Abstract
In Brazil, around 80% of snakebites are caused by snakes of the genus Bothrops. A three-dimensional culture model was standardized and used to perform treatments with Bothrops erythromelas venom (BeV) and its antivenom (AV). The MRC-5 and L929 cell lines were cultured at increasing cell densities. Morphometric parameters were evaluated through images obtained from an inverted microscope: solidity, circularity, and Feret diameter. L929 microtissues (MT) showed better morphometric data, and thus they were used for further analysis. MT viability was assessed using the acridine orange and ethidium bromide staining method, which showed viable cells in the MT on days 5, 7, and 10 of cultivation. Histochemical and histological analyses were performed, including hematoxylin/eosin staining, which showed a good structure of the spheroids. Alcian blue staining revealed the presence of acid proteoglycans. Immunohistochemical analysis with ki-67 showed different patterns of cell proliferation. The MT were also subjected to pharmacological tests using the BeV, in the presence or absence of its AV. The results showed that the venom was not cytotoxic, but it caused morphological changes. The MT showed cell detachment, losing their structure. The antivenom was able to partially prevent the venom activities.
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Acknowledgements
This study was financially supported by grant PI-0257-2017 from Consejería de Salud y Familias, Junta de Andalucía, España and CTS-115 (Tissue Engineering Group), by Coimbra Scholarship Programme for Young Professors and Researchers from Latin American Universities and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq). The authors would like to thank the Multi-User Facility of Drug Research and Development Center (NPDM) of the Federal University of Ceara for technical support. The authors would also like to thank Prof. Dr. Nylane Maria Nunes de Alencar, coordinator of the Laboratory of Biochemical Pharmacology at NPDM - UFC, for kindly providing the L929 cell line.
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Andrade, F.R.S., da Silva, E.L., Marinho, A.D. et al. A new 3D model of L929 fibroblasts microtissues uncovers the effects of Bothrops erythromelas venom and its antivenom. Arch Toxicol 98, 3503–3512 (2024). https://doi.org/10.1007/s00204-024-03824-0
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DOI: https://doi.org/10.1007/s00204-024-03824-0