Ophiopogonin B (OP-B), a saponin compound isolated from Radix Ophiopogon japonicus, was verified to inhibit cell proliferation in numerous non-small cell lung cancer (NSCLC) cells in our previous study. However, the precise mechanisms of action have remained unclear. In the present study, we mainly investigated the effects of OP-B on adenocarcinoma A549 cells to further elaborate the underlying mechanisms of OP-B in different NSCLC cell lines. Detection by high content screening (HCS) and TUNEL assay verified that OP-B induced apoptosis in this cell line, while detection of Caspase-3, Bcl-2 and Bax showed that OP-B induced cell death was caspase and mitochondrial independent. Further experiments showed that OP-B induced cell cycle arrest in the S and G2/M phases by inhibiting the expression of Myt1 and phosphorylation of Histone H3 (Ser10), which resulted in mitotic catastrophe in the cells. Transmission electron microscopy (TEM) observation of cell micro-morphology combined with detection of Atgs by western blot analysis showed that OP-B induced autophagy in this cell line. Autophagy inhibition by the lysosome inhibitor CQ or Beclin1-siRNA knockdown both attenuated cell viability, demonstrated that autophagy also being the vital reason resulted in cell death. More importantly, the xenograft model using A549 cells provided further evidence of the inhibition of OP-B on tumor proliferation. Immunohistochemistry detection of LC3 and Tunel assay both verified that high dose of OP-B (75 mg/kg) induced autophagy and apoptosis in vivo, and western blot detection of p-Histone H3 (Ser10), Survivin and XIAP further indicated the molecular mechanism of OP-B in vivo. As our findings revealed, multiple types of cell death overlapped in OP-B treated A549 cells, it displayed multitarget characteristics of the compounds extracted from the Chinese herbal, which may be used as candidate anticancer medicine in clinic.

中文翻译：

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麦冬B诱导A549细胞凋亡，有丝分裂灾难和自噬。

麦冬B（OP-B）是从麦冬中分离得到的一种皂苷化合物，在我们先前的研究中已证实可抑制许多非小细胞肺癌（NSCLC）细胞中的细胞增殖。但是，确切的作用机制仍不清楚。在本研究中，我们主要研究OP-B对腺癌A549细胞的作用，以进一步阐明OP-B在不同NSCLC细胞系中的潜在机制。通过高含量筛选（HCS）和TUNEL分析检测证实OP-B诱导了该细胞系的凋亡，而对Caspase-3，Bcl-2和Bax的检测表明OP-B诱导的细胞死亡与caspase和线粒体无关。进一步的实验表明，OP-B通过抑制Myt1的表达和组蛋白H3（Ser10）的磷酸化来诱导S和G2 / M期的细胞周期停滞，导致细胞中的有丝分裂灾难。透射电子显微镜（TEM）观察细胞的微观形态，并通过Western印迹分析检测Atgs，表明OP-B诱导了该细胞系的自噬。溶酶体抑制剂CQ或Beclin1-siRNA敲低的自噬抑制作用均减弱了细胞活力，证明自噬也是导致细胞死亡的重要原因。更重要的是，使用A549细胞的异种移植模型提供了OP-B对肿瘤增殖的抑制作用的进一步证据。LC3的免疫组织化学检测和Tunel分析均证实高剂量的OP-B（75 mg / kg）在体内可引起自噬和细胞凋亡，p-Histone H3（Ser10），Survivin和XIAP的蛋白质印迹检测进一步表明了分子机制体内OP-B的作用。正如我们的发现所揭示的那样，