0010.1177/0394632015580907International Journal of Immunopathology and PharmacologyPałgan and Bartuzi research-article2015
Letter to the editor
Angiogenesis in bronchial asthma Krzysztof Pałgan and Zbigniew Bartuzi
International Journal of Immunopathology and Pharmacology 1–6 © The Author(s) 2015 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/0394632015580907 iji.sagepub.com
Abstract Bronchial asthma is a chronic inflammatory disease characterised by airflow obstruction that may be reversed spontaneously or in response to treatment. The airway inflammation can lead to structural changes and remodelling consisting of subepithelial layer thickening, airway smooth muscle hyperplasia and angiogenesis. Subepithelial hypervascularity and angiogenesis in the airways are part of the structural airway wall in asthma. Increased vascularity of bronchial mucosa is closely related to the expression of angiogenic factors like vascular endothelial growth factor (VEGF), angiopoietin and hypoxia-inducible factor (HIF). The scope of the present review is to summarise the roles of anagiogenic factors and treatment in vascular development. Keywords angiogenesis, angiopoietin, asthma, HIF, VEGF Date received: 5 September 2014; accepted: 16 March 2015
Introduction Historically, an interest in angiogenesis is associated with studies on cancer and factors involved in tumour progression. In 1971 a study was published by Judah Folkman (1933–2008), a pioneer in angiogenesis research in cancer, which demonstrated the major role of angiogenesis in the pathogenesis of cancer.1 Subsequent studies revealed other medical conditions in which the formation of new blood vessels plays an important role, and described numerous factors influencing angiogenesis. Angiogenesis can be defined as the emergence of new blood vessels from pre-existing vasculature. This phenomenon is observed in physiological conditions (e.g. wound healing, menstruation cycle), but can also play a role in the pathogenesis of diseases such as cancer, psoriasis, rheumatoid arthritis, proliferative retinopathy, obesity, atherosclerosis, ischaemic heart disease, endometriosis and bronchial asthma. Studies devoted to angiogenesis divided the process of new blood vessel formation into several stages. The initial stage of angiogenesis takes place in a pre-existing blood vessel located in close proximity to the wound, an ongoing malignant or inflammatory
process or angiogenic factors. In this stage, endothelial cell activation by growth factors, increased permeability and increased endothelial mitosis are observed. In the second stage, basement membrane is degraded by metalloproteinases. This stage is followed by the migration of endothelial cells towards angiogenic factors, and the formation of branch points and capillary lumen. The last stage involves modelling and stabilisation of the new capillary vessel. Finally, basement membrane is formed, interconnections between endothelial cells develop and pericytes are recruited.2 In 1960, Dunnill revealed increased vascularity in bronchial biopsy specimens from 20 subjects who died of acute asthmatic attacks. The studies on the bronchial vascular bed that followed showed a Department of Allergology, Clinical Immunology and Internal Medicine, Collegium Medicum Bydgoszcz, Nicolaus Copernicus University of Toruń, Poland Corresponding author: Krzysztof Pałgan, Nicolaus Copernicus University, Collegium Medicum in Bydgoszcz, Department of Allergology, Clinical Immunology and Internal Diseases, Ujejskiego 75, 85-168 Bydgoszcz, Poland. Email: [email protected]
2
International Journal of Immunopathology and Pharmacology
larger number of blood vessels in lamina propria and submucosa of the bronchi in patients with mild and severe bronchial asthma. The airways are supplied from blood vessels located in the smooth muscle layer of the bronchi, and through the subepithelial capillary network in lamina propria. In physiological conditions, the bronchial tree shows temporal fluctuation. Similarly, blood vessels of the bronchi undergo changes in permeability, dilation and density in the course of pathological and physiological processes. There is growing interest among scientists in neovascularisation and angiogenesis in patients with bronchial asthma.3
bronchial asthma was conducted by Grigorias et al.,4 and it demonstrated a clear relationship between the clinical stage of the disease and the number of blood vessels within the lamina propria. According to Grigorias,4 patients suffering from severe asthma have 46% more capillary vessels in bronchial submucosa compared to healthy controls. Other studies by Salvato et al.5 suggest a 30% increase in blood vessel count in patients with bronchial asthma. Positive correlation was observed between the number of blood vessels and the severity of bronchial asthma. A well-developed network of blood vessels enables migration of inflammatory cells into the bronchial wall.5
Angiogenesis and bronchial asthma
Angiogenic factors in bronchial asthma
