Oncogenic pathway signatures predict the risk of progression and recurrence in well-differentiated pancreatic neuroendocrine tumors.

BACKGROUND: Pancreatic neuroendocrine tumors (pNETs) are genomically diverse tumors. The management of newly diagnosed well-differentiated pNETs is limited by a lack of sensitivity of existing biomarkers for prognostication. Our goal was to investigate the potential utility of genetic markers as a predictor of progression-free survival (PFS) and recurrence-free survival (RFS). METHODS: Whole-exome sequencing of resected well-differentiated, low and intermediate-grade (G1 and G2) pNETs and normal adjacent tissue from patients who underwent resection from 2005 to 2015 was performed. Genetic alterations were classified using pan-genomic and oncogenic pathway classifications. Additional samples with genetic and clinicopathologic data available were obtained from the publicly available International Cancer Genome Consortium (ICGC) database and included in the analysis. The prognostic relevance of these genomic signatures on PFS and RFS was analyzed. RESULTS: Thirty-one patients who underwent resection for pNET were identified. Genomic analysis of mutational, copy number, cytogenetic, and complex phenomena revealed similar patterns to prior studies of pNETs with relatively few somatic gene mutations but numerous instances of copy number changes. Analysis of genomic and clinicopathologic outcomes using the combined data from our study as well as the ICGC pNET cohort (n = 124 patients) revealed that the recurrent pattern of whole chromosome loss (RPCL) and metastatic disease were independently associated with disease progression. When evaluating patients with local disease at the time of resection, RPCL and alterations in the TGFß oncogenic pathway were independently associated with the risk of recurrence. CONCLUSIONS: Well-differentiated pNETs are genomically diverse tumors. Pathway signatures may be prognostic for predicting disease progression and recurrence.

Pituitary hypophysitis in granulomatosis with polyangiitis (GPA): a case series.

Granulomatosis with polyangiitis (GPA) rarely involves the pituitary gland. Pituitary involvement has been reported in ~ 1% of all cases of GPA. Most commonly, pituitary swelling and inflammation results in symptoms due to pituitary mass effect and arginine vasopressin deficiency. To date, there are no pituitary-specific treatment guidelines for this rare condition. We present three patients with GPA-related hypophysitis highlighting the spectrum of pituitary involvement. All three patients were successfully treated with immunosuppressive regimens that included rituximab (RTX). Following remission induction with high-dose glucocorticoids, patients received 6 monthly RTX for remission maintenance. RTX was well tolerated without significant side effects.

Can educational video resources improve learning when used to augment traditional teaching of clinical examination? A randomized control trial of novice medical students.

BACKGROUND: Good clinical examination skills can both increase the quality of patient care and reduce its cost. A previous study by our group demonstrated that face-to-face training is the gold standard for teaching these skills. It is unclear if high quality educational videos can augment this teaching. METHODS: Forty-two Medical Students naïve to large joint examination were recruited and block randomised to two groups. The control group had face-to-face teaching alone. The intervention group had their teaching augmented with a custom educational video accessed via a web portal. Participants were assessed on their examination of a large joint using a previously standardised assessment tool at baseline and 7 days post intervention. Assessors were blinded to intervention type. RESULTS: There was no significant difference in the mean baseline scores. Mean baseline scores were 3.35 (11.2%, SD = 2.2, SE = 0.49) for the face-to-face only group and 2.65 (8.8%, SD = 1.39, SE = 0.31) for the video adjunct group [p = 0.137]. There was a significant difference in the improvement in score after intervention between each group [p = 0.005]. The mean improvement in score was 15.42 (SD = 5.64, SE = 1.29) for the face-to-face only group and 20.68 (SD = 4.33,SE = 0.99) for the video adjunct group. CONCLUSION: When used as an adjunct to more traditional face-to-face teaching methods, a custom-made educational video significantly improves the teaching of clinical examination skills and there is a role for these resources in augmenting traditional teaching methods.

Robotic versus open pancreaticoduodenectomy in elderly patients: a propensity score-matched analysis.

