Cytotoxic and genotoxic effects on human keratinocytes triggered by sphingomyelinase D from Loxosceles venom
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BIOLOGICS
Cytotoxic and genotoxic effects on human keratinocytes triggered by sphingomyelinase D from Loxosceles venom Marcelo Santos da Silva1 · Priscila Hess Lopes2 · Maria Carolina Elias1 · Denise V. Tambourgi2 Received: 27 May 2020 / Accepted: 24 June 2020 © Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract The spiders of the Loxosceles genus (called brown or violin spiders) are of medical relevance in several countries due to the many human envenomation cases reported. The main component of Loxosceles venom is the enzyme sphingomyelinase D (SMase D), which is responsible for the local and systemic effects induced by the whole venom. Here, we investigated the cytotoxic and genotoxic effects caused by Loxosceles laeta venom and SMase D on human keratinocytes to better understand the dermonecrosis development mechanism. Our findings indicate that whole venom, as well as SMase D, increases intracellular superoxide levels, leading to DNA damage. These effects appear to be dependent on the binding of SMase D to the cell surface, although the complete pathway triggered as a result of the binding still needs to be elucidated. Moreover, after SMase D treatment, we observed the presence of histone γH2AX, suggesting that the cells are undergoing DNA repair. Moreover, when ATR kinase was inhibited, the cell viability of human keratinocytes was decreased. Together, our findings strongly suggest that L. laeta venom, as well as SMase D, increases intracellular superoxide levels, leading to DNA damage in human keratinocytes. Additionally, the induced DNA damage is repaired through the activation of an apparent ATR-mediated DNA-damage response. This knowledge may contribute to a better understanding of the behaviour of human keratinocytes during cutaneous loxoscelism, a condition that affects thousands of people around the world. Keywords Loxosceles venom · Sphingomyelinase D · Reactive oxygen species · DNA damage · DNA-damage response
Introduction Loxosceles spiders, also known as brown or violin spiders, belong to a group of medically relevant organisms in several countries as many human envenomation cases have been reported (Isbister and Fan 2011; Malaque et al. 2015; Pezzi et al. 2016; Lopes et al. 2020a). Loxosceles envenomation can induce severe local and systemic effects, with extensive Marcelo Santos da Silva and Priscila Hess Lopes have contributed equally to this work. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00204-020-02830-2) contains supplementary material, which is available to authorized users. * Denise V. Tambourgi [email protected] 1
Laboratory of Cell Cycle, Butantan Institute, São Paulo, Brazil
Immunochemistry Laboratory, Butantan Institute, São Paulo, Brazil
2
tissue destruction and chronic ulcer formation (Futrell 1992; Tambourgi et al. 2010; Lopes et al. 2020a). Moreover, haemolysis, intravascular coagulation, and renal impairment, although rare, are among the main causes of death by Loxosceles
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