Bronchial asthma is a chronic disease of the airways, the predominant underlying mechanisms of which are bronchial inflammation, hyper-responsiveness and bronchial wall remodelling. Structural changes in the bronchial tree result in increasing bronchial hyper-responsiveness and a progressively more severe course of bronchial asthma. Airway remodelling in patients with bronchial asthma is defined as a sequence of morphological changes involving cells and tissues that constitute bronchioles. Structural changes involve virtually all layers of the bronchial wall, including epithelium, basal membrane, smooth muscles and blood vessels. Chronic inflammatory processes in patients with bronchial asthma, and frequent exacerbations caused mainly by allergen exposure, or viral or bacterial infections, lead to bronchial tree remodelling. Microscopic examination of bronchial biopsy specimens reveals considerable thickening of the bronchiolar walls as well as of the cartilage-containing bronchi. Although virtually all layers of the bronchial wall undergo structural remodelling, it is the bronchial basement membrane that first demonstrates morphological changes. In the course of the process, collagens are deposited in the basement membrane, leading to its thickening and a decrease in elasticity. Both hyperplasia and hypertrophy of smooth muscle cells are observed in the smooth muscle layer. Along with the bronchial wall thickening, new blood vessels emerge. A detailed histological analysis of the bronchial biopsy specimens from patients with mild, moderate or severe chronic
Mast cells The first studies on angiogenesis in cancer identified mast cells as an important source of angiogenic factors. In the initial phase of tumour development, the emergence of numerous mast cell infiltrations at the border between the tumour and the surrounding normal tissue precedes angiogenesis. In type I hypersensitivity, according to Gell and Coombs, mast cells and the mediators they release, such as histamine, platelet-activating factor (PAF), leukotriene B4 (LTB4) and prostaglandin D2 (PDG2), are the main actors of the anaphylactic reactions. Apart from cytokines, such as vascular endothelial growth factor (VEGF), angiogenin, tumour necrosis factor (TNF), interleukine-8 (IL8) and fibroblast growth factor-2 (FGF-2), mast cells release tryptase and chymase, proteolytic enzymes involved in angiogenesis.6
Hypoxia-inducible transcription factor (HIF) Inflammatory process and bronchoconstriction promote hypoxia in patients with bronchial asthma. Ischaemia, inflammatory cells and increased cellularity of the bronchi and bronchioles may lead to the initiation of angiogenesis.7 Tissue ischaemia is a strong trigger for the production of transcription factors referred to as hypoxia-inducible factor-1 (HIF-1). A whole family of proteins has been described, whose role is to minimise cell damage caused by hypoxia.8 HIF-1 is a heterodimer composed of subunits α and β. HIF activity is regulated by prolyl hydroxylase
3
Pałgan and Bartuzi
VEGF-A A
VEGFR-1 V
VEG GF-B
VEGF-C V
VEGF-D
VEGF FR-3
VEGFR-2 2
Endoth helial cells
Angiogen nesis
Lymphangiogenessis
Figure 1. Vascular endothelial growth factors (VEGF-A, VEGF-B, VEGF-C, VEGF-D).13–15
VEGF-A acts by receptors VEGFR-1, VEGFR-2, but activates mainly VEGFR-2 and induce angiogenesis. VEGF-C, VEGF-D activate VEGFR-3 and stimulate lyphangiogenesis.
domain proteins (PHD) responding to oxygen concentrations.9 It has been demonstrated that angiogenesis can occur in normoxic conditions in patients with bronchial asthma. It seems that chronic inflammatory process in the airways and the cytokines, such as transforming growth factor-β (TGF-β), tumour necrosis factor-α (TNF-α), granulocyte-macrophage colonystimulating factor (GM-CSF) or lipopolysaccharides (LPS), can stimulate the production of HIF-1α. Additionally, the animal model shows positive correlation between HIF-1α concentrations and the intensity of bronchial allergic inflammation in mice. HIF-1 is one of the most important activators for genes encoding vascular endothelial growth factor A (VEGF-A).10,11
Vascular endothelial growth factor (VEGF) VEGF was discovered in the late 1970s during research on tumour angiogenesis. VEGF is a cytokine that plays a crucial role in the creation of new blood vessels (vasculogenesis) during embryonic development.12 It was earlier referred to as vascular permeability factor (VPF). Several VEGFs have been described within the cytokine family, including VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E and the placenta growth factor (PlGF). Genes encoding VEGF are located on chromosome 6p21.3.13,14 VEGF acts mainly on the endothelial cells of blood and lymphatic vessels, inducing their migration and division (Figure 1). Inflammatory cells, such as eosinophils, mast cells
and macrophages, express VEGF receptors (VEGF Receptor-1, VEGFR-1). VEGFR-1, in turn, stimulates chemotactic migration of these cells.15 It should also be emphasised that VEGF inhibits the apoptosis of vascular endothelial cells. As mentioned before, this cytokine significantly increases the permeability of the blood vessels – according to Shulman et al.,16 it is 50,000 times more potent than histamine in doing so. Sputum examinations in patients with bronchial asthma revealed significantly elevated concentrations of VEGF in addition to cysteinyl leukotriens (Cyst-LTs).17 Studies on the role of VEGF in the pathogenesis of bronchial asthma showed that this factor can stimulate allergic inflammation in the bronchial tree and contribute to the remodelling process. Positive correlation was shown between VEGF concentrations in the sputum as well as in the lung tissue, and the severity of bronchial asthma.18 Animal model experiments of bronchial asthma proved that VEGF is an important stimulating factor for airway remodelling. According to LopezGuisa et al.,19 airway remodelling in patients with bronchial asthma is initiated by the destruction of bronchial endothelium and the release of VEGF from the endothelial cells. It was shown that Th17 lymphocytes infiltrating bronchial epithelium stimulate VEGF production. Comparative in vitro analysis of bronchial epithelium between children diagnosed with asthma and healthy controls demonstrated an excessive capacity of bronchial
4