BACKGROUND: Pancreaticoduodenectomy (PD) is complex procedure with high morbidity in the elderly. This retrospective study aimed to compare post-operative outcomes in patients ≥75 years of age who underwent robot-assisted (RA)PD and open PD. METHODS: We analyzed 2502 patients ≥75 years of age who underwent PD from 2015 to 2018 in the National Surgical Quality Improvement Program (NSQIP) database. RAPD and open PD patients were propensity score matched 1:5 to assess the 30-day outcomes of interest: postoperative complications, length of stay, discharge destination, and readmissions. RESULTS: Of 725 matched patients, 110 underwent RAPD, 615 OPD, and 12 were converted to an open operation. Post-operative outcomes were largely similar between cohorts. RAPD was associated a shorter length of stay (median 8 days, interquartile range [IQR] 6 to 11) than OPD (median 8 days, IQR 7 to 13) (p = 0.003). However, RAPD was associated with more readmissions (28.1% vs. 17.7%; p = 0.02). CONCLUSIONS: RAPD in patients ≥75 years of age appears to be safe and has a similar complication profile to open PD. Randomized or well-designed prospective matched studies are needed to confirm these findings.

Assessing tradeoffs in the design of climate change adaptation strategies for water utilities in Chile.

Climate change is a challenge to drinking water providers worldwide and to regulatory frameworks that consider long-term investment decisions. Coping with an unstable climate warrants adjustments in regulations and new investments. The investment required to maintain a selected service level needs to balance the potential for high regret stranded assets with the political and socioeconomic consequences of not meeting water demands. In recent years, the City of Santiago in Chile has seen drought events associated with climate change, which could worsen in the future. Chile's drinking water regulatory framework does not account for uncertainty in infrastructure design to cope with the potential impacts of such events. This work presents an adaptation option design process that considers multiple plausible climate change-impacted future scenarios, accommodating both structural and nonstructural measures. In our Santiago case study adaptation measures include extensions to the existing Chilean water market and traditional structural alternatives (e.g., storage infrastructure); all are represented in a simulation model of the water utility. We evaluate and optimize packages of efficient adaptation measures for various climate scenarios. This allows comparing different portfolios of combined institutional and infrastructure interventions via a range of stakeholder measures and comparing their tradeoffs under different plausible climate-impacted hydrological scenarios. Results showed that water supply performance without climate change adaptation is worse under climate scenarios with lower water availability, which are likely to be associated with higher GHG emission scenarios such as RCP 8.5. The optimized portfolios implement various combinations of adaptation strategies to reduce the impacts of this poor performance. Considering the uncertainty on future climate scenarios, the use of nonstructural adaptation measures such as option contracts exhibits the advantage of providing water in critical periods while avoiding large investments such as building reservoirs or the purchase of permanent water rights, which could end up underused if favorable climate scenarios manifest.

Coupling Nanostructured Microchips with Covalent Chemistry Enables Purification of Sarcoma-Derived Extracellular Vesicles for Downstream Functional Studies.

Tumor-derived extracellular vesicles (EVs) play essential roles in intercellular communication during tumor growth and metastatic evolution. Currently, little is known about the possible roles of tumor-derived EVs in sarcoma because the lack of specific surface markers makes it technically challenging to purify sarcoma-derived EVs. In this study, a specific purification system is developed for Ewing sarcoma (ES)-derived EVs by coupling covalent chemistry-mediated EV capture/ release within a nanostructure-embedded microchip. The purification platform-ES-EV Click Chip-takes advantage of specific anti-LINGO-1 recognition and sensitive click chemistry-mediated EV capture, followed by disulfide cleavage-driven EV release. Since the device is capable of specific and efficient purification of intact ES EVs with high purity, ES-EV Click Chip is ideal for conducting downstream functional studies of ES EVs. Absolute quantification of the molecular hallmark of ES (i.e., EWS rearrangements) using reverse transcription Droplet Digital PCR enables specific quantification of ES EVs. The purified ES EVs can be internalized by recipient cells and transfer their mRNA cargoes, exhibiting their biological intactness and potential role as biological shuttles in intercellular communication.

