Longitudinal and Comparative Measures of Serum Chitotriosidase and YKL-40 in Patients With Idiopathic Pulmonary Fibrosis.

Background: Although chitin is absent in humans, chitinases are present in healthy subjects and show dysregulated expression in a variety of diseases resulting from abnormal tissue injury and repair responses. It was shown that chitotriosidase (chitinase 1/CHIT1) and structurally-related chitinase 3-like 1 protein (CHI3L1/YKL-40) play important roles in the pathobiology of idiopathic pulmonary fibrosis (IPF), however little is known about their longitudinal serum levels and relationship to clinical measures in IPF. Methods: The present study is the first to evaluate serial measurements of serum CHIT1 activity and YKL-40 concentrations in patients with IPF starting antifibrotic treatment and followed up for 24 months. In addition, baseline serum CHIT1 and YKL-40 were compared between patients with IPF and control subjects, and possible CHIT1 and YKL-40 relationships to longitudinal clinical assessments in IPF were explored. Results: Baseline serum CHIT1 activity and YKL-40 concentrations were significantly elevated in patients with IPF compared to control subjects and showed similar discriminatory ability in distinguishing IPF from controls. No significant differences between the median serum CHIT1 activity and YKL-40 concentration measured over a study follow-up were noted. We found significantly elevated baseline serum CHIT1 activity in the progressors compared with the stables in the first year, while significantly increased baseline serum CHIT1 activity was noted in the stables compared to the progressors in the second year. Additionally, we observed a significant negative correlation between a change in serum YKL-40 concentration and a change in forced vital capacity (FVC) % predicted (% pred.) in the stables subgroup, whereas, a change in serum CHIT1 activity correlated negatively with a change in FVC% pred. in the progressors subgroup. Conclusions: This explorative study findings add further evidence that CHIT1 and YKL-40 are upregulated in patients with IPF, and suggest that longitudinally stable serum CHIT1 activity and YKL-40 concentration levels may potentially be associated with the antifibrotic treatment response. In addition, our findings are supporting the possible role of CHIT1 and YKL-40 as candidate diagnostic and prognostic biomarkers in IPF. Further research is needed to validate present study findings.

A New Biotechnology Method of Bioelements' Accumulation Monitoring in In Vitro Culture of Agaricus bisporus.

Agaricus bisporus (J.E. Lange) Imbach is one the most popular species of edible mushrooms in the world because of its taste and nutritional properties. In the research, repeatability of accumulation of bioelements and biomass yield in experimentally chosen in vitro culture medium, was confirmed. The in vitro cultures were conducted on the modified Oddoux medium enriched with bioelements (Mg, Zn, Cu, Fe). The aim of the study was to create an effective method of sampling, which enabled non-invasive monitoring of metals concentrations changes in the medium, during increase of biomass in in vitro cultures. The first, indirect method of sampling was applied. The non-invasive probe (a dipper) for in vitro culture was used; hence, the highest biomass increase and metals accumulation were gained. The method also guaranteed culture sterility. The second method, a direct one, interfered the in vitro culture conditions and growth of mycelium, and as a consequence the lower biomass increase and metals' accumulation were observed. Few cases of contaminations of mycelium in in vitro cultures were observed. The proposed method of non-invasive sampling of the medium can be used to monitor changes in the concentrations of metals in the medium and their accumulation in the mycelium in natural environment. Changes in concentrations of the selected metals over time, determined by the method of atomic absorption spectrometry, made it possible to correlate the obtained results with the specific stages of A. bisporus mycelium development and to attempt to explain the mechanism of sampling metals from the liquid substrate.

Serial Measurements of Circulating KL-6, SP-D, MMP-7, CA19-9, CA-125, CCL18, and Periostin in Patients with Idiopathic Pulmonary Fibrosis Receiving Antifibrotic Therapy: An Exploratory Study.