epithelial cells in asthmatic patients for VEGF production and release.20 Excessive VEGF production was observed also in bronchial smooth muscle cells. Genetic analyses in patients with bronchial asthma demonstrated epigenetic modifications of histone H3 and H4 involving their hyperacetylation and methylation. Such modification of histone proteins facilitates the access of VEGF promoters such as Sp1 or H3K4me3 to the effector genes, leading to their over expression. Bronchial asthma is frequently exacerbated by rhinovirus infections. These viruses also strongly stimulate VEGF production.21 Recently the role of IL-32 in the pathogenesis of bronchial asthma has been emphasised. IL-32 is produced by a variety of cells, of which the most important in the pathogenesis of asthma are dendritic, endothelial and NK cells and T lymphocytes. IL-32 significantly inhibits VEGF production and bronchial angiogenesis. It has been demonstrated that an increase in serum IL-32 concentrations in patients with bronchial asthma is indicative of effective therapy. Interestingly, IL-32 also shows antiviral and antibacterial properties.22
Angiopoietin Three types of angiopoietins have been described in humans: angiopoietin 1 (Ang-1), angiopoietin 2 (Ang-2) and angiopoietin 4 (Ang-4). Receptors binding these factors are known as Tie1 and Tie2, the latter being capable of binding all angiopoietins. Harfouche et al.23 noticed that angiopoietins are produced mainly by basophils and mast cells, which also express angiopoietin receptors. Interestingly, angiopoietins act as chemotactic agents for mast cells. A particularly high expression of genes involved in the synthesis of angiopoietins is observed in the mast cells of lung tissue.24 The crucial role of angiopoietins can be seen in the final stages of angiogenesis. Ang-1 stimulates migration of pericytes and smooth muscle cells and is therefore responsible for stabilisation of new capillary tubes. Contrary to Ang-2, Ang-1 reduces blood vessel permeability in patients with bronchial asthma. Ang-2 increases the permeability of the vascular bed through elimination of interconnections between endothelial cells and other cells constituting arterioles and venules. According to Bhandari et al.,24 Ang-2 can be responsible for lung oedema following exposure to toxic gases. Another
International Journal of Immunopathology and Pharmacology
difference between these angiopoietins is that Ang-1 is synthesised constitutively and low concentrations of this factor can be found in normal tissues, whereas Ang-2 is produced at the inflammation site only. A relationship is observed between VEGF and angiopoietins. High concentrations of VEGF and Ang-2 are noted in the initial phases of vascular remodelling, as well as angiogenesis and lymphogenesis. Hypoxia is the most important factor regulating transcription of both VEGF and Ang-2. Nomura et al.25 demonstrated high concentrations of angiopoietin and VEGF in the induced sputum of patients with bronchial asthma.
Bronchial asthma treatment and angiogenesis According to current guidelines, inhaled corticosteroids are the most effective anti-inflammatory medications recommended as a first-line treatment for chronic asthma. Corticosteroids inhibit synthesis of many cytokines known to stimulate inflammatory process in the bronchial tree. Numerous studies have emphasised the inhibiting role of inhaled corticosteroids on the migration of inflammatory cells to the bronchi in patients with bronchial asthma.26 Corticosteroids are particularly beneficial in the treatment of eosinophilic asthma. It has been demonstrated that they reduce the number of eosinophils in the bronchial epithelium. Another issue associated with inhaled corticosteroid therapy is its effect on bronchial tree remodelling. Durrani et al.27 noticed that inhaled corticosteroids inhibit structural changes of the basement membrane in patients with bronchial asthma only after 6 months of therapy with very high doses, equivalent to 1–3 mg of budesonide per day. Feltis et al.28 directly analysed angiogenesis and angiogenic factors in 35 treatment-naive patients with mild or moderate bronchial asthma. Study participants were given 750 µg fluticasone (fluticasone propionate) twice daily. After 3 months, VEGF and Ang-1 concentrations, as well as vascular density, were assessed. Fluticasone treatment significantly lowered tissue concentrations of angiogenic factors and reduced the number of capillary vessels by approximately 18.7%. The results of animal studies on angiogenesis in bronchial asthma are much more convincing. Sun et al.29 analysed the effect of budesonide on the
Pałgan and Bartuzi
angiogenesis in mice with induced asthma and proved that it not only suppressed the inflammatory process through blocking HIF-1α and VEGF expression, but also inhibited the development of new blood vessels. These findings showed considerable (over 50%) reduction in the number of blood vessels as well as in serum growth factor concentrations, compared to the control group. Other medications used in the treatment of bronchial asthma, such as beta-2 agonists, leukotriene receptor antagonists (LTRAs) or anticholinergics, do not show any significant effect on airway remodelling.27 Roth et al.30 claimed that airway remodelling in bronchial asthma is associated with overproduction of IgE, which may be responsible for thickening the bronchial basement membrane and directly stimulating proliferation of smooth muscle cells. Clinical studies focusing on the effect of omalizumab, an IgE-binding antibody, on the structural changes of the bronchial tree in patients with moderate and severe asthma seem to confirm its role in airway remodelling. As well as its anti-inflammatory activity, omalizumab also inhibited collagen deposition in the reticular layer of the basement membrane and reduced thickening of the bronchial wall. It can therefore be presumed that angiogenesis also was suppressed. Declaration of conflicting interests The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Funding This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors. References 1. Folkman J (1971) Tumor angiogenesis: Therapeutic implications. New England Journal of Medicine 285: 1182–1186. 2. Pałgan K (1993) Actual views on neoplasm angiogenesis. Postępy Higieny i Medycyny Doświadczalnej 47: 193–207. 3. Dunnill MS (1960) The pathology of asthma, with special reference to changes in the bronchial mucosa. Journal of Clinical Pathology 13: 27–33. 4. Grigoraş A, Căruntu ID, Grigoraş CC, et al. (2012) Relationship between immunohistochemical assessment of bronchial mucosa microvascularization and clinical
