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Background & AimsCholangiocarcinoma (CCA), heterogeneous biliary tumors with dismal prognosis, lacks accurate early diagnostic methods, especially important for individuals at high-risk (i.e., primary sclerosing cholangitis (PSC)). Here, we searched for protein biomarkers in serum extracellular vesicles (EVs).MethodsEVs from patients with isolated PSC (n=45), concomitant PSC-CCA (n=44), PSC who developed CCA during follow-up (PSC to CCA; n=25), CCAs from non-PSC etiology (n=56), hepatocellular carcinoma (HCC; n=34) and healthy individuals (n=56) were characterized by mass-spectrometry. Diagnostic biomarkers for PSC-CCA, non-PSC CCA or CCAs regardless etiology (Pan-CCAs) were defined and validated by ELISA. Their expression was evaluated in CCA tumors at single-cell level. Prognostic EV-biomarkers for CCA were investigated.ResultsHigh-throughput proteomics of EVs identified diagnostic biomarkers for PSC-CCA, non-PSC CCA or Pan-CCA, and for the differential diagnosis of intrahepatic CCA and HCC, that were cross-validated by ELISA using total serum. Machine learning-based algorithms disclosed CRP/FIBRINOGEN/FRIL for the diagnosis of PSC-CCA (local disease (LD)) vs isolated PSC (AUC=0.947;OR=36.9), and combined with CA19-9, overpowers CA19-9 alone. CRP/PIGR/VWF allowed the diagnosis of LD non-PSC CCAs vs healthy individuals (AUC=0.992;OR=387.5). Noteworthy, CRP/FRIL accurately diagnosed LD Pan-CCA (AUC=0.941;OR=89.4). Levels of CRP/FIBRINOGEN/FRIL/PIGR showed predictive capacity for CCA development in PSC before clinical evidences of malignancy. Multi-organ transcriptomic analysis revealed that serum EV-biomarkers were mostly expressed in hepatobiliary tissues, and scRNA-seq and immunofluorescence analysis of CCA tumors showed their presence mainly in malignant cholangiocytes. Multivariable analysis unveiled EV-prognostic biomarkers, with COMP/GNAI2/CFAI and ACTN1/MYCT1/PF4V associated negatively or positively to patients’ survival, respectively.ConclusionsSerum EVs contain protein biomarkers for the prediction, early diagnosis and prognosis estimation of CCA detectable using total serum, representing a tumor cell-derived liquid biopsy tool for personalized medicine.Impact and ImplicationsThe accuracy of current imaging tests and circulating tumor biomarkers for cholangiocarcinoma (CCA) diagnosis is far from satisfactory. Most CCAs are considered sporadic, although up to 20% of patients with primary sclerosing cholangitis (PSC) develop CCA during their lifetime, constituting a major cause of PSC-related death. This international study has proposed protein-based and etiology-related logistic models with predictive, diagnostic or prognostic capacities by combining 2-4 circulating protein biomarkers, moving a step forward into personalized medicine. These novel liquid biopsy tools may allow the: i) easy and non-invasive diagnosis of sporadic CCAs, ii) identification of patients with PSC with higher risk for CCA development, iii) establishment of cost-effective surveillance programs for the early detection of CCA in high-risk populations (e.g., PSC), and iv) prognostic stratification of patients with CCA, which, altogether, may increase the number of cases eligible for potentially curative options or to receive more successful treatments, decreasing CCA-related mortality.

Zinc finger E-box binding homeobox 1 (ZEB1) is a transcription factor that promotes metastatic and stem cell features, which has been associated with poor prognosis in cholangiocarcinoma (CCA), a desmoplastic cancer enriched in cancer-associated fibroblasts (CAFs). We aimed to define ZEB1 regulatory functions in malignant and stromal compartments of CCA.Bioinformatic and immunohistochemical analyses were performed to determine correlations between ZEB1 and markers of progressiveness in human intrahepatic CCA (iCCA). Gain-of-function and loss-of-function models were generated in CCA cells and liver myofibroblasts as a model of CAFs. Conditioned media (CM) was used to unravel tumor-stroma interplay. In vivo experiments were performed using a xenograft CCA model. ZEB1 expression in tumor cells of human iCCA was associated with undifferentiated tumor and vascular invasion. In vitro, ZEB1 promoted epithelial-mesenchymal transition and stemness in tumor cells, leading to cell migration and spheroid formation. In vivo, ZEB1-overexpressing CCA cells formed larger tumors with more abundant stroma. Expression of cellular communication network factor 2 (CCN2, encoding connective tissue growth factor [CTGF]) was increased in tumor cells from ZEB1-overexpressing xenografts and correlated with ZEB1 expression in human tumors. In vitro, CM from ZEB1-overexpressing tumor cells or recombinant CTGF induced myofibroblast proliferation. ZEB1 was also expressed by CAFs in human CCA, and its expression correlated with CCN2 in myofibroblasts and CCA stroma. In mice, cotransplantation of CCA cells with ZEB1-depleted myofibroblasts reduced CCA progressiveness compared to CCA cells/ZEB1-expressing myofibroblasts. Furthermore, ZEB1 controls the expression of paracrine signals (i.e., HGF and IL6) in tumor cells and myofibroblasts.ZEB1 plays a key role in CCA progression by regulating tumor cell-CAF crosstalk, leading to tumor dedifferentiation and CAF activation.

