ScitoVation, LLC, 6 Davis Drive, Suite 146, Durham, NC 27709, USA.
ScitoVation, LLC, 6 Davis Drive, Suite 146, Durham, NC 27709, USA.
Toxicology. 2023 Nov;499:153642. doi: 10.1016/j.tox.2023.153642. Epub 2023 Oct 18.
New Approach Methodologies (NAMs) are being widely used to reduce, refine, and replace, animal use in studying toxicology. For respiratory toxicology, this includes both in silico and in vitro alternatives to replace traditional in vivo inhalation studies. 1,3-Dichloropropene (1,3-DCP) is a volatile organic compound that is widely used in agriculture as a pre-planting fumigant. Short-term exposure of humans to 1,3-DCP can result in mucous membrane irritation, chest pain, headache, and dizziness. In our previous work, we exposed differentiated cells representing different parts of the respiratory epithelium to 1,3-DCP vapor, measured cytotoxicity, and did In Vitro to In Vivo Extrapolation (IVIVE). We have extended our previous study with 1,3-DCP vapors by conducting transcriptomics on acutely exposed nasal cultures and have implemented a separate 5-day repeated exposure with multiple endpoints to gain further molecular insight into our model. MucilAir™ Nasal cell culture models, representing the nasal epithelium, were exposed to six sub-cytotoxic concentrations of 1,3-DCP vapor at the air-liquid interface, and the nasal cultures were analyzed by different methodologies, including histology, transcriptomics, and glutathione (GSH) -depletion assays. We observed the dose-dependent effect of 1,3-DCP in terms of differential gene expression, change in cellular morphology from pseudostratified columnar epithelium to squamous epithelium, and depletion of GSH in MucilAir™ nasal cultures. The MucilAir™ nasal cultures were also exposed to 3 concentrations of 1,3-DCP using repeated exposure 4 h per day for 5 days and the histological analyses indicated changes in cellular morphology and a decrease in ciliated bodies and an increase in apoptotic bodies, with increasing concentrations of 1,3-DCP. Altogether, our results suggest that sub-cytotoxic exposures to 1,3-DCP lead to several molecular and cellular perturbations, providing significant insight into the mode-of-action (MoA) of 1,3-DCP using an innovative NAM model.
新型方法学（NAMs）正被广泛应用于减少、优化和替代动物在毒理学研究中的使用。对于呼吸毒理学，这包括替代传统体内吸入研究的体内和体外替代方法。1,3-二氯丙烯（1,3-DCP）是一种挥发性有机化合物，广泛用于农业作为种植前熏蒸剂。人类短期接触 1,3-DCP 可导致粘膜刺激、胸痛、头痛和头晕。在我们之前的工作中，我们将代表呼吸道上皮不同部位的分化细胞暴露于 1,3-DCP 蒸气中，测量细胞毒性，并进行体内外推论（IVIVE）。我们通过对急性暴露的鼻培养物进行转录组学研究，扩展了我们之前对 1,3-DCP 蒸气的研究，并进行了单独的 5 天重复暴露，以获得更多的分子信息。MucilAir™鼻细胞培养模型代表了鼻上皮，在气液界面上暴露于 6 个亚细胞毒性浓度的 1,3-DCP 蒸气中，并通过不同的方法学分析鼻培养物，包括组织学、转录组学和谷胱甘肽（GSH）耗竭测定。我们观察到 1,3-DCP 的剂量依赖性效应，表现为差异基因表达、从假复层柱状上皮到鳞状上皮的细胞形态变化，以及 MucilAir™鼻培养物中 GSH 的耗竭。MucilAir™鼻培养物还暴露于 1,3-DCP 的 3 个浓度下，每天重复暴露 4 小时，连续 5 天，组织学分析表明细胞形态发生变化，纤毛体减少，凋亡体增加，随着 1,3-DCP 浓度的增加。总之，我们的结果表明，亚细胞毒性暴露于 1,3-DCP 导致了几种分子和细胞扰动，为使用创新的 NAM 模型研究 1,3-DCP 的作用模式（MoA）提供了重要的见解。
