Peng Hongling, Wang Qiao, Qi Xiaorong, Wang Xi, Zhao Xia
Department of Gynecology and Obstetrics, West China Second Hospital, Sichuan University, Chengdu, Sichuan, China.
West China School of Medicine, Sichuan University, Chengdu, Sichuan, China.
Arch Gynecol Obstet. 2018 Sep;298(3):597-605. doi: 10.1007/s00404-018-4841-2. Epub 2018 Jul 4.
Orlistat possesses anti-tumor capacity by inducing apoptosis in ovarian cancer cells. However, the mechanism is not clearly understood. Emerging evidence indicates the overlaps between autophagy and apoptosis. In this study, we have investigated the role of autophagy in orlistat-induced apoptosis in ovarian cancer (OC) cells.
The effect of orlistat on apoptosis was evaluated in SKOV3 and A2780 cell lines by MTT and TUNEL assay. The formations of autophagosomes were observed by acridine orange and GFP-LC3 fluorescence. In addition, conversions of LC3-I to LC3-II were analyzed by western blot, as well as other autophagy-related proteins. 3-Methyladenine (3-MA) was used as an autophagy inhibitor in combined treatment with orlistat. Western blot was further conducted to investigate the molecular mechanisms of orlistat-affected apoptosis and autophagy on protein level.
The proliferation activities of OC cells were inhibited by orlistat in a dose-dependent manner. The expressions of cleaved-caspase 3 and 9 in orlistat-treated cells were increasing, which suggested that orlistat-induced apoptosis was caspase-dependent. At the same time, the average number of GFP-LC3 dots per cell was increased after 48 h of orlistat treatment. The expression levels of LC3-II were significantly up-regulated, as well as other autophagy-related proteins such as Vsp34, Atg7 and UVRAG. These results suggested orlistat-induced autophagy flux, which was further found involved in inhibiting the Akt/mTOR/p70S6K signaling pathway. However, combined treatment of orlistat and 3-MA significantly suppressed the cell viability, which indicated a pro-survival role of autophagy in OC cells.
We suggested that orlistat had anti-cancer effect in OC cells. In addition, autophagy played a pro-survival role, suppressing which the orlistat-induced anti-cancer effect would be more significant.
奥利司他通过诱导卵巢癌细胞凋亡而具有抗肿瘤能力。然而，其机制尚不清楚。新出现的证据表明自噬与凋亡之间存在重叠。在本研究中，我们调查了自噬在奥利司他诱导的卵巢癌（OC）细胞凋亡中的作用。
通过MTT和TUNEL法评估奥利司他对SKOV3和A2780细胞系凋亡的影响。通过吖啶橙和GFP-LC3荧光观察自噬体的形成。此外，通过蛋白质印迹分析LC3-I向LC3-II的转化以及其他自噬相关蛋白。3-甲基腺嘌呤（3-MA）用作自噬抑制剂与奥利司他联合处理。进一步进行蛋白质印迹以研究奥利司他影响凋亡和自噬在蛋白质水平上的分子机制。
奥利司他以剂量依赖性方式抑制OC细胞的增殖活性。奥利司他处理的细胞中裂解的半胱天冬酶3和9的表达增加，这表明奥利司他诱导的凋亡是半胱天冬酶依赖性的。同时，奥利司他处理48小时后，每个细胞中GFP-LC3点的平均数增加。LC3-II的表达水平以及其他自噬相关蛋白如Vsp34、Atg7和UVRAG显著上调。这些结果表明奥利司他诱导自噬流，进一步发现其参与抑制Akt/mTOR/p70S6K信号通路。然而，奥利司他与3-MA联合处理显著抑制细胞活力，这表明自噬在OC细胞中具有促生存作用。
我们认为奥利司他在OC细胞中具有抗癌作用。此外，自噬发挥促生存作用，抑制自噬会使奥利司他诱导的抗癌作用更显著。