Contains fulltext : 225281.pdf (Publisher’s version ) (Closed access) Contains fulltext : 225281pos.pdf (Author’s version postprint ) (Open Access) BACKGROUND & AIMS: Polycystic liver diseases (PLDs) are genetic disorders characterized by progressive development of multiple biliary cysts. Recently, novel PLD-causative genes, encoding for endoplasmic reticulum (ER)-resident proteins involved in protein biogenesis and transport, were identified. We hypothesized that aberrant proteostasis contributes to PLD pathogenesis, representing a potential therapeutic target. METHODS: ER stress was analysed at transcriptional (qPCR), proteomic (mass spectrometry), morphological (transmission electron microscopy, TEM) and functional (proteasome activity) levels in different PLD models. The effect of ER stress inhibitors [4-phenylbutyric acid (4-PBA)] and/or activators [tunicamycin (TM)] was tested in polycystic (PCK) rats and cystic cholangiocytes in vitro. RESULTS: The expression levels of unfolded protein response (UPR) components were upregulated in liver tissue from PLD patients and PCK rats, as well as in primary cultures of human and rat cystic cholangiocytes, compared to normal controls. Cystic cholangiocytes showed altered proteomic profiles, mainly related to proteostasis (ie synthesis, folding, trafficking and degradation of proteins), marked enlargement of the ER lumen (by TEM) and hyperactivation of the proteasome. Notably, chronic treatment of PCK rats with 4-PBA decreased liver weight, as well as both liver and cystic volumes, of animals under baseline conditions or after TM administration compared to controls. In vitro, 4-PBA downregulated the expression (mRNA) of UPR effectors, normalized proteomic profiles related to protein synthesis, folding, trafficking and degradation and reduced the proteasome hyperactivity in cystic cholangiocytes, reducing their hyperproliferation and apoptosis. CONCLUSIONS: Restoration of proteostasis in cystic cholangiocytes with 4-PBA halts hepatic cystogenesis, emerging as a novel therapeutic strategy.

The Epidermal Growth Factor Receptor (EGFR) has been targeted through the development of selective tyrosine kinase inhibitors (TKIs) and monoclonal antibodies (mAb). These molecules have shown effectiveness in a subset of patients with specific genetic alterations (i.e. gain-of-function EGFR mutations or EGFR gene amplification) and have been approved for their use in non-small-cell lung cancer (NSCLC), colorectal cancer (CRC), pancreatic cancer and head and neck cancer. In addition, extensive research is being performed in many other tumour types hoping for a future approval. However, the majority of the patients show no benefit from these molecules due to primary mechanisms of resistance, already present before treatment or show disease progression upon the acquisition of drug resistance mechanisms during the treatment. At present, the majority of patients display resistance due to alterations in genes related to the EGFR signalling pathway that eventually circumvent EGFR inhibition and allow cancer progression. Thus, in this review article we focus on the molecular mechanisms underlying drug resistance via genetic alterations leading to resistance to all anti-EGFR drugs approved by the FDA and/or EMA. We also discuss novel approaches to surmount these chemoresistance modalities.

One of the main objectives of Industry 4.0 (I4.0) is to generate new opportunities based on the convergence of traditionally isolated technologies such as industrial control systems (ICSs) and information and communication technology (ICT). This presents new opportunities to take advantage of ICT technologies to develop new applications and services related to industrial processes. However, there are a variety of requirements and constraints that must be addressed for the attainment of this purpose. Moreover, the large amount of existing technologies and tools that can cope with these requirements makes the definition and selection of a solution a cumbersome task for traditional industrial workers with a non-ICT focused background. This chapter analyses and describes the main requirements and technologies required to provide a data-based Industry 4.0 solution.