Cancer Modeling-on-a-Chip with Future Artificial Intelligence Integration.

Cancer is one of the leading causes of death worldwide, despite the large efforts to improve the understanding of cancer biology and development of treatments. The attempts to improve cancer treatment are limited by the complexity of the local milieu in which cancer cells exist. The tumor microenvironment (TME) consists of a diverse population of tumor cells and stromal cells with immune constituents, microvasculature, extracellular matrix components, and gradients of oxygen, nutrients, and growth factors. The TME is not recapitulated in traditional models used in cancer investigation, limiting the translation of preliminary findings to clinical practice. Advances in 3D cell culture, tissue engineering, and microfluidics have led to the development of "cancer-on-a-chip" platforms that expand the ability to model the TME in vitro and allow for high-throughput analysis. The advances in the development of cancer-on-a-chip platforms, implications for drug development, challenges to leveraging this technology for improved cancer treatment, and future integration with artificial intelligence for improved predictive drug screening models are discussed.

Motion-corrected multiparametric renal arterial spin labelling at 3 T: reproducibility and effect of vasodilator challenge.

OBJECTIVES: We investigated the feasibility and reproducibility of free-breathing motion-corrected multiple inversion time (multi-TI) pulsed renal arterial spin labelling (PASL), with general kinetic model parametric mapping, to simultaneously quantify renal perfusion (RBF), bolus arrival time (BAT) and tissue T1. METHODS: In a study approved by the Health Research Authority, 12 healthy volunteers (mean age, 27.6 ± 18.5 years; 5 male) gave informed consent for renal imaging at 3 T using multi-TI ASL and conventional single-TI ASL. Glyceryl trinitrate (GTN) was used as a vasodilator challenge in six subjects. Flow-sensitive alternating inversion recovery (FAIR) preparation was used with background suppression and 3D-GRASE (gradient and spin echo) read-out, and images were motion-corrected. Parametric maps of RBF, BAT and T1 were derived for both kidneys. Agreement was assessed using Pearson correlation and Bland-Altman plots. RESULTS: Inter-study correlation of whole-kidney RBF was good for both single-TI (r2 = 0.90), and multi-TI ASL (r2 = 0.92). Single-TI ASL gave a higher estimate of whole-kidney RBF compared to multi-TI ASL (mean bias, 29.3 ml/min/100 g; p <0.001). Using multi-TI ASL, the median T1 of renal cortex was shorter than that of medulla (799.6 ms vs 807.1 ms, p = 0.01), and mean whole-kidney BAT was 269.7 ± 56.5 ms. GTN had an effect on systolic blood pressure (p < 0.05) but the change in RBF was not significant. CONCLUSIONS: Free-breathing multi-TI renal ASL is feasible and reproducible at 3 T, providing simultaneous measurement of renal perfusion, haemodynamic parameters and tissue characteristics at baseline and during pharmacological challenge. KEY POINTS: • Multiple inversion time arterial spin labelling (ASL) of the kidneys is feasible and reproducible at 3 T. • This approach allows simultaneous mapping of renal perfusion, bolus arrival time and tissue T 1 during free breathing. • This technique enables repeated measures of renal haemodynamic characteristics during pharmacological challenge.

Regulation of fluid reabsorption in rat or mouse proximal renal tubules by asymmetric dimethylarginine and dimethylarginine dimethylaminohydrolase 1.

