Development, evaluation, and overview of standardized training phantoms for abdominal ultrasound-guided interventions.

PURPOSE: Ultrasound (US) represents the primary approach for abdominal diagnosis and is regularly used to guide diagnostic and therapeutic interventions (INVUS). Due to possible serious INVUS complications, structured training concepts are required. Phantoms can facilitate teaching, but their use is currently restricted by complex manufacturing and short durability of the materials. Hence, the aim of this study was the development and evaluation of an optimized abdominal INVUS phantom. MATERIALS AND METHODS: Phantom requirements were defined in a structured research process: Skin-like surface texture, homogeneous matrix with realistic tissue properties, implementation of lesions and abscess cavities in different sizes and depths as well as a modular production process allowing for customized layouts. The phantom prototypes were evaluated in certified ultrasound courses. RESULTS: In accordance with the defined specifications, a new type of matrix was developed and cast in multiple layers including different target materials. The phantom structure is based on features of liver anatomy and includes solid focal lesions, vessels, and abscess formations. For a realistic biopsy procedure, ultrasound-proof material was additionally included to imitate bone. The evaluation was performed by US novices (n=40) and experienced participants (n=41). The majority (73/81) confirmed realistic visualization of the lesions. The 3D impression was rated as "very good" in 64% of cases (52/81) and good in 31% (25/81). Overall, 86% (70/81) of the participants certified high clinical relevance of the phantom. CONCLUSION: The presented INVUS phantom concept allows standardized and realistic training for interventions.

Impact of Endothelial Lipase on Cholesterol Efflux Capacity of Serum and High-density Lipoprotein.

Endothelial lipase (EL) is a potent modulator of the structural and functional properties of HDL. Impact of EL on cholesterol efflux capacity (CEC) of serum and isolated HDL is not well understood and apparently contradictory data were published. Here, we systematically examined the impact of EL on composition and CEC of serum and isolated HDL, in vitro and in vivo, using EL-overexpressing cells and EL-overexpressing mice. CEC was examined in a validated assay using 3H-cholesterol labelled J774 macrophages. In vitro EL-modification of serum resulted in complex alterations, including enrichment of serum with lipid-free/-poor apoA-I, decreased size of human (but not mouse) HDL and altered HDL lipid composition. EL-modification of serum increased CEC, in line with increased lipid-free/-poor apoA-I formation. In contrast, CEC of isolated HDL was decreased likely through altered lipid composition. In contrast to in vitro results, EL-overexpression in mice markedly decreased HDL-cholesterol and apolipoprotein A-I serum levels associated with a decreased CEC of serum. HDL lipid composition was altered, but HDL particle size and CEC were not affected. Our study highlights the multiple and complex effects of EL on HDL composition and function and may help to clarify the seemingly contradictory data found in published articles.

HDL structure and function is profoundly affected when stored frozen in the absence of cryoprotectants.

Analysis of structural and functional parameters of HDL has gained significant momentum in recent years because they are stronger predictors of cardiovascular risk than HDL-cholesterol levels. Surprisingly, in most HDL studies, very low attention is paid to HDL storage, which might critically affect functional properties. In the present study, we systematically examined the impact of storage and freezing on the structural/functional properties of freshly isolated HDL. Initial damage to HDL starts between week 1 and week 4 of storage. We observed that prolonged freezing at -20°C or -70°C led to a shedding of apoA-I from HDL and to the formation of large protein-poor particles, indicating that HDL is irreversibly disrupted. These structural alterations profoundly affected key metrics of HDL function, including HDL-cholesterol efflux capacity and HDL paraoxonase activity. Flash-freezing of isolated HDL prior to storage at -70°C did not preserve HDL structure. However, addition of the cryoprotectants, sucrose or glycerol, completely preserved structure and function of HDL when stored for at least 2 years. Our data clearly indicate that HDL is a complex particle requiring special attention when stored. Addition of cryoprotectants to isolated HDL samples before storage will make biochemical and clinical HDL research studies more reproducible and comparable.

Adiponectin Predicts High-Density Lipoprotein Cholesterol Efflux Capacity in Adults Irrespective of Body Mass Index and Fat Distribution.

