Axelsson V, Holback S, Sjögren M, Gustafsson H, Forsby A
Department of Neurochemistry, The Arrhenius Laboratories for Natural Sciences, Stockholm University, Sweden.
Biochem Biophys Res Commun. 2006 Jul 7;345(3):1068-74. doi: 10.1016/j.bbrc.2006.05.019. Epub 2006 May 11.
In this study, a significant increase by 50% in intracellular free calcium concentration (Ca(2+)) was observed in differentiated human neuroblastoma (SH-SY5Y) cells after exposure to 0.25microM of the fungal metabolite gliotoxin for 72h. Further, the involvement of caspases and calpains was demonstrated to underlie the gliotoxin-induced cytotoxic and neurite degenerative effects. The caspase inhibitor Z-VAD-fmk almost completely reduced the neurite degeneration from 40% degeneration of neurites to 5% as compared to control. Inhibition of calpains with calpeptin significantly attenuated gliotoxin-induced cytotoxicity, determined as reduction in total cellular protein content, from 43% to 14% as compared to control cells. Western blot analyses of alphaII-spectrin breakdown fragments confirmed activity of the proteases, and that alphaII-spectrin was cleaved by caspases in gliotoxin-exposed cells. These results show that calpains and caspases have a role in the toxicity of gliotoxin in differentiated SH-SY5Y cells and that the process may be Ca(2+)-mediated.
在本研究中，分化的人神经母细胞瘤（SH-SY5Y）细胞在暴露于0.25微摩尔的真菌代谢产物gliotoxin 72小时后，细胞内游离钙浓度（Ca(2+)）显著增加了50%。此外，已证明半胱天冬酶和钙蛋白酶的参与是gliotoxin诱导的细胞毒性和神经突退行性变效应的基础。与对照组相比，半胱天冬酶抑制剂Z-VAD-fmk几乎完全将神经突退行性变从神经突40%的退行性变降低到5%。用钙蛋白酶抑制剂抑制钙蛋白酶可显著减轻gliotoxin诱导的细胞毒性，以总细胞蛋白含量的降低来衡量，与对照细胞相比，从43%降至14%。对αII-血影蛋白降解片段的蛋白质印迹分析证实了蛋白酶的活性，并且在暴露于gliotoxin的细胞中，αII-血影蛋白被半胱天冬酶切割。这些结果表明，钙蛋白酶和半胱天冬酶在分化的SH-SY5Y细胞中gliotoxin的毒性作用中起作用，并且该过程可能是由Ca(2+)介导的。
