Impaired autophagy increases susceptibility to endotoxin-induced chronic pancreatitis.

Chronic pancreatitis (CP) is associated with elevated plasma levels of bacterial lipopolysaccharide (LPS) and we have demonstrated reduced acinar cell autophagy in human CP tissue. Therefore, we investigated the role of autophagy in experimental endotoxin-induced pancreatic injury and aimed to identify LPS in human CP tissue. Pancreatic Atg7-deficient mice were injected with a single sub-lethal dose of LPS. Expression of autophagy, apoptosis, necroptosis, and inflammatory markers was determined 3 and 24 h later utilizing immunoblotting and immunofluorescence. The presence of LPS in pancreatic tissue from mice and from patients and healthy controls was determined using immunohistochemistry, immunoblots, and chromogenic assay. Mice lacking pancreatic autophagy exhibited local signs of inflammation and were particularly sensitive to the toxic effect of LPS injection as compared to control mice. In response to LPS, Atg7Δpan mice exhibited enhanced vacuolization of pancreatic acinar cells, increase in TLR4 expression coupled to enhanced expression of NF-κΒ, JNK, and pro-inflammatory cytokines by acinar cells and enhanced infiltration by myeloid cells (but not Atg7F/F controls). Cell death was enhanced in Atg7Δpan pancreata, but only necroptosis and trypsin activation was further amplified following LPS injection along with elevated pancreatic LPS. The presence of LPS was identified in the pancreata from all 14 CP patients examined but was absent in the pancreata from all 10 normal controls. Altogether, these results support a potential role for metabolic endotoxemia in the pathogenesis of CP. Moreover, the evidence also supports the notion that autophagy plays a major cytoprotective and anti-inflammatory role in the pancreas, and blunting metabolic endotoxemia-induced CP.

[Digital pathology in immuno-oncology-current opportunities and challenges : Overview of the analysis of immune cell infiltrates using whole slide imaging]. / Digitale Pathologie in der Immunonkologie ­ Aktuelle Chancen und Herausforderungen : Überblick zur Analyse von Immunzellinfiltraten mittels Whole Slide Imaging.

BACKGROUND: Immuno-oncology requires objective and standardized methods for measuring immune cell infiltrates for therapy selection and clinical trials. METHODS: Current approaches in applying digital pathology in immuno-oncology and developments in computational image analysis were analyzed. RESULTS: Since 2008, digital pathology has had an ever increasing importance in immuno-oncology. It is currently the only technology allowing the systematic and cost-effective quantitative spatial immune-profiling of patients. The analysis of immunological biomarkers requires integrated staining and image analysis strategies from single- to multistain on slide stacks. Statistical limits of the hypothesis to be tested have to be taken into account. Digital image analysis opens a new technological role for pathology in immuno-oncology and thereby serves as a key technological driver. CONCLUSION: Digital pathology delivers objective and quantitative data on the tumor microenvironment. But currently, a fully automatic, high-throughput analytics capability is still missing. Deep learning is the remedy for this, as it improves image analysis with increasing data availability. This requires the creation of systematic data collections but will in the end deliver standardized and automatic immunological analyses.

Altered density, composition and microanatomical distribution of infiltrating immune cells in cutaneous squamous cell carcinoma of organ transplant recipients.

BACKGROUND: The inflammatory tumour microenvironment is crucial for effective tumour control, and long-term immunosuppression has been identified as a major risk factor for skin carcinogenesis. In solid organ transplant recipients (OTRs) undergoing long-term pharmacological immunosuppression, an increased incidence of cutaneous squamous cell carcinoma (SCC) and more aggressive tumour growth compared with immunocompetent patients has been reported. OBJECTIVES: To determine the density and phenotype of immune cells infiltrating SCC and surrounding skin in OTRs, and to characterize the microanatomical distribution patterns in comparison with immunocompetent patients. METHODS: We analysed immune cell infiltrates within SCC and at defined regions of interest (ROIs) of tumour-surrounding skin in formalin-fixed paraffin-embedded tissue of 20 renal transplant patients and 18 carefully matched immunocompetent patients by high-resolution semiautomated microscopy on complete tissue sections stained for CD4, CD8, CD20 and CD68. RESULTS: The overall immune cell density of SCC arising in OTRs was significantly reduced compared with immunocompetent patients. Particularly CD4+ infiltrates at the directly invasive margin and tumour vicinity, intratumoral CD8+ T-cell densities and the overall density of CD20+ tumour-infiltrating B cells were significantly reduced in the tissue of OTRs. CONCLUSIONS: Immune cell infiltrates within SCC and at defined ROIs of tumour-surrounding skin in OTRs differ markedly in their composition and microanatomical distribution compared with tumours arising in immunocompetent patients. Our findings substantially broaden the understanding of how long-term systemic immunosuppression modulates the local inflammatory microenvironment in the skin and at the site of invasive SCC.

