CAJ | 학술논문

背景:通过动员自身或移植外来的骨髓来源肝干细胞可促进肝再生,但是,在大规模临床应用前,其安全性需要进一步研究.目的:采用含体积分数5%淤胆血清的培养基诱导大鼠骨髓间充质干细胞向肝干细胞方向分化,将这些骨髓来源肝干细胞接种到裸鼠体内,观察其是否具有成瘤性.方法:用含体积分数5%淤胆血清的培养基培养大鼠骨髓间充质干细胞;以免疫荧光法检测白蛋白、甲胎蛋白及细胞角皮素18在诱导后细胞的表达;以糖原染色及尿素合成检测细胞功能. 将培养14 d的大鼠骨髓来源肝干细胞接种于裸鼠皮下,观察局部有无新生物形成.结果与结论:用含体积分数5%淤胆血清的培养基培养大鼠骨髓间充质干细胞,4 d后出现细胞集落,细胞为圆形;7 d后集落变大,其周围开始出现多角形细胞;培养14 d后可见细胞呈多角形和排列成铺路石样,免疫荧光染色发现这些细胞表达角皮素18、甲胎蛋白和白蛋白,糖原染色显示细胞内有糖原颗粒;培养第12～15天的培养液中尿素氮浓度逐渐升高.经诱导的大鼠骨髓来源的肝干细胞接种到裸鼠皮下,30 d后局部未见新生物形成,组织结构未见异常.结果提示用体积分数5%淤胆血清培养基诱导的大鼠骨髓源性肝干细胞可能无成瘤性.
배경:통과동원자신혹이식외래적골수래원간간세포가촉진간재생,단시,재대규모림상응용전,기안전성수요진일보연구.목적:채용함체적분수5%어담혈청적배양기유도대서골수간충질간세포향간간세포방향분화,장저사골수래원간간세포접충도라서체내,관찰기시부구유성류성.방법:용함체적분수5%어담혈청적배양기배양대서골수간충질간세포;이면역형광법검측백단백、갑태단백급세포각피소18재유도후세포적표체;이당원염색급뇨소합성검측세포공능. 장배양14 d적대서골수래원간간세포접충우라서피하,관찰국부유무신생물형성.결과여결론:용함체적분수5%어담혈청적배양기배양대서골수간충질간세포,4 d후출현세포집락,세포위원형;7 d후집락변대,기주위개시출현다각형세포;배양14 d후가견세포정다각형화배렬성포로석양,면역형광염색발현저사세포표체각피소18、갑태단백화백단백,당원염색현시세포내유당원과립;배양제12～15천적배양액중뇨소담농도축점승고.경유도적대서골수래원적간간세포접충도라서피하,30 d후국부미견신생물형성,조직결구미견이상.결과제시용체적분수5%어담혈청배양기유도적대서골수원성간간세포가능무성류성.
BACKGROUND: Mobilizing autologous or extraneous bone marrow-derived liver stem cells may promote liver regeneration, however, its safety before the large scale clinical application needs further evaluation.OBJECTIVE: Bone marrow-derived liver stem cells (BDLSCs) were induced by culturing the rat bone marrow mesenchymal cells in the medium containing 5% cholestatic sera, and then were implanted into nude mice to observe the tumorigenicity. METHODS: Rat bone marrow mesenchymal cells (BMSCs) were isolated and incubated in the medium containing 5% cholestatic sera. Immunofluorescent stain was used to detect the expression of albumin, alpha-fetoprotein and cytokeratin18 by the cultured cells. Glycogen and urea synthesis by these cells were analyzed, respectively. BDLSCs following 14 days of culture were incubated in the skin of nude mice to observe neoplasia in local site. RESULTS AND CONCLUSION: Rat BMSCs survived in the medium containing 5% cholestatic serum and formed into small colonies on the fourth day after culture. Seven days later, the colonies expanded and there appeared some polygonal cells in the peripheral area. About 14 days later, these polygonal cells were confluent and presented the shape of cobblestone. Immunofluorescent stain showed that these cells expressed cytokeratin18, albumin and alpha-fetoprotein. Staining for glycogen displayed that glycogen granules were seen in cells. From 12 to15 days after culture, urea nitrogen concentrations in the medium were gradually increased. Rat BDLSCs were incubated in the skin of nude mice. Thirty days later, no neoplasia was found in the local site, and the tissue structure was normal. This result indicated that rat BDLSCs induced with the medium containing 5% cholestatic serum might have not tumorigenicity.