Cytokines and therapy in COPD: a promising combination?

COPD is a major health problem, with patients showing a progressively declining, largely irreversible, change in lung function. This is associated with chronic airways inflammation and structural remodeling, including loss of alveolar walls, and goblet cell metaplasia with mucus hypersecretion. Inflammatory cells may contribute to the airway remodeling via secretion of proteases, fibrotic or mitogenic growth factors, and cytokines. In turn, airway remodeling may contribute to the clinical symptoms of COPD. Currently available therapies are directed to improvement of clinical symptoms and reduction of the airways inflammation. The commonly used glucocorticosteroids are expected to reduce the inflammation by acting on kinases or transcription factors necessary for expression of pro-inflammatory cytokines or chemokines. However, several long-term and short-term studies showed that glucocorticosteroids are rather ineffective in improving lung function and reducing the airway inflammation in patients with COPD. New therapeutic strategies may reduce the inflammation and alleviate the clinical symptoms of COPD. Tumor necrosis factor-alpha, interleukin-8, and monocyte chemoattractant protein-1 are important chemotactic proteins for macrophages and neutrophils, the predominant inflammatory cells associated with COPD. As lung levels of these cytokines are higher in COPD compared to non-COPD patients, they may represent targets for novel therapies

Growth factors in bladder epithelium : a study on the expression and functions of growth factors in mouse urothelium

The urogenital tract is one of the major excretory paths for small metabolites and ions. The excreted liquid, urine, is produced by the kidneys, flows through the ureters into the urinary bladder and is finally excreted through the urethra. The mammalian urinary bladder has the important capacity to retain urine for some time. Normally, the bladder is resistant to toxic effects of products in the urine. But after damage to the epithelium of the bladder, bladder epithelial cells may become vulnerable to xenotoxic agents and potential toxic metabolites in the urine. In general, this damage will be repaired by physiological processes. During neoplasia, e.g. in transitional cell carcinoma, aberrations in physiologically regulated processes occur. This thesis focuses on protein factors that may be involved in the physiology of transitional epithelium of the mouse bladder, and their specific functions. The following paragraphs highlight subsequently the structure and function of the bladder (§ 1) the physiology and causes of abnormal growth of the urothelium (§2), proteins that have been shown to be involved in the physiology of normal and tumour cells (§3), what is currently known about these proteins with respect to the bladder (§3.5), and finally the aim of the thesis (§4)

An in vitro model of urothelial regeneration: Effects of growth factors and extracellular matrix proteins

Although the cellular turnover of resting urothelium is very low, its regenerative capacity is known to be outstanding. In organotypic mouse urothelial cultures closely mimicking the differentiation and multilayering of normal urothelium, we examined the cell biological mechanisms underlying urothelial regeneration and the specific role of growth factors and several extracellular matrix (ECM) components. Exposure to epidermal growth factor (EGF) and acidic fibroblast growth factor (aFGF) and culture on laminin resulted in enhanced expansion of the urothelium. Microscopy and assessment of proliferative activity revealed that enhanced urothelial expansion due to EGF could be attributed to increased proliferative activity and an increase in cell numbers, whereas aFGF-stimulated expansion must be considered the consequence of increased cellularity and migration. Laminin-enhanced urothelial expansion was shown to be the result of spreading of the entire urothelial organotypic culture. This was associated with a considerable decrease in the number of cell layers. A synergistic effect of growth factors and laminin was not found. This organotypic urothelial cell culture model seems to be very useful in studying strategies to improve urothelial regeneration

Hyperplasia of epithelium adjacent to transitional cell carcinoma can be induced by growth factors through paracrine pathways

Hyperplasia of transitional cell epithelium adjacent to human transitional cell carcinomas (TCC) is a common finding in pathology. This hyperplasia may be a precancerous aberration. Alternatively, it has been suggested that the hyperplasia is due to paracrine action of tumour-derived growth factors. In this study we tested the latter hypothesis using the mouse tumorigenic TCC cell line NUC-1. Transplantation of NUC-1 tumour cells into the urinary bladder submucosa of syngeneic mice in vivo induced hyperplasia of normal adjacent urothelium in all tested mice. Implantation of normal mouse bladder mucosa did not induce urothelial hyperplasia. In vitro, conditioned medium of NUC-1 cells induced the proliferation of the mouse urothelial cell line g/G, which closely resembles normal urothelial cells. This induction was inhibited by transforming growth factor β1 (TGFβ1). Similarly, TGFβ1 inhibited the fibroblast growth factor-1 (FGF-1) and FGF-2 induced proliferation of g/G cells. Chemico-physical examination, bioassays with conditioned media, and RNA analysis of NUC-1 cells revealed that these cells secreted a growth factor with FGF-like properties. These results indicate that epithelial hyperplasia surrounding carcinomas is not necessarily a precancerous aberration, but may result from direct paracrine action of tumour-derived growth factors

