CAJ | 학술논문

背景：脂肪源性间充质干细胞具有自我更新能力且在体外特定条件下具有多向分化潜能，在临床上具有广泛的应用前景。然而，脂肪源性间充质干细胞的分离培养仍存在诸多困难与不足。目的：体外分离培养获得兔脂肪源性间充质干细胞，对其形态学、生物学特性进行观察，并鉴定其向成骨、成脂分化的潜能。方法：切取兔颈背区皮下脂肪，用胶原酶消化法分离兔脂肪源性间充质干细胞，进行体外培养，流式细胞仪检测细胞表面标志物，CCK-8法检测细胞活性并绘制细胞生长曲线。第4代兔脂肪源性间充质干细胞向成脂、成骨诱导分化后进行油红O染色、茜素红染色、碱性磷酸酶染色，观察其成脂、成骨分化潜能。结果与结论：兔脂肪源性间充质干细胞呈长梭形漩涡样贴壁生长，能稳定传代至第10代，增殖能力强；流式细胞仪检测可高表达CD29、CD90及CD44，而CD34和CD45表达较低；成脂诱导后的兔脂肪源性间充质干细胞油红 O 染色呈阳性；成骨诱导后的兔脂肪源性间充质干细胞碱性磷酸酶和茜素红染色均呈阳性，以上结果显示实验成功分离培养出兔脂肪源性间充质干细胞，获得的细胞生长稳定，增殖活跃，高表达间充质干细胞的表面抗原以及具有向成骨、成脂等多向分化的潜能，有望为骨组织工程提供理想的种子细胞。
배경：지방원성간충질간세포구유자아경신능력차재체외특정조건하구유다향분화잠능，재림상상구유엄범적응용전경。연이，지방원성간충질간세포적분리배양잉존재제다곤난여불족。목적：체외분리배양획득토지방원성간충질간세포，대기형태학、생물학특성진행관찰，병감정기향성골、성지분화적잠능。방법：절취토경배구피하지방，용효원매소화법분리토지방원성간충질간세포，진행체외배양，류식세포의검측세포표면표지물，CCK-8법검측세포활성병회제세포생장곡선。제4대토지방원성간충질간세포향성지、성골유도분화후진행유홍O염색、천소홍염색、감성린산매염색，관찰기성지、성골분화잠능。결과여결론：토지방원성간충질간세포정장사형선와양첩벽생장，능은정전대지제10대，증식능력강；류식세포의검측가고표체CD29、CD90급CD44，이CD34화CD45표체교저；성지유도후적토지방원성간충질간세포유홍 O 염색정양성；성골유도후적토지방원성간충질간세포감성린산매화천소홍염색균정양성，이상결과현시실험성공분리배양출토지방원성간충질간세포，획득적세포생장은정，증식활약，고표체간충질간세포적표면항원이급구유향성골、성지등다향분화적잠능，유망위골조직공정제공이상적충자세포。
BACKGROUND:Adipose-derived mesenchymal stem cells have the ability to self-renew and have pluripotent potential under specific conditions in vitro, which have broad application prospects in clinical practice. However, isolation and culture of adipose-derived mesenchymal stem cells stil appear to have many difficulties and shortcomings. OBJECTIVE:To isolate and culture rabbit adipose-derived mesenchymal stem cells in vitro in order to study their morphology, cellsurface markers and biological properties as wel as to investigate the osteogenic and adipogenic potentials of adipose-derived mesenchymal stem cells in vitro. METHODS:Primary adipose-derived mesenchymal stem cells were isolated from the subcutaneous adipose tissue of posterior cervical region from New Zealand white rabbits and digested by 0.1%col agenase I. The cells were passaged and amplified by the trypsin digestion. The passage 4 adipose-derived mesenchymal stem cells were induced to differentiate after exposure to adipogenic or osteogenic medium. The oil red O staining, alkaline phosphatase and alizarin red staining were used to detect the results. The cellviability was detected by the cellcounting kit 8 method to drawn the growth curve. cellsurface markers were examined using flow cytometry. RESULTS AND CONCLUSION:The adipose-derived mesenchymal stem cells isolated from the subcutaneous adipose tissue of rabbits exhibited a fusiform adherent growth in a vortex pattern, and had a strong capability of proliferation that could be passaged stably to the 10th generation. Flow cytometry results showed that the cells highly expressed CD29, CD90, CD44, but lowly expressed CD45 and CD34. After adipogenic induction, the adipose-derived mesenchymal stem cells were positive for oil red O staining;after osteogenic induction, the cells were both positive for alkaline phosphatase and alizarin red staining. These findings suggest that the adipose-derived mesenchymal stem cells were successful y isolated and cultured from the subcutaneous adipose tissue of rabbits, and these cells are pluripotent with the potential to differentiate into adipocytes and osteoblasts, which are expected to be ideal seed cells for bone tissue engineering.