F-18 labelled PSMA-1007: biodistribution, radiation dosimetry and histopathological validation of tumor lesions in prostate cancer patients.

PURPOSE: The prostate-specific membrane antigen (PSMA) targeted positron-emitting-tomography (PET) tracer 68Ga-PSMA-11 shows great promise in the detection of prostate cancer. However, 68Ga has several shortcomings as a radiolabel including short half-life and non-ideal energies, and this has motivated consideration of 18F-labelled analogs. 18F-PSMA-1007 was selected among several 18F-PSMA-ligand candidate compounds because it demonstrated high labelling yields, outstanding tumor uptake and fast, non-urinary background clearance. Here, we describe the properties of 18F-PSMA-1007 in human volunteers and patients. METHODS: Radiation dosimetry of 18F-PSMA-1007 was determined in three healthy volunteers who underwent whole-body PET-scans and concomitant blood and urine sampling. Following this, ten patients with high-risk prostate cancer underwent 18F-PSMA-1007 PET/CT (1 h and 3 h p.i.) and normal organ biodistribution and tumor uptakes were examined. Eight patients underwent prostatectomy with extended pelvic lymphadenectomy. Uptake in intra-prostatic lesions and lymph node metastases were correlated with final histopathology, including PSMA immunostaining. RESULTS: With an effective dose of approximately 4.4-5.5 mSv per 200-250 MBq examination, 18F-PSMA-1007 behaves similar to other PSMA-PET agents as well as to other 18F-labelled PET-tracers. In comparison to other PSMA-targeting PET-tracers, 18F-PSMA-1007 has reduced urinary clearance enabling excellent assessment of the prostate. Similar to 18F-DCFPyL and with slightly slower clearance kinetics than PSMA-11, favorable tumor-to-background ratios are observed 2-3 h after injection. In eight patients, diagnostic findings were successfully validated by histopathology. 18F-PSMA-1007 PET/CT detected 18 of 19 lymph node metastases in the pelvis, including nodes as small as 1 mm in diameter. CONCLUSION: 18F-PSMA-1007 performs at least comparably to 68Ga-PSMA-11, but its longer half-life combined with its superior energy characteristics and non-urinary excretion overcomes some practical limitations of 68Ga-labelled PSMA-targeted tracers.

Patch-Based Nonlinear Image Registration for Gigapixel Whole Slide Images.

OBJECTIVE: Image registration of whole slide histology images allows the fusion of fine-grained information-like different immunohistochemical stains-from neighboring tissue slides. Traditionally, pathologists fuse this information by looking subsequently at one slide at a time. If the slides are digitized and accurately aligned at cell level, automatic analysis can be used to ease the pathologist's work. However, the size of those images exceeds the memory capacity of regular computers. METHODS: We address the challenge to combine a global motion model that takes the physical cutting process of the tissue into account with image data that is not simultaneously globally available. Typical approaches either reduce the amount of data to be processed or partition the data into smaller chunks to be processed separately. Our novel method first registers the complete images on a low resolution with a nonlinear deformation model and later refines this result on patches by using a second nonlinear registration on each patch. Finally, the deformations computed on all patches are combined by interpolation to form one globally smooth nonlinear deformation. The NGF distance measure is used to handle multistain images. RESULTS: The method is applied to ten whole slide image pairs of human lung cancer data. The alignment of 85 corresponding structures is measured by comparing manual segmentations from neighboring slides. Their offset improves significantly, by at least 15%, compared to the low-resolution nonlinear registration. CONCLUSION/SIGNIFICANCE: The proposed method significantly improves the accuracy of multistain registration which allows us to compare different antibodies at cell level.

