Role of phosphatidyl inositol 3-kinase pathway in regulating dedifferentiation and proliferation of rat pulmonary arterial vascular smooth muscle cells

Abstract

Objective To investigate the role of phosphatidyl inositol 3-kinase(PI3K) in rat pulmonary arte-rial vascular smooth muscle cell(PASMC) dedifferentiation and proliferation. Methods PASMC were isolated from Sprague-Dawley(SD) rat arteriae pulmonalis and subcultured to the third generation for experiment.PASMC were divided into blank group, platelet derived growth factor(PDGF) group, dimethylsul foxide(DMSO) group, LY294002 group, small interfening RNA(siRNA) group, control-siRNA group, empty-virus group, recombinant-adenovirus group.Real-time fluorescence quantitative polymerase chain reaction(RT-qPCR) was performed to detect the PIK3CA mRNA level in PASMC transfected with pAdxsi-GFP-PIK3CA recombinant adenovirus and PIK3CA-siRNA.Western blot analysis was used to measure the expression level of alpha smooth muscle actin(α-SM actin). The mRNA level of α-SM actin was detected by RT-qPCR.The methyl thiazolyl tetrazolium(MTT) assay and thiazole cell counting kit-8(CCK-8) method were used to assess proliferation activity of PASMC. Results The PIK3CA mRNA level was significantly increased in recombinant-adenovirus group compared with blank group(P<0.05). The mRNA level of PIK3CA in siRNA group was lower than that in blank group(P<0.05). Compared with blank group, significantly lower α-SM actin mRNA and protein levels and a significantly higher proliferation activity were detected in PDGF group(all P<0.05). PI3K inhibitor LY294002 blocked this down-regulation of α-SM actin gene expression and up-regulation of proliferation activity induced by PDGF-BB(P<0.05). PIK3CA-siRNA also attenuated these biological effects.The α-SM actin mRNA level and protein expression level in recombinant-adenovirus group were lower than those in blank group(P<0.05), while the proliferation activity in the former group was greater(P<0.05). Conclusions PI3K pathway plays a vital role in vascular remodeling through a positive regulation of dedifferentiation and proliferation activity in PASMC. Key words: Phosphatidyl inositol 3-kinase; Pulmonary arterial vascular smooth muscle cell; Dedifferentiation; Proliferation