Mendes Vanessa A, Barbaro Katia C, Sciani Juliana M, Vassão Ruth C, Pimenta Daniel C, Jared Carlos, Antoniazzi Marta M
Laboratory of Cell Biology, Butantan Institute, Av. Vital Brasil 1500, 05503-000, São Paulo, SP, Brazil.
Laboratory of Immunopathology, Butantan Institute, Av. Vital Brasil 1500, 05503-000, São Paulo, Brazil.
Toxicon. 2016 Nov;122:133-141. doi: 10.1016/j.toxicon.2016.10.004. Epub 2016 Oct 5.
Corythomantis greeningi is a tree-frog endemic of the Brazilian semi-arid (Caatinga), mainly characterized by the flat, mineralized and spiny head, which is associated with phragmotic habits. It is already known that the skin secretion of this amphibian from both head and body is quite toxic and is used as an efficient chemical defence against predators. However, the biochemical characteristics and pharmacological effects of this secretion are still very little studied. We have tested the crude skin secretion, as well as the ten major fractions obtained by RP-HPLC for nociceptive and edema activity and for in vitro cytotoxicity using murine models. SDS-PAGE analyses demonstrated that the majority of proteins ranging through the gel lie between 55 and 30 kDa. LC-MS analysis showed multiple low molecular mass molecules (200-500 Da), which are consistent with masses of alkaloids and steroids. The crude skin secretion was able to induce fast and persistent edema accompanied by intense dose-dependent nociception. From the 10 tested fractions, five induced both edema and nociception, six fractions were able to induce only edema (80-170% control), and seven fractions induced only nociception (15-30 times compared to control). In addition, inhibition of cell growth (IC50) was demonstrated in murine fibroblasts and melanoma cells. From the data obtained, we confirmed that the skin secretion of C. greeningi is very toxic and is rich in compounds able to directly provoke local inflammation and nociception. Such characteristics are important as part of the chemical defensive repertory of this species.
格林氏盔头雨蛙是一种巴西半干旱地区（卡廷加）特有的树蛙，主要特征是头部扁平、矿化且多刺，这与防御习性有关。已知这种两栖动物头部和身体的皮肤分泌物毒性很强，可作为抵御捕食者的有效化学防御手段。然而，这种分泌物的生化特性和药理作用仍鲜为人知。我们使用小鼠模型测试了粗皮肤分泌物以及通过反相高效液相色谱法获得的十种主要馏分的伤害感受和水肿活性以及体外细胞毒性。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析表明，凝胶中大多数蛋白质的分子量在55至30 kDa之间。液相色谱-质谱分析显示有多个低分子量分子（200 - 500 Da），这与生物碱和类固醇的分子量一致。粗皮肤分泌物能够引发快速且持续的水肿，并伴有强烈的剂量依赖性伤害感受。在测试的10个馏分中，5个同时引发水肿和伤害感受，6个馏分仅能引发水肿（为对照组的80 - 170%），7个馏分仅引发伤害感受（与对照组相比为15 - 30倍）。此外，在小鼠成纤维细胞和黑色素瘤细胞中证实了细胞生长抑制（IC50）。根据获得的数据，我们证实格林氏盔头雨蛙的皮肤分泌物毒性很强，富含能够直接引发局部炎症和伤害感受的化合物。这些特性作为该物种化学防御机制的一部分很重要。
