Inflammatory responses to acute elevations of carbon dioxide in mice

Abstract

Health risks are described from elevated indoor air carbon dioxide (CO₂), which often ranges from 1,000 to 4,000 ppm, but the mechanisms are unknown. Here, we demonstrate that mice exposed for 2 h to 2,000 or 4,000 ppm CO₂ exhibit, respectively, 3.4 ± 0.9-fold (SE, n = 6) and 4.1 ± 0.7-fold (n = 10) elevations in circulating microparticles (MPs); neutrophil and platelet activation, and vascular leak in brain, muscle, and distal colon. Interleukin (IL)-1β content of MPs also increases after 2,000 ppm by 3.8 ± 0.6-fold (n = 6) and after 4,000 ppm CO₂ by 9.3 ± 1.1-fold (n = 10) greater than control. CO₂-induced vascular damage is abrogated by treating mice with an antibody to IL-1β or an IL-1β receptor inhibitor. Injecting naïve mice with CO₂-induced MPs expressing a protein found on mature neutrophils recapitulates vascular damage as seen with elevated CO₂, and destruction of MPs in CO₂-exposed mice abrogates vascular injuries without altering neutrophil or platelet activation. We conclude that environmentally relevant elevations of CO₂ trigger neutrophils to generate MPs containing high concentrations of IL-1β that cause diffuse inflammatory vascular injury.NEW & NOTEWORTHY Elevated levels of CO₂ are often found in indoor air and cause adverse health effects, but the mechanisms have not been identified. In a murine model, environmentally relevant levels of CO₂ were found to cause diffuse vascular damage because neutrophils are stimulated to produce microparticles that contain high concentrations of interleukin-1β.

Keywords: indoor air quality; interleukin-1β; microparticles; neutrophil activation; vasculopathy.

Fig. 1.

Microparticles (MPs) in mice following 2-h exposures. Blood-borne MPs were quantified in control mice (air only), and mice were killed immediately after exposure for 2 h to air + 1,000 to 10,000 ppm CO₂. Where shown, mice were injected before exposure with PEG (see materials and methods). Flow cytometric measurements were made to quantify the number of all 0.3 to 1 µm diameter Annexin V-positive particles (top), as well as those expressing proteins specific to certain cells: Ly6G (mature neutrophils), CD41 (platelets), CD14 (predominantly monocytes), and CD31+/CD41-dim (endothelium). Data are means ± SE; n is shown for each sample. *P < 0.05, significantly different from control by ANOVA.

Fig. 2.

Neutrophil and platelet activation. Surface proteins on neutrophils and platelets were quantified in mice manipulated as described in the caption for Fig. 1. Frames 1–5: analyses of Ly6G-positive cells (neutrophils) where surface expression was evaluated for myeloperoxidase (MPO), Toll-like receptor 4 (TLR4), CXC chemokine receptor 4 (CXCR4), proteinase 3 (PR3), and presence of platelet-specific CD41 as reflecting platelet-neutrophil interactions. Frame 6: elevation in surface expression of thrombospondin-1 (TSP) on platelets, which were identified as CD41-positive, between 3 and 5 µm diameter and annexin V-negative. Data are means ± SE; n is shown for each sample. *P < 0.05, significantly different from control by ANOVA.

Fig. 3.

Vascular leakage of 2 × 10⁶ Da rhodamine-labeled dextran. Brain, skeletal muscle and distal colon were prepared and evaluated as described in materials and methods. Values reflect fluorescence from rhodamine as arbitrary units/mg tissue protein in homogenates from mice subjected to 2 h of air or air plus 1,000 to 10,000 ppm CO₂. Where shown, mice were injected with PEG, nonspecific control IgG, anti-IL-1β IgG, or anakinra just before exposure to 4,000 ppm CO₂. Others were injected with Ly6G-positive or -negative MPs prepared from mice exposed to 4,000 ppm CO₂ and killed 1 h later. Data are means ± SE. *P < 0.05, significantly different from control by ANOVA.