Instituto de Medicina Experimental, Facultad de Medicina, Universidad Central de Venezuela, Caracas, Venezuela.
Toxicol In Vitro. 2011 Feb;25(1):358-67. doi: 10.1016/j.tiv.2010.10.009. Epub 2010 Oct 20.
Acute lung injury following envenomation by Tityus scorpion species is due in part to activation of the inflammatory response leading to release of cytotoxic leukocyte-derived products, including cytokines and possibly reactive oxygen species (ROS). Tityus zulianus envenomation in Venezuela produces cardiorespiratory complications and death by lung injury whereas stings by Tityus discrepans produce mainly gastrointestinal and pancreatic alterations. To ascertain the role played by granulocytes in the envenomation by T. zulianus (TzV) and T. discrepans (TdV), human peripheral blood neutrophils, eosinophils, and monocytes were exposed to scorpion venoms (0.001-5 μg/mL) and the kinetics (5-15 min) of peroxide production determined by flow cytometry, using 2',7'-dichlorodihydrofluorescein diacetate (succinimidyl ester) as a fluorescent substrate. TzV induced a significantly (p<0.01) more potent increase in peroxide production in neutrophils (for 5 and 10 min of incubation), and to a lesser extent in monocytes (5-15 min), compared to TdV. TzV induced necrosis in neutrophils at doses higher than 5 μg/mL. No effect was observed on eosinophils, suggesting that TzV specifically targets neutrophil intracellular ROS production. The TzV-stimulated pathway is protein kinase C-dependent because it was almost completely (>90%) abolished by staurosporine. The stimulatory effect is associated with the lowest molecular mass venom peptides as gel filtration fractions TzII and TzIII significantly enhanced peroxide production. The combined used of the intracellular ROS agonist, phorbol myristate acetate (PMA), and TzV produced a modest but significant increase in peroxide production suggesting the possibility of overlapping signaling cascades amongst PMA and TzV. Up-regulation of intracellular neutrophil ROS production may be an important in vivo target for TzV which could have a role to play in the cardiorespiratory complications elicited after envenomation by this species.
被 Tityus 蝎物种螫伤后会引起急性肺损伤，部分原因是炎症反应的激活导致细胞毒性白细胞衍生产物的释放，包括细胞因子和可能的活性氧（ROS）。委内瑞拉的 Tityus zulianus 螫伤会导致心肺并发症和肺损伤导致的死亡，而 Tityus discrepans 的螫伤主要产生胃肠道和胰腺改变。为了确定粒细胞在 T. zulianus（TzV）和 T. discrepans（TdV）螫伤中的作用，将人外周血中性粒细胞、嗜酸性粒细胞和单核细胞暴露于蝎毒液（0.001-5 μg/mL），并用 2'，7'-二氯二氢荧光素二乙酸酯（琥珀酰亚胺酯）作为荧光底物通过流式细胞术测定过氧化物的产生动力学（5-15 分钟）。TzV 诱导中性粒细胞（孵育 5 和 10 分钟）和单核细胞（5-15 分钟）中过氧化物产生的显著（p<0.01）更强烈增加，与 TdV 相比。TzV 在高于 5 μg/mL 的剂量下诱导中性粒细胞坏死。对嗜酸性粒细胞没有观察到影响，表明 TzV 特异性靶向中性粒细胞内 ROS 产生。TzV 刺激途径依赖蛋白激酶 C，因为它几乎完全（>90%）被 staurosporine 消除。刺激作用与最低分子量的毒液肽相关，因为凝胶过滤级分 TzII 和 TzIII 显著增强过氧化物的产生。细胞内 ROS 激动剂佛波醇肉豆蔻酸酯（PMA）和 TzV 的联合使用可适度但显著增加过氧化物的产生，表明 PMA 和 TzV 之间可能存在重叠的信号级联。TzV 上调中性粒细胞内 ROS 的产生可能是体内的一个重要靶点，这可能在该物种的螫伤后引起的心肺并发症中发挥作用。
