CAJ | 학술논문

目的研究人骨髓间充质干细胞(hMSC)在肿瘤血管生成过程中的作用.方法通过梯度密度离心法及贴壁筛选法分离、培养hMSC;利用pLEGFP-N1逆转录病毒载体获得增强型绿色荧光蛋白(EGFP)标记的hMSC(hMSC-EGFP);流式细胞仪检测hMSC-EGFP的免疫表型及分化能力;通过建立BALB/C裸鼠实体瘤模型(分别皮下接种乳腺癌细胞系MCF-7及hMSC-EGFP细胞与MCF-7混悬液)来观察hMSC在肿瘤血管生成过程中的作用;检测肿瘤细胞和内皮细胞条件培养基对hMSC细胞生长和迁移的影响;体外诱导hMSC向内皮细胞分化,观察其对人脐静脉内皮细胞(HUVEC)迁移的影响.结果 hMSC-EGFP与hMSC形态相似,均呈成纤维细胞样;二者具有相似的免疫表型,在条件基质作用下,均能被诱导分化为成骨细胞及脂肪细胞;hMSC能促进肿瘤血管生成,单纯接种MCF-7组肿瘤平均血管密度为5.33±1.42,混悬液组为13.67±1.53,差异有统计学意义(P<0.05);大部分血管是由hMSC植入体内引起的宿主源性血管新生,只有少数血管的内皮细胞是由hMSC植入体内分化而来的;肿瘤细胞和内皮细胞能够通过其旁分泌作用促进hMSC的生长和迁移;经内皮诱导2周后,hMSC呈CD31阳性;hMSC能促进HUVEC的迁移.结论 MSC具有促进肿瘤血管生成的作用.
목적 연구인골수간충질간세포(hMSC)재종류혈관생성과정중적작용.방법 통과제도밀도리심법급첩벽사선법분리、배양hMSC;이용pLEGFP-N1역전록병독재체획득증강형록색형광단백(EGFP)표기적hMSC(hMSC-EGFP);류식세포의검측hMSC-EGFP적면역표형급분화능력;통과건립BALB/C라서실체류모형(분별피하접충유선암세포계MCF-7급hMSC-EGFP세포여MCF-7혼현액)래관찰hMSC재종류혈관생성과정중적작용;검측종류세포화내피세포조건배양기대hMSC세포생장화천이적영향;체외유도hMSC향내피세포분화,관찰기대인제정맥내피세포(HUVEC)천이적영향.결과 hMSC-EGFP여hMSC형태상사,균정성섬유세포양;이자구유상사적면역표형,재조건기질작용하,균능피유도분화위성골세포급지방세포;hMSC능촉진종류혈관생성,단순접충MCF-7조종류평균혈관밀도위5.33±1.42,혼현액조위13.67±1.53,차이유통계학의의(P<0.05);대부분혈관시유hMSC식입체내인기적숙주원성혈관신생,지유소수혈관적내피세포시유hMSC식입체내분화이래적;종류세포화내피세포능구통과기방분비작용촉진hMSC적생장화천이;경내피유도2주후,hMSC정CD31양성;hMSC능촉진HUVEC적천이.결론 MSC구유촉진종류혈관생성적작용.
Objective The effect of human bone marrow mesenchymal stem cells (hMSCs)on tumor neovascularization were studied. Methods hMSCs were isolated from human bone marrow by density gradient fractionation and adherence to plastic flasks. hMSCs-EGFP were obtained by pLEGFP-N1 retroviral vector. Flow cytometry was used to detect the cell surface antigen and the differentiation potential of hMSCs-EGFP was investigated under conditioned media. The effect of hMSCs on tumor neovascularization were observed by establishing solid tumor models in BALB/C nude mice. In addition,effect of the conditioned medium used for tumor cells and endothelial cells (EC)cultivation was collected,to detect its effect on the growth and migration rates of hMSC. hMSCs were induced to differentiate into EC in vitro and the migratory effect on HUVEC was also evaluated. Results hMSCs-EGFP,like hMSC,exhibited a fibroblast-like morphological feature,and both had the similar cell surface antigens. They could be induced into osteocytes or adipocytes under the conditioned media The results not only suggested that hMSCs contributed to tumor neovascularization,but also indicated that most of vessels were host-derived angiogenesis mediated by hMSCs. The mean vascular density (MVD)in suspension group (13. 67 ± 1. 53)was strikely higher than that in MCF-7 group (5.33 ± 1.42),which showed statistical significance (P < 0. 05). Only very few vessles were attributed to hMSCs transdifferentiation into ECs. Tumor cells and ECs can promote hMSCs proliferation and migration through paracrine action. Furthermore,hMSCs were positive for CD31 after 2 weeks induction and HUVEC migration can be facilitated by hMSCs. Conclusion MSCs have the effect of promoting tumor neovascularization.