Nitric oxide prevents hypertension yet enhances proximal tubule Na+ reabsorption. Nitric oxide synthase is inhibited by asymmetric dimethylarginine (ADMA) that is metabolized by dimethylarginine dimethylaminohydrolase (DDAH) whose type 1 isoform is expressed abundantly in the proximal tubule (PT). We hypothesize that ADMA metabolized by DDAH-1 inhibits fluid reabsorbtion (Jv) by the proximal tubule. S2 segments of the PT were microperfused between blocks in vivo to assess Jv in anesthetized rats. Compared with vehicle, microperfusion of ADMA or Nω-nitro-l-arginine methyl ester (l-NAME) in the proximal tubule reduced Jv dose dependently. At 10-4 mol/l both reduced Jv by ~40% (vehicle: 3.2 ± 0.7 vs. ADMA: 2.1 ± 0.5, P < 0.01 vs. l-NAME: 1.9 ± 0.4 nl·min-1·mm-1, P < 0.01; n = 10). Selective inhibition of DDAH-1 in rats with intravenous L-257 (60 mg/kg) given 2 h before and L-257 (10-5 mol/l) perfused in the proximal tubule for 5 min reduced Jv by 32 ± 4% (vehicle: 3.2 ± 0.5 vs. L-257: 2.2 ± 0.5 nl·min-1·mm-1; P < 0.01) and increased plasma ADMA by ≈50% (vehicle: 0.46 ± 0.03 vs. L-257: 0.67 ± 0.03 µmol/l, P < 0.0001) without changing plasma symmetric dimethylarginine. Compared with nontargeted control small-interference RNA, knock down of DDAH-1 in mice by 60% with targeted small-interference RNAs (siRNA) reduced Jv by 29 ± 5% (nontargeted siRNA: 2.8 ± 0.20 vs. DDAH-1 knockdown: 1.9 ± 0.31 nl·min-1·mm-1, P < 0.05). In conclusion, fluid reabsorption in the proximal tubule is reduced by tubular ADMA or by blocking its metabolism by DDAH-1. L-257 is a novel regulator of proximal tubule fluid reabsorption.

A novel multimarker assay for the phenotypic profiling of circulating tumor cells in hepatocellular carcinoma.

Current clinicopathologic staging systems and serum biomarkers poorly discriminate tumor biology in hepatocellular carcinoma (HCC), with high recurrence rates following curative-intent surgical resection and liver transplantation (LT). Identification of accurate biomarkers for improved prognostication and treatment selection is a critical unmet need. We sought to develop a novel "liquid-biopsy" assay capable of detecting HCC circulating tumor cells (CTCs) and characterizing phenotypic subpopulations with prognostic significance. Using HCC cell lines, a tissue microarray, and human blood samples, an antibody cocktail targeting the cell-surface markers asialoglycoprotein receptor (ASGPR), glypican-3, and epithelial cell adhesion molecule was optimized for HCC CTC capture using the NanoVelcro CTC Assay. The ability of HCC CTCs and vimentin (VIM)-positive CTCs (a subpopulation expressing an epithelial-to-mesenchymal phenotype) to accurately discriminate tumor stage, recurrence, progression, and overall survival (OS) was evaluated in a prospective study of 80 patients. Multimarker capture detected greater numbers of CTCs than any individual antibody alone for both cell line and patient samples (P < 0.001). HCC CTCs were identified in 59/61 (97%) patients, and HCC (median, 6 CTCs) and non-HCC patients (median, 1 CTC; area under the receiver operating characteristic curve [AUROC] = 0.92; P < 0.001; sensitivity = 84.2%; specificity = 88.5%) were accurately discriminated. VIM-positive CTCs accurately discriminated early-stage, LT eligible patients (median, 0 CTCs) from locally advanced/metastatic, LT ineligible patients (median, 6 CTCs; AUROC = 0.89; P = 0.001; sensitivity = 87.1%; specificity = 90.0%), and predicted OS for all patients (hazard ratio [HR], 2.21; P = 0.001), and faster recurrence after curative-intent surgical or locoregional therapy in potentially curable early-stage HCC (HR, 3.14; P = 0.002). In conclusion, we developed a novel multimarker CTC enrichment assay that detects HCC CTCs with high efficiency and accuracy. A phenotypic subpopulation of VIM-positive CTCs appears to signify the presence of aggressive underlying disease and occult metastases and may have important implications for treatment selection. Liver Transplantation 24 946-960 2018 AASLD.

NRF2 prevents hypertension, increased ADMA, microvascular oxidative stress, and dysfunction in mice with two weeks of ANG II infusion.