Enhanced expression of fibroblast growth factors and receptor FGFR-1 during vascular remodeling in chronic obstructive pulmonary disease

Important characteristics of chronic obstructive pulmonary disease (COPD) include airway and vascular remodeling, the molecular mechanisms of which are poorly understood. We assessed the role of fibroblast growth factors (FGF) in pulmonary vascular remodeling by examining the expression pattern of FGF-1, FGF-2, and the FGF receptor (FGFR-1) in peripheral area of lung tissues from patients with COPD (FEV(1) < or = 75%; n = 15) and without COPD (FEV(1) > or = 85%; n = 13). Immunohistochemical staining results were evaluated by digital video image analysis as well as by manual scoring. FGF-1 and FGFR-1 were detected in vascular smooth muscle (VSM), airway smooth muscle, and airway epithelial cells. FGF-2 was localized in the cytoplasm of airway epithelium and in the nuclei of airway smooth muscle, VSM, and endothelial cells. In COPD cases, an unequivocal increase in FGF-2 expression was observed in VSM (3-fold, P = 0.001) and endothelium (2-fold, P = 0.007) of small pulmonary vessels with a luminal diameter under 200 micro m. In addition, FGFR-1 levels were elevated in the intima (1.5-fold, P = 0.05). VSM cells of large (> 200 micro m) pulmonary vessels showed increased staining for FGF-1 (1.6-fold, P < 0.03) and FGFR-1 (1.4-fold, P < 0.04) in COPD. Pulmonary vascular remodeling, assessed as the ratio of alpha-smooth muscle actin staining and vascular wall area with the lumen diameter, was increased in large vessels of patients with COPD (P = 0.007) and was inversely correlated with FEV(1) values (P < 0.007). Our results suggest an autocrine role of the FGF-FGFR-1 system in the pathogenesis of COPD-associated vascular remodeling

Characterization of distinct functions for growth factors in murine transitional epithelial cells in primary organotypic culture

Although previous studies indicate that growth factors can affect several physiological processes in epithelia. their role in the biological dynamics of transitional epithelium of the bladder is not yet established. This study investigates the functional consequences of a direct action of EGF, TGFβ, FGF-1, FGF-2, PDGF-AA, and PDGF-BB on mouse urothelium in organoid-like primary cultures. Confluent and nonconfluent cultures served as a model for intact and regenerating urothelium, respectively, EGF and FGF-1 stimulated in both models under serum-free conditions the BrdU and [3H]-thymidine incorporation. This resulted in an increase in the number of cell layers, but the cultures assumed a less organoid-like morphology. In addition, EGF and FGF-1 stimulated the expansion of nonconfluent cultures. TGFβ inhibited proliferation, caused a decrease in the number of cell layers, and blocked expansion. Moreover, TGFβ induced the terminal differentiation and apoptosis of urothelial cells. In nonconfluent cultures PDGF-BB slightly stimulated the increase in the outgrowth area, but no other effect on the parameters for proliferation and differentiation was observed. FGF-2 and PDGF-AA did not affect any of the studied parameters. These data are consistent with the hypothesis that EGF and FGF-1 can promote wound healing and/or hyperplasia through direct action on the epithelial cells, while TGFβ promotes the development of a normal, differentiated transitional epithelium

Altered expression of epithelial junctional proteins in atopic asthma: Possible role in inflammation

Epithelial cells form a tight barrier against environmental stimuli via tight junctions (TJs) and adherence junctions (AJs). Defects in TJ and AJ proteins may cause changes in epithelial morphology and integrity and potentially lead to faster trafficking of inflammatory cells through the epithelium. Bronchial epithelial fragility has been reported in asthmatic patients, but little is known about the expression of TJ and AJ proteins in asthma. We studied epithelial expression of zonula occludens-1 (ZO-1) and AJ proteins E-cadherin, α-catenin, and β-catenin in bronchial biopsies from nonatopic nonasthmatic (healthy) subjects (n = 14), and stable atopic asthmatic subjects (n = 22) at baseline conditions. Immunostaining for these proteins was semi-quantified for separate cellular compartments. E-cadherin, α-catenin and β-catenin were present in the cellular membrane and less in the cytoplasm. Only β-catenin was present in the nucleus in agreement with its potential function as transcription factor. ZO-1 was present in the apicolateral membrane of superficial cells. α-Catenin expression was significantly lower in subjects with asthma than without and correlated inversely with numbers of eosinophils within the epithelium. ZO-1 and E-cadherin expression were significantly lower in asthmatic than in nonasthmatic subjects. Expression of β-catenin was not different. Our results suggest that the lower epithelial α-catenin, E-cadherin and (or) ZO-1 expression in patients with atopic asthma contributes to a defective airway epithelial barrier and a higher influx of eosinophils in the epithelium