Fine particulate matter (PM2.5) enhances allergic sensitization in BALB/c mice

Abstract

Ambient particulate matter (PM), a component of air pollution, exacerbates airway inflammation and hyperreactivity in asthmatic patients. Studies showed that PM possesses adjuvant-like properties that enhance the allergic inflammatory response; however, the mechanism (or mechanisms) by which PM enhances the allergic response remains to be determined. The aim of this study was to assess how exposure to fine PM collected from Sacramento, CA, shapes the allergic airway immune response in BALB/c mice undergoing sensitization and challenge with ovalbumin (OVA). Eight-week-old BALB/c male mice were sensitized/challenged with phosphate-buffered saline (PBS/PBS; n = 6), PM/PBS (n = 6), OVA/OVA (n = 6), or OVA + PM/OVA (n = 6). Lung tissue, bronchoalveolar lavage fluid (BALF), and plasma were analyzed for cellular inflammation, cytokines, immunoglobulin E, and heme oxygenase-1 (HO-1) expression. Mice in the OVA + PM/OVA group displayed significantly increased airway inflammation compared to OVA/OVA animals. Total cells, macrophages, and eosinophils recovered in BALF were significantly elevated in the OVA + PM/OVA compared to OVA/OVA group. Histopathological grading indicated that OVA + PM/OVA treatment induced significant inflammation compared to OVA/OVA. Both immunoglobulin (Ig) E and tumor necrosis factor (TNF) α levels were significantly increased in OVA/OVA and OVA + PM /OVA groups compared to PBS/PBS control. The number of HO-1 positive alveolar macrophages was significantly elevated in lungs of mice treated with OVA + PM /OVA compared to OVA/OVA. Our findings suggest that fine PM enhances allergic inflammatory response in pulmonary tissue through mechanisms involving increased oxidative stress.

Figure 1

Allergic sensitization and challenge protocol. Mice were sensitized (day 1, 3, and 5) and challenged (day 12-14) intranasally with PBS (30 μl/day; delivery vehicle; white triangles), PM (33.3 μg/day, light grey triangles), OVA (10 μg/day, dark grey triangles) or OVA+PM. Mice were euthanized 24 hr after the final challenge (day 15) to assess pulmonary inflammation.

Figure 2

Cellular profiles of recovered bronchoalveolar lavage fluid (BALF) from mice sensitized/challenged with PBS/PBS (control; white), PM/PBS (light grey), OVA/OVA (dark grey), or OVA+PM/OVA (black). Total cellular influx, macrophages, neutrophils, eosinophils, and lymphocytes are shown for all 4 groups in number of cells/ml. PM enhanced OVA-induced allergic inflammation with respect to total cells, macrophages, and eosinophils. Results are presented as mean ± SEM (n=6 mice per group). Bars indicate a significant difference of p < 0.05 between groups.

Figure 3

Histopathological analysis of total lung and specific lung compartments. (A-D) Micrographs of lung tissue (200x magnification) stained with hematoxylin & eosin (H&E). (E) Histopathological analysis of lung tissue stained with H&E. OVA/OVA treatment resulted in significant pulmonary inflammation compared with PBS/PBS control treatment. OVA+PM/OVA treatment significantly enhanced inflammation over OVA/OVA treatment. Data is presented as mean ± SEM (n=6 mice per group). Bars indicate a significant difference of p < 0.05 between groups. The scale bar represents a distance of 100 micrometers.

Figure 4

Immunoglobulin E (IgE) levels from blood plasma of mice sensitized/challenged with PBS/PBS, PM/PBS, OVA/OVA, or OVA+PM/OVA. Concentration is shown as nanograms (ng) of IgE per milliliter (ml) plasma. Data is presented as mean ± SEM (n = 3-5 mice per group). Bars indicate a significant difference of p < 0.05 between groups.

Figure 5

Hemeoxygenase-1 (HO-1) analysis. (A-D) Micrographs of paraffin embedded lung tissue sections immunohistochemically stained with HO-1 (brown stain, 200x magnification). The inserts show magnified macrophages within the respective image. The scale bar represents a distance of 100 micrometers. (E) The frequency of HO-1 positively stained alveolar macrophages in one total field of view (400x). Data is presented as mean ± SEM (n = 5-6 mice per group). Bars indicate a significant difference of p < 0.05 between groups.