Serial fractional exhaled nitric oxide measurements at and off work may help to identify immunologic occupational asthma in cases with complex exposures.

Serial measurements of fractional exhaled nitric oxide (FeNO) at home and at work have been described to provide complementary information for the diagnosis of occupational asthma (OA) when specific inhalation challenge (SIC) is missing or doubtful. We describe two cases in which serial FeNO measurements enabled the detection of probable OA after complex exposures. A 25-year-old industrial painter with exposure to a variety of paints suffered from work-related airway symptoms for five years. Lung function was normal, and she was not atopic. SIC with hexamethylene diisocyanate was negative. A 47-year-old sign maker (screen printing, foils) suffering from work-related dyspnoea for seven years. Moderate airway obstruction, but no atopy was detectable. Due to the complex exposures SIC was not performed. Both patients performed FeNO measurements once daily during a 2-week-holiday and a subsequent 2-week-work period. In both cases elevated baseline FeNO decreased to normal (25 ppb) during holidays and increased after resuming work (case 1: 125 ppb, case 2: 45 ppb).

Rationale and design of the iCORONARY trial: improving the cost-effectiveness of coronary artery disease diagnosis.

BACKGROUND: In patients with stable coronary artery disease (CAD), revascularisation decisions are based mainly on the visual grading of the severity of coronary stenosis on invasive coronary angiography (ICA). However, invasive fractional flow reserve (FFR) is the current standard to determine the haemodynamic significance of coronary stenosis. Non-invasive and less-invasive imaging techniques such as computed-tomography-derived FFR (FFR-CT) and angiography-derived FFR (QFR) combine both anatomical and functional information in complex algorithms to calculate FFR. TRIAL DESIGN: The iCORONARY trial is a prospective, multicentre, non-inferiority randomised controlled trial (RCT) with a blinded endpoint evaluation. It investigates the costs, effects and outcomes of different diagnostic strategies to evaluate the presence of CAD and the need for revascularisation in patients with stable angina pectoris who undergo coronary computed tomography angiography. Those with a Coronary Artery Disease-Reporting and Data System (CAD-RADS) score between 0-2 and 5 will be included in a prospective registry, whereas patients with CAD-RADS 3 or 4A will be enrolled in the RCT. The RCT consists of three randomised groups: (1) FFR-CT-guided strategy, (2) QFR-guided strategy or (3) standard of care including ICA and invasive pressure measurements for all intermediate stenoses. The primary endpoint will be the occurrence of major adverse cardiac events (death, myocardial infarction and repeat revascularisation) at 1 year. CLINICALTRIALS: gov-identifier: NCT04939207. CONCLUSION: The iCORONARY trial will assess whether a strategy of FFR-CT or QFR is non-inferior to invasive angiography to guide the need for revascularisation in patients with stable CAD. Non-inferiority to the standard of care implies that these techniques are attractive, less-invasive alternatives to current diagnostic pathways.

Expression of DNA mismatch repair proteins in melanoma patients treated with immune checkpoint inhibitors.

PURPOSE: To investigate the protein expression of DNA mismatch repair (MMR) proteins in patients with cutaneous melanoma (CM) under immune checkpoint inhibitor (ICI) therapy. METHODS: Immunohistochemistry was performed on tumor tissue for MMR proteins MLH1, MSH2, MSH6, and PMS2 in 50 metastatic CM patients treated with ICI (ipilimumab, nivolumab, pembrolizumab). RESULTS: Best overall response (BOR) rate was 48% (24/50). Reduced MMR protein expression (nuclear expression in < 80% of tumor cells) was observed in 8 patients (16%). Compared to other clinical parameters, baseline neutrophil/lymphocyte ratio and reduced intratumoral MMR protein expression (P = 0.0033) were determined as the only parameters significantly associated with favorable BOR. However, in this small study population, reduced MMR protein expression did not reach statistical significance in multivariate analysis. CONCLUSION: Reduced MMR protein expression is observed in CM and might predict favorable BOR in patients treated with ICI, as was observed for other entities. However, these findings need to be substantiated in larger patient cohorts.

Protein expression of prognostic genes in primary melanoma and benign nevi.

PURPOSE: To evaluate the protein expression characteristics of genes employed in a recently introduced prognostic gene expression assay for patients with cutaneous melanoma (CM). METHODS: We studied 37 patients with CM and 10 with benign (melanocytic) nevi (BN). Immunohistochemistry of primary tumor tissue was performed for eight proteins: COL6A6, DCD, GBP4, KLHL41, KRT9, PIP, SCGB1D2, SCGB2A2. RESULTS: The protein expression of most markers investigated was relatively low (e.g., DCD, KRT9, SCGB1D2) and predominantly cytoplasmatic in melanocytes and keratinocytes. COL6A6, GBP4, and KLHL41 expression was significantly enhanced in CM when compared to BN. DCD protein expression was significantly correlated with COL6A6, GBP4, and KLHL41. GBP4 was positively correlated with KLHL41 and inversely correlated with SCGB2B2. The latter was also inversely correlated with serum S100B levels at time of initial diagnosis. The presence of SCGB1D2 expression was significantly associated with ulceration of the primary tumor. KRT9 protein expression was significantly more likely found in acral lentiginous melanoma. The presence of DCD expression was less likely associated with superficial spreading melanoma subtype but significantly associated with non-progressive disease. The absence of SCGB2A2 expression was significantly more often observed in patients who did not progress to stage III or IV. CONCLUSIONS: The expression levels observed were relatively low but differed in part with those found in BN. Even though we detected some significant correlations between the protein expression levels and clinical parameters (e.g., CM subtype, course of disease), there was no major concordance with the protective or risk-associated functions of the corresponding genes included in a recently introduced prognostic gene expression assay.

[The Current Therapy of Asbestos-Associated Malignant Pleural Mesothelioma - An Expert Consensus Paper]. / Die aktuelle Therapie des asbestassoziierten malignen Pleuramesothelioms ­ Ein Experten-Konsensuspapier.

Asbestos-related mesotheliomas belong to the group of the most frequent occupational diseases in Germany, reaching about 1,000 new cases per year. The disease has a dismal prognosis because most tumors remain asymptomatic for a long time and therefore are diagnosed as incidental findings at later stages.During the last decade the German Social Accident Insurance (DGUV) has made considerable efforts to prepone the diagnosis in order to detect the disease at earliest possible stages. These efforts resulted in new findings showing that, in a high-risk group, a combination of the biomarkers calretinin and mesothelin was able to advance the diagnosis up to 12 months.Ideally, the diagnosis of a mesothelioma at an early stage has to be accompanied by the best possible individualized therapy. Standard therapeutic strategies are surgery and chemotherapy, added by radiotherapy and psycho-oncology. In recent years, several new therapeutic avenues are being explored. This review comprehensively presents both old and new therapeutic options in mesothelioma, based on international Leitlinien and new studies.

Expression of Hedgehog signalling molecules in microcystic adnexal carcinoma.

BACKGROUND: Microcystic adnexal carcinoma (MAC) is a rare skin neoplasm that has not been characterized on a molecular basis. AIM: To assess expression profiles of Hedgehog (HH) signalling molecules in MAC and control tumours. METHODS: Immunohistochemistry was performed for Sonic Hedgehog (SHH), Indian Hedgehog (IHH), Patched 1 (PTCH1) and Smoothened (SMO) on patient MAC tissue (n = 26) and control tumour tissue, including syringoma (SyG; n = 11), trichoepithelioma (TE; n = 11) and basal cell carcinoma (BCC; n = 12) tissues. RESULTS: Patched 1 and SMO immunoreactivity was significantly higher in BCC than in SyG, TE or MAC (P < 0.001 and P < 0.03, respectively). The highest IHH expression was observed in BCC and TE compared with SyG and MAC (P < 0.04). Notably, the highest SHH protein expression was observed in SyG compared with MAC, TE and even BCC (P < 0.001). In patients with MAC, SMO immunoreactivity significantly (r = 0.51; P < 0.01) correlated with PTCH1 expression. Further correlation studies did not show significant associations between the HH expression markers assessed (P > 0.05). CONCLUSION: Our results indicate that alterations of the HH signalling are unlikely to play a major role in the pathogenesis of MAC, which is in contrast to the morphologically similar BCC and TE. Our observation provides additional information to the limited molecular pathology knowledge on this rare tumour.

Human Inhalation Study with Zinc Oxide: Analysis of Zinc Levels and Biomarkers in Exhaled Breath Condensate.

Workers in the zinc processing, for example, welding or hot-dip galvanizing, are exposed to aerosols consisting of particles and gases, including zinc oxide (ZnO), which can affect human health. In this study, we addressed the effects of short-term controlled exposure to nano-sized ZnO on the airway inflammatory markers in healthy volunteers. To this end, we determined the influence of ZnO inhalation on the content of zinc and biomarkers (leukotriene B4 (LTB4), peptide leukotrienes (LTC4/D4/E4), 8-iso-PGF2α, pH, and prostaglandin E2 (PGE2)) in exhaled breath condensate (EBC). Sixteen non-smoking subjects (8 females, 8 men) were exposed to filtered air (sham) or ZnO nanoparticles (0.5, 1.0, and 2.0 mg/m3) for 4 h. EBC samples were collected according to specific study design. We found that the peptide leukotrienes were below the limit of quantification (LOQ) in all the EBC samples. ZnO exposure showed no detectable effect on any other parameters investigated when comparing the two groups. The content of Zn in EBC was unaffected by ZnO inhalation at any concentration used. Therefore, we conclude that the evaluation of Zn and biomarker content in EBC would not be a suitable way to assess the exposure to inhaled ZnO.

Occupational endotoxin exposure and health effects.

Lipopolysaccharide (LPS), also termed endotoxin, is an integral structural component of the outer membrane of Gram-negative bacteria and has been a focus of bioaerosol research for many years. Endotoxin is nearly ubiquitous in the environment; however, exposure at specific workplaces, such as waste collecting, livestock farming, agriculture, the textile industry has been associated with adverse health effects. The aim of this review is to summarize studies published in the last 10 years on endotoxin measurement and health effects due to endotoxin in occupational settings. The search was mainly performed using MEDLINE (Pubmed), focusing on publications related to the determination of endotoxin, inhalative occupational endotoxin exposure, and health effects. The review shows that despite the well-established methods available to measure endotoxin, a global comparison of studies still remains difficult because the details of sampling strategies and exposure assessment are variable and depend on the specific workplace situation. Thus, health-based threshold limit values still cannot be derived on the basis of available data. Since endotoxin is only one component in a heterogeneous bioaerosol mixture, the question remains open on how to evaluate and record the additional effects of the other components. In particular, there is a lack of intervention studies investigating the effectiveness of protective measures with respect to health outcome. In addition, the studies selected in this review show a wide range of endotoxin exposure, even within one industry or sector. The level of exposure seems to depend more on the specific task performed and the way it was performed rather than on the profession or industry itself. The identification of hot spots of exposure, as well as methods of communication on hazards and possible protective measures, seem to remain important tasks in occupational health protection.

Basal cell carcinoma risk and solar UV exposure in occupationally relevant anatomic sites: do histological subtype, tumor localization and Fitzpatrick phototype play a role? A population-based case-control study.

BACKGROUND: A two-fold risk increase to develop basal cell carcinoma was seen in outdoor workers exposed to high solar UV radiation compared to controls. However, there is an ongoing discussion whether histopathological subtype, tumor localization and Fitzpatrick phototype may influence the risk estimates. OBJECTIVES: To evaluate the influence of histological subtype, tumor localization and Fitzpatrick phototype on the risk to develop basal cell carcinoma in highly UV-exposed cases and controls compared to those with moderate or low solar UV exposure. METHODS: Six hundred forty-three participants suffering from incident basal cell carcinoma in commonly sun-exposed anatomic sites (capillitium, face, lip, neck, dorsum of the hands, forearms outside, décolleté) of a population-based, case-control, multicenter study performed from 2013 to 2015 in Germany were matched to controls without skin cancer. Multivariate logistic regression analysis was conducted stratified for histological subtype, phototype 1/2 and 3/4. Dose-response curves adjusted for age, age2, sex, phototype and non-occupational UV exposure were calculated. RESULTS: Participants with high versus no (OR 2.08; 95% CI 1.24-3.50; p = 0.006) or versus moderate (OR 2.05; 95% CI 1.15-3.65; p = 0.015) occupational UV exposure showed a more than two-fold significantly increased risk to develop BCC in commonly UV-exposed body sites. Multivariate regression analysis did not show an influence of phototype or histological subtype on risk estimates. The restriction of the analysis to BCC cases in commonly sun-exposed body sites did not influence the risk estimates. The occupational UV dosage leading to a 2-fold increased basal cell carcinoma risk was 6126 standard erythema doses. CONCLUSION: The risk to develop basal cell carcinoma in highly occupationally UV-exposed skin was doubled consistently, independent of histological subtype, tumor localization and Fitzpatrick phototype.

Discrimination Between Atopic, Allergic, and Asthmatic Volunteers for Human Exposure Studies on Sensory Irritation.

Atopic, allergic, and especially asthmatic subjects might be particularly susceptible to sensory irritation induced by airborne chemicals compared to healthy individuals. Therefore, a good characterization of subjects is essential in inhalation exposure studies on sensory irritants. A total of 105 volunteers, 87% of whom reported to be non-allergic, participated in a medical examination that included skin prick test (SPT), measurements of total IgE, specific IgE (sIgE) to an ubiquitous allergen mix (sx1), and fractionated exhaled nitric oxide (FeNO), as well as pulmonary function and methacholine test. The median value of sIgE to sx1 was 0.20 kU/L (0.07-91.3 kU/L) and correlated significantly with total IgE (28.8 kU/L (2-756 kU/L)) and FeNO (14 ppb (5-100 ppb)). Forty-three subjects (41%) had sIgE to sx1 ≥ 0.35 kU/L and were classified as atopic. Thirty-five subjects, all also sx1-positive, were positive in SPT. Obstruction, small airway disease, and/or bronchial hyperreactivity were diagnosed in 18 subjects. Receiver operating characteristics (ROC) were performed to check whether signs of sensitization are useful to discriminate subjects with and without airway diseases. However, sx1, total IgE, FeNO, and SPT reached only low areas under the curve (AUC: 0.57-0.66). Although predominantly young and, according to their own statements, mostly non-allergic subjects participated in the study, almost half of them were atopic, and 10% had airway disease or bronchial hyperreactivity. This indicates that the validity of self-reported data might be inaccurate. In summary, diversified investigations of the allergy-related health status appear necessary for a thorough characterization of subjects for exposure studies on sensory irritants.

The Pattern of Sensitization Influences Exhaled and Nasal Nitric Oxide Levels in Young Adults.

Nitric oxide (NO) from upper (nasal NO, nNO) or lower airways (fractional exhaled NO, FeNO) is considered a surrogate marker for Th2-type inflammation, which is influenced by atopy. The aim of this study was to analyze nNO and FeNO in regard to qualitative and quantitative aspects of sensitization. We evaluated 244 non-smoking young adults. All of them were first-year students recruited for a longitudinal study. An inhalation allergy screening tool was used for atopy definition (specific immunoglobulin E (sIgE) to sx1 ≥ 0.35 kU/L), and also sIgE response to three inhalant perennial allergens, house dust mite (HDM, d1), cat (e1), and dog (e5), was determined in the non-pollen season. With respect to sx1, 100 subjects could be classified as atopic. Sensitization to one, two, or three perennial allergens could be demonstrated in 46, 10, and 16 students, respectively. The subjects with positive IgE response to sx1, but not sensitized to HDM, cat, and/or dog, had FeNO levels comparable to those of non-atopic subjects (13.5 vs. 13.0 ppb, respectively; p = 0.485). These levels were significantly lower compared to atopic subjects being sensitized to any perennial allergen (19.0 ppb; p = 0.0003). After grouping the atopic subjects for perennial sensitization patterns, significantly higher FeNO could be detected in subjects with poly-sensitization (n = 26; 26.0 ppb) compared to the mono-sensitized ones (n = 46; 18.0 ppb; p = 0.023). Regarding nNO, no differences could be observed. Applying a two-way ANOVA, we could reveal a significant correlation of specific HDM-IgE CAP-class with FeNO (p < 0.0001) and nNO levels (p = 0.007). Finally, a significant relationship was found between nNO and FeNO for the whole cohort (p < 0.0001). In summary, our findings support the argument that atopy and perennial sensitization should be considered for the interpretation of NO.

Expression of Lefty predicts Merkel cell carcinoma-specific death.

BACKGROUND: Lefty and Nodal are transforming growth factor ß-related proteins, which, beside their role in determination of laterality during embryogenesis, have also been linked with cancer progression. OBJECTIVES: Prompted by the observed significant left-sided laterality of Merkel cell carcinoma (MCC), we addressed whether Lefty and Nodal are expressed in MCC and correlated expression patterns with clinical parameters such as MCC laterality and patient outcome. METHODS: Expression of Lefty and Nodal in primary MCC was assessed in 29 patients by immunohistochemistry. The histology (H-)score was calculated and correlated with clinical parameters. RESULTS: The median (range) H-score of Lefty and Nodal was 17.6 (0-291) and 74.9 (0.7-272), respectively. There was a significant correlation between Lefty expression and Nodal expression (correlation coefficient of 0.60, P = 0.0006). There was no significant correlation between Lefty expression and Nodal expression with either tumour laterality, gender, age, Merkel cell polyomavirus status, disease stage, anatomical localization of primary tumours or disease relapse. On univariate analysis, low Lefty expression and Nodal expression were significantly associated with MCC-specific death (P = 0.010 and P = 0.019, respectively). On univariate analysis, low Lefty expression was the only significant independent predictor for MCC-specific death (P = 0.025) as indicated by an odds ratio of 14 (95% CI: 1.43-137.33). CONCLUSIONS: Lefty and Nodal are frequently expressed in MCC, but not correlated with tumour laterality. Importantly, our data suggest that a low level of Lefty expression in primary MCC is a strong predictor of MCC-specific death.

How Healthy Is Healthy? Comparison Between Self-Reported Symptoms and Clinical Outcomes in Connection with the Enrollment of Volunteers for Human Exposure Studies on Sensory Irritation Effects.

Controlled human exposure studies on sensory irritation effects are usually performed with healthy volunteers. Therefore, in most studies pre-screening by a health questionnaire and a detailed medical examination are combined. The aim of this report is to investigate whether self-reported information about smoking and health status is sufficient or whether additional clinical tests are necessary for a successful and safe enrollment of healthy volunteers. There were 409 volunteers (55% female; 17-57 years; 79% non-smokers) who declared interest in participation in the study. However, 87 subjects failed to meet specific inclusion criteria, and further 138 had to be excluded due to the presence of chronic health problems. In effect, 184 subjects passed the initial questionnaire screening and proceed to further examination. Medical examination included electrocardiogram, blood and urine screening, and an olfactory function test. Atopy status was assessed by skin prick or specific IgE testing. Lung function and a methacholine challenge test were performed to assess respiratory health and bronchial hyperresponsiveness. Overall, only 107 non-smoking subjects (58% female; 19-40 years) who had no respiratory diseases, allergies, or chronic illnesses could be finally selected. Out of the 107 subjects, 8 were excluded due to positive cotinine tests, laboratory test results outside the reference range, or atypical ECGs. In another 12 subjects, obstruction or a bronchial hyperreactivity was diagnosed. Among the remaining 87 healthy subjects, 26 were classified as atopic and further two as hyposmic. In conclusion, although young and non-smoking volunteers considered themselves healthy by questionnaire, 20% showed signs of a heart, liver, or airway disease, and additional 24% were classified as atopics. This suggests that more detailed clinical testing may be necessary to safely exclude those who may adversely react to controlled exposure with sensory irritants.

German Environmental Specimen Bank: 24-hour urine samples from 1999 to 2017 reveal rapid increase in exposure to the para-phthalate plasticizer di(2-ethylhexyl) terephthalate (DEHTP).

The worldwide plasticizer markets are facing constant substitution processes. Many classic ortho-phthalate plasticizers like di(2-ethylhexyl) phthalate (DEHP) are phased out, due to their proven toxicity to reproduction. Assumedly less critical, less regulated plasticizers such as di(2-ethylhexyl) terephthalate (DEHTP) are increasingly applied in consumer near products like toys, food contact materials, and medical devices. With the increasing use of DEHTP, increasing exposures of the general population have to be expected likewise. Human biomonitoring is a well-established tool to determine population exposures. In the present study we investigate the time trend of exposure to DEHTP using 24-hour urine samples of the German Environmental Specimen Bank (ESB) collected from 1999 to 2017. In these samples (60 per odd-numbered year, 600 samples in total) collected from young German adults (20-29 years, equal gender distribution) we determined four specific urinary metabolites as biomarkers of DEHTP exposure. From 1999 to 2009, the main specific urinary metabolite 5cx-MEPTP was quantifiable in <10% of the samples. Thereafter, detection rates and levels constantly increased, in line with rapidly increasing DEHTP consumption volumes. In 2017, all samples had 5cx-MEPTP levels above the limit of quantification (LOQ) with a median concentration of 3.35 µg/L (95th percentile: 12.8 µg/L). The other metabolites were detected less frequently and at lower levels but correlated well with 5cx-MEPTP robustly confirming the increasing DEHTP exposure. All 5cx-MEPTP concentrations were well below the German health based guidance value (HBM-I) of 2800 µg/L for adults. Likewise, the median calculated daily intake, based on 5cx-MEPTP measured in 2017, was 0.74 µg/kg bw∗d (95th percentile: 3.86 µg/kg bw∗d), still well below the tolerable daily intake (TDI) of 1000 µg/kg bw∗d. Based on current toxicological knowledge we can hence conclude that for the population investigated, DEHTP exposure gives no reason for immediate concern. However, the steep ongoing increase of DEHTP exposure warrants further close monitoring in the future, preferably also in sub-populations with known higher exposures to plasticizers, especially children.

Low Drosha protein expression in cutaneous T-cell lymphoma is associated with worse disease outcome.

BACKGROUND: Dysregulation of microRNAs (miRNAs) key regulators may contribute to the pathogenesis of malignancies. miRNA machinery genes such Dicer and Drosha have been reported to be biomarkers in different cancer types. OBJECTIVES: We aimed to evaluate Drosha and Dicer protein expression in cutaneous T-cell lymphoma (CTCL). METHODS: We performed Drosha and Dicer immunohistochemistry in 45 patients with mycosis fungoides and subtypes. Drosha and Dicer expression scores were correlated with clinical parameters including disease-specific death (DSD), stage of disease and different laboratory data. Uni- and multivariate statistics were performed. RESULTS: On univariate analysis, elevated serum LDH and low Drosha expression were significantly associated with advanced stage (P = 0.032 and 0.0062, respectively) and lymphoma-specific death (LSD; P = 0.017 and P = 0.005, respectively). Moreover, elevated circulating CD4+/CD26- lymphocytes were significantly associated with advanced stage (P = 0.032) and DSD (P = 0.0098). On multivariate analysis, low Drosha expression remained in the logistic regression model as significant independent predictor for advanced disease stages [P = 0.013; odds ratio: 5 (confidence interval) CI 1.3-19.3]. Moreover, low Drosha expression (P = 0.026) and elevated LDH (P = 0.025) remained as significant independent predictors for DSD with odds ratios of 13.5 (CI 1.3-134.4 and 8.7 CI 1.3-57.2, respectively). CONCLUSIONS: Low Drosha expression is an independent predictor for advanced stage as well as LSD in CTCL patients indicating a tumour suppressor gene function of Drosha in this disorder.

French maritime pine bark treatment decelerates plaque development and improves spatial memory in Alzheimer's disease mice.

BACKGROUND: Plant extracts are increasingly investigated as potential drugs against Alzheimer's disease (AD) and dementia in general. Pycnogenol is an extract from the bark of the French maritime pine (Pinus pinaster Aiton subsp. atlantica) with known anti-oxidative and neuroprotective effects. HYPOTHESIS/PURPOSE: Pycnogenol is thought to improve cognitive functions in elderly. We wanted to investigate and quantify these effects in a model system of cerebral ß-amyloidosis/AD. STUDY DESIGN/METHODS: This study experimentally assessed the effects of Pycnogenol on AD-related pathology in a ß-amyloidosis mouse model. APP-transgenic mice and controls were treated orally in a pre-onset and post-onset treatment paradigm. The effects of Pycnogenol were characterized by analysing ß-amyloid (Aß) plaques, number of neurons, glia coverage, myelination pattern, and cortical coverage with axons using immunohistochemistry. Aß levels were quantified using ELISA and gene expression levels of APP-processing enzymes ADAM10, BACE1 and IDE protein levels were determined by Western blot. Behavioural changes in circadian rhythm were monitored and spatial memory / cognition was assessed using a water maze test. RESULTS: Pycnogenol significantly decreased the number of plaques in both treatment paradigms but did not alter levels of soluble Aß or the gene expression of APP-processing enzymes. The morphological analyses revealed no changes in the number of neurons, astrocytes, microglia, the myelination pattern, or the morphology of axons. Behavioural testing revealed an improvement of the spatial memory in the pre-onset treatment paradigm only. CONCLUSION: Our results suggest to evaluate clinically a potential use of Pycnogenol in the prevention or in early stages of mild cognitive impairment (MCI) and AD.

Methodological Implications and Repeatability of Nasal Nitric Oxide: Relevance for Challenge Studies.

There is an interest in assessing changes in nasal NO (nNO) levels as an effect marker of upper airways. In this study, we examined methodologic influences on short and long term repeatability of nNO levels assessed by a portable electrochemical analyzer. Nine atopic and eighteen healthy subjects were exposed for 4 h to ethyl acrylate concentration of 0.05 ppm (sham) and mean concentrations of 5 ppm (either constant 5 ppm or variable 0 to 10 ppm). Sampling of nNO was performed by using passive aspiration during both breath-holding (634 ppb) or calm tidal breathing (364 ppb, p < 0.0001). The intra-session (between-session) repeatability in terms of coefficient of variation was 16.4% (18.5%) using the tidal-breathing and 8.6% (13.0%) using the breath-holding method, respectively. Atopic subjects demonstrated a significant increase in nNO (breath-holding mean 16%, tidal-breathing mean 32%) after applying a constant ethyl acrylate concentration (5 ppm). Our findings suggest that the less elaborate tidal-breathing method might be sufficient to detect significant changes at a group level. Given a lower coefficient of variation of breath-holding we assume there is an advantage of that approach at an individual level. Further research is needed to validate the usefulness of nNO in the evaluation of irritative, non-allergic responses.

Adaptation to Occupational Exposure to Moderate Endotoxin Concentrations: A Study in Sewage Treatment Plants in Germany.

Acute or chronic inhalation of endotoxin may lead to changes of lung function and inflammatory markers in the airways. Adaptation to workplace exposure may be possible. In this study, we investigated the possible difference in inflammatory markers assessed in nasal lavage fluid (NALF) in chronical exposure compared to voluntary subjects exposed acutely to endotoxin. We sought to define the variability of inflammatory markers in NALF and the dose-related changes after moderate exposure in naïve subjects. Endotoxin exposure (4-1039 EU/m3) resulted from routine work during one shift in sewage treatment plants. Subjects were matched to pairs (8 workers escorted by 10 students). Inflammatory markers were investigated before, directly after, and 16 h after the shift end. Additional NALF samples were collected in students without any specific exposure after 3 days. In NALF, total cell count, and interleukin (IL)-8 and IL-1ß concentrations were significantly higher in workers than in students at all times pointing to workplace-related long-lasting exposure resulting in adaptation. However, concentration of inflammatory markers without specific exposure in students showed a great variability, covering the whole range of values recorded in the workers. The findings of this study make us to recommend a repeated assessment of inflammatory markers in healthy volunteers before the investigation of exposure-related changes and a sample size adequate for statistical analysis.

Methylation of L1RE1, RARB, and RASSF1 function as possible biomarkers for the differential diagnosis of lung cancer.

BACKGROUND: Lung cancer is the major cause of cancer-related deaths worldwide. Differential diagnosis can be difficult, especially when only small samples are available. Epigenetic changes are frequently tissue-specific events in carcinogenesis and hence may serve as diagnostic biomarkers. MATERIAL AND METHODS: 138 representative formalin-fixed, paraffin-embedded (FFPE) tissues (116 lung cancer cases and 22 benign controls) were used for targeted DNA methylation analysis via pyrosequencing of ten literature-derived methylation markers (APC, CDH1, CDKN2A, EFEMP1, FHIT, L1RE1, MGMT, PTEN, RARB, and RASSF1). Methylation levels were analyzed with the Classification and Regression Tree Algorithm (CART), Conditional Interference Trees (ctree) and ROC. Validation was performed with additional 27 lung cancer cases and 38 benign controls. TCGA data for 282 lung cancer cases was included in the analysis. RESULTS: CART and ctree analysis identified the combination of L1RE1 and RARB as well as L1RE1 and RASSF1 as independent methylation markers with high discriminative power between tumor and benign tissue (for each combination, 91% specificity and 100% sensitivity). L1RE1 methylation associated significantly with tumor type and grade (p<0.001) with highest methylation in the control group. The opposite was found for RARB (p<0.001). RASSF1 methylation increased with tumor type and grade (p<0.001) with strongest methylation in neuroendocrine tumors (NET). CONCLUSION: Hypomethylation of L1RE1 is frequent in tumors compared to benign controls and associates with higher grade, whereas increasing methylation of RARB is an independent marker for tumors and higher grade. RASSF1 hypermethylation was frequent in tumors and most prominent in NET making it an auxiliary marker for separation of NSCLC and NET. L1RE1 in combination with either RARB or RASSF1 could function as biomarkers for separating lung cancer and non-cancerous tissue and could be useful for samples of limited size such as biopsies.