Study Of Vascular Endothelial Cadherin On The Angiogenesis Of Non-small Cell Lung Cancer In Vitro

Objective:Vascular endothelial cadherin (VE-cadherin) is a class ofcell adhesion molecule,which specificly express in vascular endothelial cellsand is dispensable in the process of angiogenesis. The purpose of this studyis to investigate the role of VE-cadherin on the angiogenesis of non-smallcell lung cancer(NSCLC), the association with vascular endothelial growfactor(VEGF), and to provide a new target and experimental basis forNSCLC anti-angiogenic therapy.Methods:1.Expressions and significances of vascular endothelial-cadherin inNSCLC: Expressions of VE-cadherin were examined at protein level byimmunohistochemical staining in72cases of NSCLC tissues and theiradjacent non-neoplastic tissues;34of which were measured at mRNA levelby real-time quantitative polymerase chain reaction（PCR） and VEGFwas taken as positive reference.2.Effect of VE-cadherin downregulation on the proliferation,cycle andapoptosis of A549cells:Cells were divided into blank controlgroup,interference group and negative control group.VE-cadherin-siRNA was transiently transfected into A549cells,Real-time quantitative PCR andWestern blotting were used to detect the expression of VE-cadherin;the cellproliferation was detected by MTS assay;the cell cycle and apoptosis weredetected by flow cytometry（F CM）.3.Effect of VE-cadherin downregulation in A549cells, supernatant onthe proliferation,cycle and apoptosis of human umbilical vein endothelialcells(HUVECs):After72h transfected, A549cells,supernatants in eachgroup were collected,the expression of VE-cadherin in transientlytransfected A549cells, supernatant was detected by ELISA method;and theeffect of supernatant on the proliferation was detected by MTS assay;the cellcycle and apoptosis were detected by FCM, and matrigel tubule formation ofHUVECs were analyzed.Results:1.Positive rates of VE-cadherin protein in cancer cells and vascularendothelial cells were69.4%and52.8%in NSCLC tissues, which werehigher than those in alveolar epithelial cells (0%) and vascular endothelialcells (20.8%) in adjacent non-neoplastic tissues (P<0.05);Positive rates ofVE-cadherin were higher in cases with lymph node metastasis than in caseswithout lymph node metastasis (P<0.05);Positive rates of VE-cadherin werehigher at Ⅲ-Ⅳ stage than atⅠ-Ⅱstage (P<0.05).2.Positive rates of VEGF protein in cancer cells and vascularendothelial cells were84.7%and62.5%in NSCLC tissues,which were higher than those in alveolar epithelial cells (16.7%) and vascularendothelial cells (29.2%) in adjacent non-neoplastic tissues(P<0.05).3.Real-time quantitative PCR results demonstrated that relativeexpressions of VE-cadherin mRNA in NSCLC tissues and adjacentnon-neoplastic tissues were1.15±0.87and0.70±0.57; relative expression ofVEGF mRNA were1.39±1.17and0.79±0.65(P<0.05).4.VEGF and VE-cadherin were positive correlated at both protein andmRNA levels (P=0.001, P=0.008).5.The expression of VE-cadherin mRNA in interference group(0.299±0.039) was significantly lower than blank controlgroup(1.137±0.082) and negative control group (1.001±0.076)(P<0.05);theexpression of VE-cadherin protein in interference group(0.297±0.045) wassignificantly lower than control group (0.833±0.119) and negative controlgroup (0.794±0.095)(P <0.05).6.The proliferation,cell cycle and apoptosis of A549cells in blankcontrol group, interference group and negative control group had nosignificant differences.7.The expression of VE-cadherin in the supernatant of the interferencegroup was （2.89±0.07） ng/mL,which was significantly lower than blankcontrol group (11.43±0.09)ng/mLand negative control group(11.15±0.04)ng/mL(P<0.05),and there was no significant differencebetween the latter two groups. 8.Between the interference group supernatant and the negative groupsupernatant,the cycle of HUVECs had no significant difference.9.The proliferation of HUVECs in the interference group supernatantwas inhibited significantly,the inhibition rates of24h,48h and72h were29.2%,33.8%and30.1%,respectively.10.After48h,the apoptosis rate of HUVECs in the interference groupsupernatant was (27.12±5.22)%,which was significantly higher thannegative control group (13.43±3.50)%(P <0.05).11.The tube formation of HUVECs in the interference groupsupernatant was1.53±0.31,which was significantly lower than negativecontrol group（14.53±1.51）(P <0.05).Conclusions:1.VE-cadherin and VEGF are highly expressed in NSCLC tissuescancer cells and vascular endothelial cells, and their expression arepositively correlated, suggesting that VE-cadherin may play an importantrole in angiogenesis in NSCLC, and may have a synergistic effect withVEGF.2.VE-cadherin-siRNA can effectively inhibit the expression ofVE-cadherin in lung cancer cell line A549, provide a new target and methodfor anti-angiogenic therapy in NSCLC.3.VE-cadherin-siRNA transiently transfected into A549cells can notaffect its biological behavior, suggesting that VE-cadherin may not direct effect the cancer cells, but through other mechanisms involved in NSCLC.4.VE-cadherin secreted by A549cells have the effect of promoteproliferation and anti-apoptoticon to HUVECs,but have no effect on its cycle,indicating that VE-cadherin may have had an impact on the biologicalbehavior of HUVECs.5.VE-cadherin secreted by A549cells can promote the tube formationof HUVECs, indicating that VE-cadherin involved in the occurrence anddevelopment of NSCLC by promoting angiogenesis.