Katarzyna Wójcik

Lovastatin-induced decrease of intracellular cholesterol level attenuates fibroblast-to-myofibroblast transition in bronchial fibroblasts derived from asthmatic patients

Chronic inflammation of the airways and structural changes in the bronchial wall are basic hallmarks of asthma. Human bronchial fibroblasts derived from patients with diagnosed asthma display in vitro predestination towards TGF-β-induced fibroblast-to-myofibroblast transition (FMT), a key event in the bronchial wall remodelling. Statins inhibit 3-hydroxymethyl-3-glutaryl coenzyme A reductase, a key enzyme in the cholesterol synthesis pathway and are widely used as antilipidemic drugs. The pleiotropic anti-inflammatory effects of statins, independent of their cholesterol-lowering capacity, are also well established. Since commonly used anti-asthmatic drugs do not reverse the structural remodelling of the airways and statins have tentative anti-asthmatic activity, we have studied the effect of lovastatin on FMT in populations of human bronchial fibroblasts derived from asthmatic patients. We demonstrate that the intensity of FMT induced by TGF-β1 was strongly and dose-dependently attenuated by lovastatin. Furthermore, we show that neither the suppression of prenylation of signalling proteins nor the effect on reactive oxygen species formation are important for lovastatin-induced inhibition of myofibroblast differentiation. On the other hand, we show that a squalene synthase inhibitor, zaragozic acid A, reduced the TGF-β1-induced FMT to an extent comparable to lovastatin effect. Additionally we demonstrate that in bronchial fibroblast populations, both inhibitors (lovastatin and zaragozic acid A) attenuate the TGF-β1-induced Smad2 nuclear translocation in a manner dependent on intracellular cholesterol level. Our data suggest that statins can directly, by decrease of intracellular cholesterol level, affect basic cell signalling events crucial for asthmatic processes and potentially prevent perilous bronchial wall remodelling associated with intensive myofibroblast formation.

Lithium Attenuates TGF-β1-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients

Bronchial asthma is a chronic disorder accompanied by phenotypic transitions of bronchial epithelial cells, smooth muscle cells, and fibroblasts. Human bronchial fibroblasts (HBFs) derived from patients with diagnosed asthma display predestination towards TGF-β-induced phenotypic switches. Since the interference between TGF-β and GSK-3β signaling contributes to pathophysiology of chronic lung diseases, we investigated the effect of lithium, a nonspecific GSK-3β inhibitor, on TGF-β 1-induced fibroblast to myofibroblast transition (FMT) in HBF and found that the inhibition of GSK-3β attenuates TGF-β 1-induced FMT in HBF populations derived from asthmatic but not healthy donors. Cytoplasmically sequestrated β-catenin, abundant in TGF-β 1/LiCl-stimulated asthmatic HBFs, most likely interacts with and inhibits the nuclear accumulation and signal transduction of Smad proteins. These data indicate that the specific cellular context determines FMT-related responses of HBFs to factors interfering with the TGF-β signaling pathway. They may also provide a mechanistic explanation for epidemiological data revealing coincidental remission of asthmatic syndromes and their recurrence upon the discontinuation of lithium therapy in certain psychiatric diseases.

Undifferentiated Bronchial Fibroblasts Derived from Asthmatic Patients Display Higher Elastic Modulus than Their Non-Asthmatic Counterparts

During asthma development, differentiation of epithelial cells and fibroblasts towards the contractile phenotype is associated with bronchial wall remodeling and airway constriction. Pathological fibroblast-to-myofibroblast transition (FMT) can be triggered by local inflammation of bronchial walls. Recently, we have demonstrated that human bronchial fibroblasts (HBFs) derived from asthmatic patients display some inherent features which facilitate their FMT in vitro. In spite of intensive research efforts, these properties remain unknown. Importantly, the role of undifferentiated HBFs in the asthmatic process was systematically omitted. Specifically, biomechanical properties of undifferentiated HBFs have not been considered in either FMT or airway remodeling in vivo. Here, we combine atomic force spectroscopy with fluorescence microscopy to compare mechanical properties and actin cytoskeleton architecture of HBFs derived from asthmatic patients and non-asthmatic donors. Our results demonstrate that asthmatic HBFs form thick and aligned ‘ventral’ stress fibers accompanied by enlarged focal adhesions. The differences in cytoskeleton architecture between asthmatic and non-asthmatic cells correlate with higher elastic modulus of asthmatic HBFs and their increased predilection to TGF-β-induced FMT. Due to the obvious links between cytoskeleton architecture and mechanical equilibrium, our observations indicate that HBFs derived from asthmatic bronchi can develop considerably higher static tension than non-asthmatic HBFs. This previously unexplored property of asthmatic HBFs may be potentially important for their myofibroblastic differentiation and bronchial wall remodeling during asthma development.

Triterpene saponosides from Lysimachia ciliata differentially attenuate invasive potential of prostate cancer cells.

