The effect of macropore size of hydroxyapatite scaffold on the osteogenic differentiation of bone mesenchymal stem cells under perfusion culture

Previous studies have proved that dynamic culture could facilitate nutrients transport and apply mechanical stimulation to the cells within three-dimensional scaffolds,thus enhancing the differentiation of stem cells towards the osteogenic phenotype.However,the effects of macropore size on osteogenic differentiation of stem cells under dynamic condition are still unclear.Therefore,the objective of this study was to investigate the effects of macropore size of hydroxyapatite(HAp)scaffolds on osteogenic differentiation of bone mesenchymal stem cells under static and perfusion culture conditions.In vitro cell culture results showed that cell proliferation,alkaline phosphate(ALP)activity,mRNA expression of ALP,collagen-I(Col-I),osteocalcin(OCN)and osteopontin(OPN)were enhanced when cultured under perfusion condition in comparison to static culture.Under perfusion culture condition,the ALP activity and the gene expression of ALP,Col-I,OCN and OPN were enhanced with the macropore size decreasing from 1300 to 800 mm.However,with the further decrease in macropore size from 800 to 500 mm,the osteogenic related gene expression and protein secretion were reduced.Computational fluid dynamics analysis showed that the distribution areas of medium-and high-speed flow increased with the decrease in macropore size,accompanied by the increase of the fluid shear stress within the scaffolds.These results confirm the effects of macropore size on fluid flow stimuli and cell differentiation,and also help optimize the macropore size of HAp scaffolds for bone tissue engineering.

生长分化因子-5(GDF-5)促进人退行性变髓核细胞外基质表达情况的研究（英文）

目的：研究腺病毒介导的GDF-5对人退行性变椎间盘髓核细胞生长和细胞外基质表达的影响，探索椎间盘退行性变基因治疗的新途径。创新点：首次验证了GDF．5对人退行变的髓核细胞的生长和细胞外基质的分泌均具有明显的促进作用，为椎间盘退行性变疾病早期基因治疗提供了新的途径。方法：利用腺病毒介导的GDF．5转染人退变的髓核细胞，设空白对照组、阴性对照组（绿色荧光蛋白（GFP）组）和实验组（GDF-5组）三个组，3、7、14和21天四个时问点。在预定的时间点，通过蛋白质免疫印迹（Westernblotting）、番红-O染色、免疫组化染色、酶联免疫法定量检测（ELISA）、逆转录聚合酶链式反应（RT-PCR）和细胞外基质的定量分析等手段，验证GDF-5对退变髓核细胞外基质表达的影响，同时对GDF．5对髓核细胞生长情况的促进作用进行了表征。结论：Ad—GDF-5能成功转染人退变的髓核细胞（图1），能有效地促进人退变髓核细胞细胞外基质Aggreean和Ⅱ型胶原（CollagenⅡ）分泌（图4-7），RT-PCR的结果表明Aggrecan和CollagenII基因表达也得到了明显的增强（图8）。同时GDF-5对人退变的髓核细胞的生长也有一定的促进作用（图9）。因此，Ad-GDF-5是椎间盘退行性变早期基因治疗的新途径。