RNA interference of phosphatidylinositol 3-kinase signaling pathway by kinase insert domain-containing receptor promoter/enhancer influences the proliferation and apoptosis of rat vascular endothelial cells

Objective To construct the kinase insert domain-containing receptor (KDR) specific promoter/enhancer eukaryon expression vector for RNA interference of phosphatidylinositol 3-kinase,catalytic,beta polypeptide (Pik3cb) gene in rat vascular endothelial cells (VECs) and evaluate the effects on the proliferation and apoptosis of VECs.Methods Both the specific KDR promoter/enhancer eukaryon expression vector and non-specific cytomegalovirus (CMV) eukaryon expression vector were constructed and transfected into VECs,respectively.The samples were divided into 5 groups:group A,normal VECs with-out any treatment; group B,VECs transfected with 2.0 mg/L KDR plasmid; group C,VECs transfected with 2.0 mg/L CMV plasmid ; group D,VECs transfected with 2.0 mg/L empty plasmid; group E:VECs treated with 50 nmol/L wortmannin.At 24,48,and 72 h after treatments,the expression level of Pik3cb mRNA was detected by using real-time quantitative reverse transcription-polymerase chain reaction ( RT-qPCR),and proliferation and apoptosis of VECs were analyzed by cell counting Kit-8 ( CCK-8 ) and flow cytometry,respectively.Results At 24,48 and 72 h after treatments,the real-time RT-PCR revealed that the relative expression of Pik3cb mRNA in group B was (54.82 ± 2.77 ) ％,( 50.54 ± 3.98 ) ％ and (35.47 ± 4.83 ) ％ respectively,significantly lower than in groups A and D ( P ＜ 0.05 ) ; The CCK-8 analysis showed the inhibition rate of VECs proliferation in group B was ( 21.98 ± 2.25 ) ％, ( 24.32 ±3.04) ％ and ( 26.38 ± 5.06 ) ％ respectively,significantly higher than in groups A and D ( P ＜ 0.05 ) ;the results of flow cytometry indicated that the apoptosis rate in group B was (9.9 ± 1.3 ) ％,(31.0 ±7.4 ) ％ and ( 44.5 ± 8.3 ) ％ respectively,significantly higher than in group A ( P ＜ 0.05 ).Conclusion The inhibition of P13K signaling pathway by KDR specific promoter/enhancer eukaryon expression vector in rat VECs down-regulates the expression of Pik3cb mRNA,inhibits the proliferation and promotes the apoptosis of VECs specifically. Key words: RNA interference; Vascular endothelial cells; Phosphatidylinositol 3-kinase