In flexible manufacturing systems, fast feedback from simulation solutions is required for effective tool path planning and parameter optimization. In the particular sub-domain of laser heating/cutting of thin rectangular plates, current state-of-the-art methods include frequency-domain (spectral) analytic solutions that greatly reduce the required computational time in comparison to industry standard finite element based approaches. However, these spectral solutions have not been presented previously in terms of Fourier methods and Fast Fourier Transform (FFT) implementations. This manuscript presents four different schemes that translate the problem of laser heating of rectangular plates into equivalent FFT problems. The presented schemes make use of the FFT algorithm to reduce the computational time complexity of the problem from O ( M 2 N 2 ) to O ( M N log ( M N ) ) (with M &times, N being the discretization size of the plate). The test results show that the implemented schemes outperform previous non-FFT approaches both in CPU and GPU hardware, resulting in 100 &times, faster runs. Future work addresses thermal/stress analysis, non-rectangular geometries and non-linear interactions (such as material melting/ablation, convection and radiation heat transfer).

Cholangiopathies encompass a heterogeneous group of disorders affecting biliary epithelial cells (i.e. cholangiocytes). Early diagnosis, prognosis and treatment still remain clinically challenging for most of these diseases and are critical for adequate patient care. In the past decade, extensive research has emphasized microRNAs (miRs) as potential non-invasive biomarkers and tools to accurately identify, predict and treat cholangiopathies. MiRs can be released extracellularly conjugated with lipoproteins or encapsulated in extracellular vesicles (EVs). Research on EVs is also gaining attention since they are present in multiple biological fluids and may represent a relevant source of novel non-invasive biomarkers and be vehicles for new therapeutic approaches. This review highlights the most promising candidate miRs and EV-related biomarkers in cholangiopathies, as well as their relevant roles in biliary pathophysiology. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen. Research strategy PubMed search (April 2017) was done with the following terms: “microRNA”, “miRNA”, “miR”, “extracellular vesicles”, “EV”, “exosomes”, “primary biliary cholangitis”, “primary biliary cholangitis”, “PBC”, “primary sclerosing cholangitis”, “PSC”, “cholangiocarcinoma”, “CCA”, “biliary atresia”, “BA”, “polycystic liver diseases”, “PLD”, “cholangiopathies”, “cholestatic liver disease”. Most significant articles in full-text English were selected. The reference lists of selected papers were also considered.

Background and aims Cholangiocarcinoma (CCA) is an aggressive tumor type affecting cholangiocytes. CCAs frequently arise under certain cholestatic liver conditions. Intrahepatic accumulation of bile acids may facilitate cocarcinogenic effects by triggering an inflammatory response and cholangiocyte proliferation. Here, the role of bile acid receptors FXR and TGR5 in CCA progression was evaluated. Methods FXR and TGR5 expression was determined in human CCA tissues and cell lines. An orthotopic model of CCA was established in immunodeficient mice and tumor volume was monitored by magnetic resonance imaging under chronic administration of the specific FXR or TGR5 agonists, obeticholic acid (OCA) or INT-777 (0,03% in chow; Intercept Pharmaceuticals), respectively. Functional effects of FXR or TGR5 activation were evaluated on CCA cells in vitro. Results FXR was downregulated whereas TGR5 was upregulated in human CCA tissues compared to surrounding normal liver tissue. FXR expression correlated with tumor differentiation and TGR5 correlated with perineural invasion. TGR5 expression was higher in perihilar than in intrahepatic CCAs. In vitro, FXR was downregulated and TGR5 was upregulated in human CCA cells compared to normal human cholangiocytes. OCA halted CCA growth in vivo, whereas INT-777 showed no effect. In vitro, OCA inhibited CCA cell proliferation and migration which was associated with decreased mitochondrial energy metabolism. INT-777, by contrast, stimulated CCA cell proliferation and migration, linked to increased mitochondrial energy metabolism. Conclusion Activation of FXR inhibits, whereas TGR5 activation may promote, CCA progression by regulating proliferation, migration and mitochondrial energy metabolism. Modulation of FXR or TGR5 activities may represent potential therapeutic strategies for CCA.

