Phenotypic transformation of smooth muscle cells from porcine coronary arteries is associated with connexin 43

The current study aimed to investigate the relevance of the gap junction protein connexin Cx43 in coronary artery smooth muscle cell (SMC) heterogeneity and coronary artery restenosis. SMCs were isolated from the coronary artery of 3-month-old pigs using enzymatic digestion. Two distinct SMC populations were isolated: Rhomboid (R) and spindle-shaped (S) cells. S-SMCs exhibited relatively lower rates of proliferation, exhibiting a classic ''hills-and valleys'' growth pattern; R-SMCs displayed increased proliferation rates, growing as mono- or multi-layers. Immunofluorescent staining, polymerase chain reaction and western blotting were used to assess the expression of Cx40 and Cx43 in SMCs. For further evaluation, cultured SMCs were treated with 10 ng/ml platelet-derived growth factor (PDGF)-BB with or without the gap junction blocker 18α-glycyrrhetinic acid. Stent-induced restenosis was assessed in vivo. Different expression patterns were observed for Cx40 and Cx43 in R- and S-SMCs. Cx40 was the most abundant Cx in S-SMCs, whereas CX43 was identified at relatively higher levels than Cx40 in R-SMCs. Notably, PDGF-BB converted S-SMCs to R-SMCs, with increased Cx43 expression, while 18α-glycyrrhetinic acid inhibited the PDGF-BB-induced phenotypic alterations in S-SMCs. Additionally, restenosis was confirmed in pigs 1-month subsequent to stent placement. R-SMCs were the major cell population isolated from stent-induced restenosis artery tissues, and exhibited markedly increased Cx43 expression, in accordance with the in vitro data described above. In conclusion, the phenotypic transformation of coronary artery SMCs is closely associated with Cx43, which is involved in restenosis. These observations provide a basis for the use of Cx43 as a novel target in restenosis prevention.