h1-calponin基因对五指山小型猪骨髓间充质干细胞体外成骨分化的影响

【目的】通过碱性调宁蛋白（h1-calponin）基因与五指山小型猪骨髓间充质干细胞（bone marrow mesenchymal stem cells,BM MSCs）成骨分化关系的研究,探讨成骨分化机制。【方法】添加成骨分化液对BM MSCs进行成骨分化诱导,采用茜素红染色的方法鉴定细胞成骨分化的程度;通过定量PCR分析h1-calponin基因及成骨相关基因骨桥蛋白（osteopotin,OPN）、骨钙蛋白（osteocalcin,OCN）、核心结合因子α1（core binding factor a1,Cbfα1）及核心结合因子β（core binding factor beta,Cbfβ）表达谱。【结果】①随着诱导时间的延长,茜素红染色矿化结节逐渐增多;②成骨标志基因OPN和OCN表达趋势一致：诱导0至14 d,基因表达量呈增长趋势,诱导14 d表达量最高,而后下降;成骨相关基因Cbfα1和Cbfβ表达量于第7天达到最高峰,之后逐渐下降,但仍高于诱导前;③h1-calponin基因表达量随着诱导的延续逐渐下降;诱导14—21 d时,该基因基本不表达。【结论】h1-calponin基因的表达与五指山小型猪骨髓间充质干细胞体外成骨分化存在着负相关,可能是成骨分化的负调控基因。     

Some characteristics of four attenuated vaccine virus strains and a virulent strain of Aujeszky's disease virus

Summary

In the present study four attenuated virus strains, used as vaccines, and a virulent strain of Aujeszky's disease virus (AD V) were compared with respect to their virulence in mice, their ability to induce virus‐specified thymidine kinase (TK) in infected cells, and their cleavage profiles of viral DNA's after treatment with the restriction endonuclease Kpnl.

The survival time of mice inoculated with the B‐KAL or the virulent NIA‐3 strain was comparable. whereas the Hanha and BUK strains required significantly loniser periods to kill mice. Mice were resistant to the MK‐25 strain of ADV.

The strains were assayed for TK phenotype by plaque autoradiography after ³H‐thymidine labelling of infected cells. MK‐25 proved to be the only strain defective in induction of TK in pig kidney cells. Restriction endonuclease analysis of viral DNA's revealed that each vaccine strain showed a characteristic fragment pattern that could easily be differentiated from that of other vaccine and field strains of ADV. The present results demonstrate that the mouse virulence lest and the TK assay detect differences in biological properties of ADV strains, but that restriction endonuclease analysis is required for unambiguous identification of vaccine and field strains of ADV.     

骨髓间充质干细胞免疫豁免机制及其临床应用

骨髓间充质干细胞是骨髓中除造血干细胞外的一种成体干细胞,其自我更新和体外扩增的能力及其多向分化的潜能已经被广泛认识.在研究中还发现它具有免疫原性弱,不诱导免疫排斥反应等特点,还具有"免疫豁免"特性,可抑制同种异体免疫反应、降低移植物抗宿主病(GVHD)的作用.文章对其机制及其在临床上的应用进行综述.     

小鼠MSCs体外定向诱导分化神经及脂肪细胞的研究

 【目的】建立昆明小鼠骨髓间充质干细胞的分离、培养体系,并鉴定其定向诱导分化为神经及脂肪细胞的能力。【方法】采用贴壁培养法分离培养昆明小鼠骨髓间充质干细胞（MSCs）,通过相差倒置显微镜对其进行形态学特征观察;通过碱性磷酸酶（AKP）染色、Hoechst 33258染色、瑞士-姬姆萨复合染色检测MSCs的表型特征;通过β-ME诱导MSCs定向分化为神经样细胞,诱导后的细胞进行H.E染色,观察其形态学特性;通过NF-200、NSE免疫荧光细胞化学方法对已分化的神经样细胞进行鉴定和分化率分析;通过马血清诱导MSCs定向分化为脂肪细胞,观察其体外生长过程,并对诱导后的脂肪细胞进行油红O染色鉴定和分化率分析。【结果】分离得到的MSCs为成纤维样细胞,可见多个核仁,AKP染色阳性率为93.2%±1.5%;β-ME诱导后,MSCs分化为多种表型的神经细胞与原代培养的小鼠全脑细胞的外形相似,诱导后的神经细胞NF-200、NSE免疫荧光细胞化学染色均呈阳性,NF-200阳性率为85.4%±1.8%,NSE阳性率为82.7%±2.1 %;马血清诱导后,MSCs可100%分化为脂肪细胞,油红O染色为阳性。【结论】 采用贴壁培养法能够成功分离和培养昆明小鼠的MSCs,分离得到的MSCs具有定向分化成多种神经细胞和脂肪细胞的能力。