Cholangiocarcinoma (CCA) is a heterogeneous group of dysplastic disorders affecting the biliary epithelium. It is the second most common primary liver tumor which accounts for around 3% of all gastrointestinal cancers. CCA is very deadly due to its aggressiveness, late diagnosis and high chemoresistance. The incidence is increasing worldwide and the therapeutic options are very limited. Radiotherapy, chemotherapy, surgery and/or liver transplantation may be indicated in patients who meet certain criteria, but chances of success are low. There is therefore increasing interest in understanding the molecular mechanisms involved in the pathogenesis of this cancer type and in identifying new targets for therapy. Current strategies are based on targeting key signaling pathways involved in proliferation, survival, apoptosis and migration. In this review, the most relevant molecular mechanisms involved in the pathogenesis of CCA are discussed and the main preclinical and clinical studies are highlighted. Moreover, future directions in basic and clinical research are indicated.

In the context of numerical methods, the problem of frequency-domain (spectral) simulations is crucial for the solution of Partial Differential Equations. Fast Fourier Transform (FFT) algorithms significantly reduce the computational cost of such simulations and enable parallelization using Graphics Processing Units (GPUs). In the particular subdomain of laser heating/cutting of rectangular metal plates, fast simulation is required for tool path planning, parameter optimization and additive manufacturing. The currently used methods include frequency-domain analytic solutions for single-beam and multi-beam laser heating. However, the problem of formulating these spectral problems in terms of Fourier methods and implementing them in efficient manner remains. To overcome these limitations, this article presents two different schemes that translate the problem of laser beam heating of metal plates into equivalent FFT problems. The results show significant improvements in terms of executions times, being 100× faster than current state-of-the-art algorithms. Future work needed involves the inclusion of stress analysis, complex plate geometries and non-constant material properties for the plate.

Through the last decade, cold atmospheric plasma (CAP) has emerged as an innovative therapeutic option for cancer treatment. Recently, we have set up a potentially safe atmospheric pressure plasma jet device that displays antitumoral properties in a preclinical model of cholangiocarcinoma (CCA), a rare and very aggressive cancer emerging from the biliary tree with few efficient treatments. In the present study, we aimed at deciphering the molecular mechanisms underlying the antitumor effects of CAP towards CCA in both an in vivo and in vitro context. In vivo, using subcutaneous xenografts into immunocompromised mice, CAP treatment of CCA induced DNA lesions and tumor cell apoptosis, as evaluated by 8-oxoguanine and cleaved caspase-3 immunohistochemistry, respectively. The analysis of the tumor microenvironment showed changes in markers related to macrophage polarization. In vitro, the incubation of CCA cells with CAP-treated culture media (i.e., plasma-activated media, PAM) led to a dose response decrease in cell survival. At molecular level, CAP treatment induced double-strand DNA breaks, followed by an increased phosphorylation and activation of the cell cycle master regulators CHK1 and p53, leading to cell cycle arrest and cell death by apoptosis. In conclusion, CAP is a novel therapeutic option to consider for CCA in the future.

Primary hepatobiliary malignancies include a heterogeneous group of cancers with dismal prognosis, among which hepatocellular carcinoma (HCC), cholangiocarcinoma (CCA), and hepatoblastoma (HB) stand out. These tumors mainly arise from the malignant transformation of hepatocytes, cholangiocytes (bile duct epithelial cells) or hepatoblasts (embryonic liver progenitor cells), respectively. Early diagnosis, prognosis prediction and effective therapies are still a utopia for these diseases. Extracellular vesicles (EVs) are small membrane-enclosed spheres secreted by cells and present in biological fluids. They contain multiple types of biomolecules, such as proteins, RNA, DNA, metabolites and lipids, which make them a potential source of biomarkers as well as regulators of human pathobiology. In this review, the role of EVs in the pathogenesis of hepatobiliary cancers and their potential usefulness as disease biomarkers are highlighted. Moreover, the therapeutic value of EV regulation is discussed and future directions on basic and clinical research are indicated. Spanish Ministries of Economy and Competitiveness [JB (FIS PI12/00380, FIS PI15/01132 and Miguel Servet Programme CON14/00129); MP (FIS PI14/00399, FIS PI17/00022) and Ramon y Cajal Programme RYC-2015-17755] cofinanced by Fondo Europeo de Desarrollo Regional (FEDER); ISCIII [CIBERehd: JB, LB, and MP], Spain; Diputacion Foral Gipuzkoa (JB: DFG15/010, DFG16/004), BIOEF (Basque Foundation for Innovation and Health Research: EiTB Maratoia BIO15/CA/016/BD to JB); Department of Health of the Basque Country (JB: 2013111173 and 2017111010; MP: 2015111100), and AECC Scientific Foundation (JB). AL and PO were funded by the Basque Government.