Nuclear factor erythyroid factor 2 (Nrf2) transcribes genes in cultured endothelial cells that reduce reactive oxygen species (ROS) and generate nitric oxide (NO) or metabolize asymmetric dimethylarginine (ADMA), which inhibits NO synthase (NOS). Therefore, we undertook a functional study to test the hypothesis that activation of Nrf2 by tert-butylhydroquinone (tBHQ) preserves microvascular endothelial function during oxidative stress. Wild-type CB57BL/6 (wt), Nrf2 wt (+/+), or knockout (-/-) mice received vehicle (Veh) or tBHQ (0.1%; activator of Nrf2) during 14-day infusions of ANG II (to induce oxidative stress) or sham. MAP was recorded by telemetry. Mesenteric resistance arterioles were studied on isometric myographs and vascular NO and ROS by fluorescence microscopy. ANG II increased the mean arterial pressure (112 ± 5 vs. 145 ± 5 mmHg; P < 0.01) and excretion of 8-isoprostane F2α (2.8 ± 0.3 vs. 3.8 ± 0.3 ng/mg creatinine; P < 0.05) at 12-14 days. However, 12 days of ANG II reduced endothelium-derived relaxation (27 ± 5 vs. 17 ± 3%; P < 0.01) and NO (0.38 ± 0.07 vs. 0.18 ± 0.03 units; P < 0.01) but increased microvascular remodeling, endothelium-derived contractions (7.5 ± 0.5 vs. 13.0 ± 1.7%; P < 0.01), superoxide (0.09 ± 0.03 vs. 0.29 ± 0.08 units; P < 0.05), and contractions to U-46,619 (87 ± 6 vs. 118 ± 3%; P < 0.05), and endothelin-1(89 ± 4 vs. 123 ± 12%; P < 0.05). tBHQ prevented all of these effects of ANG II at 12-14 days in Nrf2+/+ mice but not in Nrf2-/- mice. In conclusion, tBHQ activates Nrf2 to prevent microvascular endothelial dysfunction, remodeling, and contractility, and moderate ADMA and hypertension at 12-14 days of ANG II infusion, thereby preserving endothelial function and preventing hypertension.

Circulating Tumor Cells Predict Occult Metastatic Disease and Prognosis in Pancreatic Cancer.

BACKGROUND: Occult metastatic tumors, below imaging thresholds, are a limitation of staging systems that rely on cross-sectional imaging alone and are a cause of the routine understaging of pancreatic ductal adenocarcinomas (PDACs). We investigated circulating tumor cells (CTCs) as a preoperative predictor of occult metastatic disease and as a prognostic biomarker for PDAC patients. EXPERIMENTAL DESIGN: A total of 126 patients (100 with cancer, 26 with benign disease) were enrolled in our study and CTCs were identified and enumerated from 4 mL of venous blood using the microfluidic NanoVelcro assay. CTC enumeration was correlated with clinicopathologic variables and outcomes following both surgical and systemic therapies. RESULTS: CTCs were identified in 78% of PDAC patients and CTC counts correlated with increasing stage (ρ = 0.42, p < 0.001). Of the 53 patients taken for potentially curative surgery, 13 (24.5%) had occult metastatic disease intraoperatively. Patients with occult disease had significantly more CTCs than patients with local disease only (median 7 vs. 1 CTC, p < 0.0001). At a cut-off of three or more CTCs/4 mL, CTCs correctly identified patients with occult metastatic disease preoperatively (area under the receiver operating characteristic curve 0.82, 95% confidence interval (CI) 0.76-0.98, p < 0.0001). CTCs were a univariate predictor of recurrence-free survival following surgery [hazard ratio (HR) 2.36, 95% CI 1.17-4.78, p = 0.017], as well as an independent predictor of overall survival on multivariate analysis (HR 1.38, 95% CI 1.01-1.88, p = 0.040). CONCLUSIONS: CTCs show promise as a prognostic biomarker for PDAC patients at all stages of disease being treated both medically and surgically. Furthermore, CTCs demonstrate potential as a preoperative biomarker for identifying patients at high risk of occult metastatic disease.

