Lowest observed adverse effect level of pulmonary pathological alterations due to nitrous acid exposure in guinea pigs

Abstract

Background: We previously demonstrated that continuous exposure to nitrous acid gas (HONO) for 4 weeks, at a concentration of 3.6 parts per million (ppm), induced pulmonary emphysema-like alterations in guinea pigs. In addition, we found that HONO affected asthma symptoms, based on the measurement of respiratory function in rats exposed to 5.8 ppm HONO. This study aimed to investigate the dose-response effects of HONO exposure on the histopathological alterations in the respiratory tract of guinea pigs to determine the lowest observed adverse effect level (LOAEL) of HONO.

Methods: We continuously exposed male Hartley guinea pigs (n = 5) to four different concentrations of HONO (0.0, 0.1, 0.4, and 1.7 ppm) for 4 weeks (24 h/day). We performed histopathological analysis by observing lung tissue samples. We examined samples from three guinea pigs in each group under a light microscope and measured the alveolar mean linear intercept (Lm) and the thickness of the bronchial smooth muscle layer. We further examined samples from two guinea pigs in each group under a scanning electron microscope (SEM) and a transmission electron microscope (TEM).

Results: We observed the following dose-dependent changes: pulmonary emphysema-like alterations in the centriacinar regions of alveolar ducts, significant increase in Lm in the 1.7 ppm HONO-exposure group, tendency for hyperplasia and pseudostratification of bronchial epithelial cells, and extension of the bronchial epithelial cells and smooth muscle cells in the alveolar duct regions.

Conclusions: These histopathological findings suggest that the LOAEL of HONO is < 0.1 ppm.

Keywords: Alveolar mean linear intercept; Asthma; LOAEL; Nitrogen dioxide; Nitrous acid; Pulmonary emphysema.

Fig. 1

The transition of the body weight in each group. No significant differences were found by ANOVA.

Fig. 2

Effect of HONO exposure on the respiratory tract observed under light microscopy using hematoxylin and eosin staining. a, b Main bronchus near the middle of right-lung middle lobe. c Alveolar duct regions. Arrows indicate smooth muscle cells in the interstitium of the alveolar duct regions

Fig. 3

Effect of HONO exposure on the respiratory tract observed under light microscopy using Elastica van Gieson and transmission electron microscopy (TEM). a The terminal bronchiole and alveolar duct regions. b The alveolar regions. c Interstitium of alveolar duct regions under TEM. Arrows in the L, M, and H groups indicate smooth muscle cells that have abundant mitochondria and intermediate microfilaments

Fig. 4

Effect of HONO exposure on the respiratory tracts, observed under scanning electron microscope. a The bronchial lumen near the main bronchus. b The alveolar duct regions. c The alveolar regions