【摘要】 背景:软骨组织修复是组织工程研究的重要领域,如何利用工程学技术有效将种子细胞定向分化成软骨细胞是组织工程的重点和难点。目前,单纯应用各种定向诱导培养试剂很难使其分化为成熟稳定的软骨细胞,正是在这一背景下,作者利用ATDC5软骨细胞的特点,除了应用有效的培养液处理外,还采用间歇净水压的压力刺激方法,对其定向诱导分化进行早期研究。目的:了解间歇静水压对ATDC5软骨细胞早期软骨方向分化成熟的影响。方法:将ATDC5软骨细胞株在单层条件下培养,3 d细胞贴壁良好,并形成复层,而后在密封条件下进行间歇静水压(施加强度10 MPa,加压频率1 Hz,4 h/d)培养,设立无间歇静水压且其他条件相同的培养细胞为对照组。在第4,7,11,14,17天,通过显微镜观察细胞形态变化,应用Real-time PCR检测Aggrecan,COL-2,SOX-9的m RNA表达水平。结果与结论:经间歇静水压作用后,ATDC5细胞表现出较明显的斑块样改变和细胞浓聚现象;Aggrecan、COL-2m RNA表达水平明显升高,SOX-9mRNA虽然与对照组变化不大,但也出现了先抑后扬的特点。结果表明,间歇静水压影响ATDC5软骨细胞向软骨方向分化的基因表达,促进软骨特征基质的分泌,利于向软骨细胞分化成熟。更多还原

【Abstract】 BACKGROUND: Cartilage tissue repair is an important field of tissue engineering. How to use engineering technology to effectively differentiate seed cells into chondrocytes is a focus and difficulty in the field of tissue engineering. At present, it is difficult to make seed cells differentiate into mature and stable chondrocytes by simple orientation-inducing culture. Thereafter, the authors preliminarily studied the induced directional differentiation using intermittent hydrostatic pressure stimulation based on the characteristics of ATDC5 chondrocytes, in addition to the use of effective culture solution. OBJECTIVE: To investigate the effects of intermittent hydrostatic pressure on the early-term chondrocyte differentiation of ATDC5 cells. METHODS: ATDC5 cell lines were cultured in multilayer. Cells adhered well with multiple-layer formation after 3 days, and were then sealed to maintain sterility. Intermittent hydrostatic pressure was applied to the cultures(10 MPa, 1 Hz, 4 h/d). Cells cultured with no intermittent hydrostatic pressure served as control group. Morphological changes of the cells were observed under microscope at 4, 7, 11, 14, and 17 days. Expression levels of Aggrecan, COL-2 and SOX-9 mRNA were detected by real-time PCR. RESULTS AND CONCLUSION: After application of intermittent hydrostatic pressure, ATDC5 cells aggregated and appeared with obvious patchy changes. The mRNA expression levels of Aggrecan and COL-2 were significantly increased. SOX-9 mRNA expression level showed no significant difference compared with the control group, but presented with fall-rise pattern. Intermittent hydrostatic pressure influences the mRNA expression related to chondrocyte differentiation and promotes the secretion of chondrogenic matrix. This method is contributive to the mature cartilage differentiation.更多还原