Context: Obesity is associated with hypoadiponectemia, dyslipidemia, and increased risk of cardiovascular disease (CVD). Mechanisms linking these conditions remain to be fully understood. Cholesterol efflux capacity (CEC) is a crucial functional property of high-density lipoprotein (HDL) that strongly predicts CVD incidence. Objective: We investigated whether age, fat distribution, and other obesity-related factors affect CEC in juvenile and adult overweight/obese participants of the STYJOBS/EDECTA cohort (NCT00482924). Design: We performed an observational study. Main Outcome Measures: CEC and its association with body measures and related metabolic parameters was assessed in 683 participants (281 juveniles, of whom 227 were overweight/obese; 402 adults, of whom 197 were overweight/obese). Results: Pearson correlation analysis showed that, after Bonferroni correction, CEC was significantly inversely correlated with body mass index (BMI), carotid diameter, waist circumference, waist-to-hip, waist-to-height ratio, oxidized low-density lipoprotein, and uric acid and with the liver markers alanine-aminotransferase and choline esterase. CEC was positively correlated with HDL cholesterol, total cholesterol, apolipoprotein A1, and adiponectin in adults, whereas in juveniles only apolipoprotein A1 showed a significant positive correlation with CEC. Age-stratified linear regression analyses with CEC as the outcome variable identified adiponectin as the most significant predictor of CEC in adults. The results did not change when either BMI or waist-to-hip ratio as a factor of fat distribution was included in the models. Conclusions: Hypoadiponectemia is a robust predictor of reduced cholesterol efflux capacity in adults irrespective of BMI and fat distribution. Further investigations are needed to assess whether adiponectin is a causal determinant of CEC.

Economic Evaluation of Pharmacologic Pre- and Postconditioning With Sevoflurane Compared With Total Intravenous Anesthesia in Liver Surgery: A Cost Analysis.

BACKGROUND: Pharmacologic pre- and postconditioning with sevoflurane compared with total IV anesthesia in patients undergoing liver surgery reduced complication rates as shown in 2 recent randomized controlled trials. However, the potential health economic consequences of these different anesthesia regimens have not yet been assessed. METHODS: An expostcost analysis of these 2 trials in 129 patients treated between 2006 and 2010 was performed. We analyzed direct medical costs for in-hospital stay and compared pharmacologic pre- and postconditioning with sevoflurane (intervention) with total IV anesthesia (control) from the perspective of a Swiss university hospital. Year 2015 costs, converted to US dollars, were derived from hospital cost accounting data and compared with a multivariable regression analysis adjusting for relevant covariables. Costs with negative prefix indicate savings and costs with positive prefix represent higher spending in our analysis. RESULTS: Treatment-related costs per patient showed a nonsignificant change by -12,697 US dollars (95% confidence interval [CI], 10,956 to -36,352; P = .29) with preconditioning and by -6139 US dollars (95% CI, 6723 to -19,000; P = .35) with postconditioning compared with the control group. Results were robust in our sensitivity analysis. For both procedures (control and intervention) together, major complications led to a significant increase in costs by 86,018 US dollars (95% CI, 13,839-158,198; P = .02) per patient compared with patients with no major complications. CONCLUSIONS: In this cost analysis, reduced in-hospital costs by pharmacologic conditioning with sevoflurane in patients undergoing liver surgery are suggested. This possible difference in costs compared with total IV anesthesia is the result of reduced complication rates with pharmacologic conditioning, because major complications have significant cost implications.

Refined purification strategy for reliable proteomic profiling of HDL2/3: Impact on proteomic complexity.

Proteomics have extended the list of high-density lipoprotein (HDL) associated proteins to about 90. One of the major issues of global protein characterization is establishing specificity of association as opposed to contamination, a fact which has never been addressed for isolated HDL. We have developed a refined purification strategy to isolate HDL by density, followed by purification by size to generate "highly purified" fractions of HDL2/3, which allow the reliable quantification of the HDL proteome for biomarker discovery. Mass spectrometry analysis revealed that the proteome of HDL2/3 is composed of 10-16 different proteins, which is in striking contrast to previous reports. Importantly, proteomic analysis revealed that many proteins which have recently been described to be associated with HDL, including α-1-antitrypsin, α-2-HS-glycoprotein, serotransferrin, apolipoprotein A-IV and others, are not associated with HDL2/3 and are exclusively found in a different molecular weight fraction containing human serum albumin, lipid-poor apolipoprotein A-I and other proteins. Interestingly, proteins found in this lower molecular weight fraction commonly share lipid-binding properties and enrichment of serum with free fatty acids/lysophophatidylcholine led to a significant increase in co-isolation of lipid-binding proteins such as albumin and α-1-antitrypsin. We propose that this refined method might become a standard in proteomic assessment of HDL2/3 making data from clinical cohorts more comparable and reproducible.

Detection of circulating tumor cells in patients with esophagogastric or pancreatic adenocarcinoma using the CellSearch® system: An observational feasibility study.

