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【目的】观察补阳还五汤对曲妥珠单抗导致的心肌细胞H9C2活性抑制及SHP-1活性的影响,通过转染SHP-1和SHPC/S-1质粒后观察补阳还五汤的干预效果,从而探讨其调控机制。【方法】构建真核表达载体pc DNA3.1(+)-SHP-1和pc DNA3.1(+)-SHPC/S-1,利用脂质体转染法转染大鼠心肌H9C2细胞,经G418筛选阳性克隆,阳性克隆细胞在曲妥株单抗中维持生长。采用定量RT-PCR法检测SHP-1基因及SHPC/S-1基因在大鼠心肌H9C2细胞中的稳定表达情况,采用磷酸酶活性分析补阳还五汤对心肌细胞中SHP-1的调控作用,并通过四甲基偶氮唑盐(MTT)法进一步观察加入补阳还五汤后细胞的凋亡情况。【结果】测序证明真核表达载体构建成功,并且可以在曲妥珠单抗干预下的心肌细胞H9C2中稳定表达,磷酸酶活性检测证实磷酸酶活性最高的是H9C2-SHP-1细胞组,经补阳还五汤干预后活性显著下降(P<0.05);MTT法观察进一步证实加入补阳还五汤后的H9C2-SHP-1细胞的凋亡率较未加补阳还五汤组的细胞显著降低(P<0.05)。【结论】补阳还五汤可以改善曲妥珠单抗对心肌细胞的抑制作用,并可影响心肌细胞中SHP-1基因的表达,为以后进一步探讨曲妥珠单抗所致的心脏毒性的治疗打下基础。
【Objective】 To observe the effect of Buyang Huanwu decoction on the inhibition of H9C2 activity and the activity of SHP-1 in cardiomyocytes induced by trastuzumab, observe the effects of Buyang Huanwu Decoction The effect of intervention, to explore its regulatory mechanism. 【Method】 The eukaryotic expression vector pcDNA3.1 (+) - SHP-1 and pcDNA3.1 (+) - SHPC / S-1 were constructed and transfected into H9C2 cells by lipofection. Positive clones were screened by G418 and positive clone cells were maintained in the trastuzumab McAb. The stable expression of SHP-1 gene and SHPC / S-1 gene in H9C2 cells was detected by quantitative RT-PCR. The regulation of SHP-1 in cardiomyocytes was analyzed by using phosphatase activity , And the cell apoptosis was further observed by MTT assay. 【Results】 Sequencing proved that the eukaryotic expression vector was successfully constructed and stably expressed in cardiomyocytes H9C2 induced by trastuzumab. H9C2-SHP-1 cells were identified by phosphatase activity assay. The activity of Buyang Huanwu Decoction significantly decreased after intervention (P <0.05). The MTT assay further confirmed that the apoptosis rate of H9C2-SHP-1 cells treated with Buyang Huanwu Decoction was higher than that of the cells treated with Buyang Huanwu Decoction Significantly lower (P <0.05). 【Conclusion】 Buyang Huanwu Decoction can improve the inhibitory effect of trastuzumab on cardiomyocytes and the expression of SHP-1 gene in cardiomyocytes, so as to further explore the cardiotoxicity of trastuzumab Treatment laid the foundation.