5 stage in asthma. Romanian Journal of Morphology and Embryology 53: 485–490. 5. Salvato G (2001) Quantitative and morphological analysis of the vascular bed in bronchial biopsy specimens from asthmatic and non-asthmatic subjects. Thorax 56: 902–906. 6. Pałgan K, Kołakowska J and Zbikowska-Gotz M (2000) The role of mast cells in inflammatory reactions. Pneumonologia i Alergologia Polska 68: 603– 609. 7. Scholz CC and Taylor CT (2013) Targeting the HIF pathway in inflammation and immunity. Current Opinion in Pharmacology 13: 646–653. 8. Mandl M and Depping R (2014) Hypoxia-inducible aryl hydrocarbon receptor nuclear translocator (ARNT) (HIF-1β): A rare exception? Molecular Medicine 20: 215–220. 9. Dehne N and Brune B (2009) HIF-1 in the inflammatory microenvironment. Experimental Cell Research 315: 1791–1797. 10. Huerta-Yepez S, Baay-Guzman GJ, Bebenek IG, et al. (2011) Hypoxia inducible factor promotes murine allergic airway inflammation and is increased in asthma and rhinitis. Allergy 66: 909–919. 11. Kuschel A, Simon P and Tug S (2012) Functional Regulation of HIF-1alpha under normoxia - is there more than posttranslational regulation? Journal of Cellular Physiology 227: 514–522. 12. Zgraggen S, Ochsenbein AM and Detmar M (2013) An important role of blood and lymphatic vessels in inflammation and allergy. Journal of Allergy 2013: 672381. 13. Papaioannou AI, Kostikas K, Kollia P, et al. (2006) Clinical implications for vascular endothelial growth factor in the lung: Friend or foe? Respiratory Research 17: 128–134. 14. Gruber BL, Marchese MJ and Kew R (1995) Angiogenic factors stimulate mast-cell migration. Blood 86: 2488–2493. 15. Feistritzer C, Kaneider NC, Sturn DH, et al. (2004) Expression and function of the vascular endothelial growth factor receptor FLT-1 in human eosinophils. American Journal of Respiratory Cell and Molecular Biology 30: 729–735. 16. Shulman K, Rosen S, Tognazzi K, et al. (1996) Expression of vascular permeability factor (VPF/ VEGF) is altered in many glomerular diseases. Journal of the American Society of Nephrology 7: 661–666. 17. Papadaki G, Bakakos P, Kostikas K, et al. (2013) Vascular endothelial growth factor and cysteinyl leukotrienes in sputum supernatant of patients with asthma. Respiratory Medicine 107: 1339–1345. 18. Meyer N, Christoph J, Makrinioti H, et al. (2012) Inhibition of angiogenesis by IL-32: Possible role in
6 asthma. Journal of Allergy and Clinical Immunology 129: 964–973. 19. Lopez-Guisa JM, Powers C, File D, et al. (2012) Airway epithelial cells from asthmatic children differentially express proremodeling factors. Journal of Allergy and Clinical Immunology 129: 990–997. 20. Wen FQ, Liu X, Manda W, et al. (2003) TH2 Cytokine- enhanced and TGF-beta-enhanced vascular endothelial growth factor production by cultured human airway smooth muscle cells is attenuated by IFN-gamma and corticosteroids. Journal of Allergy and Clinical Immunology 111: 1307–1318. 21. Clifford RL, John AE, Brightling CE, et al. (2012) Abnormal histone methylation is responsible for increased vascular endothelial growth factor 165a secretion from airway smooth muscle cells in asthma. Journal of Immunology 189: 819–831. 22. Meyer N and Akdis CA (2013) Vascular endothelial growth factor as a key inducer of angiogenesis in the asthmatic airways. Current Allergy and Asthma Reports 13: 1–9. 23. Harfouche R, Hasséssian HM, Guo Y, et al. (2002) Mechanisms which mediate the antiapoptotic effects of angiopoietin-1 on endothelial cells. Microvascular Research 64: 135–147. 24. Bhandari V, Choo-Wing R, Harijith A, et al. (2012) Increased hyperoxia-induced lung injury in nitric
International Journal of Immunopathology and Pharmacology oxide synthase 2 null mice is mediated via angiopoietin 2. American Journal of Respiratory Cell and Molecular Biology 46: 668–676. 25. Nomura S, Kanazawa H, Hirata K, et al. (2005) Relationship between vascular endothelial growth factor and angiopoietin-2 in asthmatics before and after inhaled beclomethasone therapy. Journal of Asthma 42: 141–146. 26. Papi A, Corradi M, Pigeon-Francisco C, et al. (2013) Beclometasone-formoterol as maintenance and reliever treatment in patients with asthma: A doubleblind, randomised controlled trial. Lancet Respiratory Medicine 1: 23–31. 27. Durrani SR, Viswanathan RK and Busse WW (2011) What effect does asthma treatment have on airway remodeling? Current perspectives. Journal of Allergy and Clinical Immunology 128: 439–448. 28. Feltis BN, Wignarajah D, Reid DW, et al. (2007) Effects of inhaled fluticasone on angiogenesis and vascular endothelial growth factor in asthma. Thorax 62: 314–319. 29. Sun Y, Wang J, Li H, et al. (2013) The effects of budesonide on angiogenesis in a murine asthma model. Archives of Medical Science 9: 361–367. 30. Roth M, Zhong J, Zumkeller C, et al. (2013) The role of IgE-receptors in IgE-dependent airway smooth muscle cell remodelling. PLoS One 8: e56015.