Evidence for a protective role for the rs805305 single nucleotide polymorphism of dimethylarginine dimethylaminohydrolase 2 (DDAH2) in septic shock through the regulation of DDAH activity.

BACKGROUND: Dimethylarginine dimethylaminohydrolase 2 (DDAH2) regulates the synthesis of nitric oxide (NO) through the metabolism of the endogenous inhibitor of nitric oxide synthase, asymmetric dimethylarginine (ADMA). Pilot studies have associated the rs805305 SNP of DDAH2 with ADMA concentrations in sepsis. This study explored the impact of the rs805305 polymorphism on DDAH activity and outcome in septic shock. METHODS: We undertook a secondary analysis of data and samples collected during the Vasopressin versus noradrenaline as initial therapy in septic shock (VANISH) trial. Plasma and DNA samples isolated from 286 patients recruited into the VANISH trial were analysed. Concentrations of L-Arginine and the methylarginines ADMA and symmetric dimethylarginine (SDMA) were determined from plasma samples. Whole blood and buffy-coat samples were genotyped for polymorphisms of DDAH2. Clinical data collected during the study were used to explore the relationship between circulating methylarginines, genotype and outcome. RESULTS: Peak ADMA concentration over the study period was associated with a hazard ratio for death at 28 days of 3.3 (95% CI 2.0-5.4), p < 0.001. Reduced DDAH activity measured by an elevated ADMA:SDMA ratio was associated with a reduced risk of death in septic shock (p = 0.03). The rs805305 polymorphism of DDAH2 was associated with reduced DDAH activity (p = 0.004) and 28-day mortality (p = 0.02). Mean SOFA score and shock duration were also reduced in the less common G:G genotype compared to heterozygotes and C:C genotype patients (p = 0.04 and p = 0.02, respectively). CONCLUSIONS: Plasma ADMA is a biomarker of outcome in septic shock, and reduced DDAH activity is associated with a protective effect. The polymorphism rs805305 SNP is associated with reduced mortality, which is potentially mediated by reduced DDAH2 activity. TRIAL REGISTRATION: ISRCTN Registry, ISRCTN20769191 . Registered on 20 September 2012.

The role of trimethylamine N-oxide as a mediator of cardiovascular complications in chronic kidney disease.

Patients with chronic kidney disease (CKD) have an enhanced risk of cardiovascular (CV) morbidity and mortality when compared with age- and gender-matched individuals with normal kidney function. Trimethlyamine N-oxide (TMAO) is a gut-derived amine oxide that has been implicated in the causation of CV diseases. Plasma TMAO is cleared by the kidney, and TMAO levels are elevated in CKD. Experimental studies have identified pathogenic mechanisms by which TMAO may contribute to CV disease through dysregulation of lipid metabolism, enhanced macrophage foam cell formation, and platelet dysfunction. Safe and well-tolerated therapeutic interventions such as pre- and probiotics, which modify the gut microbiome, offer the opportunity for interventional studies. This review examines the pathogenicity of TMAO, its value as a biomarker, and its potential as a therapeutic target in the context of CKD.

Determination of hepatocellular carcinoma grade by needle biopsy is unreliable for liver transplant candidate selection.

The objective of this article is to evaluate the utility of preoperative needle biopsy (PNB) grading of hepatocellular carcinoma (HCC) as a biomarker for liver transplantation (LT) candidate selection. Given the prognostic significance of HCC tumor grade, PNB grading has been proposed as a biomarker for LT candidate selection. Clinicopathologic characteristics of HCC LT recipients (1989-2014) with a PNB were analyzed, and the concordance of PNB grade to explant grade and vascular invasion was assessed to determine whether incorporation of PNB grade to accepted transplant criteria improved candidate selection. Of 965 patients undergoing LT for HCC, 234 (24%) underwent PNB at a median of 280 days prior to transplant. Grade by PNB had poor concordance to final explant pathology (κ = 0.22; P = 0.003), and low sensitivity (29%) and positive predictive value (35%) in identifying poorly differentiated tumors. Vascular invasion was predicted by explant pathologic grade (rs= 0.24; P < 0.001) but not PNB grade (rs = -0.05; P = 0.50). Increasing explant pathology grade (P = 0.02), but not PNB grade (P = 0.65), discriminated post-LT HCC recurrence risk. The incorporation of PNB grade to the established radiologic Milan criteria (MC) did not result in improved prognostication of post-LT recurrence (net reclassification index [NRI] = 0%), whereas grade by explant pathology resulted in significantly improved reclassification of risk (NRI = 19%). Preoperative determination of HCC grade by PNB has low concordance with explant pathologic grade and low sensitivity and positive predictive value in identifying poorly differentiated tumors. PNB grade did not accurately discriminate post-LT HCC recurrence and had no utility in improving prognostication compared with the MC alone. Incorporation of PNB to guide transplant candidate selection appears unjustified. Liver Transplantation 23 1123-1132 2017 AASLD.

