Berlanga-Acosta J.,Center for Genetic Engineering and Biotechnology |
Vazquez-Blomquist D.,Center for Genetic Engineering and Biotechnology |
Cibrin D.,Center for Genetic Engineering and Biotechnology |
Mendoza Y.,Center for Genetic Engineering and Biotechnology |
And 22 more authors.
Biotecnologia Aplicada | Year: 2012
Tissue fibrosis is a leading cause of morbidity and mortality. Current treatments for conditions such as hepatic fibrosis have been unsuccessful. The growth hormone relasing peptide 6 (GHRP6) is endowed with cardioprotective actions but its antifibrotic effect had not been anticipated. We examined the GHRP6 ability to prevent and revert liver cirrhosis after induction in Wistar rats by a subcutaneous administration of CCl4. GHRP6 effects were examined after concomitant and delayed administration to toxic respectively. The percentages of hepatic fat, fibrosis, nodularity and septae thickness were histologically and morphometrically determined. Ascitis and portal dilation were judged by ultrasound and serum biochemical profile and oxidative stress parameters determined. Mechanistic involvement of selective gene/proteins was assessed by RT-PCR and immunohistochemistry. Microarrays showed gene expression profiles of GHRP6-treated liver samples on CapitalBio Rat Genome Oligo Array. GHRP6 concomitant intervention prevented in more than 85% parenchymal fibrotic induration (p < 0.0001) and therapeutic administration for only 15 days allowed for 37% fibrotic clearance (p = 0.0004) with more than 30% reduction of septae thickness (p = 0.0011). The 60 days GHRP6 administration scheme produced a 75% reduction of the fibrotic area with more than 60% reduction of nodularity. GHRP6 reduced oxidative damage enhancing the activity of antioxidant enzymes. Vimentin and alpha smooth muscle actin immunodetection profile indicated GHRP6 reduced the number of activated stellate cells. GHRP6 administration reduced fibrogenic factors as TGF-β and CTGF on Kupffer cells. Differentially expressed genes in the microarray experiment indicated GHRP6 modulate the redox balance and parenchymal cells response to injury. These evidences suggest GHRP6 may control the liver's fibroplastic response.
Mendoza-Mari Y.,Direction of Biomedical Research |
Garcia-Ojalvo A.,Direction of Biomedical Research |
Fernandez-Mayola M.,Direction of Biomedical Research |
Herrera-Martinez L.,Center for Genetic Engineering and Biotechnology |
Berlanga-Acosta J.,Direction of Biomedical Research
Biotecnologia Aplicada | Year: 2014
Diabetic healing failure is the clinical expression of countless molecular and cellular disorders having hyperglycemia as the proximal trigger. The fibroblast is a critical building-block cell type for the healing process. Under high glucose concentrations fibroblasts physiology is perturbed. The epidermal growth factor receptor (EGFR) signaling system is crucial for different healing events. We examined the effect of high glucose burden on healthy-donor' cutaneous fibroblasts proliferation, so as the EGFR autophosphorylation on a critical tyrosine residue, along with the activation of downstream signaling pathways proximal to cyclin D1 expression. Fibroblasts were cultured in 15 % FBS at either 5 (normal glucose) or 35 (high glucose) mM under standard culture conditions. Concurrent osmotic control was included. After 6 days of incubation under high glucose, doubling time was calculated. Cells suspensions were plated, fixed and immunolabelled with antibodies directed to phosphorylated forms of EGFR (Y1197), AKT1 (S473) and mTOR (S2448), and native forms of PI3K p85 alpha subunit and cyclin D1. The ratio of cells positive to the diaminobenzidine/peroxidase reaction was calculated and its intensity estimated according to published methodologies. High glucose concentration significantly increased doubling time 5-fold, as compared to cells grown in physiological conditions. Hyperglycemia reduced the constitutive EGFR autophosphorylation. Accordingly, PI3K was also significantly attenuated. Downstream switches AKT1 and mTOR were also affected and very significantly on the signal intensity. Cyclin D1 expression was completely abrogated due to high glucose burden. Collectively, these data suggest that high glucose exposure hinders fibroblasts proliferation by disrupting the EGFR/PI3K/AKT1/mTOR/Cyclin D1 axis.