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Bereaved relatives often refuse to give consent for post-mortem investigation of deceased cancer patients, mainly because of the mutilation due to conventional autopsy (CA). Minimally invasive autopsy (MIA) may be a more acceptable alternative and, if implemented in clinical practice, creates an opportunity to more often obtain post-mortem tissue samples of (recurred) primary tumors and metastases for molecular research. As a measure for tissue quality for molecular studies, we hereby present a feasibility study, comparing the RNA quality of MIA and CA samples, and fresh frozen samples as reference. Tissue samples of heart, liver and kidney were prospectively collected from 24 MIAs followed by CA, and compared to corresponding archival fresh frozen tissue. After RNA isolation and RT-qPCR, RNA integrity numbers (RIN) and GAPDH expression (six amplicon sizes ranging from 71 to 530 base pairs) were measured. RIN values and GAPDH Cq values were analyzed and compared between all sample groups and post-mortem intervals (PMI). RIN values in MIA samples were significantly higher than those in CA samples. GAPDH was expressed significantly higher in MIA samples than in CA samples and 530 bp PCR products could be measured in all cases. GAPDH expression was significantly lower in samples with PMI >15 hours. As expected, the samples of the fresh frozen reference standard performed best in all analyses. MIA samples showed better RNA quality than CA samples, probably due to shorter PMI. Both had lower RNA quality and expression levels than fresh frozen tissue, however, remaining GAPDH RNA was still sufficiently intact. Therefore, other highly expressed genes are most likely also detectable. Gene array analysis should be performed to gain insight into the quality of entire post-mortem genomes. Reducing PMI will further improve the feasibility of demanding molecular research on post-mortem tissues, this is most likely more feasible with MIA than CA.

Within SPIDIA, an EC FP7 project aimed to improve pre analytic procedures, the PAXgene Tissue System (PAXgene), was designed to improve tissue quality for parallel molecular and morphological analysis. Within the SPIDIA project promising results were found in both genomic and proteomic experiments with PAXgene-fixed and paraffin embedded tissue derived biomolecules. But, for this technology to be accepted for use in both clinical and basic research, it is essential that its adequacy for preserving morphology and antigenicity is validated relative to formalin fixation. It is our aim to assess the suitability of PAXgene tissue fixation for (immuno)histological methods. Normal human tissue specimens (n = 70) were collected and divided into equal parts for fixation either with formalin or PAXgene. Sections of the obtained paraffin-embedded tissue were cut and stained. Morphological aspects of PAXgene-fixed tissue were described and also scored relative to formalin-fixed tissue. Performance of PAXgene-fixed tissue in immunohistochemical and in situ hybridization assays was also assessed relative to the corresponding formalin-fixed tissues. Morphology of PAXgene-fixed paraffin embedded tissue was well preserved and deemed adequate for diagnostics in most cases. Some antigens in PAXgene-fixed and paraffin embedded sections were detectable without the need for antigen retrieval, while others were detected using standard, formalin fixation based, immunohistochemistry protocols. Comparable results were obtained with in situ hybridization and histochemical stains. Basically all assessed histological techniques were found to be applicable to PAXgene-fixed and paraffin embedded tissue. In general results obtained with PAXgene-fixed tissue are comparable to those of formalin-fixed tissue. Compromises made in morphology can be called minor compared to the advantages in the molecular pathology possibilities.

The aim of our study was to evaluate the quality of histo- and cytomorphological features of PAXgene-fixed specimens and their suitability for histomorphological classification in comparison to standard formalin fixation. Fifteen colon cancer tissues were collected, divided into two mirrored samples and either formalin fixed (FFPE) or PAXgene fixed (PFPE) before paraffin embedding. HE- and PAS-stained sections were scanned and evaluated in a blinded, randomised ring trial by 20 pathologists from Europe and the USA using virtual microscopy. The pathologists evaluated histological grading, histological subtype, presence of adenoma, presence of lymphovascular invasion, quality of histomorphology and quality of nuclear features. Statistical analysis revealed that the reproducibility with regard to grading between both fixation methods was rather satisfactory (weighted kappa statistic ( k w ) = 0.73 (95 % confidence interval (CI), 0.41–0.94)), with a higher agreement between the reference evaluation and the PFPE samples ( k w  = 0.86 (95 % CI, 0.67–1.00)). Independent from preservation method, inter-observer reproducibility was not completely satisfactory ( k w  = 0.60). Histomorphological quality parameters were scored equal or better for PFPE than for FFPE samples. For example, overall quality and nuclear features, especially the detection of mitosis, were judged significantly better for PFPE cases. By contrast, significant retraction artefacts were observed more frequently in PFPE samples. In conclusion, our findings suggest that the PAXgene Tissue System leads to excellent preservation of histomorphology and nuclear features of colon cancer tissue and allows routine morphological diagnosis.