Idiopathic pulmonary fibrosis (IPF) is a progressive and inevitably fatal disease with a heterogeneous clinical course. This study aimed to evaluate the usefulness of circulating biomarkers in routine IPF clinical practice. We conducted an exploratory study in a cohort of 28 IPF subjects qualified for anti-fibrotic therapy with up to 24 months serial measurements of seven IPF biomarkers, including those that are well-established, Krebs von den Lungen-6 (KL-6), surfactant protein D (SP-D), matrix metalloproteinase 7 (MMP-7), and more recently introduced ones, cancer antigen 19-9 (CA19-9), cancer antigen 125 (CA-125), chemokine (C-C motif) ligand 18 (CCL18), and periostin. Among studied biomarkers, SP-D had the highest diagnostic accuracy to differentiate IPF subjects from controls, followed by MMP-7 and KL-6. At each study timepoint, KL-6 levels correlated inversely with forced vital capacity % predicted (FVC% pred.), and transfer factor of the lung for carbon monoxide % predicted (TL,CO% pred.), while SP-D levels correlated inversely with FVC% pred. and TL,CO% pred. at 24 months of anti-fibrotic therapy. Baseline KL-6 and CA19-9 concentrations were significantly elevated in patients with progressive disease in comparison to patients with stable disease. In addition, in the progressors subgroup CA19-9 concentrations significantly increased over the second year of study follow-up. In patients with progressive disease, we observed a significant inverse correlation between a change in SP-D levels and a change in FVC% pred. in the first year of treatment, whereas in the second year a significant inverse correlation between a change in KL-6 levels and a change in FVC% pred. was noted. Our study findings support the view that both well-established IPF biomarkers, including KL-6, SP-D, and MMP-7, and more recently introduced ones, like CA19-9, have the potential to support clinical practice in IPF.

The role of bronchoscopy in diagnosis of chronic cough in adults: a retrospective single-center study.

INTRODUCTION: Cough is one of the most frequent symptoms reported to pulmonologists. The role of bronchoscopy in the diagnostic work-up of chronic cough is not clearly defined. The aim of this study was to evaluate the utility of fiberoptic bronchoscopy (FOB) and additional testing of samples collected during FOB in the differential diagnosis of chronic cough in adults. MATERIAL AND METHODS: This was a single-center retrospective study. Out of 7115 conventional white light FOB examinations, we finally selected 198 with cough as the only indication. RESULTS: In 40.9% of bronchoscopic examinations, no visible cause of cough was found. Visual signs of chronic bronchitis (CB) were detected in 57.6% of reports. Only in 3 cases (1.5%) bronchoscopy revealed a potential cause of chronic cough other than CB. Mycobacterium tuberculosis or other mycobacteria were spotted in none of the samples. In 91.1% of bronchoalveolar lavage (BAL) cytologic examinations, at least one cell count abnormality was detected, but only in case of increased percentage of eosinophils, it might be considered clinically relevant. In 53% of bacteriological culture results, at least one potentially pathogenic bacterium was isolated. CONCLUSIONS: The present study results strengthen the evidence that FOB combined with additional testing of airway specimens obtained during FOB is not a powerful tool in the differential diagnosis of chronic cough, and FOB as a diagnostic tool may be overused. The appropriate timing and decision regarding referral for FOB and additional testing of achieved material requires careful clinical consideration.

A multicentre retrospective observational study on Polish experience of pirfenidone therapy in patients with idiopathic pulmonary fibrosis: the PolExPIR study.

BACKGROUND: Pirfenidone is an antifibrotic agent approved for the treatment of idiopathic pulmonary fibrosis (IPF). The drug is available for Polish patients with IPF since 2017. The PolExPIR study aimed to describe the real-world data (RWD) on the Polish experience of pirfenidone therapy in IPF with respect to safety and efficacy profiles. METHODS: This was a multicentre, retrospective, observational study collecting clinical data of patients with IPF receiving pirfenidone from January 2017 to September 2019 across 10 specialized pulmonary centres in Poland. Data collection included baseline characteristics, pulmonary function tests (PFTs) results and six-minute walk test (6MWT). Longitudinal data on PFTs, 6MWT, adverse drug reactions (ADRs), treatment persistence, and survival were also collected up to 24 months post-inclusion. RESULTS: A total of 307 patients receiving pirfenidone were identified for analysis. The mean age was 68.83 (8.13) years and 77% were males. The median time from the first symptoms to IPF diagnosis was 15.5 (9.75-30) months and from diagnosis to start of pirfenidone treatment was 6 (2-23) months. Patients were followed on treatment for a median of 17 (12-22.75) months. Seventy-four patients (24.1%) required dose adjustments and 35 (11.4%) were chronically treated with different than the full recommended dose. A total of 141 patients (45.92%) discontinued therapy due to different reasons including ADRs (16.61%), death (8.79%), disease progression (6.51%), patient's own request (5.54%), neoplastic disease (3.91%) and lung transplantation (0.33%). Over up to 24 months of follow-up, the pulmonary function remained largely stable. The median annual decline in forced vital capacity (FVC) during the first year of pirfenidone therapy was -20 ml (-200-100) and during the second year was -120 ml (-340-30). Over a study period, 33 patients (10.75%) died. CONCLUSIONS: The PolExPIR study is a source of longitudinal RWD on pirfenidone therapy in the Polish cohort of patients with IPF supporting its long-term acceptable safety and efficacy profiles and reinforce findings from the previous randomised clinical trials and observational studies.

