Abstract
Background: Pulmonary hypertension (PH) is a devastating and progressive disease characterized by excessive proliferation of pulmonary artery smooth muscle cells (PASMCs) and remodeling of the lung vasculature. Adenosine signaling through the ADORA2B receptor has previously been implicated in disease progression and tissue remodeling in chronic lung disease. In experimental models of PH associated with chronic lung injury, pharmacological or genetic inhibition of ADORA2B improved markers of chronic lung injury and hallmarks of PH. However, the contribution of ADORA2B expression in the PASMC was not fully evaluated.Hypothesis: We hypothesized that adenosine signaling through the ADORA2B receptor in PASMC mediates the development of PH.Methods: PASMCs from controls and patients with idiopathic pulmonary arterial hypertension (iPAH) were characterized for expression levels of all adenosine receptors. Next, we evaluated the development of PH in ADORA2Bf/f-Transgelin (Tagln)cre mice. These mice or adequate controls were exposed to a combination of SUGEN (SU5416, 20 mg/kg/b.w. IP) and hypoxia (10% O2) for 28 days (HX-SU) or to chronic low doses of bleomycin (BLM, 0.035U/kg/b.w. IP). Cardiovascular readouts including right ventricle systolic pressures (RVSPs), Fulton indices and vascular remodeling were determined. Using PASMCs we identified ADORA2B-dependent mediators involved in vascular remodeling. These mediators: IL-6, hyaluronan synthase 2 (HAS2) and tissue transglutaminase (Tgm2) were determined by RT-PCR and validated in our HX-SU and BLM models.Results: Increased levels of ADORA2B were observed in PASMC from iPAH patients. ADORA2Bf/f-Taglncre mice were protected from the development of PH following HX-SU or BLM exposure. In the BLM model of PH, ADORA2Bf/f- Taglncre mice were not protected from the development of fibrosis. Increased expression of IL-6, HAS2 and Tgm2 was observed in PASMC in an ADORA2B-dependent manner. These mediators were also reduced in ADORA2Bf/f- Taglncre mice exposed to HX-SU or BLM.Conclusions: Our studies revealed ADORA2B-dependent increased levels of IL-6, hyaluronan and Tgm2 in PASMC, consistent with reduced levels in ADORA2Bf/f- Taglncre mice exposed to HX-SU or BLM. Taken together, our data indicates that ADORA2B on PASMC mediates the development of PH through the induction of IL-6, hyaluronan and Tgm2. These studies point at ADORA2B as a therapeutic target to treat PH.
Highlights
Pulmonary Hypertension (PH) is a condition of the pulmonary vasculature characterized by an mPAP of ≥25 mmHg at rest (Archer et al, 2010)
We determined expression levels of CD39 and CD73, key enzymes for the catabolism of ATP to adenosine. These experiments did not show significant changes for CD39 in Idiopathic Pulmonary Arterial Hypertension (iPAH) pulmonary artery smooth muscle cells (PASMCs) compared to controls (Figure 1E)
In order to evaluate the ADORA2B-mediated mechanisms that lead to PH, we performed experiments with isolated primary pulmonary artery smooth muscle cells (PASMCs) from normal healthy donors. In these experiments we report that activation of ADORA2B by the selective agonist BAY 60-6583 leads to increased expression levels of hyaluronan synthase 2 (HAS2) under both normoxia and hypoxia (Figures 8A,B); interleukin (IL)-6 only under hypoxia but not in normoxia (Figures 8C,D) and transglutaminase 2 (TGM2) under normoxia but not hypoxia (Figures 8E,F)
Summary
Pulmonary Hypertension (PH) is a condition of the pulmonary vasculature characterized by an mPAP of ≥25 mmHg at rest (Archer et al, 2010). Group I PH, or Pulmonary Arterial Hypertension (PAH), is PH that primarily affects the pre-capillary vasculature of the lungs (Ventetuolo and Klinger, 2012; Hansdottir et al, 2013). Group III PH is associated with chronic lung diseases such as chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF) (Farkas et al, 2011; Fell, 2012; Judge et al, 2012). Pulmonary hypertension (PH) is a devastating and progressive disease characterized by excessive proliferation of pulmonary artery smooth muscle cells (PASMCs) and remodeling of the lung vasculature. Adenosine signaling through the ADORA2B receptor has previously been implicated in disease progression and tissue remodeling in chronic lung disease. Hypothesis: We hypothesized that adenosine signaling through the ADORA2B receptor in PASMC mediates the development of PH
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