Introduction Bereaved relatives often refuse to give consent for post-mortem investigation of deceased cancer patients, mainly because of the mutilation due to conventional autopsy (CA). Minimally invasive autopsy (MIA) may be a more acceptable alternative and, if implemented in clinical practice, creates an opportunity to more often obtain post-mortem tissue samples of (recurred) primary tumors and metastases for molecular research. As a measure for tissue quality for molecular studies, we hereby present a feasibility study, comparing the RNA quality of MIA and CA samples, and fresh frozen samples as reference. Materials and methods Tissue samples of heart, liver and kidney were prospectively collected from 24 MIAs followed by CA, and compared to corresponding archival fresh frozen tissue. After RNA isolation and RT-qPCR, RNA integrity numbers (RIN) and GAPDH expression (six amplicon sizes ranging from 71 to 530 base pairs) were measured. RIN values and GAPDH Cq values were analyzed and compared between all sample groups and post-mortem intervals (PMI). Results RIN values in MIA samples were significantly higher than those in CA samples. GAPDH was expressed significantly higher in MIA samples than in CA samples and 530 bp PCR products could be measured in all cases. GAPDH expression was significantly lower in samples with PMI >15 hours. As expected, the samples of the fresh frozen reference standard performed best in all analyses. Conclusion MIA samples showed better RNA quality than CA samples, probably due to shorter PMI. Both had lower RNA quality and expression levels than fresh frozen tissue, however, remaining GAPDH RNA was still sufficiently intact. Therefore, other highly expressed genes are most likely also detectable. Gene array analysis should be performed to gain insight into the quality of entire post-mortem genomes. Reducing PMI will further improve the feasibility of demanding molecular research on post-mortem tissues, this is most likely more feasible with MIA than CA.

In the nonobese diabetic (NOD) mouse model for type 1 diabetes, the inflammatory infiltration of islets starts with an influx of dendritic cells (DC) and macrophages (Mφ) at approximately 4 weeks of age. Around this time, NOD mice show endocrine abnormalities, indicated by a transient hyperinsulinemia that lasts until 8 weeks of age. Subsequently, they develop abnormally large islets of Langerhans, here designated as “mega-islets.” NODscid mice, which lack functional lymphocytes, also exhibit transient hyperinsulinemia, but to a lesser extent. First, to determine the role of lymphocytes in the morphological islet abnormalities, we compared 6-week-old (prediabetic) NOD and NODscid females regarding mega-islet development and accumulation of antigen-presenting cells (APC), particularly CD11c+ DC and ERMP23+ Mφ. In NODscid mice, early APC infiltration and mega-islets were present, but less marked compared with NOD mice, thus suggesting a role of lymphocytes in mega-islet formation. In both NOD and NODscid mice, the APC infiltration was predominantly found around the mega-islets, suggesting a relationship between both parameters. Second, to analyze the role of β-cell hyperactivity in mega-islet formation, we studied the effect of short-term prophylactic insulin treatment on these parameters. Prophylactic insulin treatment decreased the percentages of mega-islets in both NOD and NODscid mice, indicating that β-cell hyperactivity is also involved in mega-islet formation. In conclusion, mega-islet formation in mice with the NOD genetic background takes place under the influence of both β-cell hyperactivity and leukocytes.

Background Infectious mononucleosis (IM), mainly caused by the Epstein‐Barr virus, can result in prolonged symptoms. The objective of this study was to look at the length of sick leave, diagnosis of IM, treatment and comorbidities in a real‐world setting in Germany. Methods This retrospective, cross‐sectional study used electronic medical record data from office‐based practices in Germany and included patients with an initial confirmed diagnosis of IM between the 1 January 1 2016 and December 31 2018. Patients of working age (18‐65 years) with statutory health insurance were included in order to look at the working population who would need a sick note for their employers in case of illness. Results Epstein‐Barr virus was the most common cause of IM in this population of 1,596 patients with an average age of 32 years. The majority of patients were women in all cohorts (~60%). Although CFS, myocarditis and thrombocytopenia were not recorded frequently around the index date, the occurrence did increase during the follow‐up period. Around half of patients received antibiotics. About 62% of all patients were on sick leave for an average of 20 calendar days around the time of their IM diagnosis. Only 1% were still on sick leave after 6 months. Conclusions A small percentage of patients remained on continuous sick leave after 6 months, suggesting that the long‐term effect of IM on the ability to work was minor in our cohort. However, patients could still be experiencing symptoms that influence their quality of life.

Objective: The Utrecht Scale for Evaluation of Rehabilitation (USER) is a measure of functional independence that covers physical functioning (mobility, self-care), cognitive functioning and additional domains of pain, fatigue and mood. USER is implemented in the Netherlands as a generic rehabilitation outcome measure. This article reports the development and psychometric characteristics of USER. Design: (1) Inter-rater reliability study and (2) responsiveness study with measurements at admission and at discharge from clinical rehabilitation. Setting: Three rehabilitation facilities in the Netherlands. Patients: Rehabilitation inpatients (N = 319). Interventions: Not applicable. Main outcome measures: Effect size (ES) and standardized response mean (SRM) of USER scales were compared to those of the Barthel Index, Functional Independence Measure (FIM) and to relevant scales of the SF-36. Results: Inter-rater reliability of USER was satisfactory to good. Strong correlations were found between the physical and cognitive functioning scores of USER, the Barthel Index and FIM (0.84—0.94). Correlations between USER pain, fatigue and mood scores and SF-36 scores were also strong (0.58—0.84). Responsiveness of USER physical functioning (1.0—1.2) was very good and slightly better than responsiveness of the Barthel Index (0.9—1.1) and the FIM motor score (0.8—1.1). Responsiveness of the USER cognitive score was below standard (0.2—0.3), but better than responsiveness of the FIM cognitive scores (0.1—0.2). Conclusions: USER is a reliable, valid and responsive measure of functional independence that can be used as an alternative for the FIM. The pain, fatigue and mood scores were adapted after this study and need additional validation.