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| 干扰SEDL基因表达对软骨细胞生理活性的影响 |
| The influence of interferential SEDL gene in the physiological activity of human chondrocytes |
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| DOI：10.3969/j.issn.1006.7108.2015.12.006 |
| 中文关键词: X-连锁迟发性脊椎骨骺发育不良；SEDL基因；Sedlin 软骨细胞 |
| 英文关键词:X-linked spondyloepiphyseal dysplasia SEDL gene Sedlin Cartilage cell |
| 基金项目:重庆市自然科学基金计划重点项目（CSTC ,2007BA5028) |
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| 中文摘要: |
| 目的 初步研究干扰SEDL基因表达后对人软骨细胞中II型胶原蛋白表达水平、内质网位置分布及超微结构和软骨细胞活性的影响。方法 将靶向干扰SEDL基因的病毒转染入人软骨细胞中，QPCR及Western Blot分别检测SEDL基因及Sedlin蛋白表达水平，评价干扰SEDL基因的软骨细胞模型是否建立成功；免疫荧光及Western Blot检测II型胶原蛋白表达情况，荧光探针检测内质网位置分布改变，电镜检测内质网超微结构改变，MTT法检测软骨细胞增殖活性情况。结果 （1)转染干扰SEDL基因病毒的软骨细胞SEDL基因表达明显降低，具有统计学差异（P <0.05)。Sedlin蛋白表达降低，表明干扰SEDL 基因的细胞模型构建成功。（2)随着Sedlin蛋白的减少，II型胶原蛋白表达量降低，具有统计学差异（P<0. 05)。（3)细胞模型中内质网分布发生改变，其形态变圆，触角变少，颜色变淡，细胞与细胞间内质网荧光排列不规则，紊乱。（4)电镜下观察，细胞模型中内质网超微结构出现水肿扩张。（5)转染干扰SEDL基因病毒的软骨细胞活性在48 h达峰，之后细胞活性下降， 且具有统计学差异（P<0.05)。结论 Sedlin降低影响软骨细胞II型胶原蛋白表达，软骨细胞中内质网形态位置分布及超微结构发生改变，致使软骨细胞活性降低。 |
| 英文摘要: |
| Objective To preliminarily study the influence of interferential SEDL gene in the expression of collagen II，distribution and ultrastructure of endoplasmic reticulum，and the activity of hunian chondrocytes. Methods Lentivirus which can target interference SEDL gene was transfected into human cartilage cells (HC-a). The expression of SEDL gene and Sedlin protein were detected using QPCR and Western blotting. The establishment of cartilage cell model with SEDT gene was evaluated. The expression of collagen II was detected using Western blotting and immunofluorescence. The ultrastructural changes of endoplasmic reticulum were tested using electron microscope. MTT method was used to detect the proliferation of the cartilage cells. Results (l) The expression of SEDL gene in the cartilage cells was significantly reduced after lentivirus was transfected (P < 0. 05), and the expression of Sedlin protein was also decreased, indicating that the SEDT cell model was successfully established.(2) With the decrease of expression of Sedlin protein, the expression of collagen II was observably reduced (P <0. 05). (3) The distribution of endoplasmic reticulum was changed in the cell model. The cells became round in shape, less protrusion, fade in color, and the endoplasmic reticulum arranged irregularly. (4) The edematous endoplasmic reticulum was observed under transmission electron microscope. (5) Activity of the cells reached peak within 48h after lentivirus transfection. Then the cell activity significantly declined (P <0. 05) . Conclusion The reduced expression of Sedlin results in the decline of collagen II expression. The changed distribution and ultra microstructure of endoplasmic reticulum result in a decrease of cartilage cell activity. |
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