Performance and clinical applicability of machine learning in liver computed tomography imaging: a systematic review.

OBJECTIVES: Machine learning (ML) for medical imaging is emerging for several organs and image modalities. Our objectives were to provide clinicians with an overview of this field by answering the following questions: (1) How is ML applied in liver computed tomography (CT) imaging? (2) How well do ML systems perform in liver CT imaging? (3) What are the clinical applications of ML in liver CT imaging? METHODS: A systematic review was carried out according to the guidelines from the PRISMA-P statement. The search string focused on studies containing content relating to artificial intelligence, liver, and computed tomography. RESULTS: One hundred ninety-one studies were included in the study. ML was applied to CT liver imaging by image analysis without clinicians' intervention in majority of studies while in newer studies the fusion of ML method with clinical intervention have been identified. Several were documented to perform very accurately on reliable but small data. Most models identified were deep learning-based, mainly using convolutional neural networks. Potentially many clinical applications of ML to CT liver imaging have been identified through our review including liver and its lesion segmentation and classification, segmentation of vascular structure inside the liver, fibrosis and cirrhosis staging, metastasis prediction, and evaluation of chemotherapy. CONCLUSION: Several studies attempted to provide transparent result of the model. To make the model convenient for a clinical application, prospective clinical validation studies are in urgent call. Computer scientists and engineers should seek to cooperate with health professionals to ensure this. KEY POINTS: • ML shows great potential for CT liver image tasks such as pixel-wise segmentation and classification of liver and liver lesions, fibrosis staging, metastasis prediction, and retrieval of relevant liver lesions from similar cases of other patients. • Despite presenting the result is not standardized, many studies have attempted to provide transparent results to interpret the machine learning method performance in the literature. • Prospective studies are in urgent call for clinical validation of ML method, preferably carried out by cooperation between clinicians and computer scientists.

Convergent Total Synthesis of Exochelin 772SM.

A convergent total synthesis of the natural mycobacterial iron chelator desferri-exochelin 772SM (D-EXO) is described. The synthetic procedure proceeds in 11 steps in the longest linear sequence, with an overall yield of 8.6%. The described procedure uses cheap starting materials and requires a limited number of chromatographic purifications. The concise strategy divides the exochelin into five key building blocks, allowing easy alternation of each single building block. Herein, the presented synthetic strategy is well suited to facilitate the synthesis of analogues and medicinal chemistry development efforts in a time- and resource-efficient manner.

Synthesis of sp3-rich heterocyclic frameworks by a divergent synthesis strategy.

Fragment-based lead discovery (FBLD) often relies on flat, aromatic compounds which display undesirable physicochemical properties with limited exit vectors for fragment growth. Herein, we report concise synthetic strategies to sp3-rich heterocyclic fragments encompassing polar exit vectors poised for fragment-to-lead (F2L) development.

Strategies for the Diversity-Oriented Synthesis of Macrocycles.

Macrocycles have long been recognized as useful chemical entities for medicine, with naturally occurring and synthetic macrocycles clinically approved for use as prescription drugs. Despite this promise, the synthesis of collections of macrocycles has been historically challenging due to difficulties in the formation of large rings. Diversity-Oriented Synthesis (DOS) emerged in the early 2000s as a powerful strategic solution to the construction of diverse molecular libraries. This review details the various strategies developed within the field of DOS for the synthesis of macrocycle libraries, utilizing modern synthetic methodology to deliver structurally diverse collections of macrocyclic molecules, and the exploration of their therapeutic potential. Section 1 of this work details the use of algorithmic strategies and is divided into Build/Couple/Pair, Advanced Build/Couple/Pair, Initiate/Propagate/Terminate, Fragment-Based Domain Shuffling, Two-Directional Synthesis, and Successive Ring Expansion. Section 2 covers strategies based on ring distortion reactions, including Sequential Cycloaddition/Fragmentation, Ring Expansions, and Miscellaneous.

Unhindered Brownian Motion of Individual Nanoparticles in Liquid-Phase Scanning Transmission Electron Microscopy.

