Effect of extracellular signal-regulated kinase pathway on nitric oxide release by human umbilical vein endothelial cell induced by placental growth factor-1 / 中华妇产科杂志

ABSTRACT

Objective To investigate the effect of extracellular signal-regulated kinase(ERK)pathway OH nitriC oxide(NO)release by human umbilical vein endothelial cell(HUVEC)induced by placental growth factor-1(PLGF-1).Methods During January to April 2006,50 samples of umbilical vein blood were collected from newborns delivered by cesarean section due to intrauterine distress and abnormal fetal position.HUVEC were primarily cultured by trypsin digestion.Then the following procedures were performed(1)Cells were identified using the morphology andⅧfactor immunohistochemistry methods if the culture WaS satisfactory.(2)Cells were collected,and fms.1ike tyrosin kinase(Fit-1)protein and its mRNA expression were detected with immunoprints and RT-PCR methods.(3)The protein wag extractedafter cells were treated with PLGF-1(cells were collected before the treatment and 2.5,5.10,20 min after the treatment).The protein levels of ERK were determined by immunoprints.(4)The cells were cultured witll serum-free culture medim containing PLGF-1 only(culture media were collected 20,40,160,360.480.720 and 1440 rain after the treatment).The quantity of NO was detected with nitrate reductase metllod(5)The ceHs were cultured with serum.free culture medium containing PI)98059,the inhibitor of mitogen-activated protein kinase(MAPK)/MEK for 60 min Then the cells were cultured continuously with the serum-free culture medium containing PLGF-1 for 60 mira The culture media were coilected.The quantity of NO was detected by nitrate reductase method.The samples were divided into treatment group and control group.Control group was exactly the satne in treatment time,culture condition,and time to colleet the cells as the treatment group.except that it WaS not treated with PLGF-l or PD 98059.Resuits (1)By morphology and Ⅷ factor inununohistochemistry the cultured cells were identified to be HUVEC.(2)Fit-1mRNA and protein were expressed in HUVEC.(3)Expression of ERK protein started to increaSe at 2.5 min after treatment of HUVEC with PLGF-1,reaching the peak at 5 min,and decreased at 10 min.(4)Incomparison with the control group.NO started to increase at 20 rain after treatment of HUVEC with PLGF-lat 480 min(15.82±0.69)μmol/L Comparison between the two groups showed a significant difference (P<0.05).(5)ReleaSe of NO from the cells treated with PD98059 for 1 hour and PLGF WaS significantly inhibited,compared with the ceils treated with PLGF-1 only.Conclusion ERK pathways play an important role in N0 release bv HUVEC induced by PLGF.