CAJ | 학술논문

目的探讨血管内皮细胞生长因子基因用重组腺相关病毒为载体移植制备血管瘤动物模型的可行性.方法构建重组腺相关病毒(recombinant adeo-associated virus,rAAV)介导的人血管内皮细胞生长因子(human vascular endothelial growth factor,hVEGF121)基因(rAAV-hVEGF121),并进行滴度测定.取10只小鼠,每只小鼠于左耳背部给以rAAV-VEGF121 1.0×1011VG/50μl,右耳背部给以等体积磷酸盐缓冲液(PBS)作为对照;隔天观察移植处皮肤颜色变化及肿胀情况,术后2、4、6、8、12周取注射处组织,行组织学榆查.结果经酶切鉴定、聚合酶链反应(PCR)及基因测序证实rAAV包装质糙pSNAV-hVEGF121构建成功,经rAAV包装后测定滴度为2×1015VG/L.基因移植后2周所有小鼠均可见左耳背部开始发红,之后逐渐形成红色的小包块,12周时小包块面积最大,右耳背部未见变化.病理结果显示2周后移植处组织内新生血管内皮细胞逐渐增多,8周后内皮细胞排列成血窦样结构,内含红细胞,CD-34染色提示增生细胞丰要为血管内皮细胞.结论以重组腺相关病毒为载体的血管内皮细胞生长因子基因移植裸鼠可以产生稳定有效的血管瘤模型.
목적 탐토혈관내피세포생장인자기인용중조선상관병독위재체이식제비혈관류동물모형적가행성.방법 구건중조선상관병독(recombinant adeo-associated virus,rAAV)개도적인혈관내피세포생장인자(human vascular endothelial growth factor,hVEGF121)기인(rAAV-hVEGF121),병진행적도측정.취10지소서,매지소서우좌이배부급이rAAV-VEGF121 1.0×1011VG/50μl,우이배부급이등체적린산염완충액(PBS)작위대조;격천관찰이식처피부안색변화급종창정황,술후2、4、6、8、12주취주사처조직,행조직학유사.결과 경매절감정、취합매련반응(PCR)급기인측서증실rAAV포장질조pSNAV-hVEGF121구건성공,경rAAV포장후측정적도위2×1015VG/L.기인이식후2주소유소서균가견좌이배부개시발홍,지후축점형성홍색적소포괴,12주시소포괴면적최대,우이배부미견변화.병리결과현시2주후이식처조직내신생혈관내피세포축점증다,8주후내피세포배렬성혈두양결구,내함홍세포,CD-34염색제시증생세포봉요위혈관내피세포.결론 이중조선상관병독위재체적혈관내피세포생장인자기인이식라서가이산생은정유효적혈관류모형.
Objective To investigate the feasibility of establishing a marine hemangioma model with injection of recombinant adco-associated virus mediated human vascular endothelial growth factor-121 (rAAV-hVEGF121) gene.Methods rAAV-hVEGF121 was constructed, identified and then implanted to the left back ear of each mouse (1.0×1011VG in 50μl per mouse and 10 nude mice received the injection) , the rights served as controls with an injection of the same volume of phosphate buffered solution(PBS).The skin color and swelling of left back car were observed every other day.Histological examination was carried out after mice were sacrificed 2,4,6,8,12 weeks after injection.Results The rAAV-hVEGF121 was correctly constructed and confirmed by restriction endonuclease analysis, polymerase chain reaction and DNA sequencing analysis.The skin of left back ear became red 2 weeks after injection and gradually exhibited a red lump which was at its utmost 12 weeks after injection.Such phenomena were not observed in right back ear.Histological examinations showed aggregates of endothelial cells by 2 weeks and at 8 weeks the swollen tissue contained many cysts filled with a mass of red cells.CD-34 staining suggested most of the newly-formed cells were endothelial cells.Conclusions A hemangioma model was established in mice with injection of recombinant rAAV-hVEGF121 gene.