Liquid-phase transmission electron microscopy (LPTEM) offers label-free imaging of nanoparticle (NP) processes in liquid with sub-nanometer spatial and millisecond temporal resolution. However, LPTEM studies have reported only on NPs moving orders of magnitude slower than expected from bulk aqueous liquid conditions, likely due to strong interactions with the LPTEM liquid-enclosing membranes. We demonstrate how scanning transmission electron microscope (STEM) imaging can be used to measure the motion of individual NPs and agglomerates, which are not hindered by such interactions. Only at low electron flux do we find that individual NPs exhibit Brownian motion consistent with optical control experiments and theoretical predictions for unhindered passive diffusive motion in bulk liquids. For increasing electron flux, we find increasingly faster than passive motion that still appears effectively Brownian. We discuss the possible origins of this beam-sample interaction. This establishes conditions for the use of STEM as a reliable tool for imaging nanoscale hydrodynamics in situ TEM.

Regulation of Plasma Membrane Nanodomains of the Water Channel Aquaporin-3 Revealed by Fixed and Live Photoactivated Localization Microscopy.

Several aquaporin (AQP) water channels are short-term regulated by the messenger cyclic adenosine monophosphate (cAMP), including AQP3. Bulk measurements show that cAMP can change diffusive properties of AQP3; however, it remains unknown how elevated cAMP affects AQP3 organization at the nanoscale. Here we analyzed AQP3 nano-organization following cAMP stimulation using photoactivated localization microscopy (PALM) of fixed cells combined with pair correlation analysis. Moreover, in live cells, we combined PALM acquisitions of single fluorophores with single-particle tracking (spt-PALM). These analyses revealed that AQP3 tends to cluster and that the diffusive mobility is confined to nanodomains with radii of ∼150 nm. This domain size increases by ∼30% upon elevation of cAMP, which, however, is not accompanied by a significant increase in the confined diffusion coefficient. This regulation of AQP3 organization at the nanoscale may be important for understanding the mechanisms of water AQP3-mediated water transport across plasma membranes.

The effects of terlipressin and direct portacaval shunting on liver hemodynamics following 80% hepatectomy in the pig.

Liver failure is the major cause of death following liver resection. Post-resection portal venous pressure (PVP) predicts liver failure, is implicated in its pathogenesis, and when PVP is reduced, rates of liver dysfunction decrease. The aim of the present study was to characterize the hemodynamic, biochemical, and histological changes induced by 80% hepatectomy in non-cirrhotic pigs and determine if terlipressin or direct portacaval shunting can modulate these effects. Pigs were randomized (n=8/group) to undergo 80% hepatectomy alone (control); terlipressin (2 mg bolus + 0.5-1 mg/h) + 80% hepatectomy; or portacaval shunt (PCS) + 80% hepatectomy, and were maintained under terminal anesthesia for 8 h. The primary outcome was changed in PVP. Secondary outcomes included portal venous flow (PVF), hepatic arterial flow (HAF), and biochemical and histological markers of liver injury. Hepatectomy increased PVP (9.3 ± 0.4 mmHg pre-hepatectomy compared with 13.0 ± 0.8 mmHg post-hepatectomy, P<0.0001) and PVF/g liver (1.2 ± 0.2 compared with 6.0 ± 0.6 ml/min/g, P<0.0001) and decreased HAF (70.8 ± 5.0 compared with 41.8 ± 5.7 ml/min, P=0.002). Terlipressin and PCS reduced PVP (terlipressin = 10.4 ± 0.8 mmHg, P=0.046 and PCS = 8.3 ± 1.2 mmHg, P=0.025) and PVF (control = 869.0 ± 36.1 ml/min compared with terlipressin = 565.6 ± 25.7 ml/min, P<0.0001 and PCS = 488.4 ± 106.4 ml/min, P=0.002) compared with control. Treatment with terlipressin increased HAF (73.2 ± 11.3 ml/min) compared with control (40.3 ± 6.3 ml/min, P=0.026). The results of the present study suggest that terlipressin and PCS may have a role in the prevention and treatment of post-resection liver failure.

Cycloaddition Strategies for the Synthesis of Diverse Heterocyclic Spirocycles for Fragment-Based Drug Discovery.