High density of FOXP3-positive T cells infiltrating colorectal cancers with microsatellite instability.

High-level microsatellite instability (MSI-H) in colorectal cancer accounts for about 12% of colorectal cancers and is typically associated with a dense infiltration with cytotoxic CD8-positive lymphocytes. The role of regulatory T cells that may interfere with the host's antitumoural immune response in MSI-H colorectal cancers has not been analysed yet. Using an antibody directed against the regulatory T-cell marker transcription factor forkhead box P3 (FOXP3), regulatory T cells were examined in 70 colorectal cancers with known MSI status (MSI-H, n=37; microsatellite stable, n=33). In MSI-H colorectal cancers, we found a significantly higher intraepithelial infiltration with FOXP3-positive cells (median: 8.5 cells per 0.25 mm(2) vs 3.1 cells per 0.25 mm(2) in microsatellite stable, P<0.001), and a significantly elevated ratio of intraepithelial to stromal infiltration (0.05 vs 0.01 in microsatellite stable, P<0.001). CD8-positive cell counts were related positively to the number of FOXP3-positive cells (Spearman's rho=0.56 and 0.55, respectively). Our results show that the elevated number of CD8-positive lymphocytes found in MSI-H colorectal cancers is paralleled by an enhanced infiltration with CD8-negative FOXP3-positive cells. These data suggest that FOXP3-positive cells may play a role in the regulation of the immune response directed against MSI-H colorectal cancers at the primary tumour site.

[Virtual microscopy in systems pathology]. / Virtuelle Mikroskopie in der Systempathologie.

Genomics and proteomics have evolved towards systems biology. The general goal here is the construction of complex, functional models of biological systems on the basis of molecular networks. Such models enable improved quality in interpretation and evaluation of quantitative measurements and afford a substantially deeper functional understanding. Systems pathology differs from systems biology by attaching the same importance to spatial modelling of tissue alterations as to gene regulatory modelling. In this way, systems pathology is able to deploy disease models for improved diagnosis, prognosis and therapy. In the present work a generic process for systems pathology is created, integrating gene regulatory and morphological models towards molecular disease models. For this purpose, fluorescent virtual microscopy will be essential as it delivers morphological and molecular tissue data with high spatial resolution and high throughput. Using epidermal differentiation as an example, it is shown how - using virtual microscopy - the spatiotemporal expression of biomarkers can be modelled by reconstructing protein networks from fluorescent tissue sections.

[Total uterine rupture of a cesarean section scar during attempted vaginal delivery under epidural analgesia]. / Total uterusruptur af sectiocikatrice under forsøg på vaginal forløsning under epidural analgesi.

A case of complete uterine rupture and abruptio placentae during an attempt of vaginal delivery under epidural analgesia in a woman with previous caesarean section is described and discussed. When using epidural analgesia for such a patient during vaginal delivery it is imperative that newly developed pain should be ascribed to the uterine scar and the diagnosis of threatening uterine rupture be made immediately.

[Anesthesia for single-day orthopedic surgery]. / Anaestesi til ortopaedkirurgisk endagskirurgi.

During the period 1 January 1989 to 31 December 1989, a total of 1,004 patients were submitted to outpatient single-day surgery in Aalborg Hospital. As fas as possible, the patients were anaesthetized with propofol (Diprivan)/alfentanil (Rapifen) and oxygen and atmospheric air. This form of anaesthesia was found to be rapid and suitable for single-day surgery. 8.1% of the patients required admission for further observation, 2.3% of these on account of anaesthesiological complications. It is concluded that, by means of meticulous planning, it is possible to carry out ambulant anaesthesia and surgery and, employing limited staff resources, it has proved possible to reduce the waiting time for outpatient intervention to 10-12 weeks.

Contamination of subclavian vein catheters: an intraluminal culture method.