Neither androgen ablation nor chemotherapeutic agents are effective in reducing the risk of prostate cancer progression. On the other hand, multifaceted effects of phytochemicals, such as triterpene saponins, on cancer cells have been suggested. A promising safety and tolerability profile indicate their possible application in the treatment of advanced prostate cancers. We analyzed the specificity, selectivity and versatility of desglucoanagalloside B effects on human prostate cancer cells derived from prostate cancer metastases to brain (DU-145 cells) and bone (PC-3 cells). Prominent growth arrest and apoptotic response of both cell types was observed in the presence of sub-micromolar desglucoanagalloside B concentrations. This was accompanied by cytochrome c release and caspase 3/7 activation. A relatively low cytostatic and pro-apoptotic response of cancer cells to a desglucoanagalloside B analog, anagallosaponin IV, illustrated the specificity of the effects of desglucoanagalloside B, whereas the low sensitivity of normal prostate PNT2 cells to desglucoanagalloside B showed the selectivity of its action. Inhibition of cancer cell motility was observed in the presence of both saponins, however only desglucoanagalloside B attenuated cancer cell invasive potential, predominantly through an effect on cell elastic properties. These data demonstrate the versatility of its effects on prostate cancer cells. In contrast to PNT2 cells, cancer cells tested in this study were relatively resistant to mitoxantrone. The multifaceted action of desglucoanagalloside B on basic cellular traits, crucial for prostate cancer progression, opens perspectives for elaboration of combined palliative therapies and new prostate cancer prophylaxis regimens.

Trends and dynamics of changes in aortic pulse wave velocity over one-year observation period in patients treated with peritoneal dialysis.

important co-morbid condition affecting patients with chronic renal failure (1). There is an increasing body of evidence suggesting that mortality on dialysis largely depends upon initial and follow-up serum levels of proinflammatory cytokines and acute phase proteins (2, 3). Chronic inflammation is closely associated with atherosclerosis, which to a cer tain extent may be considered an inflammatory disease (4). Aortic pulse wave velocity (AoPWV) assessment proved its cl inical usefulness in patients with chronic renal disease, including those on maintenance hemodialysis (5-9). For uncertain reasons this problem was seldom studied in peritoneal dialysis (PD) patients. The aim of the present study was to assess the trends and dynamics of changes in AoPWV in the group of stable PD patients over one year and to search for possible correlations between these changes and one-year profile of selected cytokines, acute phase proteins and other variables that may be considered as risk factors for atherosclerosis progression. Forty seven patients (25M, 22F, aged mean 52.7 ± 12.8 years and on dialysis for 19.1 ± 21.6 months) were analyzed. AoPWV was measured using an automatic device with two pressure transducers (TY-306; Fukuda Denshi) placed on the carotid and femoral arteries and connected to an automatic processor (Complior® Colson). AoPWV was assessed at baseline and after 12 months. In each patient an absolute difference in PWV between two mentioned time-points was calculated (∆AoPWV). Serum Interleukin 6 (IL-6) and Tumor Necrosis Factor alpha (TNFα) were analyzed as proinflammatory cytokines (ELISA; Quantikine, R&D Systems), and C-reactive protein (high sensitive assay), lipoprotein (a) and fibrinogen – as acute phase reactants (immuno-nephelometry; Dade Boehring). In all patients total, LDL-, HDL-cholesterol and triglyceride serum levels, as well as serum calcium and phosphate were measured (Hitachi 917). All biochemical and inflammatory parameters were assessed at baseline, and after 6 and 12 months. At the end of the study mean concentrations of all analyzed biochemical parameters from three assessments was calculated. Blood pressure was measured at baseline, after 6 and 12 months, and mean arterial pressure (MAP) as well as pulse pressure (PP) were calculated. Mean AoPWV did not change significantly for the whole study group after one year (from 11.2 ± 2.26 to 11.7 ± 2.4 m/s, p= 0.07; ∆AoPWV +0.48 ± 1.77 m/s). However, in the population studied two categories of patients might be distinguished: subjects who progressed in AoPWV (29 patients, 61.7%, from 10.6 ± 2.3 to 12.3 ± 2.65 m/s; p< 0.05; ∆AoPWV +1.6 ± 1.14) and those with regression after 12 months (18 patients, 38%, from 12.2 ± 1.89 to 10.8 ± 1.67 m/s; p< 0.05; ∆AoPWV -1.31 ± 0.89). The entire group ‘averaged’ in terms of AoPWV value after 12 months, as the differences between “Pregressors” and “Regressors” were significant at the study commencement (p< 0.01) and only borderline significant (p= 0.08) at its completion. Patients who progressed were character ized by significantly higher values of mean serum phosphates, TNFα and Lp (a) levels as well as mean PP calculated from three assessments in comparison to “Regressors” (p< 0.05 for all differences). No significant difference was found between groups in terms of age, PD duration, and Short Communication The International Journal of Artificial Organs / Vol. 27 / no. 10, 2004/ pp. 904-906