Chai Lin, Huang Tingting, Wang Zhenjie, Xu Ran, Deng Chao
School of Stomatology, Wannan Medical College 22 West Wenchang Road, Wuhu 241002, Anhui Province, China.
Department of Pathophysiology, Wannan Medical College 22 West Wenchang Road, Wuhu 241002, Anhui Province, China.
Int J Clin Exp Pathol. 2019 Oct 1;12(10):3968-3974. eCollection 2019.
This study aimed to explore the role and possible mechanism of component I from agkistrodon acutus venom (AAVC-I) in promoting the apoptosis of oral squamous cell carcinoma (OSCC). HN4 cells (a human OSCC cell line) were randomly divided into four experimental groups: low AAVC-I (2.5 μg/mL) group, medium AAVC-I (5 μg/mL) group and high AAVC-I (10 μg/mL) group, as well as control group (AAVC-I, 0 μg/mL). AVVC-I was dissolved in RPMI-1640 medium and added to the culture wells at different concentrations when tumor cells had reached the logarithmic growth phase. After 24 hours, cells were harvested and the inhibitory rate of cell proliferation and the mitochondrial membrane depolarization were measured. Western blotting was used to detect the expression levels of cytochrome c, Bcl-2 associated X protein (Bax), and caspase-3 in the cellular cytoplasm either containing mitochondria or following the removal of mitochondria. Cellular apoptosis was detected by flow cytometry. Compared to the control group, AAVC-I treatment not only inhibited NH4 growth, but also upgraded the expression of caspase-3 in NH4 cells. Meanwhile, it was observed that Bax translocation to mitochondria and cytochrome c release into the cytosol increased in AAVC-I treatment. This indicated that AAVC-I could disrupt mitochondrial membrane depolarization and result in cellular apoptosis, and the apoptosis rate of NH4 increased with the concentration of AAVC-I. The data suggested that AAVC-I promotes the apoptosis of HN4 cells through the mitochondrial pathway in a dose-dependent manner, which provides experimental data and new ideas for future research and clinical treatment options for OSCC.
本研究旨在探讨尖吻蝮蛇毒成分I（AAVC-I）在促进口腔鳞状细胞癌（OSCC）细胞凋亡中的作用及可能机制。将HN4细胞（一种人OSCC细胞系）随机分为四个实验组：低剂量AAVC-I（2.5 μg/mL）组、中剂量AAVC-I（5 μg/mL）组、高剂量AAVC-I（10 μg/mL）组以及对照组（AAVC-I，0 μg/mL）。当肿瘤细胞达到对数生长期时，将AAVC-I溶解于RPMI-1640培养基中，并以不同浓度加入培养孔。24小时后，收集细胞并检测细胞增殖抑制率和线粒体膜去极化情况。采用蛋白质免疫印迹法检测含有线粒体的细胞质或去除线粒体后的细胞质中细胞色素c、Bcl-2相关X蛋白（Bax）和半胱天冬酶-3的表达水平。通过流式细胞术检测细胞凋亡情况。与对照组相比，AAVC-I处理不仅抑制了HN4细胞的生长，还提高了HN4细胞中半胱天冬酶-3的表达。同时，观察到在AAVC-I处理下，Bax向线粒体的转位以及细胞色素c释放到细胞质中的情况增加。这表明AAVC-I可破坏线粒体膜去极化并导致细胞凋亡，且HN4细胞的凋亡率随AAVC-I浓度的增加而升高。数据表明，AAVC-I通过线粒体途径以剂量依赖的方式促进HN4细胞凋亡，为OSCC未来的研究和临床治疗方案提供了实验数据和新思路。
