Effect Of Bushen Antai Granule On Expression Of VEGF/Ras/MAPK Pathway In Placental Microvascular Endothelial Cells

Objective:To observe the effect of Bushen Antai granules on the VEGF 、 VEGFR2 and downstreamRas/MAPK signal transduction pathway in placental microvascular endothelial cells under the condition of VEGFR2 inhibition.The mechanism of Bushen Antai granules in preventing and treating recurrent abortion was discussed from the angle of angiogenesis regulation.Methods:20 rats of grade SPF were randomly divided into serum group and blank group,The drug-containing serum group should be given Bushen Antai Granules,The blank group was replaced with the same amount of saline,For 7 days,Twice a day,After 1 hour of last administration,After centrifugation,it was used to prepare the serum containing drug and blank group,A MTT method was used to screen out the best serum concentration of Bushen Antai granules for subsequent experiments,In vitro culture of placental microvascular endothelial cells,And transfected VEGFR2 siRNA into placental microvascular endothelial cells,After the VEGFR2 is in the inhibition state,the follow-up experiment is carried out.The experimental groups were: normal serum group,siRNA-NC normal serum group,drug serum group,sirna normal serum group,The corresponding intervention was given respectively.Real-time fluorescence quantitative PCR、western blotting、 immunofluorescence were used to detect VEGF、VEGFR2、Ras、MAPK mRNA and protein expression.Results:1.The optimum serum concentration of Bushen Antai granule was 20,the concentration continued to increase,and the effect was no longer enhanced.2.Real-time fluorescence quantitative PCR showed no significant difference in VEGF、VEGFR2、Ras、MAPK mRNA expression between normal serum group and siRNA-NC normal serum group(p>0.05);The expression of cell VEGF、VEGFR2、Ras、MAPK mRNA in drug serum group was significantly higher than that in normal serum group(p<0.05);The expression of VEGF、VEGFR2、Ras、MAPK mRNA in siRNA1 normal serum group was significantly lower than that in normal serum group(p<0.01),and that in siRNA1 drug serum group was significantly different from that in normal serum group(p<0.01).3.Western blot results showed that there was no significant difference in VEGF 、VEGFR2、Ras、MAPK、p-MAPK protein between normal serum group and siRNA-NC normal serum group(p>0.05);The expression of VEGF、VEGFR2、Ras、MAPK、p-MAPK protein in drug serum group was significantly higher than that in normal serum group(p<0.05 or 0.01);The expression of VEGF、VEGFR2、Ras、MAPK、p-MAPK protein in siRNA1 normal serum group was significantly decreased(p<0.01)and that in siRNA1 drug serum group was significantly different from that in normal serum group(p<0.01).4.The results showed that there was no significant difference in VEGF、VEGFR2、Ras、MAPK protein between normal serum group and siRNA-NC normal serum group(p>0.05);The expression of VEGF、VEGFR2、Ras、MAPK protein in drug serum group was significantly higher than that in normal serum group(p<0.05);The expression of VEGF、VEGFR2、Ras、MAPK protein in siRNA1 normal serum group decreased significantly(p<0.01),and there was significant difference between siRNA1 drug serum group and normal serum group(p<0.01).Conclusion:Bushen Antai granules may regulate VEGF/Ras/MAPK signaling pathway by upregulating the expression of VEGF、VEGFR2、Ras、MAPK mRNA and proteins,thus improving angiogenesis at the maternal and fetal interface and treating recurrent abortion.