In recent years the pharmaceutical industry has benefited from the advances made in fragment-based drug discovery (FBDD) with more than 30 fragment-derived drugs currently marketed or progressing through clinical trials. The success of fragment-based drug discovery is entirely dependent upon the composition of the fragment screening libraries used. Heterocycles are prevalent within marketed drugs due to the role they play in providing binding interactions; consequently, heterocyclic fragments are important components of FBDD libraries. Current screening libraries are dominated by flat, sp2-rich compounds, primarily owing to their synthetic tractability, despite the superior physicochemical properties displayed by more three-dimensional scaffolds. Herein, we report step-efficient routes to a number of biologically relevant, fragment-like heterocyclic spirocycles. The use of both electron-deficient and electron-rich 2-atom donors was explored in complexity-generating [3+2]-cycloadditions to furnish products in 3 steps from commercially available starting materials. The resulting compounds were primed for further fragment elaboration through the inclusion of synthetic handles from the outset of the syntheses.

How To Characterize Individual Nanosize Liposomes with Simple Self-Calibrating Fluorescence Microscopy.

Nanosize lipid vesicles are used extensively at the interface between nanotechnology and biology, e.g., as containers for chemical reactions at minute concentrations and vehicles for targeted delivery of pharmaceuticals. Typically, vesicle samples are heterogeneous as regards vesicle size and structural properties. Consequently, vesicles must be characterized individually to ensure correct interpretation of experimental results. Here we do that using dual-color fluorescence labeling of vesicles-of their lipid bilayers and lumens, separately. A vesicle then images as two spots, one in each color channel. A simple image analysis determines the total intensity and width of each spot. These four data all depend on the vesicle radius in a simple manner for vesicles that are spherical, unilamellar, and optimal encapsulators of molecular cargo. This permits identification of such ideal vesicles. They in turn enable calibration of the dual-color fluorescence microscopy images they appear in. Since this calibration is not a separate experiment but an analysis of images of vesicles to be characterized, it eliminates the potential source of error that a separate calibration experiment would have been. Nonideal vesicles in the same images were characterized by how their four data violate the calibrated relationship established for ideal vesicles. In this way, our method yields size, shape, lamellarity, and encapsulation efficiency of each imaged vesicle. Applying this procedure to extruded samples of vesicles, we found that, contrary to common assumptions, only a fraction of vesicles are ideal.

Quality of life as a prognostic factor for survival in hepatocellular carcinoma.

BACKGROUND & AIMS: Prognostication in hepatocellular carcinoma (HCC) is demanding. Not only tumour extent and performance status are to be considered, but also liver function, which is often limiting for both survival itself and for treatment possibilities. This study was conducted to assess whether patient-reported questionnaires containing general and liver-specific questions could improve prognostication of survival. METHODS: 185 patients with hepatocellular carcinoma in Norway and Sweden were prospectively included. Patients completed the quality-of-life questionnaires EORTC QLQ C30 and HCC18, and clinical, radiological and laboratory parameters were registered. Multivariate Cox regression and Harrell's C-statistics were used to identify the model that best predicted mortality. RESULTS: Quality-of-life data were prognostic for overall survival. Fatigue and nutrition scales were prognostic in the multivariable analyses alone and in combination with clinical parameters. The prognostic value of established scoring systems was increased by the addition of QoL data. The best prognostic power was achieved by combining HCC18 nutrition scale with selected background parameters. CONCLUSION: Quality-of-life questionnaires can prognosticate mortality in HCC patients. When combined with established scoring systems, both the general cancer questionnaire EORTC QLQ C30, and the additional liver cancer-specific HCC18 increased the prognostic accuracy slightly.

Neoadjuvant chemotherapy versus surgery first for resectable pancreatic cancer (Norwegian Pancreatic Cancer Trial - 1 (NorPACT-1)) - study protocol for a national multicentre randomized controlled trial.