Letter to the editor
Angiogenesis in bronchial asthma Krzysztof Pałgan and Zbigniew Bartuzi
International Journal of Immunopathology and Pharmacology 1–6 © The Author(s) 2015 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/0394632015580907 iji.sagepub.com
Abstract Bronchial asthma is a chronic inflammatory disease characterised by airflow obstruction that may be reversed spontaneously or in response to treatment. The airway inflammation can lead to structural changes and remodelling consisting of subepithelial layer thickening, airway smooth muscle hyperplasia and angiogenesis. Subepithelial hypervascularity and angiogenesis in the airways are part of the structural airway wall in asthma. Increased vascularity of bronchial mucosa is closely related to the expression of angiogenic factors like vascular endothelial growth factor (VEGF), angiopoietin and hypoxia-inducible factor (HIF). The scope of the present review is to summarise the roles of anagiogenic factors and treatment in vascular development. Keywords angiogenesis, angiopoietin, asthma, HIF, VEGF Date received: 5 September 2014; accepted: 16 March 2015
Introduction Historically, an interest in angiogenesis is associated with studies on cancer and factors involved in tumour progression. In 1971 a study was published by Judah Folkman (1933–2008), a pioneer in angiogenesis research in cancer, which demonstrated the major role of angiogenesis in the pathogenesis of cancer.1 Subsequent studies revealed other medical conditions in which the formation of new blood vessels plays an important role, and described numerous factors influencing angiogenesis. Angiogenesis can be defined as the emergence of new blood vessels from pre-existing vasculature. This phenomenon is observed in physiological conditions (e.g. wound healing, menstruation cycle), but can also play a role in the pathogenesis of diseases such as cancer, psoriasis, rheumatoid arthritis, proliferative retinopathy, obesity, atherosclerosis, ischaemic heart disease, endometriosis and bronchial asthma. Studies devoted to angiogenesis divided the process of new blood vessel formation into several stages. The initial stage of angiogenesis takes place in a pre-existing blood vessel located in close proximity to the wound, an ongoing malignant or inflammatory
process or angiogenic factors. In this stage, endothelial cell activation by growth factors, increased permeability and increased endothelial mitosis are observed. In the second stage, basement membrane is degraded by metalloproteinases. This stage is followed by the migration of endothelial cells towards angiogenic factors, and the formation of branch points and capillary lumen. The last stage involves modelling and stabilisation of the new capillary vessel. Finally, basement membrane is formed, interconnections between endothelial cells develop and pericytes are recruited.2 In 1960, Dunnill revealed increased vascularity in bronchial biopsy specimens from 20 subjects who died of acute asthmatic attacks. The studies on the bronchial vascular bed that followed showed a Department of Allergology, Clinical Immunology and Internal Medicine, Collegium Medicum Bydgoszcz, Nicolaus Copernicus University of Toruń, Poland Corresponding author: Krzysztof Pałgan, Nicolaus Copernicus University, Collegium Medicum in Bydgoszcz, Department of Allergology, Clinical Immunology and Internal Diseases, Ujejskiego 75, 85-168 Bydgoszcz, Poland. Email: [email protected]
2
International Journal of Immunopathology and Pharmacology
larger number of blood vessels in lamina propria and submucosa of the bronchi in patients with mild and severe bronchial asthma. The airways are supplied from blood vessels located in the smooth muscle layer of the bronchi, and through the subepithelial capillary network in lamina propria. In physiological conditions, the bronchial tree shows temporal fluctuation. Similarly, blood vessels of the bronchi undergo changes in permeability, dilation and density in the course of pathological and physiological processes. There is growing interest among scientists in neovascularisation and angiogenesis in patients with bronchial asthma.3
bronchial asthma was conducted by Grigorias et al.,4 and it demonstrated a clear relationship between the clinical stage of the disease and the number of blood vessels within the lamina propria. According to Grigorias,4 patients suffering from severe asthma have 46% more capillary vessels in bronchial submucosa compared to healthy controls. Other studies by Salvato et al.5 suggest a 30% increase in blood vessel count in patients with bronchial asthma. Positive correlation was observed between the number of blood vessels and the severity of bronchial asthma. A well-developed network of blood vessels enables migration of inflammatory cells into the bronchial wall.5
Angiogenesis and bronchial asthma
Angiogenic factors in bronchial asthma
Bronchial asthma is a chronic disease of the airways, the predominant underlying mechanisms of which are bronchial inflammation, hyper-responsiveness and bronchial wall remodelling. Structural changes in the bronchial tree result in increasing bronchial hyper-responsiveness and a progressively more severe course of bronchial asthma. Airway remodelling in patients with bronchial asthma is defined as a sequence of morphological changes involving cells and tissues that constitute bronchioles. Structural changes involve virtually all layers of the bronchial wall, including epithelium, basal membrane, smooth muscles and blood vessels. Chronic inflammatory processes in patients with bronchial asthma, and frequent exacerbations caused mainly by allergen exposure, or viral or bacterial infections, lead to bronchial tree remodelling. Microscopic examination of bronchial biopsy specimens reveals considerable thickening of the bronchiolar walls as well as of the cartilage-containing bronchi. Although virtually all layers of the bronchial wall undergo structural remodelling, it is the bronchial basement membrane that first demonstrates morphological changes. In the course of the process, collagens are deposited in the basement membrane, leading to its thickening and a decrease in elasticity. Both hyperplasia and hypertrophy of smooth muscle cells are observed in the smooth muscle layer. Along with the bronchial wall thickening, new blood vessels emerge. A detailed histological analysis of the bronchial biopsy specimens from patients with mild, moderate or severe chronic
