Mold was nominated to the NTP for toxicological characterization and based on their inhalation, mycological, and immunological expertise, NIOSH was identified as a partner in 2013 to conduct multiple sub-chronic (13-week) exposure studies to individual organisms or mixtures of molds mimicking a real-world exposure. NIOSH has developed a unique delivery system that allows for precisely controlled murine exposures to particulates via nose-only inhalation. This approach overcomes a number of limitations associated with previous mold exposure studies in experimental rodent models that have used less relevant means of exposure to assess fungal toxicity. The acoustical generation system (AGS) developed as part of this partnership delivers fungal test articles to mice in a manner that simulates natural indoor fungal contaminant exposure. NIOSH continues to use the AGS and state-of-the-art methodologies to characterize toxicological endpoints following subchronic dry fungal spore and particle exposures. In FY18, a final report from the evaluation of the effects of 13-week inhalation exposure to Aspergillus fumigatus was received. Pathology samples from the study are undergoing review at the NTP. In FY 18, NIOSH completed studies to characterize pulmonary immune responses to two strains of Stachybotrys chartarum and the report for that work is in preparation. Repeated exposures to S. chartarum resulted in a Th2-mediated response in both strains at 4 weeks post-exposure and after 13 weeks, a mixed T-cell response was observed following exposure to a high fragmenting strain, compared to a Th2-mediated response to a low fragmenting strain. Histological examination of lung tissue revealed a continuum of pulmonary arteriole hyperplasia occurring earlier with the high fragmenting strain and peak remodeling was observed in both strains after 13 weeks. These histopathological datasets were supported by additional cellular, molecular, and proteomic analyses. In this series of studies, the degree of S. chartarum fragmentation appeared to modulate resulting pulmonary immune responses. In FY18, 1, 2, 4, and 13 week studies were additionally conducted to assess pulmonary and systemic toxicity following repeated exposure to Aspergillus versicolor. Preliminary data revealed an increase of cells associated with innate immunity after 1 week of repeated exposure and followed by the infiltration of innate cells, B- cells, T-cells, and type 2 innate lymphoid cells (ILC2s) by 2 weeks. Repeated exposure to A. versicolor led to the increased production of local and circulating Th2 cytokines, including IL4 and IL13, as well as an increase in ILC2s by 4 weeks. By 13 weeks, cellular infiltration was decreased for all cell types except ILC2s. Analysis of miRNA, mRNA, and proteomic datasets derived from the lung homogenates of viable, nonviable, and air-only control samples from these toxicology studies are ongoing. Similar to A. fumigatus and S. chartarum, preliminary data revealed remodeling of pulmonary arterial tissue following both 4 and 13 weeks of exposure to A. versicolor. A particularly interesting result for all of the fungal species studies to date has been the remodeling of pulmonary arterial tissue (Pulmonary Arterial Hyperplasia or PAH), highlighting the potential for cardiovascular involvement in human fungal exposures. Preliminary data suggests that this may be immune-mediated, and NIOSH will use histologic and molecular tools to investigate the potential mechanisms. Brain hemispheres derived from A. versicolor study mice have also been recently analyzed by Dr. Michelle Block at the Indiana University School of Medicine. Preliminary results demonstrate that repeated A. versicolor exposure over 4 weeks results in neuroinflammation and significant changes in the broad neuroinflammation transcriptome.