This work was supported by grants from NIH (US Department of Health and Human services)-R01AT001576 (S.C.L., J.M.M., and M.L.M.-C.), Ministerio de Economía, Industria y Competitividad: SAF2017-87301-R (M.L.M.-C.), Gobierno Vasco-Departamento de Salud 2013111114 (to M.L.M.-C.), BIOEF (Basque Foundation for Innovation and Health Research: EiTB Maratoia BIO15/CA/016/ BD (M.L.M.-C.), ELKARTEK 2016, Departamento de Industria del Gobierno Vasco (to M.L.M.-C.), Asociación Española contra el Cáncer (to T.C.D, P.F.-T. and M.L.M.-C.), Mitotherapeutix (to M.L.M.-C.), Daniel Alagille award from EASL (to T.C.D), Fundación Científica de la Asociación Española Contra el Cancer (AECC Scientific Foundation) Rare Tumor Calls 2017 (to M.L.M, J.M.B., and M.A), La Caixa Foundation Program (to M.L.M, J.M.B.), FIS: PI15/00892 (to M.F.F), FIS: PI15/01132 and FIS PI18/01075 (to J.M.B.), (to J.M.B.), FIS: PI14/00399 and PI17/ 00022 (to M.J.P.), Miguel Servet Program (CON14/00129 to J.M.B.), Ramon y Cajal Program (RYC-2015-17755 to M.J.P.), GRUPIN14-052 (to M.F.), PI14/00962 Instituto Carlos III (to J.B.), AECC Scientific Foundation (to J.B.), World Wide Cancer Research (to J.B.), Programma di Ricerca Regione-Università 2007-2009 and 2011-2012, Regione Emilia-Romagna (to E.V.), “Diputación Foral Gipuzkoa” (DFG15/010, DFG16/004 to J.M.B.), BIOEF (Basque Foundation for Innovation and Health Research: EiTB Maratoia BIO15/CA/016/BD to J.M.B.), Department of Health of the Basque Country (2013111173 to J.M.B.; 2015111100 to M.J.P.). Ciberehd_ISCIII_MINECO is funded by the Instituto de Salud Carlos III. “Hepacare Project” from Fundación La Caixa, Spain (to M.A.A and C.B.), BiO-Eusko Fundazioa project BIO15/CA/011 (to M.A.A and C.B.). We thank this work produced with the support of a 2017 Leonardo Grant for Researchers and Cultural Creators, BBVA Foundation (to M.V.R.). We thank MINECO for the Severo Ochoa Excellence Accreditation (SEV-2016-0644)., Fernández-Ramos, D., Fernández-Tussy, P., Lopitz-Otsoa, F., Gutiérrez-de-Juan, V., Navasa, N., Barbier-Torres, L., Zubiete-Franco, I., Simón, J., Fernández, A.F., Arbelaiz, A., Aransay, A.M., Lavín, J.L., Beraza, N., Perugorria, M.J., Banales, J.M., Villa, E., Fraga, M.F., Anguita, J., Avila, M.A., Berasain, C., Iruzibieta, P., Crespo, J., Lu, S.C., Varela-Rey, M., Mato, J.M., Delgado, T.C., Martínez-Chantar, M.L.

Virtual Reality training for fire fighters and managers has two main advantages. On one hand, it supports the simulation of complex scenarios like big cities, where a fire cannot be simulated in the real world. On the other hand, fire fighting VR simulators allow trainees to experience situations as similar as possible to real fire, reducing the probability of accidents when they are practising exercises with real fire. The success of the Virtual Reality training tools also depends on how close to reality the simulation process is. This work provides fire spread algorithms for forest and urban environments, which can be used at interactive rates. Due to the interactive nature of the algorithms, the users are able to fight the fire by throwing extinguishing agents. Although the algorithms assume many simplifications of the problem, their behaviour is satisfactory. This is due to the efficient management of the cell states in a 3 m×3 m cell grid. Also the variables that have more influence on fire propagation constitute the core of the algorithms. The overall system deals with user extinguishment actions, natural and artificial firebreaks, variable wind conditions (even at a cell level) and non-contiguous fire propagation (embers and spotting fires). The unified forest/urban model leads to an object oriented architecture which supports the fire propagation algorithms. This also allows the system to compute efficiently mixed forest–urban environments.