Assessment of microvascular function in vivo using flow mediated skin fluorescence (FMSF) in patients with obstructive lung diseases: A preliminary study.

BACKGROUND: Cardiovascular diseases play an important role in the morbidity and mortality of patients with obstructive lung diseases. Impaired vascular endothelial function seems to be a key element linking obstructive lung disease and cardiovascular disease. Recently developed technique named flow mediated skin fluorescence (FMSF) is a novel, non-invasive tool to study microvascular function. METHODS: Total of 69 volunteers including 26 patients with chronic obstructive pulmonary disease (COPD), 23 patients with asthma and 20 healthy subjects underwent microvascular function assessments using FMSF. FMSF assessments were composed of measurements of reduced form of nicotinamide adenine dinucleotide (NADH) fluorescence intensity signal during brachial artery occlusion - ischemic response (IRmax) and immediately after release of occlusion - hyperemic response (HRmax). Associations of microvascular function with clinical and biochemical characteristics of studied subjects were also evaluated. RESULTS: The median value of IRmax was significantly lower in COPD subjects (2.4 [1.0-6.7] %) compared with healthy subjects (9.6 [3.7-13.5] %; p < 0.01). The mean value of HRmax was also significantly reduced in COPD subjects (9.7 (4.5) %) compared with both asthma subjects (12.1 (3.5) %; p < 0.05) and healthy control subjects (13.4 (2.9) %; p < 0.01). CONCLUSIONS: The FMSF technique makes it possible to identify impairments of the microvascular function in patients with COPD, but not in asthma patients. These exploratory findings require further validation in a larger patients cohort.

Epithelial Alarmins in Serum and Exhaled Breath in Patients with Idiopathic Pulmonary Fibrosis: A Prospective One-Year Follow-Up Cohort Study.

BACKGROUND: Recently, epithelial alarmins have been shown to play important roles in non-allergen driven respiratory diseases like idiopathic pulmonary fibrosis (IPF). Little is known about the expression of the epithelial alarmins in IPF. METHODS: This study aimed to prospectively examine interleukin (IL)-25, IL-33, and thymic stromal lymphopoietin (TSLP) levels in the serum and exhaled breath condensate (EBC) in patients with IPF before and after one-year of antifibrotic treatment. A total of 82 volunteers, including 52 patients diagnosed with IPF that qualified for antifibrotic therapy as well as 30 controls, were examined. All study participants underwent baseline peripheral blood and EBC sampling. In 35 out of 52 IPF subjects, a follow-up sampling was performed after 12 months of antifibrotic treatment. Concentrations of alarmins in the serum and EBC were evaluated by means of ELISA. RESULTS: Baseline TSLP concentrations were significantly elevated in patients with IPF compared to controls both in the serum (p < 0.05) and EBC (p < 0.0001). Baseline IL-25 and IL-33 serum and EBC levels did not differ significantly between IPF subjects and controls. Prospective analysis of changes in the epithelial alarmin levels showed significantly decreased IL-25 and TSLP EBC concentrations after 12 months of antifibrotic treatment (p < 0.05), which was observed in the subgroup of IPF patients treated with pirfenidone, but not in those treated with nintedanib. In stable patients with IPF over a study period (absolute forced vital capacity (FVC) % of predicted decline/year ≤ 5%, n = 25), a significant decrease in the EBC levels of both IL-25 and TSLP after 12 months of antifibrotic treatment was noted (p < 0.05), whereas, in progressor IPF patients (absolute FVC % of predicted decline/year > 5%, n = 10), a significant decrease was noted in the IL-25 EBC levels only (p < 0.05). CONCLUSIONS: Elevated TSLP levels in patients with IPF and their significant decrease in the lung compartment during antifibrotic therapy in stable patients with IPF, but not in progressors, support its significant contribution to pro-fibrotic type 2 immune responses in IPF. Noted changes in the epithelial alarmins concentration in the lung compartment during pirfenidone therapy may suggest its possible interaction with epithelial alarmins pathways in IPF.

