CAJ | 학술논문

目的：利用RNA干扰技术下调胶质瘤细胞中癌胚抗原相关细胞黏附分子1（ CEACAM1）基因的表达情况，探讨胶质瘤细胞CEACAM1表达变化后其培养上清液诱导人脐静脉内皮细胞（ HUVEC）增殖、血管生成的情况及其可能机制。方法设计并化学合成针对CEACAM1的3对siRNA，脂质体法瞬时转染SHG44细胞，收集细胞培养上清液作为条件培养基。分别采用RT－PCR检测RNA干扰后细胞中CEACAM1及血管内皮生长因子（ VEGF） mRNA表达的变化情况，ELISA法检测上清液中VEGF蛋白的含量。 MTT比色法及Transwell侵袭实验检测共培养刺激后HUVEC增殖及迁移能力的变化，体外血管生成实验检测体外血管生成能力。结果与正常对照组和阴性对照组相比，转染CEACAM1－siRNA后胶质瘤细胞中CEACAM1 mRNA的表达水平下调（P＜0．01），以CEACAM1－siRNA3组最为显著。 RNA干扰后SHG44细胞VEGF mRNA及上清液中的VEGF蛋白含量均减少，HUVEC的增殖受到抑制，迁移及体外血管生成能力下降。结论 CEACAM1－siRNA能够下调SHG44细胞中CEACAM1 mRNA的表达，并通过减少VEGF分泌，从而影响HUVEC的增殖、迁移和体外血管生成能力。
목적：이용RNA간우기술하조효질류세포중암배항원상관세포점부분자1（ CEACAM1）기인적표체정황，탐토효질류세포CEACAM1표체변화후기배양상청액유도인제정맥내피세포（ HUVEC）증식、혈관생성적정황급기가능궤제。방법설계병화학합성침대CEACAM1적3대siRNA，지질체법순시전염SHG44세포，수집세포배양상청액작위조건배양기。분별채용RT－PCR검측RNA간우후세포중CEACAM1급혈관내피생장인자（ VEGF） mRNA표체적변화정황，ELISA법검측상청액중VEGF단백적함량。 MTT비색법급Transwell침습실험검측공배양자격후HUVEC증식급천이능력적변화，체외혈관생성실험검측체외혈관생성능력。결과여정상대조조화음성대조조상비，전염CEACAM1－siRNA후효질류세포중CEACAM1 mRNA적표체수평하조（P＜0．01），이CEACAM1－siRNA3조최위현저。 RNA간우후SHG44세포VEGF mRNA급상청액중적VEGF단백함량균감소，HUVEC적증식수도억제，천이급체외혈관생성능력하강。결론 CEACAM1－siRNA능구하조SHG44세포중CEACAM1 mRNA적표체，병통과감소VEGF분비，종이영향HUVEC적증식、천이화체외혈관생성능력。
Objective To investigate the effect of supernatant from carcinoembryonic antigen -related cell adhesion molecule 1(CEACAM1)gene RNAi glioma SHG44 cells on human umbilical vein endothelial cell proliferation and angiogenesis in vitro .Methods Three pairs of specific siRNA targeting CEACAM 1 were de-signed and synthesized , and then transiently transfected into SHG 44 cells via cathodolyte liposome transfection method.The supernatant from cultured glioma SHG 44 cells was collected as the conditional medium 48 h after transfection.RT-PCR was used to detect CEACAM1 and VEGF expression at mRNA level after CEACAM1 gene RNAi.The expression of VEGF in supernatant was measured by ELISA .The proliferation of HUVEC was assessed by MTT method after co-cultured with supernatant .The migration of HUVEC was examined by using a Transwell assay.The ability of angiogenesis was measured by capillary -like network formation assay .Results Forty -eight hours after the 3 pairs of specific CEACAM1 siRNA were transfected into SHG44 cells,RT-PCR results showed that the expression of CEACAM 1 mRNA was significantly inhibited compared with those in the normal control and the negative control groups (P<0.01).The most significant interference effect was CEACAM 1-siR-NA3.Expression of VEGF mRNA was remarkably reduced ,and expression of VEGF protein in the supernatant was also reduced after transfection of CEACAM 1-siRNA for 48 h.The proliferation of HUVEC was inhibited , HUVEC migration and tube capillary -like formation were decreased .Conclusion CEACAM1-siRNA can ef-fectively suppress the expression of CEACAM 1 mRNA in human glioma SHG44 cells,may inhibit HUVEC prolif-eration,migration and tube capillary -like formation via down -regulated the secretion of VEGF in vitro .