Objective Polycystic liver diseases (PCLDs) are genetic disorders characterised by progressive bile duct dilatation and/or cyst development. Their pathogenesis is a consequence of hyperproliferation, hypersecretion and microRNA alterations in cholangiocytes. Here we evaluate the role of matrix metalloproteases (MMPs) in the hepatic cystogenesis of PCLDs. Design Metalloprotease activity was measured by microfluorimetric assays in normal and polycystic cholangiocyte cultures from humans and rats, and gene expression by real time quantitative PCR. The role of cytokines, oestrogens and growth factors present in the cystic fluid of PCLD patients was evaluated for MMP activity. The MMP inhibitor marimastat was examined for cystic expansion in vitro and in polycystic kidney (PCK) rats. Results Polycystic human and rat cholangiocytes displayed increased MMP activity, which was associated with increased mRNA levels of different MMPs. Interleukin (IL)-6 and IL-8, and 17β-oestradiol, all stimulated MMP activity in human cholangiocytes. The presence of antibodies against IL-6 and/or IL-8 receptor/s inhibited baseline MMP hyperactivity of polycystic human cholangiocytes but had no effect on normal human cholangiocytes. MMP-3 was overexpressed in cystic cholangiocytes from PCLD human and PCK rat livers by immunohistochemistry. Marimastat reduced MMP hyperactivity of polycystic human and rat cholangiocytes and blocked the cystic expansion of PCK cholangiocytes cultured in three-dimensions. Chronic treatment of 8-week-old PCK rats with marimastat inhibited hepatic cystogenesis and fibrosis. Conclusions PCLDs are associated with cholangiocyte MMP hyperactivity resulting from autocrine/paracrine stimulation by IL-6 and IL-8. Inhibition of this MMP hyperactivity with marimastat decreased hepatic cystogenesis in vitro and in an animal model of PCLD, offering a potential therapeutic tool.

Cholangiocarcinoma (CCA) includes a heterogeneous group of biliary cancers with poor prognosis. Several conditions, such as primary sclerosing cholangitis (PSC), are risk factors. Noninvasive differential diagnosis between intrahepatic CCA and hepatocellular carcinoma (HCC) is sometimes difficult. Accurate noninvasive biomarkers for PSC, CCA, and HCC are not available. In the search for novel biomarkers, serum extracellular vesicles (EV) were isolated from CCA (n = 43), PSC (n = 30), or HCC (n = 29) patients and healthy individuals (control, n = 32); and their protein content was characterized. By using nanoparticle tracking analysis, serum EV concentration was found to be higher in HCC than in all the other groups. Round morphology (by transmission electron microscopy), size (∼180 nm diameter by nanoparticle tracking analysis), and markers (clusters of differentiation 9, 63, and 81 by immunoblot) indicated that most serum EV were exosomes. Proteome profiles (by mass spectrometry) revealed multiple differentially expressed proteins among groups. Several of these proteins showed high diagnostic values with maximum area under the receiver operating characteristic curve of 0.878 for CCA versus control, 0.905 for CCA stage I-II versus control, 0.789 for PSC versus control, 0.806 for noncirhottic PSC versus control, 0.796 for CCA versus PSC, 0.956 for CCA stage I-II versus PSC, 0.904 for HCC versus control, and 0.894 for intrahepatic CCA versus HCC. Proteomic analysis of EV derived from CCA human cells in vitro revealed higher abundance of oncogenic proteins compared to EV released by normal human cholangiocytes. Orthotopic implant of CCA human cells in the liver of immunodeficient mice resulted in the release to serum of EV containing some similar human oncogenic proteins. Conclusion: Proteomic signatures found in serum EV of CCA, PSC, and HCC patients show potential usefulness as diagnostic tools. (Hepatology 2017;66:1125-1143).

Interactive multi-beam laser machining simulation is crucial in the context of tool path planning and optimization of laser machining parameters. Current simulation approaches for heat transfer analysis (1) rely on numerical Finite Element methods (or any of its variants), non-suitable for interactive applications, and (2) require the multiple laser beams to be completely synchronized in trajectories, parameters and time frames. To overcome this limitation, this manuscript presents an algorithm for interactive simulation of the transient temperature field on the sheet metal. Contrary to standard numerical methods, our algorithm is based on an analytic solution in the frequency domain, allowing arbitrary time/space discretizations without loss of precision and non-monotonic retrieval of the temperature history. In addition, the method allows complete asynchronous laser beams with independent trajectories, parameters and time frames. Our implementation in a GPU device allows simulations at interactive rates even for a large amount of simultaneous laser beams. The presented method is already integrated into an interactive simulation environment for sheet cutting. Ongoing work addresses thermal stress coupling and laser ablation.