Wang Lei, Huang Chenglong, Li Qing, Xu Xiaomei, Liu Lin, Huang Kui, Cai Xiaoxiao, Xiao Jingang
Department of Oral and Maxillofacial Surgery, The Affiliated Stomatology Hospital of Southwest Medical University, Luzhou, China.
Orofacial Reconstruction and Regeneration Laboratory, The Affiliated Stomatology Hospital of Southwest Medical University, Luzhou, China.
Cell Prolif. 2017 Apr;50(2). doi: 10.1111/cpr.12328. Epub 2017 Jan 16.
Osteoporosis (OP) is a systemic disease caused by imbalance between bone resorption and bone formation, commonly resulting from post-menopausal oestrogen deficiency. Although osteogenic differentiation potential of adipose-derived stem cells (ASCs) has been demonstrated, the effect of OP on osteogenic differentiation of ASCs remains unclear. Here, our work has been designed to compare proliferative capacity and osteogenic differentiation ability of ASCs obtained from osteoporotic mice and normal control mice.
Twenty 14-week-old female C57BL/6 mice were randomly divided into two groups: one, the ovariectomy (OVX) group (n=10), the other being the sham operated (Sham) group (n=10). ASCs and OP-ASCs were obtained from subcutaneous fat of female inguinal sites. Cells were passaged three times prior to subsequent experimentation. The xCELLigence system was used to monitor cell adhesion and proliferation. Mineralized nodules of differentiated ASCs and OP-ASCs were analysed using Alizarin red staining after osteogenic induction. Expressions of osteogenic-specific genes including osteopontin (Opn) and runt-related transcription factor 2 (Runx2) were assessed by real-time PCR and expression of bone-related proteins was detected by Western blotting.
Numbers of cells in all groups increased steadily for 6 days; rate of cell proliferation in the Sham group was found to be higher than in the OVX group after 48 hours. Mineralized bone nodular structures were significantly more concentrated in the Sham group than in the OVX group by day 21, and mRNA levels of Runx2 in the OVX group were significantly lower than in the Sham group. Transcript levels of genes coding for Opn showed a similar pattern to those of Runx2. Western blot results indicated that protein expression levels of OPN and RUNX2 in the OVX group were lower than those in the Sham group, at each time point.
These results indicated that the proliferative capacity and osteogenic potential of ASCs were significantly impaired in osteoporotic mice compared to normal controls. However, use of autologous transplantation of modified OP-ASCs for treatment of OP, or combination of composite scaffolds and modified OP-ASCs for repair of osteoporotic bone defects, can overcome shortcomings of other methods.
骨质疏松症（OP）是一种由骨吸收与骨形成失衡引起的全身性疾病，通常由绝经后雌激素缺乏所致。尽管已证实脂肪来源干细胞（ASC）具有成骨分化潜能，但OP对ASC成骨分化的影响仍不清楚。在此，我们的研究旨在比较从骨质疏松小鼠和正常对照小鼠获得的ASC的增殖能力和成骨分化能力。
将20只14周龄雌性C57BL/6小鼠随机分为两组：一组为卵巢切除术（OVX）组（n = 10），另一组为假手术（Sham）组（n = 10）。从雌性腹股沟部位的皮下脂肪中获取ASC和OP - ASC。在后续实验前，细胞传代三次。使用xCELLigence系统监测细胞黏附和增殖。成骨诱导后，采用茜素红染色分析分化的ASC和OP - ASC的矿化结节。通过实时PCR评估包括骨桥蛋白（Opn）和 runt相关转录因子2（Runx2）在内的成骨特异性基因的表达，并通过蛋白质免疫印迹法检测骨相关蛋白的表达。
所有组的细胞数量在6天内稳步增加；48小时后发现Sham组的细胞增殖率高于OVX组。到第21天，Sham组矿化骨结节结构明显比OVX组更密集，且OVX组Runx2的mRNA水平明显低于Sham组。编码Opn的基因转录水平呈现与Runx2相似的模式。蛋白质免疫印迹结果表明，在每个时间点，OVX组中OPN和RUNX2的蛋白表达水平均低于Sham组。
这些结果表明，与正常对照相比，骨质疏松小鼠中ASC的增殖能力和成骨潜能显著受损。然而，使用经修饰的OP - ASC自体移植治疗OP，或使用复合支架与经修饰的OP - ASC联合修复骨质疏松性骨缺损，可克服其他方法的缺点。