BACKGROUND: Pancreatic cancer is the fourth leading cause of cancer-related death. While surgical resection remains the foundation for potentially curative treatment, survival benefit is achieved with adjuvant oncological treatment. Thus, completion of multimodality treatment (surgical resection and (neo)adjuvant chemotherapy) to all patients and early treatment of micrometastatic disease is the ideal goal. NorPACT-1 aims to test the hypothesis that overall mortality at one year after allocation of treatment can be reduced with neoadjuvant chemotherapy in surgically treated patients with resectable pancreatic cancer. METHODS/DESIGN: The NorPACT- 1 is a multicentre, randomized controlled phase III trial organized by the Norwegian Gastrointestinal Cancer Group for Hepato-Pancreato-Biliary cancer. Patients with resectable adenocarcinoma of the pancreatic head are randomized to receive either surgery first (Group 1: SF/control) or neoadjuvant chemotherapy (Group 2: NT/intervention) with four cycles FOLFIRINOX followed by resection. Both groups receive adjuvant chemotherapy with gemicitabine and capecitabine (six cycles in Group 1, four cycles in Group 2). In total 90 patients will be randomized in all the five Norwegian university hospitals performing pancreatic surgery. Primary endpoint is overall mortality at one year following commencement of treatment for those who ultimately undergo resection. Secondary endpoints are overall survival after date of randomization (intention to treat), overall survival after resection, disease-free survival, histopathological response, complication rates after surgery, feasibility of neoadjuvant and adjuvant chemotherapy, completion rates of all parts of multimodal treatment, and quality-of-life. Bolt-on to the study is a translational research program that aims at identifying factors that are predictive of response to NT, the risk of distant cancer spread, and patient outcome. DISCUSSION: NorPACT- 1 is designed to investigate the additional benefit of NT compared to standard treatment only (surgery + adjuvant chemotherapy) for resectable cancer of the pancreatic head to decrease early mortality (within one year) in resected patients. TRIAL REGISTRATION: Trial open for accrual 01.02.2017. ClinicalTrials.gov Identifier: NCT02919787 . Date of registration: September 14, 2016.

Predicting colorectal surgical complications using heterogeneous clinical data and kernel methods.

OBJECTIVE: In this work, we have developed a learning system capable of exploiting information conveyed by longitudinal Electronic Health Records (EHRs) for the prediction of a common postoperative complication, Anastomosis Leakage (AL), in a data-driven way and by fusing temporal population data from different and heterogeneous sources in the EHRs. MATERIAL AND METHODS: We used linear and non-linear kernel methods individually for each data source, and leveraging the powerful multiple kernels for their effective combination. To validate the system, we used data from the EHR of the gastrointestinal department at a university hospital. RESULTS: We first investigated the early prediction performance from each data source separately, by computing Area Under the Curve values for processed free text (0.83), blood tests (0.74), and vital signs (0.65), respectively. When exploiting the heterogeneous data sources combined using the composite kernel framework, the prediction capabilities increased considerably (0.92). Finally, posterior probabilities were evaluated for risk assessment of patients as an aid for clinicians to raise alertness at an early stage, in order to act promptly for avoiding AL complications. DISCUSSION: Machine-learning statistical model from EHR data can be useful to predict surgical complications. The combination of EHR extracted free text, blood samples values, and patient vital signs, improves the model performance. These results can be used as a framework for preoperative clinical decision support.

Caulobacter chromosome in vivo configuration matches model predictions for a supercoiled polymer in a cell-like confinement.

We measured the distance between fluorescent-labeled DNA loci of various interloci contour lengths in Caulobacter crescentus swarmer cells to determine the in vivo configuration of the chromosome. For DNA segments less than about 300 kb, the mean interloci distances, , scale as n(0.22), where n is the contour length, and cell-to-cell distribution of the interloci distance r is a universal function of r/n(0.22) with broad cell-to-cell variability. For DNA segments greater than about 300 kb, the mean interloci distances scale as n, in agreement with previous observations. The 0.22 value of the scaling exponent for short DNA segments is consistent with theoretical predictions for a branched DNA polymer structure. Predictions from Brownian dynamics simulations of the packing of supercoiled DNA polymers in an elongated cell-like confinement are also consistent with a branched DNA structure, and simulated interloci distance distributions predict that confinement leads to "freezing" of the supercoiled configuration. Lateral positions of labeled loci at comparable positions along the length of the cell are strongly correlated when the longitudinal locus positions differ by <0.16 µm. We conclude that the chromosome structure is supercoiled locally and elongated at large length scales and that substantial cell-to-cell variability in the interloci distances indicates that in vivo crowding prevents the chromosome from reaching an equilibrium arrangement. We suggest that the force causing rapid transport of loci remote from the parS centromere to the distal cell pole may arise from the release at the polar region of potential energy within the supercoiled DNA.

A sensitive method for the analysis of glutathione in porcine hepatocytes.