Epithelial alarmin levels in exhaled breath condensate in patients with idiopathic pulmonary fibrosis: A pilot study.

INTRODUCTION: Interleukin (IL)-25, IL-33 and thymic stromal lymphopoietin (TSLP) are epithelial alarmins involved in innate immune responses and have been shown to play an important role in chronic lung diseases. No data are available regarding their levels in exhaled breath condensate (EBC) in idiopathic pulmonary fibrosis (IPF). OBJECTIVES: To examine IL-25, IL-33 and TSLP levels in the EBC obtained from patients with IPF and compare them to those in healthy controls, patients with asthma and chronic obstructive pulmonary disease (COPD). METHODS: Twenty-three patients with asthma, 25 patients with COPD, 15 patients with IPF and 16 healthy controls were studied. Concentrations of alarmins in the EBC were evaluated by means of ELISA. RESULTS: IL-25 EBC levels were numerically lowest in IPF (25.33 ± 8.84 pg/ml). However, they did not differ significantly from healthy subjects (43.18 ± 5.53 pg/ml), but were significantly lower compared to asthma (72.07 ± 6.03 pg/ml; P < .001). IL-33 EBC levels were significantly increased in IPF (3.41 ± 0.55 pg/ml) compared to healthy controls (1.20 ± 0.60 pg/ml; P < .01) but did not differ from asthma (3.68 pg/ml) and COPD levels (2.47 ± 0.34 pg/ml). There were significant correlations between IL-33 EBC levels and lung diffusion capacity of carbon monoxide (DLco ) absolute (r = .63; P < .05) and % of predicted values (r = .67; P < .01) as well as with time since diagnosis (r = -.59; P < .05) in IPF subjects. TSLP was undetectable in examined samples. CONCLUSION: IL-25 and IL-33 are detectable in the EBC obtained from IPF subjects. Increased levels of IL-33 compared to healthy controls indicate its possible role in the pathobiology of IPF.

Overexpression of chitotriosidase and YKL-40 in peripheral blood and sputum of healthy smokers and patients with chronic obstructive pulmonary disease.

Background: Despite the absence of endogenous chitin in humans, chitinases are present in the serum of healthy subjects and their levels are increased in a variety of chronic inflammatory conditions. It has been shown that chitotriosidase and structurally related chitinase-like protein-YKL-40 contribute to the pathogenesis of COPD. However, details regarding the relation of their systemic and local airways levels remain unknown. Objectives: To examine peripheral blood and sputum chitotriosidase and YKL-40 expression in smokers and patients with COPD. Methods: Forty patients with COPD, 20 healthy smokers and 10 healthy never-smokers were studied. Serum and induced sputum chitotriosidase protein and activity levels, YKL-40 concentrations, and their gene expression in sputum cells and peripheral blood mononuclear cells (PBMC) were evaluated. Results: Both chitotriosidase protein levels and activity were higher in sputum obtained from COPD subjects compared to healthy never-smokers (P<0.05 and P<0.01, respectively). A similar pattern was observed for PBMC chitotriosidase mRNA expression (P<0.001). YKL-40 serum concentrations were elevated in healthy smokers and COPD subjects compared to healthy never-smokers (P<0.001 and P<0.01, respectively). In sputum, YKL-40 levels were increased in COPD compared to healthy never-smokers (P<0.01). PBMC YKL-40 mRNA expression was increased in COPD and healthy smokers compared to healthy never-smokers (P<0.0001). No associations were found between chitotriosidase or YKL-40 peripheral blood levels and sputum levels. Conclusions: Our results demonstrate that chitotriosidase and YKL-40 are overexpressed in peripheral blood and airways in both healthy smokers and COPD subjects which may indicate smoking-related activation of macrophages, neutrophils, and epithelial cells.

The usefulness of soluble receptor for advanced glycation end-products in the identification of COPD frequent exacerbator phenotype.

