Orbeli Institute of Physiology, Orbely str. 22, Yerevan 0028, Armenia.
Toxins (Basel). 2022 Oct 23;14(11):724. doi: 10.3390/toxins14110724.
Increasing concern about the use of animal models has stimulated the development of in vitro cell culture models for analysis of the biological effects of snake venoms. However, the complexity of animal venoms and the extreme synergy of the venom components during envenomation calls for critical review and analysis. The epithelium is a primary target for injected viper venom's toxic substances, and therefore, is a focus in modern toxinology. We used the Vero epithelial cell line as a model to compare the actions of a crude (Levantine viper) venom with the actions of the same venom with two key enzymatic components inhibited (specifically, phospholipase A2 (PLA2) and metalloproteinases) in the bioenergetic cellular response, i.e., oxygen uptake and reactive oxygen species generation. In addition to the rate of free-radical oxidation and lipid peroxidation, we measured real-time mitochondrial respiration (based on the oxygen consumption rate) and glycolysis (based on the extracellular acidification rate) using a Seahorse analyzer. Our data show that viper venom drives an increase in both glycolysis and respiration in Vero cells, while the blockage of PLA2 or/and metalloproteinases affects only the rates of the oxidative phosphorylation. PLA2-blocking in venom also increases cytotoxic activity and the overproduction of reactive oxygen species. These data show that certain components of the venom may have a different effect within the venom cocktail other than the purified enzymes due to the synergy of the venom components.
人们对动物模型使用的担忧日益增加，这刺激了用于分析蛇毒生物学效应的体外细胞培养模型的发展。然而，动物毒液的复杂性以及毒液成分在注毒时的极端协同作用，需要进行批判性的审查和分析。上皮组织是注射性毒蛇毒液中有毒物质的主要靶标，因此也是现代毒素学的研究重点。我们使用 Vero 上皮细胞系作为模型，比较了一种粗制（Levantine 蝰蛇）毒液与同一种毒液的作用，该毒液的两种关键酶成分（即磷脂酶 A2 (PLA2) 和金属蛋白酶）受到抑制，作用于细胞的生物能量反应，即氧摄取和活性氧的生成。除了自由基氧化和脂质过氧化的速率外，我们还使用 Seahorse 分析仪测量了实时线粒体呼吸（基于耗氧量）和糖酵解（基于细胞外酸化率）。我们的数据表明，蝰蛇毒液可促进 Vero 细胞中糖酵解和呼吸作用的增加，而 PLA2 或/和金属蛋白酶的阻断仅影响氧化磷酸化的速率。在毒液中阻断 PLA2 还会增加细胞毒性活性和活性氧的过度产生。这些数据表明，由于毒液成分的协同作用，毒液混合物中的某些成分可能会产生不同于纯化酶的效果。
