Connective tissue growth factor dependent collagen gene expression induced by MAS agonist AR234960 in human cardiac fibroblasts
Fig 2
Agonist activated MAS receptor induces CTGF through ERK1/2 and regulates collagen expression in HEK293 cells stably expressing MAS.
(A) Real-time PCR analysis shows significant upregulation of CTGF expression in response to MAS receptor agonist (AR234960; 10μM); while MAS inverse-agonist (AR244555; 10μM) along with agonist suppresses the expression of CTGF below the basal level. (B) Western-blot showing significant upregulation of CTGF in MAS agonist (AR234960) activated samples whereas CTGF expression decreases in presence of inverse-agonist (AR244555). MAS activated by AR234960 induces phosphorylation of ERK1/2, MAS inhibition by the inverse-agonist (AR244555) reduces ERK1/2 activation. MAS expressing HEK293 cells also show significant down-regulation of CTGF in presence of MEK1 inhibitor (PD98059). CTGF and p-ERK1/2 expression were normalized by GAPDH and ERK1/2 respectively. The western blot image shown is a representative of all the experiments done under similar experimental conditions and data from multiple experiments quantitated and cumulative data were presented as bar graph, (*p<0.05; **p<0.01). (C) Bar graphs showing real-time PCR analysis of different collagen sub-types (Col1A1, Col1A2 and Col4A1) [represented as fold increase (2‒ΔΔCt)] in HEK293-MAS cell line. Activated MAS induces collagen synthesis while repression of MAS receptor by its inverse agonist (AR244555) shows significant down-regulation of the same collagen sub-types (*p<0.05; **p<0.01). RT-qPCR was normalized by GAPDH.