An intraluminal culture method was evaluated for central venous catheters and compared to conventional catheter tip cultures and cultures from the insertion site. The colonization-rate of the catheters was 43.7% using conventional tip culture, 34.4% with the intraluminal method and 40.1% at the skin puncture site. Only 37.5% of the catheters showed identical bacteria at the skin puncture site and catheter-tip, compared with 87.5% between intraluminal culture and catheter-tip culture (P less than 0.05).

Contamination of intravenous infusion systems--the effect of changing administration sets.

Intravenous administration sets were changed at varying time intervals between every 24 h and 120 h in 387 patients. The rates of intraluminal contamination of the cannulae and of local inflammation were measured in relation to the time interval between changing sets. There was no correlation between phlebitis and intraluminal contamination, but a significant association was found between phlebitis and fever, infusion of potassium at greater than 10 mmol l-1, Venflon type 140 and infusion of blood or intralipid. No correlation was found between septicaemia and intraluminal contamination of the infusion systems. Contamination of cannulae increased slightly with time, but this was not statistically significant. We conclude that there will be no clinical benefit by daily changing of administration sets, compared with changing up to every fifth day.

Oral lormetazepam in premedication. A comparison with diazepam.

Lormetazepam, a relatively new benzodiazepine was compared in a randomised, double blind trial with diazepam for its effectiveness as an oral premedicant drug. A scoring system was used to assess sedation, relief of anxiety, nausea, dizziness and cardiovascular effects in two groups of patients having orthopaedic operations. Some statistical indication that lormetazepam has a greater anxiolytic effect than diazepam was found, but in assessing total effect using a known scoring system, no difference was demonstrated between the two drugs.

A trial of povidone-iodine for prevention of contamination of intravenous cannulae.

A prospective randomized study of 227 intravenous cannulae was undertaken to evaluate the effects of two different skin disinfectants before intravenous cannulation. Patients were divided into two groups. In one povidone-iodine was used as disinfectant while in the other 70% isopropylalcohol was used. Intraluminal contamination rates, colony counts and organisms isolated are presented. We found no correlation between use of cannulae sideport and contamination. It was found that povidone-iodine as a skin disinfectant reduced the intraluminal contamination from 22 to 7.25% (P less than 0.01).

Metoprolol decreases the amount of halothane required to induce hypotension during general anaesthesia.

A double-blind, randomized and placebo-controlled study was undertaken to assess the effects of metoprolol on controlled hypotension during halothane anaesthesia. Nineteen patients under-going middle ear or nasal septum surgery were studied. The aim was to achieve a 25% reduction of the mean arterial pressure in both groups. Metoprolol was given by mouth on two occasions in the 10-12 h before operation. In the group pretreated with metoprolol the halothane concentration was reduced by 35.5%. The duration of bleeding and its degree were reduced significantly. The time to awakening was shorter and of better quality after pretreatment with metoprolol. Rate-pressure product was less after pretreatment with metoprolol, and rebound hypertension was well controlled.

Skin disinfection before intravenous cannulation. Intraluminal contamination after disinfection with 70% isopropyl alcohol.

A prospective randomized study evaluating the effects of skin disinfectants before intravenous cannulation was undertaken. 187 intravenous cannulae were examined. Patients were divided into two groups: in one, 70% isopropyl alcohol was used as a skin disinfectant while in the other, no skin disinfectant was used. Intraluminal contamination rates, colony counts and organisms isolated were analysed. No significant differences were found between the two groups. There was no correlation between the use of the sideport and contamination. It appears, therefore, that 70% isopropyl alcohol cannot prevent or even reduce the intraluminal contamination.

The contaminated skin-knife--fact or fiction.

Three hundred and fifty-eight patients undergoing clean orthopaedic surgery were included in this prospective study. The knives used during surgery, i.e. the knife used for skin incision (the skin-knife) and the knife used to finish the operation were bacteriologically examined. We found no evidence that the practise of discarding the knife after skin incision reduced the incidence of wound infection after clean orthopaedic surgery. Laminar air flow significantly reduced the rate of knife contamination.