鹿茸角软骨细胞体外培养方法的建立及生长特性分析

【研究目的】建立梅花鹿鹿茸软骨细胞体外分离培养和扩增方法,观察鹿茸软骨细胞在体外培养的生物学特性,为鹿茸再生机理的研究奠定基础;【方法】体外分离培养鹿茸软骨细胞,采用组织学、MTT比色法、免疫组化等多种手段动态检测细胞生长增殖情况;【结果】核心部分,约150字鹿茸软骨细胞贴壁生长,原代细胞比传代生长速度快,原代及传代4代内的鹿茸软骨细胞均有活跃的增殖能力,软骨细胞呈三角形,多边形等多种形态,II型胶原免疫组化,甲苯胺蓝染色为阳性;【结论】体外分离培养鹿茸软骨细胞具有较强的增殖能力,传代培养4代内细胞生长旺盛并维持其生物学特性,可满足后续实验研究要求。     

肉鸡胫骨软骨症发病趋势及组织学研究

本试验采用４８０只１日龄Ａｒｂｒｏ Ａｃｒｅｓ商品代仔公鸡，研究胫骨软骨症的发生特征，胫骨软骨症随肉鸡生长的动态发展趋势以及与日粮氯和镁水平之间的关系。试验为２×２×４因子设计，日粮氯，镁分别采用２个水平，氯为０．１１５％和０．３５％，镁为０．１５％和０．４０％。时间因素为第三个因子，１０天为一个阶段，共４个阶段。试验期为４０天，实验结果表明，肉鸡胫骨软骨症的发生与发展具有明显的时间性并与肉鸡的生     

Effect of glucocorticoid induced autophagy on senescence in chondrocytes

AIM: To explore the effect of glucocorticoid on autophagy and senescence in the chondrocytes. METHODS: The collagen II in the normal chondrocytes isolated from the SD rats was checked. After stimulation with glucocorticoid, LysoTracker Red staining, MDC staining and Western blot were used to detect the level of autophagy in the chondrocytes. The mTOR pathway related molecules were investigated by Western blot. Cell senescence was analyzed by SA-β-gal staining. RESULTS: A dose-dependent increase in the number of autophagic vacuoles was observed in the dexamethasone-treated chondrocytes, which was demonstrated by the LysoTracker Red and MDC staining. The expression of LC3-II and beclin-1 was increased by dexamethasone, especially in the cells treated with dexamethasone for 4 d. However, P62 expression was decreased. SA-β-gal staining showed that the percentage of cell senescence was increased by dexamethasone. Surprisingly, the cell senescence induced by dexamethasone was exacerbated by the autophagic inhibitor 3-MA. CONCLUSION: Autophagy induced by dexamethasone protects chondrocyte from senescence. The mTOR pathway may be involved in the autophagy activation.     

Primary Cilia in Chondrogenic Differentiation of Equine Bone Marrow Mesenchymal Stem Cells: Ultrastructural Study

Locomotor system disorders in equine species, such as tendon lesions, osteoarthritis, or ligament injuries, are some of the most frequent causes of dramatic reduction in horse performance. Traditional therapies are aimed at the inflammatory process and pain, but they do not regenerate normal tendon or ligament matrix and do not reduce re-injured rates. Mesenchymal stem cells have started to use as therapeutic option to repair these injured tissues. Most studies have focused on their isolation, in vitro culture and phenotyping. However, mesenchymal stem cell ultrastructure has been disregarded in the last years. We investigate the ultrastructural characteristics of these cells once differentiated into chondrocytes. Ultrastructural analysis was conducted on suspension cultures of differentiated chondrocytes from bone marrow mesenchymal stem cells by means of transmission electron microscopy. The morphologic characteristics of these cells, their ability to produce the extracellular matrix, and the presence of a single cilium could be indicative of the mesenchymal cells differentiation into chondrocyte phenotype. This study provides essential data to evaluate the degree of suitable phenotypic stability, for these cells can be used with repair purposes.     

鹿茸角软骨细胞体外培养方法的建立及生长特性分析

【研究目的】建立梅花鹿鹿茸软骨细胞体外分离培养和扩增方法，观察鹿茸软骨细胞在体外培养的生物学特性，为鹿茸再生机理的研究奠定基础；【方法】体外分离培养鹿茸软骨细胞，采用组织学、MTT比色法、免疫组化等多种手段动态检测细胞生长增殖情况；【结果】核心部分，约150字鹿茸软骨细胞贴壁生长，原代细胞比传代生长速度快，原代及传代4代内的鹿茸软骨细胞均有活跃的增殖能力，软骨细胞呈三角形，多边形等多种形态，II型胶原免疫组化，甲苯胺蓝染色为阳性；【结论】体外分离培养鹿茸软骨细胞具有较强的增殖能力，传代培养4代内细胞生长旺盛并维持其生物学特性, 可满足后续实验研究要求。     

梅花鹿鹿茸软骨细胞的培养及X型胶原的克隆和序列分析

型胶原是肥大软骨细胞的标志物,据GenBank中收录的人、鼠、牛、猪型胶原基因的序列,进行同源性分析,在同源性高的相对保守区域设计了1对特异性引物。建立梅花鹿鹿茸软骨细胞的分离培养方法,培养鹿茸软骨细胞。提取细胞的总RNA,以总RNA为模板,用RT-PCR方法克隆了梅花鹿的型胶原基因序列为877的片段,此片段全部位于编码区,与已报道的牛的型胶原基因的序列比较,同源性达到96%。共有35个碱基发生变异。DNAStar软件分析表明,二者推导的氨基酸序列同源性为95.5%。     

梅花鹿鹿茸软骨细胞的培养及X型胶原的克隆和序列分析

X型胶原是肥大软骨细胞的标志物，据GenBank 中收录的人、鼠、牛、猪X型胶原基因的序列，进行同源性分析，在同源性高的相对保守区域设计了1对特异性引物。建立梅花鹿鹿茸软骨细胞的分离培养方法，培养鹿茸软骨细胞。提取细胞的总RNA，以总RNA为模板，用RT-PCR方法克隆了梅花鹿的X型胶原基因序列为877的片段，此片段全部位于编码区，与已报道的牛的X型胶原基因的序列比较，同源性达到96%。共有35个碱基发生变异。DNAStar 软件分析表明，二者推导的氨基酸序列同源性为95.5%。