Assessment of Inhalation Exposures to Indoor and Occupational Aerosols

反复真菌吸入暴露的鼠类模型-有害健康影响与微生物生长和潮湿的室内环境有关;然而，与真菌暴露有关的毒性机制仍未得到充分研究。霉菌被提名给NTP进行毒理学表征，根据方法和专业知识，NIOSH被确定为合作者，对NTP提名的真菌物种进行亚慢性吸入暴露研究。NIOSH为这些研究开发的声学发生器系统（AGS）模拟了在真菌污染环境中遇到的自然人体暴露。在该子项目下进行的研究描述了与反复真菌暴露相关的毒理学和肺部免疫反应。这些研究分为两个阶段-第一阶段表征目标真菌物种的培养和气溶胶优化，第二阶段包括亚慢性研究。 该分项目还包括对受污染的室内环境中发现的真菌多样性进行评价。20财年期间，发表了两篇论文，描述了产生单端孢菌素的葡萄穗霉的培养和雾化，以及暴露后的肺部免疫反应。在这些研究中，两株S.使用命名为菌株A和菌株B的chartarum，其中菌株A产生的片段和霉菌毒素的量高于菌株B。 这些出版物的第一个特点是培养可行的和不可行的S。以及AGS的优化，包括S. chartarum给小鼠的鼻子只暴露吊舱。然后在进一步的研究中利用暴露系统的表征和优化来测量重复真菌暴露后的肺免疫应答。各组小鼠吸入活的或热灭活的S。chartarum分生孢子，或HEPA过滤的空气，每周两次，持续4周和13周。在暴露后4周，观察到T辅助细胞2型（Th 2）介导的反应。13周后，与菌株B暴露后Th 2介导的应答相比，暴露于菌株A后观察到混合T细胞应答。两种菌株在暴露后13周诱导肺动脉重塑;然而，菌株A暴露的小鼠比菌株B暴露的小鼠进展更快。灌洗液由炎性细胞群组成，包括嗜酸性粒细胞、中性粒细胞和巨噬细胞。总体而言，在暴露于高片段产生菌株A的小鼠中，免疫病理学反应发生得更早，这表明真菌片段的存在增加了暴露，并促成了观察到的反应。 NIOSH目前正在评估一项大型研究的结果，该研究评估了重复暴露于A后的肺和全身毒性。变色。 重复暴露1周后观察到先天免疫细胞数量增加，随后在4周内观察到额外B细胞、T细胞和2型先天淋巴细胞（ILC 2）浸润增加。反复暴露于A。在4周时，杂色导致局部和循环Th 2细胞因子（包括IL 4和IL 13）的产生增加，以及ILC 2增加。到13周时，除ILC 2外，所有细胞类型的细胞浸润均减少。来自活的、非活的和仅空气对照样品的肺匀浆的miRNA、mRNA和蛋白质组数据集的分析仍在进行中。与A. fumigatus和S. chartarum，初步数据显示，肺动脉组织重塑后，反复亚慢性暴露于A。versicolor，突出了人类真菌暴露中心血管受累的可能性。 对来自A.变色暴露仍在继续。这些数据集正在使用Inconsistency Pathway Analysis进行分析，以确定生物系统和疾病之间的已知关联，并确定与生物功能，途径和疾病的新相互作用和关联。

Murine Models of Repeated Fungal Inhalation Exposure - Adverse health effects have been associated with microbial growth and damp indoor environments; however, the mechanisms of toxicity associated with fungal exposure remain understudied. Mold was nominated to the NTP for toxicological characterization and based on the methodologies and expertise, NIOSH was identified as a collaborator to conduct sub-chronic inhalation exposures studies with NTP-nominated fungal species. The Acoustical Generator System (AGS) developed by NIOSH for these studies models a natural human exposure one would encounter in a fungal contaminated environment. The studies conducted under this subproject characterize the toxicological and pulmonary immune responses associated with repeated fungal exposure. These studies are separated into two phases – the first phase characterizing the cultivation and aerosol optimization of the targeted fungal species and the second phase consists of subchronic studies. Also included in this subproject is the evaluation of fungal diversity found within contaminated indoor environments. During FY20, two manuscripts characterizing the cultivation and aerosolization of trichothecene-producing Stachybotrys chartarum, as well as the pulmonary immune response following exposure were published. For these studies, two strains of S. chartarum designated as strain A and strain B were utilized with strain A producing a higher amount of fragments and mycotoxin compared to strain B. The first of these publications characterized the cultivation of viable and non-viable S. chartarum, as well as the optimization of the AGS including the aerosolization and delivery of S. chartarum to mice housed in nose-only exposure pods. The characterization and optimization of the exposure system was then utilized in further studies to measure the pulmonary immune responses following repeated fungal exposure. Groups of mice inhaled viable or heat-inactivated S. chartarum conidia, or HEPA–filtered air twice per week for 4 and 13 weeks. At 4 weeks after exposure, a T-helper cell type 2 (Th2)–mediated response was observed. After 13 weeks, a mixed T-cell response was observed following exposure to strain A compared with a Th2–mediated response after strain B exposure. Both strains induced pulmonary arterial remodeling at 13 weeks following exposure; however, strain A–exposed mice progressed more quickly than strain B–exposed mice. Lavage fluid was composed of inflammatory cell populations including eosinophils, neutrophils, and macrophages. Overall, the immunopathological responses occurred earlier in mice exposed to high fragment-producing strain A, suggesting that the presence of fungal fragments increased exposure and contributed to the observed responses. NIOSH is currently evaluating the results from a large study that assessed pulmonary and systemic toxicity following repeated exposure to A. versicolor. An increased number of innate immune cells were observed after 1 week of repeated exposure followed by the increasing infiltration of additional B-cells, T-cells, and type 2 innate lymphoid cells (ILC2s) over 4 weeks. Repeated exposure to A. versicolor led to the increased production of local and circulating Th2 cytokines, including IL4 and IL13, as well increased ILC2s by 4 weeks. By 13 weeks, cellular infiltration was decreased for all cell types except ILC2s. The analysis of miRNA, mRNA, and proteomic datasets derived from the lung homogenates of viable, nonviable, and air-only control samples are still ongoing. Similar to A. fumigatus and S. chartarum, preliminary data revealed pulmonary arterial tissue remodeling following repeated subchronic exposure to A. versicolor, highlighting the potential for cardiovascular involvement in human fungal exposures. An analysis of miRNA, mRNA, and proteomic datasets derived from the large toxicology study following A. versicolor exposure is ongoing. The datasets are being analyzed using Ingenuity Pathway Analysis to identify known associations within biological systems and diseases, as well as to identify novel interactions and associations with biological functions, pathways and diseases.