Precision oncology using a limited number of cells: optimization of whole genome amplification products for sequencing applications.

BACKGROUND: Sequencing analysis of circulating tumor cells (CTCs) enables "liquid biopsy" to guide precision oncology strategies. However, this requires low-template whole genome amplification (WGA) that is prone to errors and biases from uneven amplifications. Currently, quality control (QC) methods for WGA products, as well as the number of CTCs needed for reliable downstream sequencing, remain poorly defined. We sought to define strategies for selecting and generating optimal WGA products from low-template input as it relates to their potential applications in precision oncology strategies. METHODS: Single pancreatic cancer cells (HPAF-II) were isolated using laser microdissection. WGA was performed using multiple displacement amplification (MDA), multiple annealing and looping based amplification (MALBAC) and PicoPLEX. Quality of amplified DNA products were assessed using a multiplex/RT-qPCR based method that evaluates for 8-cancer related genes and QC-scores were assigned. We utilized this scoring system to assess the impact of de novo modifications to the WGA protocol. WGA products were subjected to Sanger sequencing, array comparative genomic hybridization (aCGH) and next generation sequencing (NGS) to evaluate their performances in respective downstream analyses providing validation of the QC-score. RESULTS: Single-cell WGA products exhibited a significant sample-to-sample variability in amplified DNA quality as assessed by our 8-gene QC assay. Single-cell WGA products that passed the pre-analysis QC had lower amplification bias and improved aCGH/NGS performance metrics when compared to single-cell WGA products that failed the QC. Increasing the number of cellular input resulted in improved QC-scores overall, but a resultant WGA product that consistently passed the QC step required a starting cellular input of at least 20-cells. Our modified-WGA protocol effectively reduced this number, achieving reproducible high-quality WGA products from ≥5-cells as a starting template. A starting cellular input of 5 to 10-cells amplified using the modified-WGA achieved aCGH and NGS results that closely matched that of unamplified, batch genomic DNA. CONCLUSION: The modified-WGA protocol coupled with the 8-gene QC serve as an effective strategy to enhance the quality of low-template WGA reactions. Furthermore, a threshold number of 5-10 cells are likely needed for a reliable WGA reaction and product with high fidelity to the original starting template.

Etelcalcetide, A Novel Calcimimetic, Prevents Vascular Calcification in A Rat Model of Renal Insufficiency with Secondary Hyperparathyroidism.