Mast cells The first studies on angiogenesis in cancer identified mast cells as an important source of angiogenic factors. In the initial phase of tumour development, the emergence of numerous mast cell infiltrations at the border between the tumour and the surrounding normal tissue precedes angiogenesis. In type I hypersensitivity, according to Gell and Coombs, mast cells and the mediators they release, such as histamine, platelet-activating factor (PAF), leukotriene B4 (LTB4) and prostaglandin D2 (PDG2), are the main actors of the anaphylactic reactions. Apart from cytokines, such as vascular endothelial growth factor (VEGF), angiogenin, tumour necrosis factor (TNF), interleukine-8 (IL8) and fibroblast growth factor-2 (FGF-2), mast cells release tryptase and chymase, proteolytic enzymes involved in angiogenesis.6
Hypoxia-inducible transcription factor (HIF) Inflammatory process and bronchoconstriction promote hypoxia in patients with bronchial asthma. Ischaemia, inflammatory cells and increased cellularity of the bronchi and bronchioles may lead to the initiation of angiogenesis.7 Tissue ischaemia is a strong trigger for the production of transcription factors referred to as hypoxia-inducible factor-1 (HIF-1). A whole family of proteins has been described, whose role is to minimise cell damage caused by hypoxia.8 HIF-1 is a heterodimer composed of subunits α and β. HIF activity is regulated by prolyl hydroxylase
3
Pałgan and Bartuzi
VEGF-A A
VEGFR-1 V
VEG GF-B
VEGF-C V
VEGF-D
VEGF FR-3
VEGFR-2 2
Endoth helial cells
Angiogen nesis
Lymphangiogenessis
Figure 1. Vascular endothelial growth factors (VEGF-A, VEGF-B, VEGF-C, VEGF-D).13–15
VEGF-A acts by receptors VEGFR-1, VEGFR-2, but activates mainly VEGFR-2 and induce angiogenesis. VEGF-C, VEGF-D activate VEGFR-3 and stimulate lyphangiogenesis.
domain proteins (PHD) responding to oxygen concentrations.9 It has been demonstrated that angiogenesis can occur in normoxic conditions in patients with bronchial asthma. It seems that chronic inflammatory process in the airways and the cytokines, such as transforming growth factor-β (TGF-β), tumour necrosis factor-α (TNF-α), granulocyte-macrophage colonystimulating factor (GM-CSF) or lipopolysaccharides (LPS), can stimulate the production of HIF-1α. Additionally, the animal model shows positive correlation between HIF-1α concentrations and the intensity of bronchial allergic inflammation in mice. HIF-1 is one of the most important activators for genes encoding vascular endothelial growth factor A (VEGF-A).10,11
Vascular endothelial growth factor (VEGF) VEGF was discovered in the late 1970s during research on tumour angiogenesis. VEGF is a cytokine that plays a crucial role in the creation of new blood vessels (vasculogenesis) during embryonic development.12 It was earlier referred to as vascular permeability factor (VPF). Several VEGFs have been described within the cytokine family, including VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E and the placenta growth factor (PlGF). Genes encoding VEGF are located on chromosome 6p21.3.13,14 VEGF acts mainly on the endothelial cells of blood and lymphatic vessels, inducing their migration and division (Figure 1). Inflammatory cells, such as eosinophils, mast cells
and macrophages, express VEGF receptors (VEGF Receptor-1, VEGFR-1). VEGFR-1, in turn, stimulates chemotactic migration of these cells.15 It should also be emphasised that VEGF inhibits the apoptosis of vascular endothelial cells. As mentioned before, this cytokine significantly increases the permeability of the blood vessels – according to Shulman et al.,16 it is 50,000 times more potent than histamine in doing so. Sputum examinations in patients with bronchial asthma revealed significantly elevated concentrations of VEGF in addition to cysteinyl leukotriens (Cyst-LTs).17 Studies on the role of VEGF in the pathogenesis of bronchial asthma showed that this factor can stimulate allergic inflammation in the bronchial tree and contribute to the remodelling process. Positive correlation was shown between VEGF concentrations in the sputum as well as in the lung tissue, and the severity of bronchial asthma.18 Animal model experiments of bronchial asthma proved that VEGF is an important stimulating factor for airway remodelling. According to LopezGuisa et al.,19 airway remodelling in patients with bronchial asthma is initiated by the destruction of bronchial endothelium and the release of VEGF from the endothelial cells. It was shown that Th17 lymphocytes infiltrating bronchial epithelium stimulate VEGF production. Comparative in vitro analysis of bronchial epithelium between children diagnosed with asthma and healthy controls demonstrated an excessive capacity of bronchial
4
