同种和异种血管内皮细胞的培养及其在抗肿瘤血管治疗中的作用  

CULTURE OF AUTOLOGOUS AND XENOGENEIC ENDOTHELIAL CELLS AND DISCUSSION OF ITS ROLE IN ANTI-ANGIOGENIC THERAPY OF CANCER

作　　者：赵军[1,2] 李珊[1] 路静[1] 杨洪艳[1] 赵继敏[1] 翟景明[1] 张曦[3] 董子明[1] 

机构地区：[1]郑州大学基础医学院病理生理学教研室,郑州450052 [2]长治医学院附属和济医院呼吸科 [3]郑州大学第三附属医院妇产科

出　　处：《现代预防医学》2009年第5期922-925,941,共5页Modern Preventive Medicine

基　　金：河南省卫生厅科技攻关项目(20060011);河南省教育厅自然科学研究计划项目(2008A3310018)

摘　　要：[目的]建立简单高效的同种和异种血管内皮细胞的培养以及探讨其在抗肿瘤血管治疗中的作用。[方法]1周龄内BALB/C小鼠的肺组织运用组织块法得到原代新生小鼠肺微血管内皮细胞(Pulmonary Microvascular Endothelial Cells,PMVECs),作为同种抗原;人新鲜脐静脉内灌注0.25%胰蛋白酶消化获得原代人脐静脉内皮细胞(Human Umbilical Vein Endothelial Cell,HUVECs),作为异种抗原,均用内皮细胞培养基(Endothelial Cell Medium,ECM)进行培养。通过倒置显微镜,免疫组织化学、RT-PCR鉴定,并用液氮冻融、-80℃冻融、煮沸、超声等方法比较制备抗原。[结果]成功原代培养出新生小鼠PMVECs和人HUVECs,vWF(Von Willebrand factor)、CD144(VE-Cadherin)和CD31(PECAM21)免疫组化表达阳性,RT-PCR检测vWF和CD144有特异性条带产生,通过几种方法比较得出液氮冻融制备抗原方法简单,浓度高。[结论]新生小鼠PMVECs和人HUVECs的原代培养方法简便、可靠,高效,保持了血管内皮细胞的结构和功能,且运用液氮冻融法获得纯度较高的抗原,为进一步研究同种PMVECs和异种HUVECs在抗肿瘤血管治疗中奠定了基础。[Objective] To construct a simple and effective culture method of autologous and xenogeneic endothelial cells and discuss its role in anti-angiogenic therapy of cancer. [Methods] The lung tissue blocks of neogenetic BALB / C mice were employed for PMVECs cultivation as autologous antigen. 0.25% Trypsin perfusion method was used to isolate human HUVECs as xenogeneic antigen. Two cells were cultured with endothelial cell medium （ECM）. The cell morphology was observed using inverted microscope and inverted phase contrast microscope. Immunohistochemical methods and RT-PCR were employed to identify the two cells. Then, the cultured mice PMVECs and human HUVECs were prepared to antigen through four rapid freeze （liquid nitrogen） -thaw （37℃ ） cycles, freeze （-80℃ ） -thaw （37℃ ） cycles, boiled and ultrasonic disruption methods. [Results] Mice PMVECs and human HUVECs were cultured successfully in vitro. Expression of vWF, CD144 and CD31 in immunohistochemical methods were positive. There were special straps of vWF and CD144 in RT-PCR. The antigen preparation through freeze and thaw method in liquid nitrogen was better than the other three methods. [ Conclusion] The pure mice PMVECs and human HUVECs can be attained rapidly and easily by the described methods above. Preparation of the antigen is simple too. These methods will be a strong foundation for anti-angiogenic therapy in cancer.

关 键 词：同种和异种血管内皮细胞 细胞培养 抗原制备 抗肿瘤血管治疗 

分 类 号：R45[医药卫生—治疗学]