[Triptolide reverses apatinib resistance in gastric cancer cell line MKN45 via inhibition of heat shock protein 70].

Objective: To investigate the effect of triptolide, a specific inhibitor of heat shock protein 70 (HSP70), on apatinib resistance in gastric cancer cells line MKN45. Methods: The apatinib-resistant cells (MKN45/AR) and MKN45 parental cells were treated with apatinib, triptolide and apatinib combined with triptolide, respectively. CCK-8 assay was performed to determine the half maximal inhibitory concentration (IC(50)) of MKN45/AR and MKN45 cells in the presence of different treatment. The mRNA expression of heat shock protein gene (HSPA1A and HSPA1B) was detected by RT-PCR, while the protein expression of heat shock protein 70 was analyzed using Western blot in MKN45/AR and MKN45 cells. Results: The IC(50) values of apatinib-sensitive and apatinib-resistant MKN45 cells were 10.411 μmol/L and 70.527 μmol/L, respectively, showing a significant difference (P<0.05). The mRNA expression of HSPA1A and HSPA1B in MKN45/AR cells was significantly higher than that in MKN45 cells (P<0.001). The protein expression of heat shock protein 70 was significantly decreased after 0.25 μmol/L triptolide treatment in MKN45/AR cells (P<0.01). When heat shock protein 70 was inhibited by triptolide, the IC(50) value of apatinib in MKN45/AR cells was reduced to 11.679 μmol/L, which was significantly lower than cells treated with apatinib alone (P<0.05). Conclusions: The apatinib-resistant MKN45 cells have high levels of heat shock protein 70. Low doses of triptolide can significantly inhibit heat shock protein 70, leading to reverse the resistance phenotype of MKN45/AR cells. Therefore, inhibition of heat shock protein 70 provides a new therapy strategy for patients with apatinib resistance.目的： 探讨热休克蛋白70(HSP70)特异性抑制剂雷公藤内酯逆转胃癌细胞MKN45阿帕替尼耐药的作用。 方法： 以阿帕替尼、雷公藤内酯、阿帕替尼联合雷公藤内酯处理胃癌耐药细胞MKN45/AR及胃癌敏感细胞MKN45，采用CCK－8实验检测半数抑制浓度(IC(50))，比较阿帕替尼单独及联合雷公藤内酯对MKN45和MKN45/AR细胞活性的影响。采用逆转录聚合酶链反应(RT－PCR)技术检测MKN45细胞耐药前后的HSP70基因(HSPA1A和HSPA1B)的mRNA表达情况，采用Western blot法检测HSP70的蛋白表达水平。 结果： 阿帕替尼对MKN45细胞的IC(50)为10.411 μmol/L，对MKN45/AR细胞的IC(50)为70.527 μmol/L，两者差异有统计学意义(P<0.05)。HSPA1A mRNA和HSPA1B mRNA在MKN45/AR细胞中的表达水平远高于MKN45细胞(P<0.001)。0.25 μmol/L雷公藤内酯作用于MKN45/AR细胞后，HSP70的表达水平明显下降(P<0.01)，阿帕替尼对MKN45/AR细胞的IC(50)降为11.679 μmol/L，与单纯使用阿帕替尼时相比，差异有统计学意义(P<0.05)。 结论： 在阿帕替尼耐药的胃癌细胞MKN45/AR中，HSP70表达明显升高。低剂量的雷公藤内酯可以明显抑制MKN45/AR细胞中HSP70的表达，MKN45/AR细胞对阿帕替尼的耐药性明显下降，抑制HSP70的表达是克服阿帕替尼耐药性的方法之一。.