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.

由于报道了许多人类中毒病例，Loxosceles属的蜘蛛（称为棕色或小提琴蜘蛛）在几个国家具有医学意义。Loxosceles毒液的主要成分是鞘磷脂酶D（SMase D），它负责整个毒液诱导的局部和全身作用。在这里，我们研究了Loxosceles laeta引起的细胞毒性和基因毒性作用毒液和 SMase D 在人角质形成细胞上的作用，以更好地了解皮肤坏死的发展机制。我们的研究结果表明，全毒液以及 SMase D 会增加细胞内超氧化物水平，导致 DNA 损伤。这些效应似乎取决于 SMase D 与细胞表面的结合，尽管仍需要阐明由结合引发的完整途径。此外，在 SMase D 处理后，我们观察到组蛋白 γH2AX 的存在，表明细胞正在进行 DNA 修复。此外，当 ATR 激酶被抑制时，人角质形成细胞的细胞活力会降低。总之，我们的研究结果强烈表明L. laeta毒液以及 SMase D 会增加细胞内超氧化物水平，导致人类角质形成细胞的 DNA 损伤。此外，通过激活明显的 ATR 介导的 DNA 损伤反应来修复诱导的 DNA 损伤。这些知识可能有助于更好地了解人类角质形成细胞在皮肤 loxoscelism 期间的行为，这种情况影响了世界各地的数千人。