Introduction: Exacerbations of COPD (ECOPDs) are important events in the course of COPD, accelerating the rate of decline in lung function and increasing the mortality risk. A growing body of evidence suggests the significance of the "frequent exacerbator" phenotype. This phenotype seems to be associated with a more severe airflow limitation, symptoms, health-related quality of life impairment, and higher mortality. However, there is no described biomarker that would help to identify this group of patients. Patients and methods: Patients with COPD in "D" GOLD category were monitored for 3 years according to events of ECOPD. Serum samples were collected from the patients. Circulating level of plasma soluble receptor for advanced glycation end-products (sRAGE) was measured using commercially available high sensitivity kits. The receiver operating characteristic (ROC) curve analysis was used to assess the usefulness of sRAGE to identify frequent exacerbator phenotype. Log-rank test was used in the analysis of time to the subsequent exacerbation. Pearson (R) or Spearman's rank (RS) correlation coefficients were used for correlation analysis. Results: Nineteen patients were enrolled. The area under the ROC curve (AUROC) for sRAGE for the identification of frequent exacerbator phenotype was 0.81. Analysis identified the cutoff point as 850.407 pg/mL, characterized by a sensitivity of 0.80 (95% CI: 0.28-1.0) and specificity of 0.93 (95% CI: 0.66-1.0). Additionally, in the group with sRAGE ≤850.407 pg/mL, we observed significantly shorter time to the subsequent exacerbation: median of 32 vs 105.5 days (P=0.03). Correlation analysis revealed significant negative correlation between sRAGE and the number of exacerbations requiring hospitalization during the whole time of follow-up (RS=-0.53; P=0.02) and significant positive correlation with FEV1 expressed as the percentage of reference value (R=0.6; P=0.006). Conclusion: sRAGE seems to be useful in the identification of frequent exacerbator phenotype. This parameter may also be used in the prediction of time to ECOPD. Our findings should be confirmed in a sufficiently powered larger sample.

Morphometric analysis of mitochondria in lymphocytes of patients with exacerbations of chronic obstructive pulmonary disease - pilot study.

Introduction: Exacerbations of chronic obstructive pulmonary disease (ECOPD) are important events in the course of the disease, negatively influencing health status and disease progression. Therefore, there is a strong need for deeper understanding of the pathology of ECOPD to elaborate new therapeutic approaches and ameliorate prognoses. Contributions of mitochondria to pathobiology of COPD are still under investigation, although growing evidence suggests their important role in this disease. The aim of our study was to assess the morphometric parameters of mitochondria in lymphocytes of patients with ECOPD. Patients and methods: Lymphocytes were isolated from the peripheral blood of patients with COPD. Transmission electron microscopy was used to assess absolute number of mitochondria per cell, mitochondrial content, and morphometric parameters of individual mitochondria. We also counted indexes for elongation and interconnectivity. Results: Eighteen patients (9 with ECOPD and 9 in the stable period of the disease) were analyzed. We observed significantly lower length of mitochondrion (P=0.03) and significant decrease both in elongation (P=0.03) and interconnectivity indexes (P=0.04) in ECOPD patients. Conclusions: The morphometric parameters of mitochondria in lymphocytes derived from patients during the early period of ECOPD requiring hospitalization are altered in comparison to patients in the stable period of the disease. This suggests their contribution to pathobiology of ECOPD. These preliminary outcomes should be further validated in larger size samples.

The association between airway eosinophilic inflammation and IL-33 in stable non-atopic COPD.

