| 引用本文： |
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顾羊林,朱文辉,王予彬,等.犬成肌细胞体外向纤维软骨细胞表型分化的实验研究[J].同济大学学报（医学版）,2012,33(2):1-6. [点击复制]
- GU Yang-lin,ZHU Wen-hui,WANG Yu-bin,et al.Phenotype transformation from canine skeletal myoblast to fibrochondrocyte in vitro[J].同济大学学报（医学版）,2012,33(2):1-6. [点击复制]
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| 摘要: |
| 目的观察体外培养的犬成肌细胞在诱导培养液作用下向纤维软骨细胞表型分化的可行性。方法取成年比格犬大腿后群肌肉,以机械分解法与二步酶消化法相结合,分离获取成肌细胞,取第4代成肌细胞,实验组以2.0×104/cm2细胞密度接种于24孔板,在含有人源性软骨源性形态发生蛋白-2(human cartilage-derived morphogenic protein-2,hCDMP-2)的高糖DMEM完全培养液中诱导分化,对照组以相同细胞密度接种于24孔板,加入不含诱导因子的高糖DMEM完全培养液.诱导21 d后,倒置显微镜下观察诱导细胞形态学的改变,H-E染色、甲苯胺蓝染色、Ⅱ型胶原蛋白免疫组织化学方法检测诱导细胞的分泌功能,RT-PCR检测Ⅰ型、Ⅱ型胶原蛋白和Sox9的mRNA表达情况。结果诱导后成肌细胞的形态由长梭形向多角形类、圆形转变.甲苯胺蓝染色呈明显异染,Ⅱ型胶原蛋白免疫细胞化学检测阳性,对照组阴性.RT-PCR检测显示经诱导后细胞Ⅰ型、Ⅱ型胶原蛋白和Sox9的mRNA在对应区域均有目的条带出现,而对照组未见明显的目的条带亮带。结论在hCDMP-2诱导下,犬成肌细胞可定向分化为软骨细胞. |
| 关键词: 成肌细胞 软骨分化 种子细胞 人源性软骨源性形态发生蛋白-2 软骨组织工程 |
| DOI：10.3969/j.issn1008-0392.2012.02.001 |
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| 基金项目:上海市自然科学基金(09ZR1425500) |
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| Phenotype transformation from canine skeletal myoblast to fibrochondrocyte in vitro |
| GU Yang-lin,ZHU Wen-hui,WANG Yu-bin,LU Liang-yu,CHENG Yan,LI Jing-long |
| (Dept.of Sports Medicine,East Hospital,Tongji University,Shanghai 200120,China) |
| Abstract: |
| Objective To induce the phenotype transformation of canine skeletal myoblast into fibrochondrocyte in vitro.Methods The muscle from the hindlimb of adult Beagle canine was obtained,from which skeletal myoblasts were isolated and harvested by mechanical decomposition and two-step enzyme digestion.The cultured skeletal myoblasts of the fourth passage were digested and cultured in a medium containing human cartilage-derived morphogenic protein-2(hCDMP-2) in a 24-well plate at a density of 2.0×104/cm2 in the experimental group;while in the control group the DMEM-LG complete culture medium without hCDMP-2 was used.After 21days of incubation the cellular morphology,secretion function and expression of type Ⅱ collagen in cultured cells were examined by HE and toluidine blue staining,and immunohistochemistry,respectively.The expression of collagen Ⅰ,Ⅱ and Sox9 mRNA was detected by reverse transcription-polymerase chain reaction(RT-PCR).Results The skeletal myoblasts changed from a spindle-like appearance into a polygonal shape after induction.The extracellular matrix was positive stained by H-E and toluidine blue staining in experimental group,and the cells were positively stained by immunohistochemistry for type Ⅱ collagen.RT-PCR revealed that the expression of collagenⅠ,Ⅱ and Sox9 mRNA were significantly displayed in the experimental group.No positive expression was observed in the control group.Conclusion The canine skeletal myoblasts can be induced for phenotype transformation to chondrocy-tes by hCDMP-2 in vitro. |
| Key words: skeletal myoblasts chondrogenic differentiation seeding cells human cartilage-derived morphogenic protein-2 cartilaginous tissue engineering |