Circulating tumor cells (CTCs) in the blood of cancer patients have been demonstrated to be of prognostic value regarding metastasis and survival. The CellSearch® system has been certified for the detection of CTCs and as a prognostic tool in patients with metastatic breast, colon and prostate cancer. Few studies have evaluated the detection of CTCs originating from esophagogastric or pancreatic cancer with the CellSearch® system. In the present small pilot study, a total of 16 patients with either esophagogastric (n=8) or pancreatic (n=8) adenocarcinomas at various disease stages were randomly screened and included. A total of 7.5 ml of blood was drawn from each patient and analyzed for CTCs using the CellSearch® device. CTCs could be detected in 1 out of 8 patients (12.5%) with esophagogastric and in 7 out of 8 patients (87.5%) with pancreatic cancer. The preliminary data obtained from this observational feasibility study suggested that the CellSearch® system may become a valuable tool for the detection of CTCs in patients with pancreatic adenocarcinoma, whereas the usefulness in patients with early-stage esophagogastric adenocarcinoma may be limited. This study clearly points towards a requirement for larger studies focusing on patients with pancreatic adenocarcinoma at various disease stages and assessing CTCs, whereas patients with esophagogastric adenocarcinomas should be part of further pilot studies.

Correction: High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration.

[This corrects the article DOI: 10.1371/journal.pone.0154397.].

High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration.

PURPOSE: High-density lipoproteins (HDL) have long been implicated in the pathogenesis of age-related macular degeneration (AMD). However, conflicting results have been reported with regard to the associations of AMD with HDL-cholesterol levels. The present study is the first to assess HDL composition and metrics of HDL function in patients with exudative AMD and control patients. METHODS: Blood samples were collected from 29 patients with exudative AMD and 26 age-matched control patients. Major HDL associated apolipoproteins were determined in apoB-depleted serum by immunoturbidimetry or ELISA, HDL-associated lipids were quantified enzymatically. To get an integrated measure of HDL quantity and quality, we assessed several metrics of HDL function, including cholesterol efflux capacity, anti-oxidative and anti-inflammatory activities using apoB-depleted serum from study participants. RESULTS: In our study, we observed that the HDL associated acute phase protein serum amyloid A (SAA) was significantly increased in AMD patients (p<0.01), whereas all other assessed apolipoproteins including ApoA-I, apoA-II, apoC-II, apoC-III and apoE as well as major HDL associated lipids were not altered. HDL efflux capacity, anti-oxidative capacity and arylesterase activity were not different in AMD patients when compared with the control group. The ability of apoB-depleted serum to inhibit monocyte NF-κB expression was significantly improved in AMD patients (mean difference (MD) -5.6, p<0.01). Moreover, lipoprotein-associated phospholipase A2 activity, a marker of vascular inflammation, was decreased in AMD subjects (MD -24.1, p<0.01). CONCLUSIONS: The investigated metrics of HDL composition and HDL function were not associated with exudative AMD in this study, despite an increased content of HDL associated SAA in AMD patients. Unexpectedly, anti-inflammatory activity of apoB-depleted serum was even increased in our study. Our data suggest that the investigated parameters of serum HDL function showed no significant association with exudative AMD. However, we cannot exclude that alterations in locally produced HDL may be part of the AMD pathogenesis.

Biomarker discovery in heterogeneous tissue samples -taking the in-silico deconfounding approach.

BACKGROUND: For heterogeneous tissues, such as blood, measurements of gene expression are confounded by relative proportions of cell types involved. Conclusions have to rely on estimation of gene expression signals for homogeneous cell populations, e.g. by applying micro-dissection, fluorescence activated cell sorting, or in-silico deconfounding. We studied feasibility and validity of a non-negative matrix decomposition algorithm using experimental gene expression data for blood and sorted cells from the same donor samples. Our objective was to optimize the algorithm regarding detection of differentially expressed genes and to enable its use for classification in the difficult scenario of reversely regulated genes. This would be of importance for the identification of candidate biomarkers in heterogeneous tissues. RESULTS: Experimental data and simulation studies involving noise parameters estimated from these data revealed that for valid detection of differential gene expression, quantile normalization and use of non-log data are optimal. We demonstrate the feasibility of predicting proportions of constituting cell types from gene expression data of single samples, as a prerequisite for a deconfounding-based classification approach.Classification cross-validation errors with and without using deconfounding results are reported as well as sample-size dependencies. Implementation of the algorithm, simulation and analysis scripts are available. CONCLUSIONS: The deconfounding algorithm without decorrelation using quantile normalization on non-log data is proposed for biomarkers that are difficult to detect, and for cases where confounding by varying proportions of cell types is the suspected reason. In this case, a deconfounding ranking approach can be used as a powerful alternative to, or complement of, other statistical learning approaches to define candidate biomarkers for molecular diagnosis and prediction in biomedicine, in realistically noisy conditions and with moderate sample sizes.