Role of macrophages in pulmonary blood-air barrier impairment induced by PM2.5 exposure

Abstract

Objective To investigate the role of macrophages in the process of fine particulate matter (PM2.5) exposure induced damage to pulmonary blood-air barrier. Methods Eighteen male BALB/C mice (aged of 10 weeks, weighing 24~27 g) were randomly divided into control group and low- and high-dose PM2.5 exposure groups (receiving 1.8 and 16.2 mg/kg, respectively), with 6 mice in each group. The control group received tracheal instillations of normal saline on days 1, 4, and 7, whereas the exposure groups were administered corresponding dose of PM2.5 exposure at the same time points. In 24 h after last exposure, pathological changes in the lung tissues were observed, and the contents of total protein (TP), lactate dehydrogenase (LDH), and alkaline phosphatase (AKP) in bronchoalveolar lavage fluid (BALF), and F4/80 protein level in lung tissue were measured to evaluate the blood-air barrier damage and macrophage infiltration within the lung tissues. Additionally, an in vitro model of the blood-air barrier was established using A549 alveolar epithelial cells and EA.hy926 vascular endothelial cells. In combination with a THP-1 macrophage model, the supernatant PM2.5 supernatant, macrophage supernatant, and PM2.5-macrophage supernatant were incubated with the barrier model for 24 h, respectively. Transmembrane electrical resistance (TEER), sodium fluorescein permeability of the barrier model, and LDH release from the barrier cells were measured to ascertain the extent of macrophage-mediated enhancement in barrier damage induced by PM2.5 exposure. Furthermore, the expression of inflammatory cytokines, such as TNF-α, IL-1β, IL-6, and IL-8 in the macrophages after PM2.5 exposure was analyzed with quantitative real-time PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA). Results PM2.5 exposure induced lung tissue damage in mice in a dose-dependent manner, significantly elevated the contents of TP, LDH and AKP in the BALF and caused marked infiltration of macrophages into the lung tissue, especially the high-dose exposure when compared with the mice from the control group (P<0.01). In vitro barrier model exposure experiments showed that in comparison with the treatment of 150 and 300 μg/mL PM2.5 and macrophage supernatant, the same doses of PM2.5-macrophage supernatant resulted in notably decreased TEER and significantly enhanced permeability in the barrier model (P<0.01), and markedly increased LDH release from epithelial and endothelial barrier cells (P<0.01). Additionally, the exposure of 150 and 300 μg/mL PM2.5 led to a significant up-regulation of TNF-α, IL-1β, IL-6, and IL-8 in the macrophages (P<0.01). Conclusion Macrophages deteriorate PM2.5-induced functional impairment of the pulmonary blood-air barrier.

Keywords

• macrophages
• pm2.5
• blood-air barrier
• pulmonary alveoli