CAJ | 학술논문

背景:利用基因转染技术诱导脂肪间充质干细胞成软骨细胞已有报道,但腺病毒和腺相关病毒转染脂肪间充质干细胞各有不同,且腺相关病毒载体能否转染脂肪间充质干细胞众说不一.目的:观察Ad5-增强型绿色荧光蛋白(enhanced green fluorescent protein,EGFP)和Raav2-EGFP转染脂肪间充质干细胞后增强型绿色荧光蛋白的表达及转染后细胞增值能力的变化.方法:脂肪组织来源于6月龄新西兰大白兔颈背部.机械消化和酶消化法分取脂肪间充质干细胞,体外培养扩增.分别采用腺病毒载体(Ad5-EGFP)和腺相关病毒载体(Raav2-EGFP)转染脂肪间充质干细胞,并观察EGFP的表达.Raav2-EGFP转染后,需加入2 Μl丁酸钠(1 mol/L).采用MTT法检测基因转染对脂肪间充质干细胞增殖能力的影响.结果与结论:体外培养的脂肪间充质干细胞呈扁平状和长梭形,细胞形态均一,传代稳定.Ad5-EGFP组和Raav2-EGFP组均能观察到较多荧光细胞,转染效率分别达到88%和83%.腺病毒和腺相关病毒载体均能转染脂肪间充质干细胞,且转染效率很高.腺相关病毒需要借助丁酸钠来提高其基因表达水平.
배경:이용기인전염기술유도지방간충질간세포성연골세포이유보도,단선병독화선상관병독전염지방간충질간세포각유불동,차선상관병독재체능부전염지방간충질간세포음설불일.목적:관찰Ad5-증강형록색형광단백(enhanced green fluorescent protein,EGFP)화Raav2-EGFP전염지방간충질간세포후증강형록색형광단백적표체급전염후세포증치능력적변화.방법:지방조직래원우6월령신서란대백토경배부.궤계소화화매소화법분취지방간충질간세포,체외배양확증.분별채용선병독재체(Ad5-EGFP)화선상관병독재체(Raav2-EGFP)전염지방간충질간세포,병관찰EGFP적표체.Raav2-EGFP전염후,수가입2 Μl정산납(1 mol/L).채용MTT법검측기인전염대지방간충질간세포증식능력적영향.결과여결론:체외배양적지방간충질간세포정편평상화장사형,세포형태균일,전대은정.Ad5-EGFP조화Raav2-EGFP조균능관찰도교다형광세포,전염효솔분별체도88%화83%.선병독화선상관병독재체균능전염지방간충질간세포,차전염효솔흔고.선상관병독수요차조정산납래제고기기인표체수평.
BACKGROUND:There are reports concerning differentiation of adipose-derived mesenchymal stem cells(ADSCs)into chondrocytes using gene transfection technique.However,the transfection of adenovirus and adeno-associated virus into ADSCs is various.It is controversial whether adeno-associated virus(AAV)can transfect ADSCs.OBJECTIVE:To observe the enhanced green fluorescent protein(EGFP)expression following Ad5-EGFP and rAAV2-EGFP transfection into ADSCs,and investigate the cell proliferation ability following transfection.METHODS:ADSCs were isolated from the adipose tissue,which was from 6-month-old New Zealand albino rabbit back and neck by mechanical digestion and enzyme digestion,then ADSCs were cultured and amplified in vitro.ADSCs were infected with Ad5-EGFP and rAAV2-EGFP,and the EGFP expression was observed.A total of 2 μL sodium butyrate(1 mol/L)was added into the medium after rAAV2-EGFP transfection.MTT assay was used to detect the gene transfection effects on reproductive activity of ADSCs.RESULTS AND CONCLUSION:ADSCs isolated and cultured in vitro were flat,long-spindle and amplified stabry.The cell morphology was uniform.Many green fluorescent cells were observed in Ad5-EGFP and rAAV2-EGFP groups.Transfection efficiencies were about 88% and 83%.Adenovirus and adeno-associated virus vector can be transfected with ADSCs,and transfection efficiency is high.Adeno-associated virus needs sodium butyrate to increase its level of gene expression.