内质网应激介导的自噬反应在β,β-二甲基丙烯酰紫草素诱导人肺腺癌细胞凋亡中的作用及其机制研究

β,β-Dimethylacrylshikonin (DMAS) is a major component of Lithospermum erythrorhizon. To date, the inhibitory effects of DMAS on human lung adenocarcinoma has not been reported. Our previous findings had shown that DMAS could inhibit the proliferation and induce the apoptosis of A549 cells, and up-regulate the expression of GRP78、CHOP and LC3-II. Taken together, our results suggested that endoplasmic reticulum stress (ERS) and autophagy may be involved in DMAS-induced apoptosis in A549 cells. According to the literature, autophagy and apoptosis interrelate, ERS can induce an autophagic response. Therefore, we postulate that autophagy mediated by ERS may be involved in DMAS-induced apoptosis in lung adenocarcinoma cells. To determine the role of autophagy in DMAS-induced apoptosis in human lung adenocarcinoma cells, we intend to interfere the autophagic activity by using autophagy inhibitor, or silencing Atg5 and Atg7, and then observe the changes of autophagy and apoptosis. To further explore the effect and molecular mechanism of ERS response on DMAS-induced autophagy, we prepare to observe the changes of autophagy and ERS after treatment with ERS protective agent, or silencing ERS-related gene. By revealing the crosstalk between ERS, autophagy and apoptosis, we provide evidence that DMAS could be developed as a potential therapeutic agent for the treatment of lung adenocarcinoma.

β,β-二甲基丙烯酰紫草素（β,β-Dimethylacrylshikonin，DMAS）作为紫草素的重要衍生物，主要发挥抗肿瘤作用，但未见其抗肺腺癌作用机制的研究报道。我们前期的实验发现DMAS不仅可抑制A549细胞的增殖并诱导其凋亡，还能增加内质网应激相关蛋白GRP78、CHOP以及自噬体标志蛋白LC3-II的表达，提示内质网应激和自噬可能参与了DMAS诱导人肺腺癌A549细胞的凋亡。根据文献报道，自噬和凋亡相互关联，内质网应激可以激活自噬。因此，我们推测内质网应激介导的自噬参与了DMAS诱导的人肺腺癌细胞的凋亡。为了证实这一假说，我们拟利用细胞和裸鼠模型为研究对象，应用小分子抑制剂、siRNA等方法观察自噬在DMAS诱导人肺腺癌细胞凋亡中的作用以及内质网应激相关信号通路在自噬激活中的作用。本项目通过揭示内质网应激、自噬和凋亡之间的内在联系，为DMAS治疗肺腺癌提供新的思路和靶点。

β,β-二甲基丙烯酰紫草素（β,β-Dimethylacrylshikonin，DMAS）是紫草素的一种衍生物，主要发挥抗肿瘤作用。在本课题经费的资助下，依据项目计划书的内容，我们对内质网应激介导的自噬参与了DMAS诱导的人肺腺癌细胞的凋亡作用进行了研究。我们的研究发现DMAS在体外可显著抑制人肺腺癌细胞增殖，并且能够诱导其发生凋亡，且这种凋亡依赖于线粒体介导的caspase家族的激活。进一步的研究发现DMAS可诱导人肺腺癌细胞发生自噬，且自噬是作为一种保护性应激反应来对抗细胞凋亡；单用DMAS可部分抑制裸鼠皮下移植瘤生长，而联合自噬阻滞剂CQ后可显著抑制移植瘤生长并促进其凋亡。最后，我们的研究还证实内质网应激反应参与了DMAS诱导的人肺腺癌细胞自噬；DMAS诱导人肺腺癌细胞发生自噬依赖于内质网应激激活的PERK- eIF2α-ATF4-CHOP通路、IRE1-TRAF2-JNK通路以及Ca2+相关信号通路。本研究揭示了内质网应激、自噬和凋亡三者之间的关系，将为中药单体及联合用药治疗肺腺癌提供新的思路和治疗靶点。