Lung remodeling in a mouse model of asthma involves a balance between TGF-β1 and BMP-7.
PLoS One. 2014 Apr 29;9(4):e95959.
1 Department of Immunology, University of Sao Paulo, Sao Paulo, SP, Brazil, 2 Department of Biochemistry, University of Sao Paulo, Sao Paulo, SP, Brazil, 3 Department of Biological Sciences, Federal University of Sao Paulo, Diadema, SP, Brazil, 4 Division of Nephrology, Federal University of Sao Paulo, Sao Paulo, SP, Brazil
A key event in chronic allergic asthma is the TGF-β-induced activation of fibroblasts into α-SMA-positive myofibroblasts which synthesize type-I collagen. In the present study we investigated the effect of the anti-fibrotic molecule BMP-7 in asthma. Balb/c mice were immunized i.p. with ovalbumin in alum and challenged every 2 days with ovalbumin aerosol (two or six challenges for acute and chronic protocols, respectively). The lung was evaluated for: α-SMA and type-I collagen by immunohistochemistry; BMP-7 and TGF- β1 gene expression by qRT-PCR; type-I collagen and Smads 2 and 3 by immunoblotting; mucus by PSA staining. Type-I collagen around bronchi, α-SMA, mucus secretion, TGF- β1 and BMP-7 gene expression were all increased in asthma. The TGF- β1/BMP-7 ratio was higher in the chronic group and correlated with higher levels of collagen. Fibroblasts isolated from asthmatic and healthy lungs produced type-I collagen upon stimulation with TGF- β1 via phosphorylation of Smad-2, Smad-3. Pre-treatment of the fibroblasts with BMP-7 reduced collagen production and Smads phosphorylation. Intranasal treatment of asthmatic mice with recombinant BMP-7 during the immunization protocol reduced lung inflammation and type I collagen deposition. These results suggest a protective role for BMP-7 in lung allergic inflammation, opposing the pro-fibrotic effects of TGF- β1.
Allergic asthma is by chronic inflammation of the airways, which involves infiltration of effector Th2 lymphocytes and eosinophils to the airways. The activation of these cells leads to their degranulation, and the liberation of the granules’ contents causes tissue damage. During this process, several growth factors are released form the extracellular matrix (ECM), where they originally attach in an inactive form after secretion. TGF-b1 is one of these growth factors, and one of its roles in asthma is the activation of myofibroblasts, which actively synthesize and secrete type I collagen, leading to fibrosis of the affected areas. Overall, the characteristic structural changes which occur in the asthmatic lung are: thickening of the bronchial smooth muscle wall, high number of infiltrating cells, and deposition of an abnormal amount of extracellular matrix (ECM) components beneath the bronchial epithelium. These alterations end up replacing the normal lung parenchyma by scar tissue, diminishing the gas exchange surface and, consequently, determining the decreased respiratory capacity of the patients.
Even though the inflammatory process in asthma have been extensively studied, much less is known about regulation of this process. BMP-7 has been shown to counteract the effects of TGF-b1 in different contexts, particularly in kidney fibrosis. Further studies showed that it is also involved in the regulation of a wide spectrum of cell functions, such as proliferation, differentiation, and apoptosis.
In this study, we investigated the role of TGF-b1 and BMP-7 in lung remodeling in a mouse model of asthma, and the mechanisms underlying collagen production by isolated lung fibroblasts.
First, we established a mouse model of asthma and characterized it thoroughly through the morphologic analysis of collagen deposition, mucus production and the expression of a-SMA and type I collagen in situ, all hallmarks of airway inflammation and fibrosis.
Next, we cultivated the mouse lung fibroblasts of control and asthmatic animals in order to study if BMP-7 would interfere with TGF-b1 signaling and collagen production in vitro. We saw that the treatment of the primary lung fibroblasts with TGF-b1 caused p38 and SMAD 2/3 phosphorylation, as expected. When the cells were concomitantly treated with TGFb-1 and BMP-7, however, this phosphorylation decreased. This was true for the lung fibroblasts of both control and asthmatic animals. The addition of BMP-7 to the treatment was not able, in our experience, to influence the phosphorylation of ERK 1/2.
The next step was to evaluate the role of BMP-7 in vivo. We then treated the animals with recombinant human BMP-7 during the asthma protocol, starting after the fibrotic process had already begun, therefore being of therapeutic relevance. Administration of BMP-7 was performed via the intranasal route. Morphometric analysis of the pulmonary tissue from these animals, showed that this treatment significantly decrease collagen deposition around small bronchi and also impairs the influx of inflammatory cells to the airways.
Our work brings novel and important information about the balance between TGF-b1 and BMP-7 in the remodeling process in the lung in a model of allergic asthma and proposes that the TGF-b1/BMP-7 ratio could be used as a marker of disease severity. Additionally, it suggests the possibility of interfering with the fibrotic process in asthma by intranasal instillation of BMP-7.
Figure 1 LEGEND: BMP-7 inhibits TGF-b1 signaling in lung fibroblasts, reducing p38 and SMAD 2/3 phosphorylation. Treatment of asthmatic mice with rhBMP-7 reduced the influx of inflammatory cells to the airways and remodeling/fibrosis, as measured by collagen deposition around small bronchi. We propose the TGF-b1/BMP-7 ratio as a marker of asthma severity and intranasal instillation of BMP-7 to reduce the fibrotic process in asthma.