epithelial cells in asthmatic patients for VEGF production and release.20 Excessive VEGF production was observed also in bronchial smooth muscle cells. Genetic analyses in patients with bronchial asthma demonstrated epigenetic modifications of histone H3 and H4 involving their hyperacetylation and methylation. Such modification of histone proteins facilitates the access of VEGF promoters such as Sp1 or H3K4me3 to the effector genes, leading to their over expression. Bronchial asthma is frequently exacerbated by rhinovirus infections. These viruses also strongly stimulate VEGF production.21 Recently the role of IL-32 in the pathogenesis of bronchial asthma has been emphasised. IL-32 is produced by a variety of cells, of which the most important in the pathogenesis of asthma are dendritic, endothelial and NK cells and T lymphocytes. IL-32 significantly inhibits VEGF production and bronchial angiogenesis. It has been demonstrated that an increase in serum IL-32 concentrations in patients with bronchial asthma is indicative of effective therapy. Interestingly, IL-32 also shows antiviral and antibacterial properties.22
Angiopoietin Three types of angiopoietins have been described in humans: angiopoietin 1 (Ang-1), angiopoietin 2 (Ang-2) and angiopoietin 4 (Ang-4). Receptors binding these factors are known as Tie1 and Tie2, the latter being capable of binding all angiopoietins. Harfouche et al.23 noticed that angiopoietins are produced mainly by basophils and mast cells, which also express angiopoietin receptors. Interestingly, angiopoietins act as chemotactic agents for mast cells. A particularly high expression of genes involved in the synthesis of angiopoietins is observed in the mast cells of lung tissue.24 The crucial role of angiopoietins can be seen in the final stages of angiogenesis. Ang-1 stimulates migration of pericytes and smooth muscle cells and is therefore responsible for stabilisation of new capillary tubes. Contrary to Ang-2, Ang-1 reduces blood vessel permeability in patients with bronchial asthma. Ang-2 increases the permeability of the vascular bed through elimination of interconnections between endothelial cells and other cells constituting arterioles and venules. According to Bhandari et al.,24 Ang-2 can be responsible for lung oedema following exposure to toxic gases. Another
International Journal of Immunopathology and Pharmacology
difference between these angiopoietins is that Ang-1 is synthesised constitutively and low concentrations of this factor can be found in normal tissues, whereas Ang-2 is produced at the inflammation site only. A relationship is observed between VEGF and angiopoietins. High concentrations of VEGF and Ang-2 are noted in the initial phases of vascular remodelling, as well as angiogenesis and lymphogenesis. Hypoxia is the most important factor regulating transcription of both VEGF and Ang-2. Nomura et al.25 demonstrated high concentrations of angiopoietin and VEGF in the induced sputum of patients with bronchial asthma.
Bronchial asthma treatment and angiogenesis According to current guidelines, inhaled corticosteroids are the most effective anti-inflammatory medications recommended as a first-line treatment for chronic asthma. Corticosteroids inhibit synthesis of many cytokines known to stimulate inflammatory process in the bronchial tree. Numerous studies have emphasised the inhibiting role of inhaled corticosteroids on the migration of inflammatory cells to the bronchi in patients with bronchial asthma.26 Corticosteroids are particularly beneficial in the treatment of eosinophilic asthma. It has been demonstrated that they reduce the number of eosinophils in the bronchial epithelium. Another issue associated with inhaled corticosteroid therapy is its effect on bronchial tree remodelling. Durrani et al.27 noticed that inhaled corticosteroids inhibit structural changes of the basement membrane in patients with bronchial asthma only after 6 months of therapy with very high doses, equivalent to 1–3 mg of budesonide per day. Feltis et al.28 directly analysed angiogenesis and angiogenic factors in 35 treatment-naive patients with mild or moderate bronchial asthma. Study participants were given 750 µg fluticasone (fluticasone propionate) twice daily. After 3 months, VEGF and Ang-1 concentrations, as well as vascular density, were assessed. Fluticasone treatment significantly lowered tissue concentrations of angiogenic factors and reduced the number of capillary vessels by approximately 18.7%. The results of animal studies on angiogenesis in bronchial asthma are much more convincing. Sun et al.29 analysed the effect of budesonide on the
Pałgan and Bartuzi
angiogenesis in mice with induced asthma and proved that it not only suppressed the inflammatory process through blocking HIF-1α and VEGF expression, but also inhibited the development of new blood vessels. These findings showed considerable (over 50%) reduction in the number of blood vessels as well as in serum growth factor concentrations, compared to the control group. Other medications used in the treatment of bronchial asthma, such as beta-2 agonists, leukotriene receptor antagonists (LTRAs) or anticholinergics, do not show any significant effect on airway remodelling.27 Roth et al.30 claimed that airway remodelling in bronchial asthma is associated with overproduction of IgE, which may be responsible for thickening the bronchial basement membrane and directly stimulating proliferation of smooth muscle cells. Clinical studies focusing on the effect of omalizumab, an IgE-binding antibody, on the structural changes of the bronchial tree in patients with moderate and severe asthma seem to confirm its role in airway remodelling. As well as its anti-inflammatory activity, omalizumab also inhibited collagen deposition in the reticular layer of the basement membrane and reduced thickening of the bronchial wall. It can therefore be presumed that angiogenesis also was suppressed. Declaration of conflicting interests The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Funding This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors. References 1. Folkman J (1971) Tumor angiogenesis: Therapeutic implications. New England Journal of Medicine 285: 1182–1186. 2. Pałgan K (1993) Actual views on neoplasm angiogenesis. Postępy Higieny i Medycyny Doświadczalnej 47: 193–207. 3. Dunnill MS (1960) The pathology of asthma, with special reference to changes in the bronchial mucosa. Journal of Clinical Pathology 13: 27–33. 4. Grigoraş A, Căruntu ID, Grigoraş CC, et al. (2012) Relationship between immunohistochemical assessment of bronchial mucosa microvascularization and clinical
