CAJ | 학술논문

背景：脂肪干细胞作为一种新的成体干细胞，具有来源丰富、取材容易、增殖能力强等优点，受到人们越来越多地关注。目的：体外分离培养小鼠附睾脂肪组织中的脂肪干细胞，并鉴定其生物学特性。方法：无菌切取小鼠附睾处脂肪组织，采用胶原酶进行消化，利用1次消化多次收集与差速贴壁法分离纯化脂肪干细胞。倒置显微镜及透射电子显微镜观察脂肪干细胞的形态。绘制脂肪干细胞的生长曲线。流式细胞术检测脂肪干细胞的免疫表型。加入细胞诱导剂对脂肪干细胞进行成骨及成脂肪的诱导分化。扫描电子显微镜观察脂肪干细胞与胶原支架材料的相容性。结果与结论：倒置显微镜下可见脂肪干细胞呈长梭形或成纤维细胞样，密集排列成漩涡状或成束状交织，可在体外稳定传至第9代。透射电子显微镜下可见脂肪干细胞表面有丰富的微绒毛结构，细胞核体积大，细胞质内可见线粒体、粗面内质网等细胞器。脂肪干细胞表达CD44、CD29，不表达CD34。经成脂肪诱导剂诱导后，多数脂肪干细胞胞质内可见小脂滴，油红O染色可将小脂滴染成红色。经成骨诱导剂诱导后，在脂肪干细胞生长密集区域内的细胞结构模糊、细胞间界限不清楚。经茜素红染色后，可见较多大小不一的折光率较强颗粒状结构沉积。扫描电子显微镜观察到脂肪干细胞呈铺展状生长于胶原支架材料上。结果表明该方法分离出的脂肪干细胞能在体外扩增并稳定传代，在一定诱导条件下，能够分化为成骨细胞和脂肪细胞并且与胶原支架具有良好的相容性。
배경：지방간세포작위일충신적성체간세포，구유래원봉부、취재용역、증식능력강등우점，수도인문월래월다지관주。목적：체외분리배양소서부고지방조직중적지방간세포，병감정기생물학특성。방법：무균절취소서부고처지방조직，채용효원매진행소화，이용1차소화다차수집여차속첩벽법분리순화지방간세포。도치현미경급투사전자현미경관찰지방간세포적형태。회제지방간세포적생장곡선。류식세포술검측지방간세포적면역표형。가입세포유도제대지방간세포진행성골급성지방적유도분화。소묘전자현미경관찰지방간세포여효원지가재료적상용성。결과여결론：도치현미경하가견지방간세포정장사형혹성섬유세포양，밀집배렬성선와상혹성속상교직，가재체외은정전지제9대。투사전자현미경하가견지방간세포표면유봉부적미융모결구，세포핵체적대，세포질내가견선립체、조면내질망등세포기。지방간세포표체CD44、CD29，불표체CD34。경성지방유도제유도후，다수지방간세포포질내가견소지적，유홍O염색가장소지적염성홍색。경성골유도제유도후，재지방간세포생장밀집구역내적세포결구모호、세포간계한불청초。경천소홍염색후，가견교다대소불일적절광솔교강과립상결구침적。소묘전자현미경관찰도지방간세포정포전상생장우효원지가재료상。결과표명해방법분리출적지방간세포능재체외확증병은정전대，재일정유도조건하，능구분화위성골세포화지방세포병차여효원지가구유량호적상용성。
BACKGROUND:As a new kind of adult stem cells, adipose-derived stem cells get more and more attention, because of rich source, drawing materials easily and powerful proliferation. OBJECTIVE:To isolate and culture adipose-derived stem cells from the epididymal adipose tissue in mice, and to identify their biological characteristics. METHODS:Adipose tissue was obtained from epididymis in mice by aseptical y cutting. The tissue was digested using col agenase. Adipose-derived stem cells were separated and purified by using one digestion, multiple col ection method and differential adhesion method. The morphology of adipose-derived stem cells was observed using inverted microscopy and transmission electron microscopy. Growth curve of adipose-derived stem cells was drawn. Immunophenotype of adipose-derived stem cells was identified by flow cytometry. Adipose-derived stem cells were induced to differentiate into adipocytes and osteocytes using cellinductors. Compatibility of adipose-derived stem cells and col agen scaffold material was observed using scanning electron microscope. RESULTS AND CONCLUSION:Adipose-derived stem cells exhibited long spindle-like or fibroblast-like appearance, grew intensively and arranged in scrol and fascicular shape. In vitro, adipose-derived stem cells could be passaged to passage 9 under the inverted microscope. Under the transmission electron microscope, adipose-derived stem cells showed abundant microvil i on the cellsurface. The nuclei were big in size. Some organel es were seen in cytoplasma, such as mitochondria and rough endoplasmic reticulum. Adipose-derived stem cells expressed CD44 and CD29, did not express CD34. After inducing by inductor, many smal lipid droplets were seen in the cytoplasm of adipose-derived stem cells. The smal lipid droplets were dyed red with oil red O. After induction of osteogenic inductor, the boundary line among adipose-derived stem cells was not clear and the structure of cells was fuzzy in the growth-intensive areas. There were many strong refractive granular material deposits at that field after dyeing with alizarin red. Scanning electron microscope revealed that adipose-derived stem cells were spread on the col agen scaffold. Results suggested that adipose-derived stem cells isolated by this method could amplify in vitro and stably subcultured. Under a certain inducing condition, adipose-derived stem cells could differentiate into osteoblasts and adipocytes, which showed a good compatibility with col agen scaffold.