组织块培养法扩增人脂肪源性干细胞的生物学特征鉴定

背景：组织块培养法分离人脂肪源性干细胞并进行体外培养，培养体系简单，节约时间，较好地保持了干细胞的生物学特性，并据此向多方向诱导分化，为组织工程学种子细胞的临床应用提供理论依据和参考路线。 目的：观察组织块培养法分离的人脂肪源性干细胞的基本生物学特性、表面抗原特点及其向成骨细胞和脂肪细胞诱导分化潜能。 方法：取健康人腹部皮下脂肪组织，组织块培养法分离出脂肪源性干细胞，进行体外培养，观察其形态特征。流式细胞仪检测第3代脂肪干细胞的细胞周期及表面抗原。取第3代人脂肪干细胞，分别用成骨和成脂诱导培养液诱导其向成骨细胞和脂肪细胞分化，Von Kossa染色、油红O染色定性鉴定。 结果与结论：原代及传代的人脂肪干细胞形态均类似成纤维细胞，生长能力旺盛。第3代人脂肪干细胞中，超过88%的细胞都处于G0/G1期。第3代脂肪干细胞表面抗原表达：CD29、CD44、CD90、CD105表达阳性，CD34、CD45、CD106表达阴性。人脂肪干细胞经成脂诱导后21 d，油红O染色阳性；成骨诱导培养后，茜素红染色和Von Kossa染色阳性。因此，实验证实人脂肪源性干细胞能向成骨细胞和脂肪细胞诱导分化。

Amplification of human adipose-derived stem cells using tissue culture technique: An evaluation of biological properties

BACKGROUND: Tissue culture technique was used to isolate human adipose-derived stem cells (ADSCs) for in vitro culture. This method is simple, can save time, and keeps biological properties of stem cells. According to this, the multiple-directional differentiation provides theoretic evidences and reference line for clinical application of tissue engineered seed cells. OBJECTIVE: To investigate the basic biological characteristics of human ADSCs by tissue culture techniques and their surface protein expression, and to explore their osteogenic and adipogenic potential in vitro. METHODS: Human adipose tissue was obtained from the abdominal subcutaneous adipose tissue of health patients, and then ADSCs were isolated with tissue culture techniques and cultured in vitro. Morphology of ADSCs was observed. Cell cycle and surface proteins of the third passage of ADSCs were analyzed by flow cytometry technique. ADSCs from the third passages were induced into osteogenic and adipogenic lineages by different revulsants. Cells were examined by von Kossa staining and Oi1 Red O staining. RESULTS AND CONCLUSION: Primary and passage ADSCs were fibroblast-like and could rapidly expand. The majority of the third passage of cells was in G0/G1 phase (88%). The third passage of ADSCs was positive for CD29, CD44, CD90 and CD105, while negative for CD34, CD45 and CD106. Human ADSCs were positive for Oi1 Red O staining at 21 days of adipogenic induction, and positive for Alizarin Reds staining and Von Kossa staining after osteogenic induction. Therefore, human ADSCs have the potential to differentiate into osteoblasts and adipocytes.