Konstantakopoulos Nicki, Isbister Geoffrey K, Seymour Jamie E, Hodgson Wayne C
Department of Pharmacology, Monash University, Victoria, 3800 Australia.
J Pharmacol Toxicol Methods. 2009 May-Jun;59(3):166-70. doi: 10.1016/j.vascn.2009.02.003. Epub 2009 Feb 28.
Chironex fleckeri is a large box jellyfish that has been labelled the 'most venomous animal' in the world. We have recently shown that the primary effect of C. fleckeri venom in vivo is cardiovascular collapse. This study utilised a cell-based assay to examine the effects of C. fleckeri venom on the proliferation of a rat aortic smooth muscle cell line. In addition, the ability of CSL box jellyfish antivenom and/or various potential treatment strategies to neutralise the effects of the venom was examined.
A7r5 cells were cultured in media containing venom. The effect of CSL box jellyfish antivenom (5 U/mL), CSL polyvalent snake antivenom (5 U/mL), lanthanum (5 microM), MgSO(4) (50 mM), verapamil (5 microM) or felodipine (5 microM) was examined. Cell viability was determined using a Cell titer 96 AQueous One Solution cell proliferation assay.
Incubation of A7r5 cells with serially diluted venom (2-0.004 microg/mL) caused a concentration-dependent inhibition of cell proliferation with an IC(50) value of 0.056 microg/mL. This response was not affected by the absence of calcium or the presence of lanthanum in the media. Box jellyfish antivenom (5 U/mL) prevented the inhibition of cell proliferation caused by the venom. Verapamil (5 microM) had no significant effect on the inhibition. In contrast, felodipine (5 microM) or MgSO(4) (50 mM) potentiated the effects of the venom and partially negated the protective effect of the antivenom.
This study displayed the ability to utilise a cell-based assay to determine the effects of C. fleckeri venom on vascular cell viability. It showed that CSL box jellyfish can neutralise the effects of the venom but only if added prior to the venom. In addition, potential adjunct therapies verapamil, felodipine and MgSO(4) were found to be ineffective, with felodipine and MgSO(4) potentiating the detrimental effects of the venom.
箱形水母是一种大型水母，被称为世界上“最毒的动物”。我们最近发现，箱形水母毒液在体内的主要作用是导致心血管衰竭。本研究利用细胞试验来检测箱形水母毒液对大鼠主动脉平滑肌细胞系增殖的影响。此外，还检测了英联邦血清实验室（CSL）箱形水母抗蛇毒血清和/或各种潜在治疗策略中和毒液作用的能力。
将A7r5细胞培养于含有毒液的培养基中。检测了CSL箱形水母抗蛇毒血清（5 U/mL）、CSL多价蛇抗蛇毒血清（5 U/mL）、镧（5 microM）、硫酸镁（50 mM）、维拉帕米（5 microM）或非洛地平（5 microM）的作用。使用Cell titer 96 AQueous One Solution细胞增殖试验测定细胞活力。
用系列稀释的毒液（2 - 0.004 microg/mL）孵育A7r5细胞会导致细胞增殖受到浓度依赖性抑制，IC50值为0.056 microg/mL。培养基中无钙或存在镧对此反应无影响。箱形水母抗蛇毒血清（5 U/mL）可防止毒液引起的细胞增殖抑制。维拉帕米（5 microM）对这种抑制无显著影响。相比之下非洛地平（5 microM）或硫酸镁（50 mM）增强了毒液的作用，并部分抵消了抗蛇毒血清的保护作用。
本研究展示了利用细胞试验来确定箱形水母毒液对血管细胞活力影响的能力。结果表明，CSL箱形水母抗蛇毒血清可以中和毒液的作用，但前提是要在毒液之前添加。此外，发现潜在的辅助治疗药物维拉帕米、非洛地平和硫酸镁无效，非洛地平和硫酸镁反而增强了毒液的有害作用。