BACKGROUND: Interleukin(IL)-33 is an epithelial alarmin important for eosinophil maturation, activation and survival. The aim of this study was to examine the association between IL-33, its receptor expression and airway eosinophilic inflammation in non-atopic COPD. METHODS: IL-33 concentrations were measured in exhaled breath condensate (EBC) collected from healthy non-smokers, asthmatics and non-atopic COPD subjects using ELISA. Serum and sputum samples were collected from healthy non-smokers, healthy smokers and non-atopic COPD patients. Based on sputum eosinophil count, COPD subjects were divided into subgroups with airway eosinophilic inflammation (sputum eosinophils > 3%) or without (sputum eosinophils ≤3%). IL-33 and soluble form of IL-33 receptor (sST2) protein concentrations were measured in serum and sputum supernatants using ELISA. ST2 mRNA expression was measured in peripheral mononuclear cells and sputum cells by qPCR. Hemopoietic progenitor cells (HPC) expressing ST2 and intracellular IL-5 were enumerated in blood and induced sputum by means of flow cytometry. RESULTS: IL-33 levels in EBC were increased in COPD patients to a similar extent as in asthma and correlated with blood eosinophil count. Furthermore, serum and sputum IL-33 levels were higher in COPD subjects with sputum eosinophilia than in those with a sputum eosinophil count ≤3% (p < 0.001 for both). ST2 mRNA was overexpressed in sputum cells obtained from COPD patients with airway eosinophilic inflammation compared to those without sputum eosinophilia (p < 0.01). Similarly, ST2 + IL-5+ HPC numbers were increased in the sputum of COPD patients with airway eosinophilia (p < 0.001). CONCLUSIONS: Our results indicate that IL-33 is involved in the development of eosinophilic airway inflammation in non-atopic COPD patients.

Skin condition and its relationship to systemic inflammation in chronic obstructive pulmonary disease.

BACKGROUND: The systemic (extrapulmonary) effects and comorbidities of chronic obstructive pulmonary disease (COPD) contribute substantially to its burden. The supposed link between COPD and its systemic effects on distal organs could be due to the low-grade systemic inflammation. The aim of this study was to investigate whether the systemic inflammation may influence the skin condition in COPD patients. MATERIALS AND METHODS: Forty patients with confirmed diagnosis of COPD and a control group consisting of 30 healthy smokers and 20 healthy never-smokers were studied. Transepidermal water loss, stratum corneum hydration, skin sebum content, melanin index, erythema index, and skin temperature were measured with worldwide-acknowledged biophysical measuring methods at the volar forearm of all participants using a multifunctional skin physiology monitor. Biomarkers of systemic inflammation, including high-sensitivity C-reactive protein (hsCRP), interleukin-6 (IL-6), and tumor necrosis factor α (TNF-α), were measured in serum using commercially available enzyme-linked immunosorbent assays. RESULTS: There were significant differences between COPD patients and healthy never-smokers in skin temperature, melanin index, sebum content, and hydration level (P<0.05), but not for transepidermal water loss and erythema index. No significant difference was noted between COPD patients and smokers in any of the biophysical properties of the skin measured. The mean levels of hsCRP and IL-6 in serum were significantly higher in COPD patients and healthy smokers in comparison with healthy never-smokers. There were significant correlations between skin temperature and serum hsCRP (R=0.40; P=0.02) as well as skin temperature and serum IL-6 (R=0.49; P=0.005) in smokers. Stratum corneum hydration correlated significantly with serum TNF-α (R=0.37; P=0.01) in COPD patients. CONCLUSION: Differences noted in several skin biophysical properties and biomarkers of systemic inflammation between COPD patients, smokers, and healthy never-smokers may suggest a possible link between smoking-driven, low-grade systemic inflammation, and the overall skin condition.

Analysis of changes in expression of IL-4/IL-13/STAT6 pathway and correlation with the selected clinical parameters in patients with atopic asthma.

INTRODUCTION: Asthma is associated with activation of interleukin-4 (IL-4)/interleukin-13 (IL-13)/signal transducer and activator of transcription factor-6(STAT6) inflammatory response via overexpression of all pathway components: IL-4, IL-13, and STAT6. OBJECTIVES: To evaluate the association of IL-4, IL-13, and STAT6 expression and immunoexpression with atopic asthma development. PATIENTS AND METHODS: Fifty patients with atopic asthma and 20 healthy controls were enrolled into the study. Relative gene expression was analyzed by qPCR method. Immunoexpression was assessed by ELISA method. RESULTS: The expression levels of IL-4, IL-13, and STAT6 were higher in patients compared to the controls, but a statistically significant difference was observed only for IL-13 (P = 0.03). In immunoexpression analysis, a statistically significant difference between patients and controls was found for IgE (P = 0.03). Significant positive correlations in the patient group were found between IL-13 gene expression and total level of serum IgE (rho = 0.230, P = 0.033), STAT6 gene/STAT6 protein and total level of serum IgE (STAT6: rho = 0.077, P = 0.038; STAT6: rho = 0.049, P = 0.042), IL-4, and STAT6 expression (rho = 0.098, P = 0.048). Any significant correlations were found between expression/immunoexpression levels of the studied genes and clinical classification, clinical features, or lung function parameters. CONCLUSIONS: Our data support the role of Th2 cytokines (IL-4, IL-13) and STAT6 in Th1/Th2 imbalance and highlight the etiological relationship between IL-4/IL-13/STAT6 signaling and atopy and asthma.

