Apoptosis inhibitor of macrophage (AIM)/CD5L is involved in the pathogenesis of COPD

Abstract

Background: Alveolar macrophages (AMs) and AM-produced matrix metalloprotease (MMP)-12 are known to play critical roles in the pathogenesis of chronic obstructive pulmonary disease (COPD). The apoptosis inhibitor of the macrophages (AIM)/CD5 molecule-like (CD5L) is a multifunctional protein secreted by the macrophages that mainly exists in the blood in a combined form with the immunoglobulin (Ig)M pentamer. Although AIM has both facilitative and suppressive roles in various diseases, its role in COPD remains unclear.

Methods: We investigated the role of AIM in COPD pathogenesis using porcine pancreas elastase (PPE)-induced and cigarette smoke-induced emphysema mouse models and an in vitro model using AMs. We also analyzed the differences in the blood AIM/IgM ratio among nonsmokers, healthy smokers, and patients with COPD and investigated the association between the blood AIM/IgM ratio and COPD exacerbations and mortality in patients with COPD.

Results: Emphysema formation, inflammation, and cell death in the lungs were attenuated in AIM^-/- mice compared with wild-type (WT) mice in both PPE- and cigarette smoke-induced emphysema models. The PPE-induced increase in MMP-12 was attenuated in AIM^-/- mice at both the mRNA and protein levels. According to in vitro experiments using AMs stimulated with cigarette smoke extract, the MMP-12 level was decreased in AIM^-/- mice compared with WT mice. This decrease was reversed by the addition of recombinant AIM. Furthermore, an analysis of clinical samples showed that patients with COPD had a higher blood AIM/IgM ratio than healthy smokers. Additionally, the blood AIM/IgM ratio was positively associated with disease severity in patients with COPD. A higher AIM/IgM ratio was also associated with a shorter time to the first COPD exacerbation and higher all-cause and respiratory mortality.

Conclusions: AIM facilitates the development of COPD by upregulating MMP-12. Additionally, a higher blood AIM/IgM ratio was associated with poor prognosis in patients with COPD.

Trial registration: This clinical study, which included nonsmokers, healthy smokers, and smokers with COPD, was approved by the Ethics Committee of the Hokkaido University Hospital (012-0075, date of registration: September 5, 2012). The Hokkaido COPD cohort study was approved by the Ethics Committee of the Hokkaido University School of Medicine (med02-001, date of registration: December 25, 2002).

Keywords: Alveolar macrophage; Apoptosis inhibitor of macrophage; Chronic obstructive lung disease; Matrix metalloprotease-12.

Fig. 1

Lung inflammation assessed BALF from PPE-treated mice. (A) The number of total cells, (B) the number of macrophages, (C) the number of neutrophils, (D) the number of lymphocytes, and (E) the number of eosinophils in the BALF. Data are expressed as mean ± SEM (n = 4–6). ^*p < 0.05 by the Welch’s t test

Fig. 2

Gene expressions and protein levels in the lungs from PPE-treated mice. Gene expression in the lungs and protein levels in the BALF during PPE treatment were assessed using qRT-PCR or ELISA. (A) MMP-12 mRNA, (B) IL-33 mRNA, (C) MMP-12 protein, and (D) IL-33 protein. Data are expressed as mean ± SEM (n = 3–4). ^*p < 0.05 by the Mann–Whitney U test

Fig. 3

Emphysema formation and cell death in PPE-treated mice. (A) Representative photomicrographs of lungs of the AIM^−/− or WT mice on day 21 after PPE or vehicle treatment. Scale bar: 500 μm. (B) Quantification of MLI on day 21 after PPE or vehicle treatment. (C) Representative photomicrographs of TUNEL and Hoechst staining of lungs of the AIM^−/− or WT mice on day 21 after PPE or vehicle treatment. Scale bar: 100 μm. (D) Quantification of TUNEL-positive cells on day 21 after PPE or vehicle treatment. Data are expressed as mean ± SEM (n = 3–6). ^*p < 0.05, ^****p < 0.001 by the Welch’s t test with a Holm adjustment for (B) and the Tukey–Kramer test for (D)

Fig. 4

Lung inflammation, emphysema formation, and cell death in CS-exposed mice. (A) The numbers of total cells, (B) number of macrophages, (C) number of neutrophils, and (D) number of lymphocytes in the BALF. (E) Representative photomicrographs of lungs of the AIM^−/− or WT mice 16 weeks after CS or air exposure. Scale bar: 500 μm. (F) Quantification of MLI 16 weeks after CS or air exposure. (G) Representative photomicrographs of TUNEL and Hoechst staining of lungs of the AIM^−/− or WT mice after 16 weeks of CS or air exposure. Scale bar: 100 μm. (H) Quantification of TUNEL-positive cells 16 weeks after CS or air exposure. Data are expressed as mean ± SEM (n = 4–8). ^*p < 0.05, ^**p < 0.01, ^***p < 0.005 by the Welch’s t test for (A)–(D), Welch’s t test with a Holm adjustment for (F), and Tukey–Kramer test for (H)

Fig. 5

MMP-12 levels in the CSE-stimulated AMs. MMP-12 mRNA (A) and MMP-12 protein (B) levels in the CSE-stimulated AMs from the WT and AIM^−/− mice and CSE-stimulated AMs from AIM^−/− mice cultured with rAIM. Data are expressed as mean ± SEM (n = 3–5). ^**p < 0.01, ^****p < 0.001 by the Dunnett test

Fig. 6

Blood AIM/IgM ratios in humans. (A) Plasma AIM/IgM ratio in nonsmokers, healthy smokers, and smokers with COPD. (B) Serum AIM/IgM ratio in patients with COPD classified by the degree of airflow limitation (GOLD classification). The Kaplan–Meier curves for time to first exacerbation (C), all-cause mortality (D), and respiratory-cause mortality (E) among patients with COPD classified by serum AIM/IgM ratio. Data are expressed as mean ± SEM. ^*p < 0.05, ^**p < 0.01, ^***p < 0.005 by the Tukey–Kramer test for (A), Jonckheere–Terpsta test for (B), and log-rank trend test for (C)–(E).