Over-expression of VEGF165 in the adipose tissue-derived stem cells via the lentiviral vector

作     者：SUN Xiang-zhou LIU Gui-hua WANG Zhuo-qing ZHENG Fu-fu BIAN Jun HUANG Yan-ping GAO Yong ZHANG Ya-dong DENG Chun-hua 

作者机构：Department of Urology First Affiliated Hospital of Sun Yet-sen University Guangzhou Guangdong 510080 China Department of Scientific Research First Affiliated Hospital of Sun Yet-sen University Guangzhou Guangdong 510080 China 

出 版 物：《Chinese Medical Journal》 (中华医学杂志（英文版）)

年 卷 期：2011年第124卷第19期

页      面：3093-3097页

核心收录：

学科分类：0710[理学-生物学] 1002[医学-临床医学] 07[理学] 08[工学] 09[农学] 071007[理学-遗传学] 0901[农学-作物学] 0836[工学-生物工程] 090102[农学-作物遗传育种] 

基　　金：国家自然科学基金 National Youth Natural Science Fundation of China Natural Science Fund Program of Guangdong Province Natural Science Fund Program of Guangdong Province Guangdong Province Science & Technology Project 

主　　题：adipose tissue-derived stem cells VEGF16s lentivirus over-expression 

摘      要：Background Many researchers studied the possibility of using stem cells as gene therapeutic vector. But few related reports on the adipose tissue-derived stem cells （ADSCs） are available. Therefore we intended to construct a lentiviral VEGF165 expression vector and then infect the ADSCs to produce therapeutic seed *** EHS1001-68950485313912 clone was mutated by PCR method to produce consensus fragment of VEGF165 transcript （NM_001025368）. Lentivirus was enveloped with pGC-FU, pHelper 1.0 and pHelper 2.0 plasmids in 293T *** then the ADSCs （multiplicity of infection=20） were transfected with the vectors after titer determination. Stable expression of VEGF165 in ADSCs was confirmed by immunofluorescence staining, enzyme-linked immunosorbent assay （ELISA） and Western blotting *** DNA sequencing and 293T transfection verified VEGF165 was linked to the GFP fused vector. The virus titer is up to 2x10a determined by quantitative PCR. VEGF165 transduced cells could show green fluorescence confirmed by immunofluorescence staining （almost 95%）. ELISA analyses could detect out the density of VEGF was 850.86-1202.13pg/ml （mean （923.00±31.22） pg/ml） in the supernatant of VEGF16s-transduced cells but not detected in the GFP-transduced cells （P 〈0.001） and the Western blotting analyses also confirmed VEGF165 expression in VEGF165-transduced *** The VEGF165 over-expression ADSCs were obtained and may be used as a cell therapeutic tool and may be applied for vascular regeneration, especially in the treatment of erectile dysfunction.