Expression of transduced HSP70 gene protects chondrocytes from stress.
Expression of transduced HSP70 gene protects chondrocytes from stress.
复制标题
转导的 HSP70 基因的表达可保护软骨细胞免受压力。
作者：
T. Kubo;Y. Arai;K. Takahashi;T. Ikeda;S. Ohashi;I. Kitajima;O. Mazda;M. Takigawa;J. Imanishi;Y. Hirasawa
客观化
目的观察腺病毒载体介导的热休克蛋白70(HSP70)基因转导人软骨细胞样细胞(HCS-2/8)的抗热应激作用。
方法
构建了受SRpha启动子调控的野生型(AxSHEwt)和突变型(AxSHEmt)HSP70基因的腺病毒载体。突变型缺乏表达压力持久性的区域。将其中一种腺病毒载体加入人软骨细胞样细胞(HCS-2/8)培养中。将热应激(48℃)作用于转导细胞2 h，采用Alamar蓝和四甲基偶氮唑盐比色法观察腺病毒载体介导的HSP70基因转导对软骨细胞的抗热应激作用。
结果
在48℃时，对照组的吸光度分别为300.3+/-51.9和1.173+/-0.011，转导细胞的吸光度分别为278.5+33.8和1.217+/-0.018，转导细胞的吸光度为1.371+/-0.033。转导AxSHEwt的细胞表达水平显著高于其他两组(p<；0.05)。在37摄氏度的处理下，没有观察到显著差异。
结论
转导HSP70基因的软骨细胞在热应激条件下具有较高的代谢活性和活力。
OBJECTIVE
To investigate the efficacy of adenovirus vector mediated transduction of heat shock protein 70 (HSP70) gene to human chondrocyte-like cell (HCS-2/8) against heat stress.
METHODS
Two adenovirus vectors that contain wild-type (AxSHEwt) or mutant-type (AxSHEmt) HSP70 gene, and that are regulated by SRalpha promoter, were constructed. The mutant-type lacks the area that expresses stress durability. One of the 2 adenovirus vectors was added to the cultures of human chondrocyte-like cells (HCS-2/8). Heat stress (48 degrees C) was applied to the transduced cells for 2 h, and the efficacy of adenovirus vector mediated transduction of HSP70 gene against heat stress in the chondrocytes was investigated using alamar blue assay and MTT assay.
RESULTS
Absorbance levels at 48 degrees C were 300.3 +/- 51.9 and 1.173 +/- 0.011 in the controls, 278.5 + 33.8 and 1.217 +/- 0.018 in the AxSHEmt transduced cells, and 349 +/- 14.7 and 1.371 +/- 0.033 in the AxSHEwt transduced cells. The level in the AxSHEwt transduced cells was significantly higher than in the other 2 groups (p < 0.05). With 37 degrees C treatment, no significant difference was observed.
CONCLUSION
Chondrocytes to which HSP70 gene was transduced had a significantly higher metabolic activity and viability under heat stress.
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