Synergistic interactions between anticancer chemotherapeutics and phenolic compounds and anticancer synergy between polyphenols.

Chemoprevention has recently gained a new dimension due to the possibility of studying the mechanisms of action of chemopreventive agents at the molecular level. Many compounds have been proved to inhibit early stages of carcinogenesis in experimental models. These compounds include both recognized drugs (such as tamoxifen and nonsteroidal anti-inflammatory drugs) and natural constituents of edible and therapeutic plants, particularly polyphenols. Phenolics are characterized by high structural diversity and, consequently, a very broad spectrum of biological activities. They are increasingly looked upon as a valuable alternative or a support for synthetic drugs, as evidenced by a growing number of clinical trials regarding the use of phenolic compounds and polyphenol-rich extracts in chemoprevention and therapy. In the present work, we discuss the effectiveness of natural polyphenols as cancer preventive and therapeutic agents resulting from their synergy with synthetic or semisynthetic anticancer drugs as well as with other phenolic compounds of plant origin.

Influence of polyphenol extract from evening primrose (Oenothera paradoxa) seeds on human prostate and breast cancer cell lines.

There is growing interest in plant polyphenols which exhibit pleiotropic biological activities, including anti-inflammatory, antioxidant, and anticancer effects. The objective of our study was to evaluate the influence of an evening primrose extract (EPE) from defatted seeds on viability and invasiveness of three human cell lines: PNT1A (normal prostate cells), DU145 (prostate cancer cells) and MDA-MB-231 (breast cancer cells). The results revealed that after 72 h of incubation the tested extract reduced the viability of DU 145 and MDA-MB-231 with IC50 equal to 14.5 µg/mL for both cell lines. In contrast, EPE did not inhibit the viability of normal prostate cells. Furthermore, EPE reduced PNT1A and MDA-MB-231 cell invasiveness; at the concentration of 21.75 µg/mL the suppression of invasion reached 92% and 47%, respectively (versus control). Additionally, zymographic analysis revealed that after 48 h of incubation EPE inhibited metalloproteinase-2 (MMP-2) and metalloproteinase-9 (MMP-9) activities in a dose-dependent manner. For PNT1A the activities of MMP-2 and MMP-9 decreased 4- and 2-fold, respectively, at EPE concentration of 29 µg/mL. In the case of MDA-MB-231 and DU 145 the decrease in MMP-9 activity at EPE concentration of 29 µg/mL was 5.5-fold and almost 1.9-fold, respectively. In conclusion, this study suggests that EPE may exhibit antimigratory, anti-invasive and antimetastatic potential towards prostate and breast cancer cell lines.

Overview of metabolism and bioavailability enhancement of polyphenols.

A proper diet is one of major factors contributing to good health and is directly related to general condition of the organism. Phenolic compounds are abundant in foods and beverages (fresh and processed fruits and vegetables, leguminous plants, cereals, herbs, spices, tea, coffee, wine, beer) and their pleiotropic biological activities result in numerous health beneficial effects. On the other hand, high reactivity and very large diversity in terms of structure and molecular weight renders polyphenols one of the most difficult groups of compounds to investigate, as evidenced by ambiguous and sometimes contradictory results of many studies. Furthermore, phenolics undergo metabolic transformations, which significantly change their biological activities. Here, we discuss some aspects of metabolism and absorption of phenolic compounds. On the basis of information reported in the literature as well as in summaries of clinical trials and patent applications, we also give an overview of strategies for enhancing their bioavailability.

Procyanidins from evening primrose (Oenothera paradoxa) defatted seeds inhibit invasiveness of breast cancer cells and modulate the expression of selected genes involved in angiogenesis, metastasis, and apoptosis.

