Adiponectin deficiency increases allergic airway inflammation and pulmonary vascular remodeling

Abstract

Obesity is associated with an increased incidence and severity of asthma, as well as other lung disorders, such as pulmonary hypertension. Adiponectin (APN), an antiinflammatory adipocytokine, circulates at lower levels in the obese, which is thought to contribute to obesity-related inflammatory diseases. We sought to determine the effects of APN deficiency in a murine model of chronic asthma. Allergic airway inflammation was induced in APN-deficient mice (APN(-/-)) using sensitization without adjuvant followed by airway challenge with ovalbumin. The mice were then analyzed for changes in inflammation and lung remodeling. APN(-/-) mice in this model develop increased allergic airway inflammation compared with wild-type mice, with greater accumulation of eosinophils and monocytes in the airways associated with elevated lung chemokine levels. Surprisingly, APN(-/-) mice developed severe pulmonary arterial muscularization and pulmonary arterial hypertension in this model, whereas wild-type mice had only mild vascular remodeling and comparatively less pulmonary arterial hypertension. Our findings demonstrate that APN modulates allergic inflammation and pulmonary vascular remodeling in a model of chronic asthma. These data provide a possible mechanism for the association between obesity and asthma, and suggest a potential novel link between obesity, inflammatory lung disease, and pulmonary hypertension.

Figure 1.

Increased chronic airway inflammation in adiponectin (APN)-deficient (−/−) mice. (A) Hematoxylin and eosin–stained lung sections of wild-type (top panels: i, 100× magnification; iii, 200× magnification; v, 400× magnification) and APN^−/− mice (bottom panels: ii, 100× magnification; iv, 200× magnification; vi, 400× magnification) harvested after 4 weeks of ovalbumin (OVA) challenges. Extensively remodeled vessels are indicated with closed arrows. Scale bars, 100 μm. Further enlarged views of lungs from wild-type mice (vii) and APN^−/− mice (viii) demonstrating eosinophils around the airways (open arrows). (B) Blinded histologic scoring of inflammation (scale of 0–4 [0 = normal, 4 = severe]) from wild-type and APN^−/− mice harvested after 4 weeks of OVA challenges (*P < 0.001 compared with wild-type mice; n = 10 mice per group from two experiments). (C) Percentage and number of lymphocytes (Lym), monocytes/macrophages (Mono), neutrophils (PMN), and eosinophils (Eos) in the bronchoalveolar lavage (BAL) fluid of wild-type and APN^−/− mice (*P < 0.05 compared with wild-type mice; n = 7 mice per group from two experiments). (D) Percentage and number of CD4⁺, CD8⁺, CD4⁺/CD69⁺, and CD8⁺/CD69⁺ lymphocytes in the BAL of wild-type and APN^−/− mice (*P < 0.05 compared with wild-type mice; n = 13 mice per group from three experiments).

Figure 2.

Decreased dynamic lung compliance in APN^−/− mice. Airway resistance (A) and dynamic lung compliance (B) in wild-type and APN^−/− mice in response to methacholine inhalation, measured after 4 weeks of OVA or PBS challenges (P = 0.11 by ANOVA for resistance between wild-type and APN^−/− OVA-challenged mice; P = 0.002 by ANOVA for compliance between wild-type and APN^−/− OVA-challenged mice; n = 10 mice per group from two experiments).

Figure 3.

Increased chemokine production in APN^−/− mice. (A and B) Chemokine RNA expression in lungs from APN^−/− mice and wild-type mice harvested after 4 weeks of OVA or PBS challenges (*P < 0.05 compared with wild-type mice; n = 10 mice per group from two experiments). (C and D) CCL11 and CCL24 protein measured by ELISA in the BAL from wild-type and APN^−/− mice after 4 weeks of OVA challenges (P = 0.011 for CCL11 and P = 0.071 for CCL24 wild-type versus APN^−/− mice; n = 10 mice per group from two experiments). (E) CCL11 and CCL24 RNA expression in bone marrow–derived macrophages treated with IL-4 (50 ng/ml) and TNF-α (100 ng/ml) with or without APN pretreatment (10 μg/ml) (P = 0.048 for CCL11 and P = 0.18 for CCL24, comparing IL-4/TNF-α with and without APN pretreatment; n = 3 samples per group, repeated twice).

Figure 4.

Changes in fibrosis and pulmonary arteries in APN^−/− mice by proliferating smooth muscle cells (SMCs). (A) The percentage area of lung sections that stained positive for Picrosirius red is not different in wild-type and APN^−/− mice after OVA immunization and challenge (n = 9 mice per group from two experiments). (B) The concentration of hydroxyproline in the right lung is not different in wild-type and APN^−/− mice after OVA immunization and challenge (n = 10 mice per group from two experiments). (C) Blinded histologic scoring of vascular obliteration in lung sections (scale of 0–4 [0 = normal, 4 = severe]) from wild-type and APN^−/− mice harvested after 4 weeks of OVA challenges (*P < 0.001 compared with wild-type mice; n = 10 mice per group from two experiments). (D) Trichrome staining of lung sections from wild-type (i, 400×) and APN^−/− mice (ii, 400×) (scale bars, 100 μm; pictures are representative from n = 10 mice per group from two experiments). (E) Vascular wall thickening in preacinar blood vessels in lung sections from wild-type and APN^−/− mice (P < 0.001 by ANOVA between all four groups; P = 0.023 between wild-type and APN^−/− mice after OVA challenge; n = 9 mice per OVA group; n = 3 mice per naive group; from two experiments). (F) Representative staining for α-SMC actin in lung sections from APN^−/− mice (i, 100× magnification; ii, 400× magnification), demonstrating positivity within the obliterated pulmonary arteries. Representative staining for proliferating cell nuclear antigen in a lung section from APN^−/− mice (iii, 400× magnification), demonstrating positivity in nuclei in cells (closed arrows) surrounding and within the obliterated pulmonary artery. Scale bars, 100 μm.

Figure 5.

Changes in right ventricular systolic pressure (RVSP) and growth factor expression in APN^−/− mice. (A) RVSP in wild-type and APN^−/− mice measured after 4 weeks of OVA or PBS challenges (P < 0.001 by ANOVA; P = 0.0007 between wild-type and APN^−/− mice after OVA challenge; n = 8 mice per group from two experiments). RVSP in wild-type and APN^−/− mice measured after 3 weeks of hypoxia (n = 6 mice per group from one experiment). (B) Growth factor RNA expression in the lungs from APN^−/− mice and wild-type mice harvested after 4 weeks of OVA or PBS challenges (n = 5 mice per group from two experiments).