In vitro assessment of cytotoxic activities of Lachesis muta muta snake venom.
In vitro assessment of cytotoxic activities of Lachesis muta muta snake venom.
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DOI：
10.1371/journal.pntd.0006427
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发表时间：
2018-04
影响因子：
3.8
通讯作者：
Braga VMM
中科院分区：
文献类型：
--
作者：
Stransky S;Costal-Oliveira F;Lopes-de-Souza L;Guerra-Duarte C;Chávez-Olórtegui C;Braga VMM
被毒蛇拉克西斯穆塔穆塔的毒液蛰伤被认为是严重的，其特征是局部影响，包括坏死，这是永久残疾的主要原因。然而，与蛇毒引发的细胞死亡和组织破坏相关的细胞机制却很少被探索。本研究的目的是探讨L. M.穆塔毒液在正常人角质形成细胞中的作用，并鉴定参与细胞毒液蛰入的细胞过程。为了研究毒液对不同细胞类型的影响，进行Alcohol Blue测定以定量细胞代谢水平作为细胞活力的读数。通过流式细胞术评估细胞凋亡、坏死和线粒体膜电位的变化，同时通过GFP-LC 3的表达评估自噬的诱导，并使用荧光显微镜进行分析。毒液的细胞毒性潜力表现为以浓度依赖性方式降低细胞活力。还观察到经历自噬（6小时）、凋亡和坏死（12和24小时）的细胞的顺序出现。形态学上，与L. M.穆塔毒液导致显著的细胞收缩和细胞聚集体的形成。这些结果表明，L. M.穆塔毒液对正常人角质形成细胞和其它细胞系具有细胞毒性，并且这种毒性涉及不同细胞死亡模式的整合。自噬作为一种细胞死亡机制，除了细胞凋亡和坏死，可以帮助解开由毒液触发的细胞途径和机制。了解细胞损伤和组织破坏的机制将有助于开发针对蛇咬伤的替代疗法。在这项工作中，我们调查的细胞事件和机制，参与细胞envenomation的Lachesis穆塔穆塔蛇，这是负责蛇在巴西的事故之一。由于毒液的病理作用与局部症状，如水肿和组织坏死，我们评估毒液对正常人角质形成细胞，在envenomation直接受影响的皮肤细胞的作用。我们的数据显示了由L. M.刺激自噬的穆塔毒液影响线粒体膜电位并激活通过凋亡和坏死导致细胞死亡的机制。粗毒液也会引起细胞形态和上皮细胞集落的改变。这些发现首次证明了L. M.皮肤细胞中的穆塔毒液。了解细胞损伤的潜在途径将有助于解释在局部毒液蛰入中观察到的一些病理效应。
Envenomation by the bushmaster snake Lachesis muta muta is considered severe, characterized by local effects including necrosis, the main cause of permanent disability. However, cellular mechanisms related to cell death and tissue destruction, triggered by snake venoms, are poorly explored. The purpose of this study was to investigate the cytotoxic effect caused by L. m. muta venom in normal human keratinocytes and to identify the cellular processes involved in in cellulo envenomation. In order to investigate venom effect on different cell types, Alamar Blue assay was performed to quantify levels of cellular metabolism as a readout of cell viability. Apoptosis, necrosis and changes in mitochondrial membrane potential were evaluated by flow cytometry, while induction of autophagy was assessed by expression of GFP-LC3 and analyzed using fluorescence microscopy. The cytotoxic potential of the venom is shown by reduced cell viability in a concentration-dependent manner. It was also observed the sequential appearance of cells undergoing autophagy (by 6 hours), apoptosis and necrosis (12 and 24 hours). Morphologically, incubation with L. m. muta venom led to a significant cellular retraction and formation of cellular aggregates. These results indicate that L. m. muta venom is cytotoxic to normal human keratinocytes and other cell lines, and this toxicity involves the integration of distinct modes of cell death. Autophagy as a cell death mechanism, in addition to apoptosis and necrosis, can help to unravel cellular pathways and mechanisms triggered by the venom. Understanding the mechanisms that underlie cellular damage and tissue destruction will be useful in the development of alternative therapies against snakebites. In this work, we investigate cellular events and mechanisms involved in in cellulo envenomation by Lachesis muta muta snake, which is one of the snakes responsible for the ophidic accidents in Brazil. Since the venom pathological effects are related to local symptoms such as edema and tissue necrosis, we evaluate venom action on normal human keratinocytes, skin cells directly affected during envenomation. Our data show the chronological analysis of cellular events triggered by L. m. muta venom, which stimulates autophagy, affects mitochondrial membrane potential and activates mechanisms that lead to cell death by apoptosis and necrosis. Crude venom also causes cell morphology and epithelial colony alterations. These findings give the first evidence about which cell death mechanisms are elicited by L. m. muta venom in skin cells. Understanding the pathways that underlie cellular damage will be useful to explain some of the pathological effects observed in local envenomation.
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影响因子：
3.1
作者：
Correa, C. R.;Bertollo, C. M.;Goes, A. M.
通讯作者：
Goes, A. M.
影响因子：
2.8
作者：
Bhowmik, Tanmoy;Gomes, Antony
通讯作者：
Gomes, Antony
影响因子：
3.8
作者：
Baldo, Cristiani;Jamora, Colin;Moura-da-Silva, Ana M.
通讯作者：
Moura-da-Silva, Ana M.
影响因子：
8.2
作者：
Han, Rong;Liang, Hui;Liu, Chun-yu
通讯作者：
Liu, Chun-yu
影响因子：
3.3
作者：
Junqueira-de-Azevedo, Inacio L. M.;Ching, Ana T. C.;Diniz, Marcelo R. V.
通讯作者：
Diniz, Marcelo R. V.