Adiponectin deficiency: a model of pulmonary hypertension associated with pulmonary vascular disease

Abstract

Adiponectin (APN) is an adipocyte-derived factor that exists at high concentrations in serum and has anti-inflammatory and systemic vascular-protective properties. In this study, we investigated the role of APN in pulmonary vascular homeostasis. We found that APN localizes to the luminal side of blood vessels in lung and acts in vitro to block TNF-alpha-induced E-selectin upregulation in pulmonary artery endothelial cells. Targeted deletion of the APN gene in mice leads to a vascular phenotype in lung characterized by E-selectin upregulation and age-dependent increases in perivascular inflammatory cell infiltration and pulmonary arterial pressures. Taken together, these findings demonstrate an important role for APN in lung vascular homeostasis and suggest that APN-deficient states may contribute to the pathogenesis of inflammatory pulmonary vascular disease and to the development of pulmonary hypertension.

Fig. 1.

Adiponectin (APN) localizes to vascular sites in lung. A and B: representative sections showing that APN protein (brown stain) localizes to blood vessels (v) in lung. Staining is absent in proximal (AW) and distal airway structures. APN staining is not detected in lung sections of APN-deficient mice (C) and in wild-type (WT) lung sections stained with isotype control antibody (D). E: confocal microscopic analysis of APN protein localization was detected on the luminal side of blood vessels in lung where it colocalized with the vascular endothelium. APN (green) colocalizes to F-actin (red) on the endothelial side of the blood vessel wall. Scale bar = 200 μm.

Fig. 2.

Perivascular inflammation is present in lungs of 3-mo-old APN-deficient mice. CD45 staining (brown) was performed in WT and APN-deficient lung sections. A and B: low- and high-power images of WT lungs, respectively. Scale bars = 200 μm. C and D: low- and high-power images of APN-deficient lung sections, respectively. Scale bars = 100 μm. E: bar graph showing the average number of perivascular CD45-positive cells per blood vessel (×40) in WT and APN-deficient lungs (*P < 0.05).

Fig. 3.

E-selectin is upregulated in pulmonary vascular endothelium of 3-mo-old APN-deficient mice. E-selectin staining is not detected by immunohistochemistry in lungs of WT mice (A), but staining is present throughout the endothelium of APN-deficient mice (B). Staining was not detected in APN-deficient sections stained with isotype control antibody (data not shown). C: Western blot showing E-selectin concentration in WT and APN-deficient lungs. D: bar graph showing relative E-selectin concentration when controlled for GAPDH.

Fig. 4.

APN blocks TNF-α-induced E-selectin upregulation in human pulmonary artery endothelial cells (HPAEC). HPAEC were pretreated with APN for 18 h followed by stimulation with TNF-α for 6 h. Data are expressed as E-selectin mRNA levels relative to internal control gene 36B4 levels (n = 5; *P < 0.05).

Fig. 5.

Perivascular inflammation increases with age. CD45 staining (brown) at 1 yr demonstrates the presence of perivascular inflammatory cell infiltrates in WT and APN-deficient lungs. Mild inflammation is observed in lungs of WT mice. A and B: low- and high-power images, respectively. Perivascular inflammation is readily apparent in lungs of APN-deficient mice. C and D: low- and high-power images, respectively. E: bar graph showing the average number of perivascular CD45-positive cells per blood vessel (×40) in WT and APN-deficient lungs (*P < 0.05). Scale bars: A and C, 200 μm; B and D, 100 μm.

Fig. 6.

Inflammatory vascular phenotype is present in lungs of 1-yr-old APN-deficient mice. E-selectin expression is detected in lungs of 1-yr-old APN-deficient mice (B) but not in lungs of age-matched WT mice (A). Perivascular inflammation and proteinaceous exudates are readily apparent in lungs of 1-yr-old APN-deficient mice (E, low-power; F, high-power view). Similar findings were not observed in lungs of 1-yr-old WT mice (C, low-power; D, high-power view). Scale bars: A, B, C, and E, 250 μm; D and F, 100 μm.