BACKGROUND: Reduced glutathione (γ-glutamylcysteinylglycine), GSH, is essential when protecting cells from oxidative stress and also an indicator of disease risk. Reported concentrations of GSH and its oxidized form, glutathione disulfide (GSSG), varies considerably, primarily due to the instability of GSH and various analytical methods. METHODS: We designed a sensitive method to analyze GSH and GSSG in porcine hepatocytes using liquid chromatography-tandem mass spectrometry (LC-MS/MS) after stabilization with N-ethylmaleimide (NEM). This method includes stable isotope labeled internal standards and simple synthesis of labeled GSSG which commercial sources rarely offer. GSH and GSSG were analyzed in porcine liver biopsies giving a reference interval based on a large number of samples (26 pigs; 3 parallels). RESULTS: The LC-MS/MS results revealed excellent linearity for both GSH and GSSG, with interday coefficient of variation (%CV) for GSH-NEM and GSSG <10 %. Accuracy for recovery tests was between 95.6% and 106.7% (n = 3) for GSH and between 92.3% and 107.7% (n = 3) for GSSG. The limits of quantification were 0.1 µM for GSH-NEM and 0.08 µM for GSSG. The mean concentration of GSH was 3.5 (95% CI = 1.5-8.1) mmol/liter and of GSSG 0.0023 (95% CI = 0.0003-0.019) mmol/liter. CONCLUSION: For the first time GSH and GSSG are analyzed in porcine hepatocytes by LC-MS/MS yielding a reference level of GSH and GSSG. The method is reproducible in any laboratory with LC-MS/MS service and will probably be applicable in all soft tissues and cell suspensions, essentially with no modification.

Liver sinusoidal cells eliminate blood-borne phage K1F.

Phage treatment has regained attention due to an increase in multiresistant bacteria. For phage therapy to be successful, phages must reach their target bacteria in sufficiently high numbers. Blood-borne phages are believed to be captured by macrophages in the liver and spleen. Since liver sinusoids also consist of specialized scavenger liver sinusoidal endothelial cells (LSECs) and Kupffer cells (KCs), this study investigated the contribution of both cell types in the elimination of Escherichia coli phage K1Fg10b::gfp (K1Fgfp) in mice. Circulatory half-life, organ, and hepatocellular distribution of K1Fgfp were determined following intravenous administration. Internalization of K1Fgfp and effects of phage opsonization on uptake were explored using primary mouse and human LSEC and KC cultures. When inoculated with 107 virions, >95% of the total K1Fgfp load was eliminated from the blood within 20 min, and 94% of the total retrieved K1Fgfp was localized to the liver. Higher doses resulted in slower elimination, possibly reflecting temporary saturation of liver scavenging capacity. Phage DNA was detected in both cell types, with a KC:LSEC ratio of 12:1 per population following cell isolation. Opsonization with plasma proteins increased time-dependent cellular uptake in both LSECs and KCs in vitro. Internalized phages were rapidly transported along the endocytic pathway to lysosomal compartments. Reduced viability of intracellular K1Fgfp corroborated inactivation following endocytosis. This study is the first to identify phage distribution in the liver at the hepatocellular level, confirming clearance of K1Fgfp performed mostly by KCs with a significant uptake also in LSECs.IMPORTANCEFaced with the increasing amounts of bacteria with multidrug antimicrobial resistance, phage therapy has regained attention as a possible treatment option. The phage field has recently experienced an emergence in commercial interest as research has identified new and more efficient ways of identifying and matching phages against resistant superbugs. Currently, phages are unapproved drugs in most parts of the world. For phages to reach broad clinical use, they must be shown to be clinically safe and useful. The results presented herein contribute to increased knowledge about the pharmacokinetics of the T7-like phage K1F in the mammalian system. The cell types of the liver that are responsible for rapid phage blood clearance are identified. Our results highlight the need for more research about appropriate dose regimens when phage therapy is delivered intravenously and advise essential knowledge about cell systems that should be investigated further for detailed phage pharmacodynamics.

Tuning the double lipidation of salmon calcitonin to introduce a pore-like membrane translocation mechanism.

