Pyrrolidine dithiocarbamate inhibits the production of interleukin-6, interleukin-8, and granulocyte-macrophage colony-stimulating factor by human endothelial cells in response to inflammatory mediators: modulation of NF-kappa B and AP-1 transcription factors activity

Abstract

Endothelial cells (EC) play a key role in the inflammatory response, both by the production of proinflammatory cytokines and by their interaction with leukocytes. Molecular genetic analysis has demonstrated that functional NF-kappa B sites are involved in the transcription of interleukin-6 (IL-6), IL-8, and granulocyte-macrophage colony-stimulating factor (GM-CSF) genes in response to inflammatory mediators. Thus, we have explored the effect of two inhibitors of the NF-kappa B activation, pyrrolidine dithiocarbamate (PDTC) and N-acetylcysteine (NAC), on the production of these cytokines by EC. Both PDTC and NAC inhibited, in a dose-dependent manner, the synthesis of IL-6, IL-8, and GM-CSF induced by tumor necrosis factor (TNF)-alpha or bacterial lipopolysaccharides (LPS) in human umbilical vein endothelial cells (HUVEC). PDTC appeared to prevent IL-6, IL-8, and GM-CSF gene transcription, as it blocked the induction of specific mRNA by TNF-alpha or LPS. The TNF-alpha mediated transcriptional activation of a chloramphenicol acetyltransferase (CAT) plasmid containing three copies of the -72 kappa B binding site from the IL-6 promoter was abrogated by PDTC. According to transfection experiments, electrophoretic mobility shift assays (EMSA) demonstrated that the antioxidant prevented the induction of NF-kappa B DNA-binding activity by TNF-alpha. Under the same conditions, PDTC by itself or in combination with TNF-alpha, enhanced the DNA-binding activity of AP-1, as well as c-fos and c-jun mRNA levels. Altogether, these results indicate that the antioxidant PDTC specifically inhibits the transcription of IL-6, IL-8, and GM-CSF genes through the inhibition of the NF-kappa B activation, while increasing the expression of AP-1. Our data make evident the antiinflammatory and immunoregulatory potential of the pharmacological inhibition of the NF-kappa B activation. In addition, PDTC and related molecules may be a useful tool to explore the expression of genes involved in the inflammatory response.

Pyrrolidine dithiocarbamate protects mice from lethal shock induced by LPS or TNF-α

“…We found that PDTC strongly inhibits the in vivo activation of NF-O B in lung and liver of LPS-challenged mice. A number of antioxidants, including PDTC, efficiently inhibit NF-O B activation induced by TNF- § or LPS in a number of cell systems [13,20,23,24]. These effects appear to be mediated by the powerful radical scavenger properties of this antioxidant, which apparently counteracts ROI signaling generated by NF-O B activators [18,23,25].…”

“…Despite these considerations, mice protected from LPS-or TNF- § -induced septic shock remain alive for months after LPS challenge, and no differences were observed with respect to untreated control mice during this time. In addition, PDTC activates the transcription factors AP-1 and C/EBP g [18,24,43,44]. Whereas the PDTCmediated activation of C/EBP g appears to account for important beneficial effects in colon cancer therapy, enhancing the cytotoxic efficacy of the conventional anti-tumoral drugs such as 5-fluouracil and doxorubicin [43], the effects of PDTC activating AP-1 in a number of cells types have not been evaluated in vivo.…”

p120‐Catenin modulating nuclear factor‐κB activation is partially RhoA/ROCKdependent in scratch injury

“…As an important pro‐inflammatory chemokine, IL‐8 is one of the well‐established target genes for NF‐κB and synthesized by many different cell types including bronchial epithelium after stimulated by bacterial, viral products or chemicals . To further confirm scratch injury can induce inflammatory responses in BECs, IL‐8 was examined by ELISA and real‐time PCR.…”

“…On cellular stimulation, IκBα is rapidly ubiquitinated and degraded through the 26S proteasome pathway and exposed the nuclear localization sequence of NF‐κB, which allows NF‐κB entering the nucleus . Activation of NF‐κB could enhance the expression of genes encoding inflammatory cytokines, such as IL‐8 . It is the core of inflammatory reactions, closely related to cellular stimulation such as LPS and scratch‐induced inflammation .…”

“…It is the core of inflammatory reactions, closely related to cellular stimulation such as LPS and scratch‐induced inflammation . It is well established that NF‐κB involves in the regulation of airway inflammatory processes after stimulation and inflammatory responses could be reduced by blocking NF‐κB signaling pathway …”