缺氧诱导过表达骨形态发生蛋白-2的兔脂肪来源干细胞成骨。

Hypoxia induces osteogenesis in rabbit adipose-derived stem cells overexpressing bone morphogenic protein-2.

作者信息

Xu L, Sun X, Cao K, Wu Y, Zou D, Liu Y, Zhang X, Zhang X, Wang G, Huang Q, Jiang X

机构信息

Department of Prosthodontics, Ninth People's Hospital, Shanghai Jiao Tong University, School of Medicine, Shanghai, China; Oral Bioengineering and Regenerative Medicine Lab, Shanghai Research Institute of Stomatology, Ninth People's Hospital, Shanghai Key Laboratory of Stomatology, Shanghai Jiao Tong University, School of Medicine, Shanghai, China.

出版信息

Oral Dis. 2014 Jul;20(5):430-9. doi: 10.1111/odi.12148. Epub 2013 Jul 19.

DOI:10.1111/odi.12148

PMID:23865899

Abstract

OBJECTIVE

Hypoxic culture potentiates mesenchymal stem cells (MSCs) to survive and secrete various growth factors. Genetically modified stem cells overexpressing bone morphogenic protein-2 (BMP-2) demonstrate strong osteogenic ability. Hence, we investigated the coeffect of hypoxic culture conditions and BMP-2 overexpression on the osteogenic ability of rabbit adipose-derived stem cells (rASCs) in vitro.

MATERIALS AND METHODS

Rabbit adipose-derived stem cells with or without adenoviral-BMP-2 transduction were cultured in hypoxic (1%) and normoxic (21%) conditions. Cell viability, attachment, and proliferation were compared. Real-time PCR amplification of osteogenic and angiogenic genes including alkaline phosphatase (ALP), osteocalcin (OCN), HIF-1α, and vascular endothelial growth factor (VEGF) was performed. Moreover, ALP activity, immunofluorescent staining of OCN, and mineralization assay by alizarin red S quantification and von Kossa staining were conducted.

RESULTS

Cells under hypoxic conditions attached better within 12 h and proliferated faster. While BMP-2 overexpression and hypoxic condition separately elevated the transcription of key osteogenic and angiogenic genes, a cooperative effect was observed to enhance the upregulation of osteogenic as well as angiogenic genes. Identical changes were observed in ALP activity, immunofluorescent staining of OCN, and mineralization assay.

CONCLUSIONS

Hypoxic culture can enhance the osteogenic ability of BMP-2 gene-modified rASCs, which provides a strategy to improve the osteogenesis of rASCs for in vivo bone regeneration.

摘要

目的

缺氧培养可增强间充质干细胞（MSCs）的存活能力并促使其分泌多种生长因子。过表达骨形态发生蛋白-2（BMP-2）的基因修饰干细胞具有很强的成骨能力。因此，我们研究了缺氧培养条件和BMP-2过表达对兔脂肪来源干细胞（rASCs）体外成骨能力的协同作用。

材料与方法

将转导或未转导腺病毒-BMP-2的兔脂肪来源干细胞在缺氧（1%）和常氧（21%）条件下培养。比较细胞活力、贴壁情况和增殖能力。对包括碱性磷酸酶（ALP）、骨钙素（OCN）、低氧诱导因子-1α（HIF-1α）和血管内皮生长因子（VEGF）在内的成骨和血管生成基因进行实时PCR扩增。此外，进行了ALP活性检测、OCN免疫荧光染色以及茜素红S定量和冯科萨染色的矿化分析。