hTERT转染对人海绵体平滑肌细胞的生物学行为-第三军医大学学报.DOC

hTERT 转染对人阴茎海绵体平滑肌细胞生物学行为的作用 吴小军 宋波 张恒 王永权 何鹏 周晓洲 李为兵 周占松 （第三军医大学西南医院全军泌尿外科研究所 重庆 400038） [摘要]目的：探讨hTERT基因转染对人海绵体平滑肌细胞生物学行为的作用。方法：原代培养人海绵体平滑肌细胞，构建hTERT基因荧光真核表达载体，采用脂质体法首次将hTERT-IRES2-EGFP正义重组质粒转染人海绵体平滑肌细，对转染正确性鉴定并测定转染后平滑肌细胞端粒酶活性、细胞分裂指数、细胞凋亡、细胞生长曲线及转染后细胞是否存在恶性表型。结果：成功构建人正义hTERT基因荧光真核表达载体并正确转染人海绵体平滑肌细胞，转染后细胞端粒酶活性、细胞分裂指数、细胞生长曲线显著高于未转染细胞及EGFP空载转染细胞，而细胞凋亡发生率低。hTERT转染后细胞染色体仍为23对,且均为正常二倍体细胞，裸鼠皮下不具有成瘤的能力。结论：首次将正义hTERT基因转染人海绵体平滑肌细胞，其有助于增加细胞内端粒酶活性，而使细胞凋亡减少，细胞衰老变慢。hTERT基因转染的平滑肌细胞不具有恶性表型。 关键词：勃起功能障碍，衰老，端粒酶，人端粒酶蛋白催化亚单位，平滑肌细胞 The Effects of hTERT Transfection on Human Corpus Cavernosum Smooth Muscles WU Xiaojun， SONG Bo，ZHANG Heng，WANG Yong Quan，HE Peng，ZHOU Xiao Zhou，LI Weibing，ZHOU Zhansong (Institute of Urinary Surgery, Southwest Hospital, Third Military Medical University Chongqing,400038,China) Abstract: Objective: To explore effects of hTERT gene transfection on biological behaviors of human corporal smooth muscle cells.Methods: Human corporal smooth muscle cells were grown in primary culture. A fluorescent eukaryotic expression vector, named hTERT-IRES2-EGFP, carrying the hTERT gene was constructed and transfected into human corporal smooth muscle cells using Lipofectin reagent.The telomerase activity, mitotic index, cell apoptosis, cell growth curves of transfected smooth muscle cells were determined; the potential formation of malignant phenotypes in these transfected cells was investigated. Results: We constructed a eukaryotic expression vectors carrying hTERT and EGFP genes and correctly transfected human corporal smooth muscle cells. Telomerase activity, mitotic index, and cell growth of hTERT-transfected cells were significantly higher than those of non-transfected cells and cells transfected with the empty EGFP vector, while apoptosis rates [作者简介] 吴小军（1970-），男，博士、博士后，副教授，硕士研究生导师，研究方向为男性勃起功能障碍的基础与临床研究。 [通信作者] 周占松，电话（023E-mail：zhouzhans@ [基金项目] 国家自然科学基金面上项目81270692） were lowest in hTERT-transfected cells. No changes in cell morphology, chromosom