the endotoxin (ET/LPS [lipopolysaccharide])-induced release of IL-1 beta, but not IL-6 or TNF-alpha. Furthermore, Zn2+, an anti-inflammatory antioxidant, which is required for the biological activity of thymulin, independently reduced the secretion of IL-1 beta, TNF-alpha and, to a lesser extent, at a supraphysiologic dose (1 mM), IL-6. The underlying cellular and molecular pathways associated with the anti-inflammatory effect of thymulin and Zn2+ in the alveolar epithelium are not well established. Further in this study, the role of cyclic AMP (cAMP) in the anti-inflammatory effect of thymulin was investigated, in addition to unraveling the possible involvement of the NF-kappa B pathway. Interestingly, thymulin upregulated, in a dose- and time-dependent manner, the release of the nucleotide cAMP. To understand whether the inhibitory effect of thymulin on cytokine release is cAMP-dependent, Forskolin, a labdane diterpene known to elevate intracellular cAMP, was shown to reduce the secretion of IL-1 beta and TNT-alpha, but not IL-6, an effect mimicked by dibutyryl-cAMP (dbcAMP), an analog of cAMP. Alveolar epithelial cells treated with thymulin markedly showed a downregulation of the nuclear translocation of Re1A (p65), the major transactivating member of the NF-kappa B family, in addition to NF-kappa B-1 (p50) and c-Rel (p75), an effect mildly substantiated with Zn2+. Furthermore, thymulin/Zn2+ reduced, in a dose-dependent manner, the DNA-binding activity of NF-kappa B (Re1A/p65). These results indicate that the anti-inflammatory effect of thymulin, which is mediated by cAMP, is NF-kappa B-dependent and involves the downregulation of the release of proinflammatory cytokines, particularly IL-1 beta, an effect synergistically amplified, at least in part, by Zn2+. The molecular regulation of thymulin via a NF-kappa B-dependent pathway is critical to understanding the anti-inflammatory alleviating role of this nonapeptide in regulating proinflammatory signals. (C) 2009 Elsevier Ltd. All rights reserved.