cyclic-gmp and Keloid

Keloids are characterized by excessive proliferation of fibroblasts and invasion of surrounding healthy skin. High levels of Nitric Oxide (NO) are thought to be the crucial factor within the micro-environment in promoting keloid formation. However, the effects and mechanisms of NO on the proliferation of Keloid Fibroblasts (KDFs) remain unclear. In this study, we investigated the effect of NO on KDFs proliferation by Sodium Nitroprusside (SNP), an NO donor. Our results show that SNP significantly enhanced KDFs proliferation. Moreover, with prolonged treatment with SNP after cell confluence, the growth of KDFs escape contact inhibition and experience significant pile up growth. Furthermore, PTIO, an NO scavenger, attenuated SNP-enhanced cell proliferation effectively. The mechanism involved in SNP-induced KDFs proliferation was soluble Guanylyl Cyclase (sGC) and cGMP independent. ODQ, a specific sGC inhibitor, failed to suppress SNP-enhanced KDFs proliferation. 8-Bromo-c GMP, a cell-permeable cGMP analogue, could not stimulate KDFs proliferation. Erk and Akt provide important signaling for cell growth. U0126 and LY294002, inhibitors of Erk and Akt respectively, block SNP-enhanced KDFs proliferation effectively. As expected, a Western blot showed that SNP up-regulated the phosphorylation levels of Erk and Akt. Moreover, it decreased the expression of p

Topics: Cell Proliferation; Cyclic GMP; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Humans; Keloid; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; Proto-Oncogene Proteins c-akt; Signal Transduction

Keloids arise from the aberrant wound healing process and nitric oxide (NO) plays an important role in the inflammation stage of wound healing. In order to better define the potential effect of NO/cGMP signal pathway in the keloid pathogenesis, the enhancing effect of exogenous NO (released from NO donor) on collagen expression in the keloid fibroblast (KF) as well as on the induction of collagen type I protein and TGF-beta1 expression in the KF were studied in this investigation. The DETA NONOate, an NO donor, was added to the KF, as the exogenous NO, to release NO in the culture medium. The expression of collagens was then determined by assaying the total soluble collagens and collagen type I in the KF. The cellular concentration of cGMP was measured by EIA in the KF. Exogenous NO was found to enhance the expression of collagens and elevate the cellular levels of cGMP. Moreover, to evaluate the effect of the elevated cellular cGMP levels on the expression of collagen and TGF-beta1, both cGMP and TGF-beta1 were measured by ELISA. The inhibitors for phosphodiesterase (PDE), such as IBMX (3-isobutyl-1-methylxanthine), Vinpocetine, EHNA, Milrinone and Zapriast, which have been reported to reduce the ability of PDE and subsequently produce an increase of cellular cGMP, induce the production of autocrine TGF-beta1 as well as the synthesis of collagen in the KF. In this investigation, the inhibition of the PDE enzyme activity was observed to enhance the effect on the collagen synthesis, and was induced by exogenous NO. Taken together, these results have suggested that the NO/cGMP pathway could positively influence the progression of keloid formation, via the TGF-beta1 expression in the KF.

Topics: 1-Methyl-3-isobutylxanthine; Adult; Aged; Blotting, Western; Cells, Cultured; Collagen Type I; Cyclic GMP; Female; Fibroblasts; Humans; Keloid; Male; Middle Aged; Nitric Oxide; Transforming Growth Factor beta

The excessive accumulation of extracellular matrix is a hallmark of many fibrotic diseases, including the hypertrophic scar and keloid. Recent reports from this research team had shown that exogenous nitric oxide (NO) participates in the keloid formation; however, its role on the synthesis of fibrotic factor (TGF-beta1, TIMP-1 and HSP47) in the keloid fibroblasts (KF) remained unclear.. In this study, to better define the potential effect of exogenous NO on the expression of fibrotic factors in KF, the enhancing effect of exogenous NO, released from a NO donor, on the synthesis of fibrotic factors in KF was investigated.. The seven primary KF cultures were set up to measure the effect of exogenous NO on enhancing the expression of fibrotic factor.. Elevation of cellular cGMP levels was observed to be induced by NO or blocked by the hydrolysis activity of phosphodiesterase (PDE) by the PDE inhibitor. The elevated levels of cellular cGMP were noted to enhance the expression of TIMP-1 and HSP47 in KF. Exogenous NO was found to significantly accelerate the production of TIMP-1 and HSP47 in the seven primary KFs with a corresponding increase in the production of TGF-beta1.. The results have led to a conclusion, that is: the excess collagen formations in the keloid lesion may be attributed to the NO/cGMP signal pathway by initiating a rapid increase in the expression of TGF-beta1, TIMP-1 and HSP47 in the KF cells.

Topics: Adult; Aged; Cells, Cultured; Cyclic GMP; Dose-Response Relationship, Drug; Female; Fibroblasts; HSP47 Heat-Shock Proteins; Humans; Keloid; Male; Middle Aged; Nitric Oxide; Nitric Oxide Donors; Nitroso Compounds; Recombinant Proteins; Signal Transduction; Smad2 Protein; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta1