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Supplementary Tables S1 & S2 from Sustained Expression of Early Growth Response Protein-1 Blocks Angiogenesis and Tumor Growth

Transient induction of the transcription factor early growth response protein-1 (EGR-1) plays a pivotal role in the transcriptional response of endothelial cells to the angiogenic growth factors vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), which are produced by most tumors and are involved in the angiogenic switch. We report here that sustained expression of EGR-1 by recombinant adenoviruses in endothelial cells, however, leads to the specific induction of potent feedback inhibitory mechanisms, including strong up-regulation of transcriptional repressors, negative cell cycle check point effectors, proteins with established antiangiogenic activity, and several proapoptotic genes. Sustained EGR-1 expression consistently leads to an antiangiogenic state characterized by an altered responsiveness to VEGF and bFGF and a striking inhibition of sprouting and tubule formation in vitro. Furthermore, EGR-1–expressing viruses potently inhibit cell invasion and vessel formation in the murine Matrigel model and repress tumor growth in a murine fibrosarcoma model. We propose that gene therapy involving sustained EGR-1 expression may constitute a novel therapeutic principle in the treatment of cancer due to the simultaneous induction of multiple pathways of antiangiogenesis, growth arrest, and apoptosis induction in proliferating cells leading to preferential inhibition of angiogenesis and tumor growth. (Cancer Res 2006; 66(13): 6708-13)

ObjectivesVarious commercial anti-Spike SARS-CoV-2 antibody tests are used for studies and in clinical settings after vaccination. An international standard for SARS-CoV-2 antibodies has been established to achieve comparability of such tests, allowing conversions to BAU/ml. This study aimed to investigate the comparability of antibody tests regarding the timing of blood collection after vaccination.MethodsFor this prospective observational study, antibody levels of 50 participants with homologous AZD1222 vaccination were evaluated at 3 and 11 weeks after the first dose and 3 weeks after the second dose using two commercial anti-Spike binding antibody assays (Roche and Abbott) and a surrogate neutralization assay.ResultsThe correlation between Roche and Abbott changed significantly depending on the time point studied. Although 3 weeks after the first dose, Abbott provided values three times higher than Roche, 11 weeks after the first dose, the values for Roche were twice as high as for Abbott, and 3 weeks after the second dose even 5-6 times higher.ConclusionsThe comparability of quantitative anti-Spike SARS-CoV-2 antibody tests is highly dependent on the timing of blood collection after vaccination. Therefore, standardization of the timing of blood collection might be necessary for the comparability of different quantitative SARS-COV-2 antibody assays.Abstract Figure

Changes in circulating cell populations may promote ischemic events that occur soon after discontinuation of P2Y12-inhibition. The aim of the study was to track the course of thrombopoietic and ery...

ObjectivesOur objective was to determine whether SARS-CoV-2 antibody levels after the first dose can predict the final antibody response and whether this is dependent on the vaccine type.Methods69 BNT162b2 (Pfizer/BioNTech) and 55 AZD1222 (AstraZeneca) vaccinees without previous infection or immunosuppressive medication were included. Anti-body levels were quantified 3 weeks after dose 1, in case of AZD1222 directly before boostering (11 weeks after dose 1) and 3 weeks after dose 2, with the Roche SARS-CoV-2 S total antibody assay.ResultsPre-booster (BNT162b2: 80.6 [25.5-167.0] BAU/mL, AZD1222: 56.4 [36.4-104.8] BAU/mL, not significant) and post-booster levels (BNT162b2: 2,092.0 [1,216.3-4,431.8] BAU/mL, AZD1222: 957.0 [684.5-1,684.8] BAU/mL, pConclusionsIn conclusion, our data suggest that antibody levels quantified by the Roche Elecsys SARS-CoV-2 S assay before the booster shot could infer post-booster responses to BNT162b2, but not to AZ1222. In addition, we found a vaccine-dependent effect on antibody responses, suggesting a possible link between vaccine response and vector immunity.

ObjectivesSARS-CoV-2 infection induces the formation of different antibodies. However, not all of which might prevent the virus from entering the cell, although their concentrations correlate with the titers of viral neutralization tests (NTs). Antibodies against the viral nucleocapsid (NC), e.g., can be classified as such. We aimed to prove the hypothesis that the apparent correlation between NC-antibody levels and NT-titers is mediated by simultaneously occurring antibodies against viral spike-protein components.MethodsWe included 64 individuals with previous SARS-CoV-2 infection (>14d after symptom onset). SARS-CoV-2 antibodies against the NC (Roche total antibody ECLIA, Abbott IgG CMIA) and spike-protein (Technozym RBD ELISA, DiaSorin S1/S2 CLIA) were measured, and neutralization tests were performed. The effect of spike-protein antibodies on the correlation between NC-antibodies and NT-titers was evaluated by partial correlation and mediation analyses.ResultsBoth tested assays assessing antibodies against the NC correlated significantly with NT titers: Abbott ρ=0.742, PConclusionsOur data suggest that the apparent correlation between NC antibodies and NT titers is strongly mediated by co-occurring RBD antibody concentrations. To avoid falsely implied causal relationships, all correlation analyses of non-spike-associated antibody assays and neutralization assays should include a partial correlation analysis to exclude a possible mediator effect of spike-associated antibodies.

Reliable quantification of the antibody response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is highly relevant, e.g., for identifying possible vaccine failure and estimating the time of protection. Therefore, we evaluated five different anti-SARS-CoV-2 antibody assays regarding the quantification of anti-spike (S) antibodies. Sera from 69 SARS-CoV-2-naive individuals 21 ± 1 days after vaccination with a single dose of BNT162b2 (Pfizer/BioNTech) were tested using the following quantitative assays: Roche S total antibody, DiaSorin trimeric spike IgG, DiaSorin S1/S2 IgG, Abbott II IgG, and Serion/Virion IgG. Results were further compared to the percent inhibition calculated from a surrogate virus neutralization test (sVNT). Individual values were distributed over several orders of magnitude for all assays. Although the assays were in good overall agreement (ρ = 0.80 to 0.94), Passing-Bablok regression revealed systematic constant and proportional differences, which could not be eliminated by converting the results to binding antibody units (BAU) per milliliter, as suggested by the manufacturers. Seven (10%) individuals had negative sVNT results (i.e.

Background There is preliminary evidence that individuals with previous SARS‐CoV‐2 infections exhibit a more pronounced antibody response. However, these assumptions have not yet been supported by data obtained through various CE‐marked tests. This study aimed to close this gap. Methods Sixty‐nine seronegatives and 12 individuals post‐SARS‐CoV‐2 infection (tested by CE‐labelled Roche NC immunoassay or PCR‐confirmed assay) were included 21 ± 1 days after receiving the first dose of the Pfizer/BioNTech BNT162b2 vaccine. Antibody response to viral spike protein (S) was assessed by CE‐labelled Roche S and DiaSorin S1/S2 assays and by a surrogate virus neutralization test (sVNT). Results After a single dose of BNT162b2, individuals after natural SARS‐CoV‐2 infection presented with markedly higher anti‐S levels than naïve individuals (Roche S: 9078.5 BAU/mL [5267.0‐24 298.5] vs 79.6 [24.7‐142.3]; and DiaSorin S1/S2: 1465.0 AU/mL [631.0‐5365.0] vs 63.7 [47.8‐87.5]) and showed all the maximum observed inhibition activity in the sVNT (98%), without overlaps between groups. There was a trend for higher responses in those with a more distant infection, although not statistically significant. The relative antibody increase after dose 2 was significantly higher among naïve individuals (25‐fold), but antibody levels remained below that of seropositives. Conclusions Compared with naïve individuals, seropositives after natural SARS‐CoV‐2 infection presented with a substantially higher antibody response already after dose 1 of BNT162b2, as measured by two CE‐marked in vitro diagnostic tests and a sVNT. These results should stimulate discussion and research on whether individuals after previous SARS‐CoV‐2 infection would benefit from a two‐part vaccination schedule or whether these currently much‐needed second doses could be saved.

Purpose: To explore differences in baseline characteristics following three weeks of semi-standardized in-patient care between patients with obesity without and with type 2 diabetes (T2D). Patients and methods: Patients without or with T2D were matched according to age, sex, and BMI. Food intake was restricted to 1,200-1,600 kcal/d to which a 400-600 kcal/d exercise load was added, and data were compared using Student's t-test, general linear models, and Spearman-rank correlations. Results: At baseline, patients with obesity and T2D displayed, besides elevated blood glucose and HbA1c values, higher serum liver enzymes (p

BackgroundReliable quantification of the antibody response to SARS-CoV-2 vaccination is highly relevant for identifying possible vaccine failure and estimating the time of protection. Therefore, we aimed to evaluate the performance of five different Anti-SARS-CoV-2 antibody assays regarding the quantification of anti-spike (S) antibodies induced after a single dose of BNT162b2.MethodsSera of n=69 SARS-CoV-2 naïve individuals 21±1 days after vaccination with BNT162b2 (Pfizer/BioNTech) were tested using the following quantitative SARS-CoV-2 antibody assays: Roche S total antibody, DiaSorin trimeric spike IgG, DiaSorin S1/S2 IgG, Abbott II IgG, and Serion/Virion IgG. Test agreement was assessed by Passing-Bablok regression. Results were further compared to the percent inhibition calculated from a surrogate virus neutralization test (sVNT) by correlation and ROC (receiver-operating-characteristics) analysis.ResultsIndividual values were distributed over several orders of magnitude for all assays evaluated. Although the assays were in good overall agreement (ρ=0.80-0.94), Passing-Bablok regression revealed systematic and proportional differences, which could not be eliminated by converting the results to BAU/mL as suggested by the manufacturers. 7 (10%) individuals had a negative sVNT results (i.e. ConclusionsAlthough all assays evaluated showed good correlation, readings from different assays were not interchangeable, even when converted to BAU/mL using the WHO international standard for SARS-CoV-2 immunoglobulin. This highlights the need for further standardization of SARS-CoV-2 serology.

In an aging society, late‐life depression has become an increasing problem. There is evidence that physical activity ameliorates depressive symptoms and increases the quality of life (QoL). However, the underlying mechanisms are still poorly understood. Myokines are molecules secreted in response to muscle contraction. Some of them can cross the blood‐brain barrier, making them promising candidates for mediating the beneficial effects of physical activity on mood. The present study aims to compare circulating myokine levels to depression/QoL in older athletes and controls. 55 athletes, 57 controls >59 years were enrolled. The assessment included ergometry, magnetic resonance imaging, blood withdrawal, and neuropsychological testing. Serum interleukin‐6 (IL‐6), irisin, brain‐derived neurotrophic factor (BDNF), kynurenine, and cathepsin B were analyzed and compared to surrogates of depression and quality of life. Athletes presented with higher levels of Cathepsin B. Among controls, all myokines but irisin were associated with age. Also, among controls, kynurenine and IL‐6 correlated inversely with specific dimensions of quality of life questionnaires, and IL‐6 further with depressive symptoms and decreased physical performance. No such associations could be found among athletes. Irisin levels were inversely associated with mild depression and low‐grade white matter‐lesions in the brain and predicted impaired QoL. The circulating levels of several myokines/muscle activity‐related factors appear to be associated with depressive symptoms and impaired QoL among older adults. However, in athletes, some of these connections seem ameliorated, suggesting additional stressors (as f.e. age) or a different pathomechanism among athletes., Impact Statement Physical exercise positively affects mood and quality of life, however, the underlying mechanisms are not yet fully understood. Myokines are released in response to muscle contraction. They might be promising candidates for mediating muscle‐brain‐crosstalk. Our results suggest that basal kynurenine and interleukin‐6 levels are correlated with poor quality of life and/or depressive symptoms. In contrast, low irisin levels were associated with mild depression and low‐grade white matter‐hyperintensities in brain MRIs and they predicted impaired quality of life.

Background Serological tests are widely used in various medical disciplines for diagnostic and monitoring purposes. Unfortunately, the sensitivity and specificity of test systems is often poor, leaving room for false positive and false negative results. However, conventional methods used to increase specificity decrease sensitivity and vice versa. Using SARS-CoV-2 serology as an example, we propose here a novel testing strategy: the "Sensitivity Improved Two-Test" or "SIT2" algorithm. Methods SIT2 involves confirmatory re-testing of samples with results falling in a predefined retesting-zone of an initial screening test, with adjusted cut-offs to increase sensitivity. We verified and compared the performance of SIT2 to single tests and orthogonal testing (OTA) in an Austrian cohort (1,117 negative, 64 post-COVID positive samples) and validated the algorithm in an independent British cohort (976 negatives, 536 positives). Results The specificity of SIT2 was superior to single tests and non-inferior to OTA. The sensitivity was maintained or even improved using SIT2 when compared to single tests or OTA. SIT2 allowed correct identification of infected individuals even when a live virus neutralization assay could not detect antibodies. Compared to single testing or OTA, SIT2 significantly reduced total test errors to 0.46% (0.24-0.65) or 1.60% (0.94-2.38) at both 5% or 20% seroprevalence. Conclusion For SARS-CoV-2 serology, SIT2 proved to be the best diagnostic choice at both 5% and 20% seroprevalence in all tested scenarios. It is an easy to apply algorithm and can potentially be helpful for the serology of other infectious diseases.

Abstract Obesity and body fat distribution are important risk factors for the development of type 2 diabetes and metabolic syndrome. Evidence has accumulated that this risk is related to intrinsic differences in behavior of adipocytes in different fat depots. We recently identified LIM domain only 3 (LMO3) in human mature visceral adipocytes; however, its function in these cells is currently unknown. The aim of this study was to determine the potential involvement of LMO3-dependent pathways in the modulation of key functions of mature adipocytes during obesity. Based on a recently engineered hybrid rAAV serotype Rec2 shown to efficiently transduce both brown adipose tissue (BAT) and white adipose tissue (WAT), we delivered YFP or Lmo3 to epididymal WAT (eWAT) of C57Bl6/J mice on a high-fat diet (HFD). The effects of eWAT transduction on metabolic parameters were evaluated 10 weeks later. To further define the role of LMO3 in insulin-stimulated glucose uptake, insulin signaling, adipocyte bioenergetics, as well as endocrine function, experiments were conducted in 3T3-L1 adipocytes and newly differentiated human primary mature adipocytes, engineered for transient gain or loss of LMO3 expression, respectively. AAV transduction of eWAT results in strong and stable Lmo3 expression specifically in the adipocyte fraction over a course of 10 weeks with HFD feeding. LMO3 expression in eWAT significantly improved insulin sensitivity and healthy visceral adipose tissue expansion in diet-induced obesity, paralleled by increased serum adiponectin. In vitro, LMO3 expression in 3T3-L1 adipocytes increased PPARγ transcriptional activity, insulin-stimulated GLUT4 translocation and glucose uptake, as well as mitochondrial oxidative capacity in addition to fatty acid oxidation. Mechanistically, LMO3 induced the PPARγ coregulator Ncoa1, which was required for LMO3 to enhance glucose uptake and mitochondrial oxidative gene expression. In human mature adipocytes, LMO3 overexpression promoted, while silencing of LMO3 suppressed mitochondrial oxidative capacity. LMO3 expression in visceral adipose tissue regulates multiple genes that preserve adipose tissue functionality during obesity, such as glucose metabolism, insulin sensitivity, mitochondrial function, and adiponectin secretion. Together with increased PPARγ activity and Ncoa1 expression, these gene expression changes promote insulin-induced GLUT4 translocation, glucose uptake in addition to increased mitochondrial oxidative capacity, limiting HFD-induced adipose dysfunction. These data add LMO3 as a novel regulator improving visceral adipose tissue function during obesity. Key messages LMO3 increases beneficial visceral adipose tissue expansion and insulin sensitivity in vivo. LMO3 increases glucose uptake and oxidative mitochondrial activity in adipocytes. LMO3 increases nuclear coactivator 1 (Ncoa1). LMO3-enhanced glucose uptake and mitochondrial gene expression requires Ncoa1.

Changes in circulating cell populations may promote ischemic events that occur soon after discontinuation of P2Y

Background In the context of the COVID-19 pandemic, numerous new serological test systems for the detection of anti-SARS-CoV-2 antibodies rapidly have become available. However, the clinical performance of many of these is still insufficiently described. Therefore, we compared 3 commercial CE-marked, SARS-CoV-2 antibody assays side by side. Methods We included a total of 1154 specimens from pre-COVID-19 times and 65 samples from COVID-19 patients (≥14 days after symptom onset) to evaluate the test performance of SARS-CoV-2 serological assays by Abbott, Roche, and DiaSorin. Results All 3 assays presented with high specificities: 99.2% (98.6–99.7) for Abbott, 99.7% (99.2–100.0) for Roche, and 98.3% (97.3–98.9) for DiaSorin. In contrast to the manufacturers’ specifications, sensitivities only ranged from 83.1% to 89.2%. Although the 3 methods were in good agreement (Cohen’s Kappa 0.71–0.87), McNemar tests revealed significant differences between results obtained from Roche and DiaSorin. However, at low seroprevalences, the minor differences in specificity resulted in profound discrepancies of positive predictive values at 1% seroprevalence: 52.3% (36.2–67.9), 77.6% (52.8–91.5), and 32.6% (23.6–43.1) for Abbott, Roche, and DiaSorin, respectively. Conclusion We found diagnostically relevant differences in specificities for the anti-SARS-CoV-2 antibody assays by Abbott, Roche, and DiaSorin that have a significant impact on the positive predictive values of these tests.

Childhood obesity is an increasing health care problem associated with insulin resistance and low-level systemic inflammation, which can ultimately lead to diabetes. Evidence for efficacy of therapeutic intervention programs on the early development of obesity associated sequelae is moderate. This paper investigates the effect of a multidisciplinary short-term intervention program on insulin resistance and metaflammation in childhood obesity. Two hundred and thirty-six overweight or obese children and adolescents between the ages of 10 and 14 were included in a prospective five-month intervention study, which included sports, psychotherapy, and nutritional counseling. Primary endpoints were the effects on body mass index standard deviation score (BMI-SDS) and homeostatic model assessment of insulin resistance (HOMA-IR), key secondary endpoints were the levels of C-reactive protein (CRP), leptin, and adiponectin. At baseline, a substantial proportion of participants showed signs of insulin resistance (mean HOMA-IR 5.5  3.4) despite not meeting the diagnostic criteria for diabetes, and low-level inflammation (mean CRP 3.9 mg/l  3.8 mg/l). One hundred and ninety-five participants (83%) completed the program resulting in a significant reduction in BMI-SDS, HOMA IR, CRP, and leptin and a significant increase in adiponectin (mean change compared to baseline -0.14, -0.85, -1.0 mg/l, -2.8 ng/ml, and 0.5µg/ml respectively; p < .001 each). Effects on BMI SDS, HOMA IR, CRP, and adiponectin were largely independent whereas leptin was positively correlated with BMI-SDS and total fat mass before and after intervention (r = 0.56 and 0.61, p < .001 each). Short-term multidisciplinary intervention successfully improved body composition, insulin sensitivity, low-level systemic inflammation, and metabolic profile of adipose tissue in childhood obesity. Our findings highlight the immediate connection between obesity and the pathophysiology of its sequelae, and emphasize the importance of early intervention. Continued lifestyle modification is likely necessary to consolidate and augment the long-term effects.

Introduction Fractures of the proximal femur are a significant cause of mortality and morbidity in the elderly population. Yet predictive marker of unfavourable prognosis are still lacking. Calcium phosphate product is a marker of osteo-renal dysregulation. This study investigated the role of serum calcium phosphate product (SCPP) levels as a prognostic parameter for outcome in those patients. Patients and methods A total of 3577 consecutive patients with diagnosed fractures of the proximal femur were included in our study (72.5% females). SCPP was divided into tertiles: 2 /l 2 , 1.93–2.38 mmol 2 /l 2 and >2.39 mmol 2 /l 2 . Data collection was performed prospectively and statistical evaluation was performed retrospectively. Results Mean follow up in our study group was 11.0 ± 0.3 months. The mean age of our study group was 79.0 years (SEM ±14 years). To facilitate analysis, patients were divided in two groups: ≤84 years (64.4%) and ≥85 years (35.6%), and mortality n = 445). In our study population higher SCPP levels ad admission were associated with a markedly elevated mortality. In a multivariate logistic regression model adjusted for age and sex, plasma creatinine and haemoglobin at admission caused a 1.3 (CI: 1.01–1.6) for SCPP 1.93–2.38 mmol 2 /l 2 , and a 1.6 (CI: 1.2–2.0) for SPP >2.39 mmol 2 /l 2 fold increase in overall mortality compared to patients with baseline SCPP levels ( 2 /l 2 ) as reference category. Conclusion Those findings in our study population with 3577 patients over a period of 20 years proved to be, that serum Ca levels may be a good predictor for mortality in patients with fracture of the proximal femur. Further studies are required to evaluate whether these high risk patients might benefit from specific therapeutic measurements. This prognostic factor may help to increase the outcome of elderly patients with a fracture of the proximal femur.

Background and Aim: It is increasingly recognized that biomedical research has serious reproducibility issues, which could be overcome at least in part by standardized processing of biomaterials. Therefore, professional biobanks have emerged, positively influencing sample and data quality. However, quantitative data about a biobank's contribution to published results are still hard to find, although they could serve as valuable benchmark figures for the community. We therefore aimed to report usage data from the MedUni Wien Biobank facility regarding its prospective fluid cohorts. Methods: Input and access statistics and publication output were reported for the years 2010–2017. Performance dynamics were tested by correlation analyses according to Spearman. Additionally, virtual costs per sample were calculated. Results: The amount of annually collected aliquots rose significantly from 68,500 in 2010 to 151,966 in 2017 (p = 0.015), although no further increase was recorded after 2012 (p = 0.266). In the same period, the quotient of requested to stored aliquots increased from 3.5% to 6.1% (p = 0.001), as the yearly number of requested aliquots nearly quadrupled from 2401 to 9342. Likewise, the number of published research articles per year to which the MedUni Wien Biobank contributed increased from 2 (total impact factor: 8.6) in 2010 to 16 (total impact factor: 69.0) in 2017, resulting in a total of 69 identified publications. Currently, the biobank operates at 15- to 20-fold overproduction, leading to virtual costs per accessed sample of ∼€20. Conclusion: The reported usage data might serve as a benchmark for other hospital-integrated biobanks, and implies that academic biobanks are able to produce considerable scientific impact at comparable moderate costs.

Introduction Hip fractures are a significant cause of mortality and morbidity in the elderly. This study investigated the relationship between initial haemoglobin (Hb) levels and a prognostic parameter for outcome in those patients. Patients and methods A total of 3595 consecutive patients with diagnosed hip fractures were included in our study (72.2% females). Anaemia was defined according to WHO criteria, with according subgroups mild, moderate and severe anaemia. Data collection was performed prospectively and statistical evaluation was performed retrospectively. Results Mean follow up in our study group was 11.2±0.3 months. The mean age of our study group was 78.5 years (SEM±0.2 years). To facilitate analysis, patients were divided in two groups: ≤84 years (60.1%) and ≥85 years (39.9%). Mortality n =439). In our study population lower Hb levels ad admission were associated with a markedly elevated short-term mortality. In a multivariate logistic regression model adjusted for age and sex, mild anaemia at admission caused a 1.5 (CI: 1.1–1.9), moderate anaemia a 2.6 (95 CI: 2.0–3.4), and severe anaemia a 3.6 (CI: 1.8–6.9) fold increase in three months mortality compared to patients without anaemia. Total lymphocyte count (1.2±0) did not show any differences between the subgroups. Conclusion Those findings in our study population with 3595 patients over a period of twenty years have proven that initial Hb levels are a useful and cost effective parameter to predict mortality in elderly patients with a hip fracture. This prognostic factor may help to increase the outcome of elderly patients with a hip fracture.

SummaryCellular adhesion molecules also known as selectins promote recruitment of inflammatory cells into the arterial wall where they interact with lipid particles leading subsequently to plaque formation. The intercellular adhesion molecule-1 (ICAM-1), the vascular cell adhesion molecule-1 (VCAM-1) and the endothelial-leukocyte adhesion molecule 1 (ELAM-1) also known as E-selectin mediate the attachment of leukocytes and have been implicated in the destabilisation of atherosclerotic plaques. Therefore, we hypothesised that plasma selectin levels are associated with adverse clinical outcome. We prospectively studied 855 patients with sonographically confirmed carotid atherosclerosis. During a median follow-up of 6.2 years, corresponding to 5,551 overall person-years, 275 patients (26 %) died. We detected a significant association between cardiovascular mortality and ICAM-1 (adjusted hazard ratio [HR]: 3.43, 95 % confidence interval [CI] 2.00–5.88, p< 0.001) as well as VCAM-1 (adjusted HR: 2.51, 95 % CI 1.45–4.34, p=0.001) when comparing the fourth with the first quartile. Comparable results were obtained for all-cause mortality. In contrast, we could not detect a significant association between E-selectin and all-cause or cardiovascular mortality. We identified the selectins ICAM-1 and VCAM-1 as strong and independent predictors of all-cause and cardiovascular mortality in patients with stable carotid atherosclerosis. These molecules are elevated in states of endothelial activation and might assist to monitor anti-atherosclerotic therapy and select those patients with carotid atherosclerosis, who are at higher risk for cardiovascular events.

Over the past years, plenty of evidence has emerged illustrating how metabolism supports many aspects of cellular function and how metabolic reprogramming can drive cell differentiation and fate. Here, we present a method to assess the metabolic configuration of single cells within their native tissue microenvironment via the visualization and quantification of multiple enzymatic activities measured at saturating substrate conditions combined with subsequent cell type identification. After careful validation of the approach and to demonstrate its potential, we assessed the intracellular metabolic configuration of different human immune cell populations in healthy and tumor colon tissue. Additionally, we analyzed the intercellular metabolic relationship between cancer cells and cancer-associated fibroblasts in a breast cancer tissue array. This study demonstrates that the determination of metabolic configurations in single cells could be a powerful complementary tool for every researcher interested to study metabolic networks in situ.

Endurance sports are enjoying greater popularity, particularly among new target groups such as the elderly. Predictors of future physical capacities providing a basis for training adaptations are in high demand. We therefore aimed to estimate the future physical performance of elderly marathoners (runners/bicyclists) using a set of easily accessible standard laboratory parameters. To this end, 47 elderly marathon athletes underwent physical examinations including bicycle ergometry and a blood draw at baseline and after a three-year follow-up period. In order to compile a statistical model containing baseline laboratory results allowing prediction of follow-up ergometry performance, the cohort was subgrouped into a model training (n = 25) and a test sample (n = 22). The model containing significant predictors in univariate analysis (alanine aminotransferase, urea, folic acid, myeloperoxidase and total cholesterol) presented with high statistical significance and excellent goodness of fit (R2 = 0.789, ROC-AUC = 0.951±0.050) in the model training sample and was validated in the test sample (ROC-AUC = 0.786±0.098). Our results suggest that standard laboratory parameters could be particularly useful for predicting future physical capacity in elderly marathoners. It hence merits further research whether these conclusions can be translated to other disciplines or age groups.

Background Moderately elevated unconjugated bilirubin concentrations protect against inflammatory diseases and are present in individuals with Gilbert's syndrome. This study examined the relationship between circulating haem oxygenase catabolites, unconjugated bilirubin, carboxy haemoglobin, iron and inflammatory parameters. Materials and methods Seventy-six matched individuals were allocated to Gilbert's syndrome (GS) or control group (unconjugated bilirubin ≥ or

Increased lipid availability reduces insulin-stimulated glucose disposal in skeletal muscle, which is generally explained by fatty acid–mediated inhibition of insulin signaling. It remains unclear whether lipids also impair transcapillary transport of insulin and glucose, which could become rate controlling for glucose disposal. We hypothesized that lipid-induced insulin resistance is induced by inhibiting myocellular glucose uptake and not by interfering with the delivery of insulin or glucose. We measured changes in interstitial glucose and insulin in skeletal muscle of healthy volunteers during intravenous administration of triglycerides plus heparin or glycerol during physiologic and supraphysiologic hyperinsulinemia, by combining microdialysis with oral glucose tolerance tests and euglycemic-hyperinsulinemic clamps. Lipid infusion reduced insulin-stimulated glucose disposal by ∼70% (P < 0.05) during clamps and dynamic insulin sensitivity by ∼12% (P < 0.05) during oral glucose loading. Dialysate insulin and glucose levels were unchanged or even transiently higher (P < 0.05) during lipid than during glycerol infusion, whereas regional blood flow remained unchanged. These results demonstrate that short-term elevation of free fatty acids (FFAs) induces insulin resistance, which in skeletal muscle occurs primarily at the cellular level, without impairment of local perfusion or transcapillary transport of insulin and glucose. Thus, vascular effects of FFAs are not rate controlling for muscle insulin-stimulated glucose disposal.

Background:Irreproducibility of scientific results constitutes an undesirably onerous economic burden and is in many cases caused by low-quality materials. Therefore, researchers are increasingly devoting their attention to the bioresources they use. In turn, those bioresources are required to validate their preanalytical processes in order to ensure best possible quality. The present study thus aimed to evaluate the impact of repeated temperature fluctuations, as they occur in most research biobanks due to repetitive opening and closing of freezer doors, on the stability of 26 biochemical analytes.Methods:Serum of 43 individuals was randomly assigned to a fluctuation (n=21) and a control group (n=22). Serum of the fluctuation group underwent controlled temperature fluctuations (30 fluctuations Results:Sixteen biomarkers showed statistically significant changes over time, whereas only seven of those presented with diagnostically/clinically relevant changes at certain time points (aspartate aminotransferase, amylase, calcium, uric acid, creatinine, inorganic phosphate and total protein). However, there was no difference between the fluctuation and the control group.Conclusions:Some serum analytes are influenced by storage, even at temperatures as low as

Background and Purpose— Inflammatory responses play a key role in atherogenesis. The aim of this study was to assess the prognostic value of hsCRP (high-sensitivity C-reactive protein) and to evaluate whether degree of carotid stenosis and serum levels of hsCRP jointly predict long-term mortality in asymptomatic patients with carotid atherosclerosis. Methods— One thousand sixty-five patients with neurological asymptomatic carotid atherosclerosis as evaluated by duplex sonography were prospectively followed for cause-specific mortality. Results— During a median of 11.81 years, a total of 549 deaths, including 362 cardiovascular deaths, were recorded. The risk of all-cause and cardiovascular mortality significantly increased in patients with elevated serum levels of hsCRP (the adjusted hazard ratio for cardiovascular mortality per increase of 1 mg/dL of hsCRP levels was 1.47; P 0.29 mg/dL (=median) had nearly twice as high a risk of cardiovascular mortality compared with patients with carotid stenosis of P P Conclusions— Measurement of hsCRP in combination with ultrasound investigations of the carotid arteries at a single time point provides additional prognostic information for patients with asymptomatic carotid atherosclerosis.

A single nucleotide variant within the promoter of the 5-hydroxytryptamine1A (5HT1A) receptor, rs6295, is part of a binding site for the transcription factor NUDR (nuclear-DEAF-1-related). We aimed to ascertain whether the rs6295 mediates the effect of exercise on depressive mood in elderly endurance athletes. We prospectively enrolled 55 elderly athletes (marathon runners/bicyclists) and 58 controls. In a controlled, univariate model, an interaction between the [C]-allele and physical activity indicated that only among athletes, the variant resulting in an imperfect NUDR binding site was associated with a lower depression score. Hence, athletes presented with a significantly lower relative risk of achieving a suspicious depression score among carriers of at least one [C]-allele. Our results suggest that the positive effect of physical exercise on depressive mood might be mediated by the 5HT1A receptor and the extent of this protective effect seems to be enhanced by the [C]-allele of the rs6295 variant.

Background and Aim Recent studies revealed a link between hypovitaminosis D3 and the risk for hyperglycemia. Further mechanistic and interventional investigations suggested a common reason for both conditions rather than a causal relationship. Exposure to sunlight is the most relevant source of vitamin D3 (25(OH)D), whereas adipose tissue is able to store relevant amounts of the lipophilic vitamin. Since running/bicycling leads to increased out-door time and alters physiological response mechanisms, it can be hypothesized that the correlation between hypovitaminosis D3 and hyperglycemia might be disturbed in outdoor athletes. Methods 47 elderly marathoners/bicyclists and 47 age/sex matched controls were studied in a longitudinal setting at baseline and after three years. HbA1c as a surrogate for (pre-)diabetic states was quantified via HPLC, 25(OH)D levels were measured by means of chemiluminescent assays. Physical performance was assessed by ergometry. Results When adjusted for seasonal variations, 25(OH)D was significantly higher in athletes than in controls. 25(OH)D levels inversely correlated with triglycerides in both groups, whereas only in controls an association between high BMI or low physical performance with hypovitaminosis D3 had been found. Likewise, the presence of hypovitaminosis D3 at baseline successfully predicted hyperglycemia at the follow up examinations within the control group (AUC = 0.85, 95% CI [0.74, 0.96], p < .001, statistically independent from BMI), but not in athletes. Conclusion Our data suggest that mechanisms of HbA1c elevation might differ between athletes and controls. Thus, intense physical activity must be taken into account as a potential pre-analytic confounder when it is aimed to predict metabolic risk by vitamin D3 levels.

Atherosclerotic lesions are characterized by the accumulation of oxidized LDL (OxLDL) and the infiltration of macrophages and T cells. Cytokine expression in the microenvironment of evolving lesions can profoundly contribute to plaque development. While the pro-atherogenic effect of T helper (Th) 1 cytokines, such as IFN-γ, is well established, the role of Th2 cytokines is less clear. Therefore, we characterized the role of the Th2 cytokine interleukin (IL)-13 in murine atherosclerosis. Here, we report that IL-13 administration favourably modulated the morphology of already established atherosclerotic lesions by increasing lesional collagen content and reducing vascular cell adhesion molecule-1 (VCAM-1)-dependent monocyte recruitment, resulting in decreased plaque macrophage content. This was accompanied by the induction of alternatively activated (M2) macrophages, which exhibited increased clearance of OxLDL compared to IFN-γ-activated (M1) macrophages in vitro. Importantly, deficiency of IL-13 results in accelerated atherosclerosis in LDLR(-/-) mice without affecting plasma cholesterol levels. Thus, IL-13 protects from atherosclerosis and promotes a favourable plaque morphology, in part through the induction of alternatively activated macrophages.

ObjectiveProgranulin (PGRN) was recently introduced as a novel marker of chronic inflammatory response in obesity and type 2 diabetes capable of directly affecting the insulin signaling pathway. This study aimed to investigate the role of PGRN in gestational diabetes mellitus (GDM), which is regarded as a model for early type 2 diabetes.MethodsPGRN serum levels were measured in 90 pregnant women (45 GDM and 45 normal glucose tolerance (NGT)). In addition, PGRN was measured during a 2-h, 75 g oral glucose tolerance test in 20 pregnant women (ten GDM and ten NGT) and in 16 of thempost partum(ten GDM and six NGT).ResultsPGRN concentrations were significantly higher in pregnant women compared withpost partumlevels (536.79±31.81 vs 241.53±8.86,PR=0.47,P=0.034;R=0.63,P=0.012). HbA1c and the oral glucose insulin sensitivity index showed significantpost partumassociations with PGRN (R=0.43,P=0.049;R=−0.65,P=0.009).ConclusionsPGRN concentrations are markedly lower after pregnancy regardless of the gestational glucose tolerance state. PGRN levelsper sedo not discriminate between mild GDM and NGT in pregnant women. Therefore, the development of GDM appears to be due to impaired β-cell function that is not related to PGRN effect.

The main purpose of the study was to investigate whether baseline myeloperoxidase (MPO) levels are associated with executive cognitive function in individuals with high physical activity. Baseline serum MPO levels of 56 elderly marathon runners and 58 controls were assessed by ELISA. Standardized tests were applied to survey domain-specific cognitive functions. Changes in brain morphology were visualized by magnetic resonance imaging (MRI). High baseline serum MPO levels correlated with worse outcome in tests assessing executive cognitive function in athletes but not in the control group (NAI maze test p0.05, Trail Making Test ratio p0.01). In control participants, subcortical white matter hyperintensities were associated with higher scores on the Geriatric Depression Scale (p0.05), whereas athletes seem to be protected from this effect. During strenuous exercising, MPO as well as its educts may be elevated due to increased oxygen intake and excretion of pro-inflammatory mediators inducing host tissue damage via oxidative stress. This outweighs the potential benefits of physical activity on cognitive function.

Autoantibodies specific for malondialdehyde-modified LDL (MDA-LDL) represent potential biomarkers to predict cardiovascular risk. However, MDA-LDL is a high variability antigen with limited reproducibility. To identify peptide mimotopes of MDA-LDL, phage display libraries were screened with the MDA-LDL-specific IgM monoclonal Ab LRO4, and the specificity and antigenic properties of MDA mimotopes were assessed in vitro and in vivo. We identified one 12-mer linear (P1) and one 7-mer cyclic (P2) peptide carrying a consensus sequence, which bound specifically to murine and human anti-MDA monoclonal Abs. Furthermore, MDA mimotopes were found to mimic MDA epitopes on the surface of apoptotic cells. Immunization of mice with P2 resulted in the induction of MDA-LDL-specific Abs, which strongly immunostained human atherosclerotic lesions. We detected IgG and IgM autoAbs to both MDA mimotopes in sera of healthy subjects and patients with myocardial infarction and stable angina pectoris undergoing percutaneous coronary intervention, and the titers of autoAbs correlated significantly with respective Ab titers against MDA-LDL. In conclusion, we identified specific peptides that are immunological mimotopes of MDA. These mimotopes can serve as standardized and reproducible antigens that will be useful for diagnostic and therapeutic applications in cardiovascular disease.

BACKGROUND Low serum butyrylcholinesterase activity was associated with all-cause and cardiovascular mortality in a community-based study; however, there are no data from investigations of the long-term effects of butyrylcholinesterase on mortality in patients with diagnosed coronary artery disease (CAD). We therefore assessed the effect of butyrylcholinesterase activity on the outcomes of patients with CAD. METHODS AND RESULTS We prospectively included 720 patients in our study: 293 patients with stable CAD and 427 patients with acute coronary syndrome. During a median follow-up of 11.3 years corresponding to 6469 overall person-years, 278 deaths (38.6%) were recorded. We detected a significant and independent protective effect of butyrylcholinesterase on all-cause mortality [adjusted hazard ratio (HR) for a 1-SD increase, 0.62; 95% CI, 0.54–0.71; P < 0.001] and cardiovascular mortality (adjusted HR, 0.64; 95% CI, 0.54–0.76; P < 0.001) in a Cox proportional hazards regression analysis. The 10-year survival rates were 42%, 74%, and 87% in the first, second, and third tertiles of butyrylcholinesterase activity. The presentation of CAD affected the effect of butyrylcholinesterase on mortality (P for interaction = 0.012), with a stronger association found in patients with stable CAD (adjusted HR, 0.56; 95% CI, 0.45–0.70; P < 0.001). CONCLUSIONS Our study demonstrates a strong inverse association between butyrylcholinesterase activity and long-term outcome in patients with known CAD. Because butyrylcholinesterase added predictive information after adjustment for established cardiovascular risk factors, additional underlying pathophysiological mechanisms and the potential applicability of butyrylcholinesterase activity for secondary risk prediction needs to be addressed in future studies.

Accurate assessment of injury severity is critical for decision making related to the prevention, triage, and treatment of several injured patients. Early estimation of mortality risk of critically injured patients is mandatory for adequate therapeutic strategies. Current risk stratification relies on clinical diagnosis and scoring systems. In our study, we hypothesized whether a simple laboratory test, the CK/CK-MB ratio, could help improving risk prediction in severely traumatized patients.In a 9-year period, 328 nonselected trauma patients were included in our retrospective study at a Level I Trauma Center up to September 2002. Data for this study were obtained from our computerized trauma database, established in September 1992.In our study population, we could show a negative correlation between Injury Severity Score (ISS) and leukocytes. A positive correlation was detected for liver enzymes and CK-MB. The correlation between ISS and Na(+) was significant. No correlation between ISS, K(+), and Hb/Ht could be observed. Exitus was associated with ISS, alteration in thrombocytes, CK, CK-MB, CRP, Crea, and Na(+).In our study population, CK-MB levels showed a significant correlation with overall surveillance in polytraumatized patients. In our opinion, this might suggest that CK-MB levels could be taken as an indirect predictor for survival. Our findings need to be proven in further prospective clinical trials.

Eur J Clin Invest 2012; 42 (9): 921–926 Abstract Background Humoral mediators of inflammation, in particular the complement system, have been described to play an important role in atherogenesis. Previously, we found a single-nucleotide polymorphism (SNP) in the complement 5 gene (C5 rs17611, A>G) independently associated with stroke. Up to now, the impact of C5 rs17611 on the progression of atherosclerosis and cardiovascular outcome in patients with asymptomatic atherosclerosis was unclear. Materials and Methods We investigated C5 rs17611 in a cohort of 1065 consecutive patients with asymptomatic carotid atherosclerosis. All patients were prospectively followed for the progression of carotid atherosclerosis and the development of a first major cardiovascular event (MACE), respectively. Results Three hundred and thirty-seven patients (31·6%) experienced a MACE during a median follow-up of 3·0 years. The homozygous GG genotype of the C5 rs17611 was significantly associated with adverse cardiovascular outcome (adjusted HR: 1·36 [95% CI, 1·07–1·73]; P = 0·01). After stratification for sex, C5 rs17611 CC was found to be an independent risk factor for MACE in men (HR 1·50 [95% CI, 1·12–1·83]). No association of C5 rs17611 with progression of carotid stenosis, observed in 93 (8·7%) patients, was detectable. Performance of ELISA indicated a significant association of the C5 rs17611 variant with C5a plasma levels. Conclusion The C5 rs17611 GG genotype is associated with increased C5a plasma levels and represents a risk factor for adverse cardiovascular outcome in male patients with carotid atherosclerosis.

Systemic activation of hemostasis is frequently observed in cancer patients, even in the absence of thrombosis. Moreover, this activation has been implicated in tumor progression, angiogenesis and metastatic spread. Increased levels of D-dimer, which is a degradation product of cross-linked fibrin, indicate a global activation of hemostasis and fibrinolysis.In a prospective and observational cohort study, we assessed the prognostic value of D-dimer levels for overall survival and mortality risk in 1178 cancer patients included in the Vienna Cancer and Thrombosis Study (CATS). Patients were followed over 2 years at regular intervals until occurrence of symptomatic venous thromboembolism or death. D-dimer levels were measured with a quantitative D-dimer latex agglutination assayThe main solid tumors were malignancies of the lung (n=182), breast (n=157), lower gastrointestinal tract (n=133), pancreas (n=74), stomach (n=50), kidney (n=37), prostate (n=133), and brain (n=148); 201 of the patients had hematologic malignancies; 63 had other tumors. During a median follow-up of 731 days, 460 (39.0%) patients died. The overall survival probabilities for patients with D-dimer levels categorized into four groups based on the 1(st), 2(nd) and 3(rd) quartiles of the D-dimer distribution in the total study population were 88%, 82%, 66% and 53% after 1 year, and 78%, 66%, 50% and 30% after 2 years, respectively (P0.001). The univariate hazard ratio of D-dimer (per double increase) for mortality was 1.5 (95% confidence interval: 1.4-1.6, P0.001) and remained increased in multivariable analysis including tumor subgroups, age, sex and venous thromboembolism.High D-dimer levels were associated with poor overall survival and increased mortality risk in cancer patients.

Eur J Clin Invest 2012; 42 (5): 463–469 Abstract Background Myeloperoxidase (MPO) is involved in a multitude of inflammatory processes involving oxidative modification of soluble components and cellular surfaces. Thus, MPO plays a key role in promoting atherosclerosis via oxidative stress by modification of both high- and low-density lipoprotein and production of other bioactive molecules. A polymorphism (MPO 463G>A, rs2333227) results in different expression rates of MPO. We aimed to assess whether MPO could be of clinical use as a risk marker for vascular disease in a high-risk group. Material and methods Plasma MPO levels of 406 patients suffering from peripheral arterial disease (PAD) were measured on an Abbott Architect i2000sr and grouped into patients with high (>115 ng/mL) and low (< 115 ng/mL) MPO levels. Genotyping of rs2333227 was performed on an ABI TaqMan 7900HT RT-PCR thermocycler. Results The relative risk of major adverse cardiovascular events (MACE) for patients with high plasma MPO is 1·2 (95%CI: 1·038–1·377, P A is not an independent risk factor for MACE in patients suffering from PAD.

OBJECTIVE T cells and level of the cytokine interferon-γ (IFN-γ) are increased in adipose tissue in obesity. Hedgehog (Hh) signaling has been shown to potently inhibit white adipocyte differentiation. In light of recent findings in neurons that IFN-γ and Hh signaling cross-talk, we examined their potential interaction in the context of adipogenesis. RESEARCH DESIGN AND METHODS We used Hh reporter cells, cell lines, and primary adipocyte differentiation models to explore costimulation of IFN-γ and Hh signaling. Genetic dissection using Ifngr1−/− and Stat1−/− mouse embryonic fibroblasts, and ultimately, anti–IFN-γ neutralization and expression profiling in obese mice and humans, respectively, were used to place the findings into the in vivo context. RESULTS T-cell supernatants directly inhibited hedgehog signaling in reporter and 3T3-L1 cells. Intriguingly, using blocking antibodies, Ifngr1−/− and Stat1−/− cells, and simultaneous activation of Hh and IFN-γ signaling, we showed that IFN-γ directly suppresses Hh stimulation, thus rescuing adipogenesis. We confirmed our findings using primary mouse and primary human (pre)adipocytes. Importantly, robust opposing signals for Hh and T-cell pathways in obese human adipose expression profiles and IFN-γ depletion in mice identify the system as intact in adipose tissue in vivo. CONCLUSIONS These results identify a novel antagonistic cross-talk between IFN-γ and Hh signaling in white adipose tissue and demonstrate IFN-γ as a potent inhibitor of Hh signaling.

BACKGROUND AND PURPOSE Haem oxygenase 1 (HO-1) is an inducible protein that plays a major protective role in conditions such as ischaemia-reperfusion injury and inflammation. In this study, we have investigated the role of haem arginate (HA) in human male subjects in the modulation of HO-1 expression and its correlation with the GT length polymorphism (GTn) in the promoter of the HO-1 gene. EXPERIMENTAL APPROACH In a dose-escalation, randomized, placebo-controlled trial, seven healthy male subjects with a homozygous short (S/S) and eight with a long (L/L) GTn genotype received intravenous HA. HO-1 protein expression and mRNA levels in peripheral blood monocytes, bilirubin, haptoglobin, haemopexin and haem levels were analysed over a 48 h observation period. KEY RESULTS We found that the baseline mRNA levels of HO-1 were higher in L/L subjects, while protein levels were higher in S/S subjects. HA induced a dose-dependent increase in the baseline corrected area under the curve values of HO-1 mRNA and protein over 48 h. The response of HO-1 mRNA was more pronounced in L/L subjects but the protein level was similar across the groups. CONCLUSIONS AND IMPLICATION HA is an effective inducer of HO-1 in humans irrespective of the GTn genotype. The potential therapeutic application of HA needs to be evaluated in clinical trials.

Eur J Clin Invest 2011; 41 (4): 365–371 Abstract Background A single-nucleotide polymorphism (SNP) in the palladin gene (PALLD, rs7439293) has recently been reported to be associated with coronary heart disease (CHD) in two case–control studies as well as in a large population-based cohort (Atherosclerosis Risk in Communities study, ARIC). Its clinical relevance, however, has not been evaluated prospectively. We investigated whether the risk allele (A) of PALLD rs7439293 (G>A) is associated with the occurrence of future major cardiovascular events (MACE) in a cohort of patients with prevalent carotid atherosclerosis. Materials and methods A total of 1283 consecutive patients with neurologically asymptomatic carotid atherosclerosis were included in the study and prospectively followed for a median of 3·5 years (interquartile range 3–4 years). We analysed whether the risk allele is associated with progression of carotid atherosclerosis after a 6–9-month period as measured by duplex Doppler sonography. Patients were then followed for the occurrence of a first MACE, a composite of myocardial infarction, stroke, coronary revascularization and death. Results After a median of 7·5 months (interquartile range 6–9 months), progression of carotid stenosis was observed in 103 (8·1%) patients. Cardiovascular events occurred in 337 (30%) patients after a median follow-up of 3·5 years. The risk allele of PALLD was neither associated with progressive carotid atherosclerosis (P = 0·21) nor with MACE (P = 0·58). Adjusted hazard ratios for a first MACE in heterozygous and homozygous carriers were 0·83 (95% CI 0·58–1·18) and 0·94 (95% CI 0·65–1·35) compared to wild type, respectively. Conclusions The A-allele of PALLD rs7439293 was not associated with progressive carotid atherosclerosis as measured by duplex Doppler sonography nor did it represent a risk factor for adverse cardiovascular outcome among patients with prevalent carotid atherosclerosis.

Invasive, extravillous trophoblasts (EVT) of the human placenta are critically involved in successful pregnancy outcome since they remodel the uterine spiral arteries to increase blood flow and oxygen delivery to the placenta and the developing fetus. To gain more insights into their biological role different primary cell culture models are commonly utilised. However, access to early placental tissue may be limited and primary trophoblasts rapidly cease proliferation in vitro impairing genetic manipulation. Hence, trophoblastic cell lines have been widely used as surrogates to study EVT function. Although the cell lines share some molecular markers with their primary counterpart, it is unknown to what extent they recapitulate the invasive phenotype of EVT. Therefore, we here report the first thorough GeneChip analyses of SGHPL-5, HTR-8/SVneo, BeWo, JEG-3 and the novel ACH-3P trophoblast cells in comparison to previously analysed primary villous cytotrophoblasts (CTBs) and extravillous trophoblasts (EVTs). Analyses of approximately 14,000 commonly expressed genes revealed that EVTs most closely resemble CTBs with considerable differences to the group of choriocarcinoma cells (JEG-3, BeWo, ACH-3P) and the group of SV40 Large T Antigen-selected cell types (SGHPL-5, HTR-8/SVneo). Similarly, analyses of 912 genes discriminating EVT from CTB, or 370 EVT-specific genes did not unravel a particular cell line with close similarity to any of the primary cell types, although molecular signatures common to EVT and each group of cell lines could be identified. Considering the diversity of mRNA expression patterns it is suggested that molecular studies in trophoblast cell lines require verification of the critical steps in an appropriate primary model system.

Introduction Vitamin D deficiency, as reflected by low 25-hydroxyvitamin D blood levels (25D), is a prevalent correctable risk factor for death in most populations around the globe. The evidence ranges from numerous association studies and meta-analyses thereof, over Mendelian randomization studies, to randomized controlled trials (RCTs). However, most studies reported to date were performed in rather older populations and some of the largest association studies may have been confounded by increased vitamin D supplementation at old age, especially in women, and by the use of vitamin D2, which is fully measured by 25D immunoassays but biologically considerably less active. In addition, cause-specific mortalities and the impact of age on the 25D association with the risk of death have not been reported in detail, yet. Methods Data of all patients who had a 25D measurement at the Department of Laboratory Medicine, General Hospital of Vienna between 1991 and 2011 were retrieved and matched with the Austrian national register of deaths. First 3 years of mortality since 25D measurement were excluded in the analyses. Fine-Gray regression models adjusting for competing risks were used to estimate the survival time in dependence on 25D, adjusting for sex, age, year and month of blood draw. 25D was represented using a spline with 5 knots placed on the corresponding 1/6th quantiles. Age group (0– Results Data from 78,581 patients (mean age = 51.0 years, men 31.5%) were used for analyses. During 20 years (median = 10.5) of follow-up, 11877 deaths were observed. Among these patients, 25D ≤ 10 nmol/L had 2-3 fold increased risk of death ( Conclusions Our survival data from a large cohort, covering all age groups, from a population with minimal vitamin D supplementation at old age and negligible intake of vitamin D2, confirm a strong association of vitamin D deficiency (25D

Elevated coagulation factor VIII activity has been associated with increased risk for both venous and arterial thrombosis. The current study evaluated the influence of Factor VIII levels and interactions with gender on all cause mortality in a large Austrian cohort.During 1991 and 2003, 11203 individuals, first ever request for laboratory analyses of FVIII: C, ageor =18 years, were included in this study. The median observation period was 5 years covering a total of 46000 person-years. The death rate was 17.1%.Compared to individuals within the reference category (FVIII: C94%) hazard ratios gradually increased from 1.4 (95% CI: 1.1-1.8) in the 152-170% category (5th decile) to finally 4.4 (95% CI: 3.5-5.5) in the313% category (highest decile, all p0.05). The association between FVIII: C levels and mortality remained essentially unchanged when considering non-cancer mortality, all cause vascular mortality or mortality due to ischemic heart disease. Compared to males females with elevated FVIII: C had a worse outcome resulting in higher hazard ratios reaching 6.8 (95% CI: 4.6-9.9) within the highest decile compared to males (HR: 3.4 (95% CI: 2.6-4.5)).In our large patient cohort we might be able to demonstrate for the first time that FVIII: C plasma activity is strongly associated with all cause mortality. Additionally, FVIII: C appears to interact with gender. Especially in women FVIII: C might help identifying high-risk cohorts, which might benefit from individualized prevention strategies.

Background Outcome of type 2 diabetes care depends on the acceptance of self-responsibility by informed patients, as treatment goals will otherwise be missed. Aims and methods This pre/post-observational report describes the clinical outcome of type 2 diabetes care in patients with type 2 diabetes (N =930) admitted consecutively to a diabetes rehabilitation clinic (DRC) between June 2013, and June 2016, where they were exposed to standardized lifestyle modification with meals low in salt and rich in vegetables and fruits, totaling 1,200 to 1,600 kcal/d, and an add-on exercise load equivalent to 400–600 kcal/d. Results At admission, patients presented with multiple treatment modes, elevated HbA1c levels (7.6±1.5%, 60±16 mmol/mol), a high prevalence of co-morbidities dominated by obesity (79%), a low rate of influenza and pneumococcal immunization (

Over 1 billion people are estimated to be overweight, placing them at risk for diabetes, cardiovascular disease, and cancer. We performed a systems-level genetic dissection of adiposity regulation using genome-wide RNAi screening in adult Drosophila. As a follow-up, the resulting approximately 500 candidate obesity genes were functionally classified using muscle-, oenocyte-, fat-body-, and neuronal-specific knockdown in vivo and revealed hedgehog signaling as the top-scoring fat-body-specific pathway. To extrapolate these findings into mammals, we generated fat-specific hedgehog-activation mutant mice. Intriguingly, these mice displayed near total loss of white, but not brown, fat compartments. Mechanistically, activation of hedgehog signaling irreversibly blocked differentiation of white adipocytes through direct, coordinate modulation of early adipogenic factors. These findings identify a role for hedgehog signaling in white/brown adipocyte determination and link in vivo RNAi-based scanning of the Drosophila genome to regulation of adipocyte cell fate in mammals.

Moderate alcohol consumption is associated with increased insulin sensitivity and reduced cardiovascular risk. We hypothesized that this relates to a direct effect of alcohol and therefore investigated whether acute alcohol intake altered insulin sensitivity or endothelial function in patients with type 2 diabetes. In an open-label two period design, the effect of a single oral dose of 40 g of alcohol (168 ml 40% vodka) on an insulin-modified frequently sampled intravenous glucose tolerance test (FSIGT) and on endothelium-dependent (flow mediated, FMD) or endothelium-independent (glyceroltrinitrate (GTN)-induced) vasodilation of the brachial artery measured by ultrasound was studied. Experiments were carried out in twelve male patients with type 2 diabetes mellitus (64+/-6 years, body mass index 28.4+/-5.7 kg/m (2)). Baseline insulin sensitivity index (S (I)) was 1.10+/-0.34 min (-1).microU (-1).ml, baseline FMD was +4.1+/-3.0%, and GTN-induced vasodilation +7.4+/-2.3% from resting brachial artery diameter. Acute alcohol intake increased alcohol plasma levels to 0.33+/-0.04 per thousand, S (I) to 1.86+/-0.45 min (-1).microU (-1).ml (p

Recent developments in proteomics technology offer new opportunities for clinical applications in hospital or specialized laboratories including the identification of novel biomarkers, monitoring of disease, detecting adverse effects of drugs, and environmental hazards. Advanced spectrometry technologies and the development of new protein array formats have brought these analyses to a standard, which now has the potential to be used in clinical diagnostics. Besides standardization of methodologies and distribution of proteomic data into public databases, the nature of the human body fluid proteome with its high dynamic range in protein concentrations, its quantitation problems, and its extreme complexity present enormous challenges. Molecular cell biology (cytomics) with its link to proteomics is a new fast moving scientific field, which addresses functional cell analysis and bioinformatic approaches to search for novel cellular proteomic biomarkers or their release products into body fluids that provide better insight into the enormous biocomplexity of disease processes and are suitable for patient stratification, therapeutic monitoring, and prediction of prognosis. Experience from studies of in vitro diagnostics and especially in clinical chemistry showed that the majority of errors occurs in the preanalytical phase and the setup of the diagnostic strategy. This is also true for clinical proteomics where similar preanalytical variables such as inter- and intra-assay variability due to biological variations or proteolytical activities in the sample will most likely also influence the results of proteomics studies. However, before complex proteomic analysis can be introduced at a broader level into the clinic, standardization of the preanalytical phase including patient preparation, sample collection, sample preparation, sample storage, measurement, and data analysis is another issue which has to be improved. In this report, we discuss the recent advances and applications that fulfill the criteria for clinical proteomics with the focus on cellular proteomics (cytoproteomics) as related to preanalytical and analytical standardization and to quality control measures required for effective implementation of these technologies and analytes into routine laboratory testing to generate novel actionable health information. It will then be crucial to design and carry out clinical studies that can eventually identify novel clinical diagnostic strategies based on these techniques and validate their impact on clinical decision making.

Purpose Venous thromboembolism (VTE) is a well-recognized complication of cancer. Laboratory parameters might be useful to assess the VTE risk in patients with cancer. The aim of this study was to investigate D-dimer and prothrombin fragment 1 + 2 (F 1 + 2), which reflect activation of blood coagulation and fibrinolysis, for prediction of cancer-associated VTE. Patients and Methods In a prospective, observational, cohort study of 821 patients with newly diagnosed cancer or progression of disease who did not recently receive chemotherapy, radiotherapy, or surgery were enrolled and followed for a median of 501 days (interquartile range, 255 to 731 days). The malignancies in these patients were as follows: breast (n = 132), lung (n = 119), stomach (n = 35), lower gastrointestinal tract (n = 106), pancreas (n = 46), kidney (n = 22), and prostate (n = 101) cancers; high-grade glioma (n = 102); malignant lymphoma (n = 94); multiple myeloma (n = 17); and other tumor types (n = 47). The study end point was occurrence of objectively confirmed symptomatic or fatal VTE. Results VTE occurred in 62 patients (7.6%). The cutoff level for elevated D-dimer and elevated F 1 + 2 was set at the 75th percentile of the total study population. In multivariable analysis that included elevated D-dimer, elevated F 1 + 2, age, sex, surgery, chemotherapy, and radiotherapy, the hazard ratios (HRs) of VTE in patients with elevated D-dimer (HR, 1.8; 95% CI, 1.0 to 3.2; P = .048) and elevated F 1 + 2 (HR, 2.0; 95% CI, 1.2 to 3.6; P = .015) were statistically significantly increased. The cumulative probability of developing VTE after 6 months was highest in patients with both elevated D-dimer and elevated F 1 + 2 (15.2%) compared with patients with nonelevated D-dimer and nonelevated F 1 + 2 (5.0%; P < .001). Conclusion High D-dimer and F 1 + 2 levels independently predict occurrence of VTE in patients with cancer.