There is a growing interest in plant polyphenols (including flavanols) that exhibit pleiotropic biological activities such as antiinflammatory, antioxidant, and anticancer effects. Here, we report for the first time the inhibition of MDA-MB-231 breast cancer cell viability and invasiveness by an evening primrose flavanol preparation (EPFP). We observed a decrease in MDA-MB-231 viability of 50% vs. a control after 72 h of incubation with EPFP at a concentration of 58 µM gallic acid equivalents (GAE) and an inhibition of their invasiveness of 65% vs. a control at 75 µM GAE after 48 h of incubation. EPFP caused a 10-fold reduction in matrix metalloproteinase-9 (MMP-9) activity at 100 µM GAE. Furthermore, through modulation of mRNA expression, EPFP reduced the expression levels of the following proteins: antiapoptotic Bcl-2, angiogenic vascular endothelial growth factor (VEGF), and 2 transcription factors (c-Jun, c-Fos). Moreover, analysis by flow cytometry revealed that EPFP induced apoptosis in MDA-MB-231 cells. In conclusion, our data shows that EPFP inhibits cell viability by increasing apoptosis and decreases cell invasiveness by decreasing angiogenesis.

Anticancer Activity of New Synthetic α-Methylene-δ-Lactones on Two Breast Cancer Cell Lines.

Natural products are important leads in drug discovery. The search for effective plant-derived anticancer agents or their synthetic analogues has continued to be of interest to biologists and chemists for a long time. In this report, cytotoxicity and anticancer activity of new synthetic α-methylene-δ-lactones was tested against two breast cancer cell lines, invasive, hormone-independent MDA-MB-231 and hormone-dependent MCF-7. Cytotoxicity was examined using MTT assay. The ability to induce apoptosis and changes in mitochondrial membrane potential was studied by flow cytometry. The expression levels of pro- and anti-apoptotic genes were determined by quantitative real-time PCR. Cancer cell migration and invasion were assessed by wound healing and Matrigel assays. Additionally, secretion of proteins associated with invasiveness, metalloproteinase-9 (MMP-9) and urokinase plasminogen activator (uPA) was investigated using commercial ELISA kits and MMP-9 activity by gelatin zymography. A natural sesquiterpene lactone, parthenolide, was used as a positive control. Screening results showed all four analogues to be highly cytotoxic. The most potent compound of the series, 1-isopropyl-2-methylene-1,2-dihydrobenzochromen-3-one, designated DL-3, which reduced the number of viable MDA-MB-231 and MCF-7 cells with the IC50 values of 5.3 µM and 3.54 µM, respectively, was selected for further research. DL-3 activated the intrinsic pathway of apoptosis, associated with the loss of mitochondrial membrane potential and changes in Bax/Bcl-2 ratio. DL-3 also inhibited the movement of both types of breast cancer cells. Suppression of cell migration and invasion was the result of the decreased secretion of enzymes responsible for the degradation of the extracellular matrix, MMP-9 and uPA. These findings show that the synthetic α-methylene-δ-lactone, DL-3, displays potential to be further explored in the development of new anticancer agents.

Flavanols from evening primrose (Oenothera paradoxa) defatted seeds inhibit prostate cells invasiveness and cause changes in Bcl-2/Bax mRNA ratio.

In this study, we assessed the influence of an evening primrose flavanol preparation (EPFP) on proliferation and invasiveness of human prostate cancer cells (DU 145) and immortalized prostate epithelial cells (PNT1A). We report for the first time that EPFP reduces DU 145 cell proliferation (IC50 = 97 µM GAE for 72 h incubation) and invasiveness (by 24% versus control at 75 µM GAE). EPFP strongly inhibited PNT1A invasiveness in a concentration-dependent manner (by 67% versus control at 75 µM GAE) and did not cause a reduction in their proliferation. Furthermore, EPFP inhibited the activities of MMP-2 and MMP-9 secreted to culture medium by PNT1A cells by 84% and 34% versus control at 100 µM GAE, respectively. In the case of DU 145, MMP-9 activity at 100 µM GAE was reduced by 37% versus control. Moreover, the evening primrose seed flavanols suppressed the expression of selected genes (MMP-1, MMP-9, MMP-14, c-Fos, c-Jun, and VEGF) and also caused favorable changes in Bcl-2/Bax mRNA ratio which render DU 145 cells more sensitive to apoptosis-triggering agents. An additional confirmation of the proapoptotic activity of EPFP toward DU 145 was visualization of characteristic apoptotic bodies by DAPI staining. In conclusion, this study suggests that EPFP may increase apoptosis and reduce angiogenesis of prostate cancer cells.