Etelcalcetide, a novel peptide agonist of the calcium-sensing receptor, prevents vascular calcification in a rat model of renal insufficiency with secondary hyperparathyroidism. Vascular calcification occurs frequently in patients with chronic kidney disease (CKD) and is a consequence of impaired mineral homeostasis and secondary hyperparathyroidism (SHPT). Etelcalcetide substantially lowers parathyroid hormone (PTH) and fibroblast growth factor-23 (FGF23) levels in SHPT patients on hemodialysis. This study compared the effects of etelcalcetide and paricalcitol on vascular calcification in rats with adenine-induced CKD and SHPT. Uremia and SHPT were induced in male Wistar rats fed a diet supplemented with 0.75% adenine for 4 weeks. Rats were injected with vehicle, etelcalcetide, or paricalcitol for 4 weeks from the beginning of adenine diet. Rats fed an adenine-free diet were included as nonuremic controls. Similar reductions in plasma PTH and parathyroid chief cell proliferation were observed in both etelcalcetide- and paricalcitol-treated rats. Serum calcium and phosphorus were significantly lower in etelcalcetide-treated uremic rats and was unchanged in paricalcitol-treated rats. Both serum FGF23 and aortic calcium content were significantly lower in etelcalcetide-treated uremic rats compared with either vehicle- or paricalcitol-treated uremic rats. The degree of aortic calcium content for etelcalcetide-treated rats was similar to that in nonuremic controls and corroborated findings of lack of histologic aortic mineralization in those groups. In conclusion, etelcalcetide and paricalcitol similarly attenuated progression of SHPT in an adenine rat model of CKD. However, etelcalcetide differentially prevented vascular calcification, at least in part, due to reductions in serum FGF23, calcium, and phosphorus levels.

"Take-home" box trainers are an effective alternative to virtual reality simulators.

BACKGROUND: Practice on virtual reality simulators (VRSs) has been shown to improve surgical performance. However, VRSs are expensive and usually housed in surgical skills centers that may be inaccessible at times convenient for surgical trainees to practice. Conversely, box trainers (BT) are inexpensive and can be used anywhere at anytime. This study assesses "take-home" BTs as an alternative to VRS. METHODS: After baseline assessments (two simulated laparoscopic cholecystectomies, one on a VRS and one on a BT), 25 surgical trainees were randomized to two groups. Trainees were asked to practice three basic laparoscopic tasks for 6 wk (BT group using a "take-home" box trainer; VR group using VRS in clinical skills centers). After the practice period, all performed two laparoscopic cholecystectomy, one on a VRS and one on a BT; (i.e., posttraining assessment). VRS provided metrics (total time [TT], number of movements instrument tip path length), and expert video assessment of cholecystectomy in a BT (Global Operative Assessment of Laparoscopic Skills [GOALS] score) were recorded. Performance during pretraining and posttraining assessment was compared. RESULTS: The BT group showed a significant improvement for all VRS metrics (P = 0.008) and the efficiency category of GOALS score (P = 0.03). Only TT improved in the VRS group, and none of the GOALS categories demonstrated a statistically significant improvement after training. Finally, the improvement in VRS metrics in the BT group was significantly greater than in the VR group (TT P = 0.005, number of movements P = 0.042, path length P = 0.031), although there were no differences in the GOALS scores between the groups. CONCLUSIONS: This study suggests that a basic "take-home" BT is a suitable alternative to VRS.

Gender as a risk factor for adverse intraoperative and postoperative outcomes of elective pancreatectomy.

BACKGROUND AND OBJECTIVES: Patient selection remains paramount when developing and adopting quality-based assessment and reimbursement models, and enhanced recovery protocols. Gender is a patient characteristic known before surgery which can inform risk stratification. Our aim was to evaluate the effect of gender on intraoperative blood transfusions, operative time, length of hospital stay, estimated blood loss (EBL) as well as postoperative surgical site infections (SSIs), and mortality. METHODS: Patients undergoing elective pancreatectomy from 2005 to 2013 were identified in the American College of Surgeons National Surgical Quality Improvement Program (ACS-NSQIP) and Northwestern institutional databases. Multivariable analyses were conducted to identify the association between gender and these outcomes. RESULTS: Analyses demonstrated that male gender was independently associated with blood transfusion (OR 1.23), operative time >6 hr (OR 1.76), length of stay greater than 11 days (OR 1.17), and all-type SSIs (OR 1.17), especially superficial SSIs (OR 1.15) and organ space SSIs (OR 1.18). Analysis of the institutional cohort found that male gender was independently associated with increased odds of EBL > 1 L for Whipple procedures (OR 2.85). CONCLUSIONS: Male gender is a significant predictor of increased operative time, length of stay, transfusions, EBL > 1L, as well as postoperative organ space surgical site infections in these patients. J. Surg. Oncol. 2017;115:131-136. © 2016 Wiley Periodicals, Inc.