[人、兔和大鼠脂肪组织来源的基质细胞体外生物学特性比较]

[Comparison of biological characteristics of human, rabbit and rat adipose tissue-derived stromal cells in vitro].

作者信息

Ni Yong-wei, Zhou Yong-sheng, Liu Yun-song, Zeng Bai-jin, Xu Yong-wei

机构信息

Department of Prosthodontics, Peking University School and Hospital of Stomatology, Beijing 100081, China.

出版信息

Beijing Da Xue Xue Bao Yi Xue Ban. 2009 Feb 18;41(1):95-9.

DOI:

PMID:19221574

Abstract

OBJECTIVE

This study aims to investigate the difference of proliferation patterns and osteogenic and adipogenic differentiation capability of adipose-derived stromal cells (ADSCs) obtained from human lipoaspirates, rat and rabbit inguinal subcutaneous adipose tissues in vitro.

METHODS

Adipose tissues of healthy adults were obtained by liposuction. Human ADSCs were isolated from these adipose tissues and cultured in DMEM containing 10% fetal bovine serum (FBS). Rat and rabbit ADSCs were obtained from inguinal subcutaneous adipose tissues and cultured with the same methods. These cells were observed under inverted microscope each day and cell growth was measured with MTT assay. Adipogenic differentiation was induced by culturing ADSCs for 1 or 2 weeks in adipogenic medium (AM) containing 1 micromol/L dexamethasone, 10 micromol/L insulin, 200 micromol/L indomethacin, 0.5 mmol/L isobutyl-methylxanthine (IBMX), and assessed by Oil Red O staining as an indicator of intracellular lipid accumulation. Osteogenic differentiation was induced by culturing ADSCs in osteogenic medium (OM) containing 0.1 micromol/L dexamethasone, 50 micromol/L ascorbate-2-phosphate, 10 mmol/L beta-glycerophosphate, and examined via alkaline phosphatase (AP) activity and extracellular matrix (ECM) calcification by alizarin red S staining and quantification of matrix calcification.

RESULTS

Fibroblast-like cells were digested from both inguinal subcutaneous adipose tissues of rabbit or rat and human lipoaspirates obtained from subcutaneous adipose tissues. Lipid-filled droplets were accumulated in human, rat and rabbit ADSCs upon treatment with adipogenic medium and were stained by Oil Red O. No lipid droplets were observed in the control undifferentiated ADSCs. After exposure to osteogenic differentiation medium, human and rat ADSCs were found to possess greater osteogenic potentials than cells isolated from rabbit inguinal subcutaneous adipose tissues, which was evidenced by significantly different osteogenic markers including alkaline phosphatase and mineral deposition.

CONCLUSION

Rabbit ADSCs obtained from inguinal subcutaneous adipose tissues poorly possess osteogenic potentials compared with ADSCs of human lipoaspirates obtained from subcutaneous adipose tissues or ADSCs of rat from inguinal subcutaneous adipose tissues, although they all possess comparable adipogenic capacity.

摘要

目的

本研究旨在探讨从人抽脂物、大鼠和兔腹股沟皮下脂肪组织获取的脂肪来源基质细胞（ADSCs）在体外增殖模式、成骨分化及成脂分化能力的差异。

方法

通过抽脂获取健康成年人的脂肪组织。从这些脂肪组织中分离出人ADSCs，并在含10%胎牛血清（FBS）的DMEM中培养。从大鼠和兔腹股沟皮下脂肪组织获取ADSCs，并采用相同方法培养。每天在倒置显微镜下观察这些细胞，并用MTT法检测细胞生长情况。将ADSCs在含1 μmol/L地塞米松、10 μmol/L胰岛素、200 μmol/L吲哚美辛、0.5 mmol/L异丁基甲基黄嘌呤（IBMX）的成脂培养基（AM）中培养1或2周以诱导成脂分化，并用油红O染色评估细胞内脂质积累情况作为指标。将ADSCs在含0.1 μmol/L地塞米松、50 μmol/L抗坏血酸-2-磷酸酯、10 mmol/L β-甘油磷酸的成骨培养基（OM）中培养以诱导成骨分化，并通过碱性磷酸酶（AP）活性、茜素红S染色检测细胞外基质（ECM）钙化情况及基质钙化定量来进行检测。

结果

从兔或大鼠的腹股沟皮下脂肪组织以及人皮下脂肪组织抽脂物中均消化出成纤维样细胞。用成脂培养基处理后人、大鼠和兔的ADSCs中均积累了充满脂质的小滴，且用油红O染色。在未分化的对照ADSCs中未观察到脂质小滴。在暴露于成骨分化培养基后，发现人与大鼠的ADSCs比从兔腹股沟皮下脂肪组织分离的细胞具有更强的成骨潜能，这通过碱性磷酸酶和矿物质沉积等显著不同的成骨标志物得以证明。