Zhao Xin-Yue, Tian Ying-Ying, Liu Chuang, Li Yi-Lin, Lyu Ying-Nan, Yu Shang-Yue, Tian Shi-Qiu, Pei Hai-Luan, Zuo Ze-Ping, Wang Zhi-Bin
School of Chinese Materia Medica,Beijing University of Chinese Medicine Beijing 102488,China.
Beijing Tongrentang Technologies Co.,Ltd.,Pharmaceutical Factory Beijing 100079,China.
Zhongguo Zhong Yao Za Zhi. 2023 Aug;48(16):4475-4482. doi: 10.19540/j.cnki.cjcmm.20230413.704.
This study investigated the effect and mechanism of morin in inducing autophagy and apoptosis in hepatocellular carcinoma cells through the protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/signal transducer and activator of transcription protein 3(STAT3) pathway. Human hepatocellular carcinoma SK-HEP-1 cells were stimulated with different concentrations of morin(0, 50, 100, 125, 200, and 250 μmol·L(-1)). The effect of morin on the viability of SK-HEP-1 cells was detected by Cell Counting Kit-8(CCK-8). The effect of morin on the proliferation and apoptosis of SK-HEP-1 cells was investigated using colony formation assay, flow cytometry, and BeyoClick(TM) EdU-488 with different concentrations of morin(0, 125, and 250 μmol·L~(-1)). The changes in the autophagy level of cells treated with morin were examined by transmission electron microscopy and autophagy inhibitors. The impact of morin on the expression levels of proteins related to the Akt/mTOR/STAT3 pathway was verified by Western blot. Compared with the control group, the morin groups showed decreased viability of SK-HEP-1 cells in a time-and concentration-dependent manner, increased number of apoptotic cells, up-regulated expression level of apoptosis marker PARP, up-regulated phosphorylation level of apoptosis-regulating protein H2AX, decreased number of positive cells and the colony formation rate, an upward trend of expression levels of autophagy-related proteins LC3-Ⅱ, Atg5, and Atg7, and decreased phosphorylation levels of Akt, mTOR, and STAT3. These results suggest that morin can promote apoptosis, inhibit proliferation, and induce autophagy in hepatocellular carcinoma cells, and its mechanism of action may be related to the Akt/mTOR/STAT3 pathway.
本研究通过蛋白激酶B（Akt）/雷帕霉素哺乳动物靶蛋白（mTOR）/信号转导子与转录激活子3（STAT3）通路，探讨了桑色素诱导肝癌细胞自噬和凋亡的作用及机制。用不同浓度的桑色素（0、50、100、125、200和250 μmol·L⁻¹）刺激人肝癌SK-HEP-1细胞。采用细胞计数试剂盒-8（CCK-8）检测桑色素对SK-HEP-1细胞活力的影响。使用集落形成试验、流式细胞术以及不同浓度（0、125和250 μmol·L⁻¹）的桑色素处理后的BeyoClick™ EdU-488，研究桑色素对SK-HEP-1细胞增殖和凋亡的影响。通过透射电子显微镜和自噬抑制剂检测桑色素处理后细胞自噬水平的变化。通过蛋白质印迹法验证桑色素对Akt/mTOR/STAT3通路相关蛋白表达水平的影响。与对照组相比，桑色素处理组的SK-HEP-1细胞活力呈时间和浓度依赖性降低，凋亡细胞数量增加，凋亡标志物PARP的表达水平上调，凋亡调节蛋白H2AX的磷酸化水平上调，阳性细胞数量和集落形成率降低，自噬相关蛋白LC3-Ⅱ、Atg5和Atg7的表达水平呈上升趋势，Akt、mTOR和STAT3的磷酸化水平降低。这些结果表明，桑色素可促进肝癌细胞凋亡、抑制增殖并诱导自噬，其作用机制可能与Akt/mTOR/STAT3通路有关。
