Department of Laboratory Animal Science, Hebei Medical University, Key Lab of Laboratory Animal Science of Hebei Province, Shijiazhuang 050017, China.
Department of Dermatology, the Second Hospital of Hebei Medical University, Shijiazhuang 050051, Hebei Province, China.
J Ethnopharmacol. 2018 Oct 28;225:169-177. doi: 10.1016/j.jep.2018.06.022. Epub 2018 Jun 19.
Spiders and spider venoms have been used in traditional Chinese medicine to treat various ailments for more than 1000 years. For instance, several large spiders have been utilized by the Li People, who mainly live in Hainan Island of China, in their own unique traditional Chinese medicine therapy. Recent studies have indicated that spider venoms may be an important source of bioactive compounds for anti-tumor treatments. However, the specific mechanisms underlying these activities are not yet completely understood.
The present study investigated how the venom of the spider Haplopelma hainanum regulate proliferation and apoptosis in HepG2 cells via the underlying molecular mechanisms.
We treated HepG2 cells with various concentrations of the spider venom (0, 10, 50, 100 and 200 μg/mL) for 48 h, and then analyzed anti-proliferation activity, apoptosis-inducing effects, mitochondrial membrane potential (Δψm) and changes in the pro-apoptotic pathway. The anti-proliferation activity was detected by the MTT assay and Western blotting. Flow cytometry was used to analyze both apoptosis and mitochondrial membrane potential. The key pro-apoptotic molecules in the caspase-3 and -9 dependent mitochondrial pathway, including Bcl2 family, were assessed through realtime PCR, Western blotting and enzymatic test.
Obvious morphological changes induced by the spider venom included decreased cell numbers, shorter cell length and reduced cell adhesion. MTT and Western blotting demonstrated that the spider venom potently suppressed cell proliferation in a dose- and time-dependent manner with IC of 126.00 μg/mL for 48 h. In addition, the spider venom caused a reduction in the mitochondrial membrane potential and cytochrome c release from mitochondria to cytoplasm under the participation of Bax. Finally, cytochrome c activated caspase-3 and caspase-9, and induced the apoptosis in the HepG2 cells.
The results indicated that the venom of H. hainanum exhibited potent inhibition effects in HepG2 cells through suppressing proliferation, reducing the mitochondrial membrane potential, activating caspase-3 and caspase-9, and inducing the apoptosis through a mitochondrial-dependent pathway.
蜘蛛及其毒液在中国传统医学中已被用于治疗各种疾病超过 1000 年。例如，几种大型蜘蛛已被主要居住在中国海南岛的黎族人民用于他们自己独特的传统中医治疗中。最近的研究表明，蜘蛛毒液可能是抗肿瘤治疗的生物活性化合物的重要来源。然而，这些活性的具体机制尚不完全清楚。
本研究旨在通过潜在的分子机制，研究海南捕鸟蛛毒液如何调节 HepG2 细胞的增殖和凋亡。
我们用不同浓度的蜘蛛毒液（0、10、50、100 和 200μg/ml）处理 HepG2 细胞 48 小时，然后分析抗增殖活性、诱导凋亡作用、线粒体膜电位（Δψm）和促凋亡途径的变化。用 MTT 法和 Western blot 法检测抗增殖活性。用流式细胞术分析凋亡和线粒体膜电位。通过实时 PCR、Western blot 和酶测定法评估 caspase-3 和 caspase-9 依赖的线粒体途径中的关键促凋亡分子，包括 Bcl2 家族。
蜘蛛毒液引起的明显形态变化包括细胞数量减少、细胞长度变短和细胞黏附减少。MTT 和 Western blot 法表明，蜘蛛毒液以剂量和时间依赖的方式强烈抑制细胞增殖，48 小时的 IC 为 126.00μg/ml。此外，蜘蛛毒液在 Bax 的参与下导致线粒体膜电位降低和细胞色素 c 从线粒体释放到细胞质。最后，细胞色素 c 激活 caspase-3 和 caspase-9，诱导 HepG2 细胞凋亡。
结果表明，海南捕鸟蛛毒液通过抑制增殖、降低线粒体膜电位、激活 caspase-3 和 caspase-9，以及通过线粒体依赖性途径诱导凋亡，对 HepG2 细胞表现出强大的抑制作用。