A widespread strategy to increase the transport of therapeutic peptides across cellular membranes has been to attach lipid moieties to the peptide backbone (lipidation) to enhance their intrinsic membrane interaction. Efforts in vitro and in vivo investigating the correlation between lipidation characteristics and peptide membrane translocation efficiency have traditionally relied on end-point read-out assays and trial-and-error-based optimization strategies. Consequently, the molecular details of how therapeutic peptide lipidation affects it's membrane permeation and translocation mechanisms remain unresolved. Here we employed salmon calcitonin as a model therapeutic peptide and synthesized nine double lipidated analogs with varying lipid chain lengths. We used single giant unilamellar vesicle (GUV) calcein influx time-lapse fluorescence microscopy to determine how tuning the lipidation length can lead to an All-or-None GUV filling mechanism, indicative of a peptide mediated pore formation. Finally, we used a GUVs-containing-inner-GUVs assay to demonstrate that only peptide analogs capable of inducing pore formation show efficient membrane translocation. Our data provided the first mechanistic details on how therapeutic peptide lipidation affects their membrane perturbation mechanism and demonstrated that fine-tuning lipidation parameters could induce an intrinsic pore-forming capability. These insights and the microscopy based workflow introduced for investigating structure-function relations could be pivotal for optimizing future peptide design strategies.

Optimized localization analysis for single-molecule tracking and super-resolution microscopy.

We optimally localized isolated fluorescent beads and molecules imaged as diffraction-limited spots, determined the orientation of molecules and present reliable formulas for the precision of various localization methods. Both theory and experimental data showed that unweighted least-squares fitting of a Gaussian squanders one-third of the available information, a popular formula for its precision exaggerates beyond Fisher's information limit, and weighted least-squares may do worse, whereas maximum-likelihood fitting is practically optimal.

Quantitative live-cell imaging of lipidated peptide transport through an epithelial cell layer.

Oral drug delivery increases patient compliance and is thus the preferred administration route for most drugs. However, for biologics the intestinal barrier greatly limits the absorption and reduces their bioavailability. One strategy employed to improve on this is chemical modification of the biologic through the addition of lipid side chains. While it has been established that lipidation of peptides can increase transport, a mechanistic understanding of this effect remains largely unexplored. To pursue this mechanistic understanding, end-point detection of biopharmaceuticals transported through a monolayer of fully polarized epithelial cells is typically used. However, these methods are time-consuming and tedious. Furthermore, most established methods cannot be combined easily with high-resolution live-cell fluorescence imaging that could provide a mechanistic insight into cellular uptake and transport. Here we address this challenge by developing an axial PSF deconvolution scheme to quantify the transport of peptides through a monolayer of Caco-2 cells using single-cell analysis with live-cell confocal fluorescence microscopy. We then measure the known cross-barrier transport of several compounds in our model and compare the results with results obtained in an established microfluidic model finding similar transport phenotypes. This verifies that already after two days the Caco-2 cells in our model form a tight monolayer and constitute a functional barrier model. We then apply this assay to investigate the effects of side chain lipidation of the model peptide drug salmon calcitonin (sCT) modified with 4­carbon and 8­carbon-long fatty acid chains. Furthermore, we compare that with experiments performed at lower temperature and using inhibitors for some endocytotic pathways to pinpoint how lipidation length modifies the main avenues for the transport. We thus show that increasing the length of the lipid chain increases the transport of the drug significantly but also makes endocytosis the primary transport mechanism in a short-term cell culture model.

Practice patterns in diagnostics, staging, and management strategies of gallbladder cancer among Nordic tertiary centers.

BACKGROUND AND OBJECTIVE: Gallbladder cancer (GBC) is a rare malignancy in the Nordic countries and no common Nordic treatment guidelines exist. This study aimed to characterize the current diagnostic and treatment strategies in the Nordic countries and disclose differences in these strategies. METHODS: This was a survey study with a cross-sectional questionnaire of all 19 university hospitals providing curative-intent surgery for GBC in Sweden, Norway, Denmark, and Finland. RESULTS: In all Nordic countries except Sweden, neoadjuvant/downstaging chemotherapy was used in GBC patients. In T1b and T2, majority of the centers (15-18/19) performed extended cholecystectomy. In T3, majority of the centers (13/19) performed cholecystectomy with resection of segments 4b and 5. In T4, majority of the centers (12-14/19) chose palliative/oncological care. The centers in Sweden extended lymphadenectomy beyond the hepatoduodenal ligament, whereas all other Nordic centers usually limited lymphadenectomy to the hepatoduodenal ligament. All Nordic centers except those in Norway used adjuvant chemotherapy routinely for GBC. There were no major differences between the Nordic centers in diagnostics and follow-up. CONCLUSIONS: The surgical and oncological treatment strategies of GBC vary considerably between the Nordic centers and countries.