Inhibitory effect of heat shock proteins in apoptosis in acquisition of anticancer drug resistance

根据我们之前的研究，热休克蛋白（HSPs）在抗肿瘤药物的耐药获得中起着重要作用。本研究旨在通过RT-PCR和gel-shift两种方法阐明热休克蛋白的表达调控机制以及热休克蛋白在细胞凋亡通路中的作用。此外，还检测了HSPs和HSE的反义寡聚探针，以估计克服耐药性的效果。无论热休克蛋白的表达如何分化，耐药细胞系中HSE的水平在每个细胞系中都没有改变。这一发现表明HSF而不是HSE在HSPs的表达中起作用，尽管在本研究中无法得出确切的机制。为了评估反义寡聚探针对HSP或HSE的克服作用，合成的三种反义寡聚探针，尤其是对HSP27的反义寡聚探针，对耐药表现出明显的克服作用。HSP27反义寡聚探针抑制了HSP27 mRNA的表达，部分克服了HSP27的耐药。有趣的是，该探针可以克服对TGF-β和各种抗癌药物耐药的HACC-TG细胞株的TGF-β耐药和抗癌耐药。进一步将该探针与脂肪酸snythetase抑制剂cerulenin结合，提示HSP27在脂质合成中起一定作用。另外两种均能抑制热休克蛋白mRNA的表达，但对耐药缺乏克服作用。转染各种热休克蛋白基因后，细胞凋亡相关基因均未被诱导。然而，与亲本细胞相比，转染HSP70或HSP27基因的细胞中baxmRNA减少。然而，在细胞凋亡途径中，热休克蛋白的确切靶点尚不清楚。

Based on the our previous research, heat shock proteins (HSPs) play a role in acquisition of drug resistance, regardless of spieces of anticancer drugs. The aim of this research is to elucidate the regulation mechanism of expression of HSPs and the role of HSPs in apoptosis pathway by RT-PCR and gel-shift methods. Furthermore anti-sense oligoprobes to HSPs and HSE have been examined to estimate the effect of overcome of drug resistance. The levels of HSE in drug resistance cell lines have not been altered in each cell line, regardless of differentiated expression of HSPs. This finding suggests that HSF but not HSE plays a role in expression of HSPs, although the exact mechanism can not be elicited in this study. To estimate the overcome effect of anti-sense oligoprobe for HSP or HSE, three of synthesized anti-sense oligoprobes, especially anti-sense oligoprobe to HSP27, showed an evident overcome effect on drug resistance. The anti-sense oligoprobe to HSP27 inhibited the expression of mRNA of HSP27 and partilly overcame the drug resistance. Interestingly this probe could overcome the TGF-β resistance as well as anticancer resistance in HACC-TG cell line which was resistant to TGF-β as well as various anti-cancer drugs. Furthermore combination with this probe and cerulenin which was an inhibitor of fatty acid snythetase, suggesting some roles of HSP27 in lipid synthesis. The other two could inhibit of expression of mRNA of HSPs but lacked the overcome effect on drug resistance. Transfection of various HSP genes failed to induce genes associated with apoptosis. However, baxmRNA was diminished in the cells transfected with HSP70 or HSP27 genes, compared with that of the parental cells. However, the exact target points of HSPs can not be elucidated in apoptosis pathway.