益气化瘀补肾方及其拆方调控大鼠颈椎间盘基因表达谱的研究

Study of Yiqi Huayu Bushen Recipe and its decomposed formulas in regulating gene expressions in degenerated cervical intervertebral discs of rats

目的建立大鼠颈椎间盘退变模型,观察益气化瘀补肾方及其拆方调控退变大鼠颈椎间盘模型基因表达谱的变化。方法建立动静力失衡性大鼠颈椎间盘退变模型,从对照组和实验组大鼠颈椎间盘中抽提mRNA,经反转录获得大鼠椎间盘cDNA探针;cDNA探针与大鼠基因表达谱芯片杂交,由激光扫描仪得到表达谱图片,然后进行图像分析、标准化处理、ratio值分析、聚类分析。最后得出这些基因在对照组和实验组间的表达差异。结果聚类分析结果(1)1号(益气化瘀组)、2号(益气补肾组)、3号(化瘀补肾组)芯片为一类;(2)4号(模型组)和5号(益气化瘀补肾组)芯片为一类。4号芯片与其他芯片的差异最大,2号和3号结果最接近。3例以上差异表达的基因共有96条,其中77条是已知基因。已知基因中48条表达上调(ratio>1.0);29条表达下调(ratio<0.5)。其中25条基因在模型组与中药组间表达存在差异。结论退变大鼠颈椎间盘基因的表达发生了改变,益气化瘀补肾方及其拆方可调控相关基因的表达,如上调细胞内信号传导类基因PI3K、PTK、ERK3、PH1B1等的表达。

Objective： To investigate the gene expression changes in the degenerated cervical intervertebral discs of rats, and to study the function of Yiqi Huayu Bushen Recipe, a compound Chinese herbal medicine, and its discomposed formulas in regulating gene expressions in the degenerated cervical intervertebral discs. Methods. The rat model with degenerated cervical intervertebral discs caused by imbalance between the dynamic and static forces was established. The mRNA was extracted from the cervical intervertebral discs of rats in the normal control and experiment groups, and the cDNA probes were obtained by inverse transcript. The cDNA probes were hybridized with the gene chips. The gene expression pattern was gained with a laser scanner. Image analysis, standardized ratio value and cluster analysis were used to investigate the differential of gene expressions between the control and experiment groups. Results： Cluster analysis showed that the gene chips of No. 1 （Yiqi Huayu group）, No. 2 （Yiqi Bushen group） and No. 3 （Huayu Bushen group） were in one class, while the gene chips of No. 4 （untreated group） and No. 5 （Yiqi Huayu Bushen group） were in the other. The gene expression of No. 4 was different from the others mostly, and the gene expressions of No. 2 and No. 3 were similar. There were 96 genes expressed differently in three cases and among them 77 genes were already known and the expression of 48 genes were up-regulated （ratio 〉1. 0）, and 29 down-regulated （ratio〈 0, 5）. There were 25 genes expressed differently between the untreated group and the herb-treated groups. Conclusions. The gene expressions of the degenerated rat intervertebral discs are changed. Yiqi Huayu Bushen Recipe and its discomposed formulas have the effect of regulating the expressions of related genes, such as PI3K, PTK, ERK3, and PH1B1.