5 stage in asthma. Romanian Journal of Morphology and Embryology 53: 485–490. 5. Salvato G (2001) Quantitative and morphological analysis of the vascular bed in bronchial biopsy specimens from asthmatic and non-asthmatic subjects. Thorax 56: 902–906. 6. Pałgan K, Kołakowska J and Zbikowska-Gotz M (2000) The role of mast cells in inflammatory reactions. Pneumonologia i Alergologia Polska 68: 603– 609. 7. Scholz CC and Taylor CT (2013) Targeting the HIF pathway in inflammation and immunity. Current Opinion in Pharmacology 13: 646–653. 8. Mandl M and Depping R (2014) Hypoxia-inducible aryl hydrocarbon receptor nuclear translocator (ARNT) (HIF-1β): A rare exception? Molecular Medicine 20: 215–220. 9. Dehne N and Brune B (2009) HIF-1 in the inflammatory microenvironment. Experimental Cell Research 315: 1791–1797. 10. Huerta-Yepez S, Baay-Guzman GJ, Bebenek IG, et al. (2011) Hypoxia inducible factor promotes murine allergic airway inflammation and is increased in asthma and rhinitis. Allergy 66: 909–919. 11. Kuschel A, Simon P and Tug S (2012) Functional Regulation of HIF-1alpha under normoxia - is there more than posttranslational regulation? Journal of Cellular Physiology 227: 514–522. 12. Zgraggen S, Ochsenbein AM and Detmar M (2013) An important role of blood and lymphatic vessels in inflammation and allergy. Journal of Allergy 2013: 672381. 13. Papaioannou AI, Kostikas K, Kollia P, et al. (2006) Clinical implications for vascular endothelial growth factor in the lung: Friend or foe? Respiratory Research 17: 128–134. 14. Gruber BL, Marchese MJ and Kew R (1995) Angiogenic factors stimulate mast-cell migration. Blood 86: 2488–2493. 15. Feistritzer C, Kaneider NC, Sturn DH, et al. (2004) Expression and function of the vascular endothelial growth factor receptor FLT-1 in human eosinophils. American Journal of Respiratory Cell and Molecular Biology 30: 729–735. 16. Shulman K, Rosen S, Tognazzi K, et al. (1996) Expression of vascular permeability factor (VPF/ VEGF) is altered in many glomerular diseases. Journal of the American Society of Nephrology 7: 661–666. 17. Papadaki G, Bakakos P, Kostikas K, et al. (2013) Vascular endothelial growth factor and cysteinyl leukotrienes in sputum supernatant of patients with asthma. Respiratory Medicine 107: 1339–1345. 18. Meyer N, Christoph J, Makrinioti H, et al. (2012) Inhibition of angiogenesis by IL-32: Possible role in
6 asthma. Journal of Allergy and Clinical Immunology 129: 964–973. 19. Lopez-Guisa JM, Powers C, File D, et al. (2012) Airway epithelial cells from asthmatic children differentially express proremodeling factors. Journal of Allergy and Clinical Immunology 129: 990–997. 20. Wen FQ, Liu X, Manda W, et al. (2003) TH2 Cytokine- enhanced and TGF-beta-enhanced vascular endothelial growth factor production by cultured human airway smooth muscle cells is attenuated by IFN-gamma and corticosteroids. Journal of Allergy and Clinical Immunology 111: 1307–1318. 21. Clifford RL, John AE, Brightling CE, et al. (2012) Abnormal histone methylation is responsible for increased vascular endothelial growth factor 165a secretion from airway smooth muscle cells in asthma. Journal of Immunology 189: 819–831. 22. Meyer N and Akdis CA (2013) Vascular endothelial growth factor as a key inducer of angiogenesis in the asthmatic airways. Current Allergy and Asthma Reports 13: 1–9. 23. Harfouche R, Hasséssian HM, Guo Y, et al. (2002) Mechanisms which mediate the antiapoptotic effects of angiopoietin-1 on endothelial cells. Microvascular Research 64: 135–147. 24. Bhandari V, Choo-Wing R, Harijith A, et al. (2012) Increased hyperoxia-induced lung injury in nitric
International Journal of Immunopathology and Pharmacology oxide synthase 2 null mice is mediated via angiopoietin 2. American Journal of Respiratory Cell and Molecular Biology 46: 668–676. 25. Nomura S, Kanazawa H, Hirata K, et al. (2005) Relationship between vascular endothelial growth factor and angiopoietin-2 in asthmatics before and after inhaled beclomethasone therapy. Journal of Asthma 42: 141–146. 26. Papi A, Corradi M, Pigeon-Francisco C, et al. (2013) Beclometasone-formoterol as maintenance and reliever treatment in patients with asthma: A doubleblind, randomised controlled trial. Lancet Respiratory Medicine 1: 23–31. 27. Durrani SR, Viswanathan RK and Busse WW (2011) What effect does asthma treatment have on airway remodeling? Current perspectives. Journal of Allergy and Clinical Immunology 128: 439–448. 28. Feltis BN, Wignarajah D, Reid DW, et al. (2007) Effects of inhaled fluticasone on angiogenesis and vascular endothelial growth factor in asthma. Thorax 62: 314–319. 29. Sun Y, Wang J, Li H, et al. (2013) The effects of budesonide on angiogenesis in a murine asthma model. Archives of Medical Science 9: 361–367. 30. Roth M, Zhong J, Zumkeller C, et al. (2013) The role of IgE-receptors in IgE-dependent airway smooth muscle cell remodelling. PLoS One 8: e56015.
