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BACKGROUND: This study aimed to ascertain the microbiology, severity stratification, and clinical outcomes of cellulitis based on our current management for comparison with international reports METHODS: A retrospective chart review was conducted of all patients with cellulitis treated by the department of surgery at Ngwelezana Hospital over an 18-month period. Severity of cellulitis was graded, and a comparison was made of the Eron and Modified Dundee classifications. Superficial swabs were taken for culture on patients who had cellulitis with open wounds or blisters. Culture results, antibiotics used, need for surgical intervention, and length of hospital stay were documented and analysed RESULTS: One hundred and thirty-four patients had cellulitis. Severity grading for classes I-IV in the Eron classification was 3%, 57%, 39% and 1%, respectively, and for the Modified Dundee classification, 47%, 11%, 38%, and 4%, respectively. Co-amoxiclav was the most used antibiotic (73%). Superficial skin swabs were taken from 49 patients and 34 cultured 44 specific organisms. The most common organism identified was Staphylococcus aureus (30%). Several gram-negative and anaerobic organisms were cultured. Fifty-three patients required surgical debridement of the infected area and one patient required an above-knee amputation. Mean hospital stay for patients who did not receive surgical intervention was 6 days (IQR 3) and 7 days (IQR 4) for those who did. There were no deaths CONCLUSION: The Dundee classification triages fewer patients as class 2 severity than the Eron system and its use has the potentail to reduce the number of patients hospitalised. Gram-positive organisms predominated in those cultured, but gram-negative cultures were frequent compared to other reported series. Co-amoxiclav is effective as first-line antimicrobial therapy in our environment

The term “biobanking” is often misapplied to any collection of human biological materials (biospecimens) regardless of requirements related to ethical and legal issues or the standardization of different processes involved in tissue collection. A proper definition of biobanks is large collections of biospecimens linked to relevant personal and health information (health records, family history, lifestyle, genetic information) that are held predominantly for use in health and medical research. In addition, the International Organization for Standardization, in illustrating the requirements for biobanking (ISO 20387:2018), stresses the concept of biobanks being legal entities driving the process of acquisition and storage together with some or all of the activities related to collection, preparation, preservation, testing, analysing and distributing defined biological material as well as related information and data. In this review article, we aim to discuss the basic principles of biobanking, spanning from definitions to classification systems, standardization processes and documents, sustainability and ethical and legal requirements. We also deal with emerging specimens that are currently being generated and shaping the so-called next-generation biobanking, and we provide pragmatic examples of cancer-associated biobanking by discussing the process behind the construction of a biobank and the infrastructures supporting the implementation of biobanking in scientific research.

Aims Microbiological culture is a key element in the diagnosis of periprosthetic joint infection (PJI). However, cultures of periprosthetic tissue do not have optimal sensitivity. One of the main reasons for this is that microorganisms are not released from the tissues, either due to biofilm formation or intracellular persistence. This study aimed to optimize tissue pretreatment methods in order to improve detection of microorganisms. Methods From December 2017 to September 2019, patients undergoing revision arthroplasty in a single centre due to PJI and aseptic failure (AF) were included, with demographic data and laboratory test results recorded prospectively. Periprosthetic tissue samples were collected intraoperatively and assigned to tissue-mechanical homogenization (T-MH), tissue-manual milling (T-MM), tissue-dithiothreitol (T-DTT) treatment, tissue-sonication (T-S), and tissue-direct culture (T-D). The yield of the microbial cultures was then analyzed. Results A total of 46 patients were enrolled, including 28 patients in the PJI group and 18 patients in the AF group. In the PJI group, 23 cases had positive culture results via T-MH, 22 cases via T-DTT, 20 cases via T-S, 15 cases via T-MM, and 13 cases via T-D. Three cases under ongoing antibiotic treatment remained culture-negative. Five tissue samples provided the optimal yield. Any ongoing antibiotic treatment had a relevant influence on culture sensitivity, except for T-DTT. Conclusion T-MH had the highest sensitivity. Combining T-MH with T-DTT, which requires no special equipment, may effectively improve bacterial detection in PJI. A total of five periprosthetic tissue biopsies should be sampled in revision arthroplasty for optimal detection of PJI. Cite this article: Bone Joint Res 2021;10(2):96–104.

Introduction Fixation is the critical step in the preservation of tissues in diagnostic pathology. The formalin is an economical and excellent fixative with the inherent property of adequate fixation. The well-established side effects of formalin include mucosal irritation, upper respiratory diseases, and corrosive injury to the gastrointestinal tract. In addition, substantial evidence exists regarding the potential role of formaldehyde as a human carcinogen. The carcinogenic and toxic effects of formalin encourage searching for alternative fixatives for tissue fixation. However, “the formalin dogma” has severely hampered the search for alternative fixatives for many years. Material and Methods Ninety tissues of liver and skeletal muscle obtained during autopsies were immersed in adequate amounts of the following fixatives: formalin (10%), methyl alcohol (70%), and acetone (100%). The comparison among the three was made based on time for fixation, preservation of tissue architecture, cell borders, cytoplasm, nuclear contours, chromatin texture, and uniformity of staining. Results The tissue preserved in formalin undergoes rapid fixation compared with alcohol and acetone. The tissue architecture, cell border characteristics of alcohol and acetone was found satisfactory compared with formalin. The cytoplasm and nuclear contour were superior with the formalin. The chromatin texture and uniformity of staining were similar with all the three fixatives. Conclusion The formalin is considered superior to most of the parameters, whereas both methyl alcohol and acetone showed nearly equivalent scores. Hence, owing to the potential human health hazards and carcinogenicity of formalin, no rational reasons hamper the complete substitution of formalin with alternative fixatives such as alcohol and acetone in diagnostic pathology and medical research.

Introduction There is no current standard by which culture specimens from revision shoulder arthroplasty should be handled, processed, cultured, and reported. Due to the relatively low numbers of cases multicenter study may provide information to form consensus recommendations. However, assimilation of multicenter data requires comparable methodologies. The objective of this study was to document and evaluate the extent of variability between surgeons and institutions. Methods An 11-question survey was sent to 20 shoulder surgeons as part of the American Shoulder and Elbow Society (ASES) Periprosthetic Joint Infection (PJI) Multicenter Workgroup. Questions addressed how samples are handled in the operating room by surgeons, processing of tissue samples and explants by laboratories, number of media, culture incubation durations, and culture reporting. Results Common practices regarding specimen handling and processing were identified including prolonged culture incubation times >13 days (94% of participants). However, substantial variation in handling of tissue and explant specimens, number and type of media used, and reporting of results were identified. The majority of surgeons reported using a sterile instrument to harvest each individual tissue specimen (10/17, 59%), more so than using any available instrument (4/17, 24%) or washing and re-using the same instrument (3/17, 18%). Half of the institutions require a time limit by which samples must be processed in the laboratory (8/16, 50%). Nine institutions (9/16, 56%) report cultures in a semi-quantitative manner, while 7 (44%) report cultures in a binary fashion. Five institutions reported having performed a negative control study, and the rate of positive cultures ranged between 0% and 17%. The majority of positive cultures from the negative controls contained Cutibacterium (92%). Discussion Specimen handling, processing, culturing, and reporting varies widely between institutions. Due to the risk of false positives as demonstrated by negative control studies, surgeons should be cognizant of potential sources of contamination at the specimen handling level in the operating room and specimen processing level in the laboratory. Given the challenges in interpretation of positive cultures in revision shoulder arthroplasty, further studies are needed to determine whether assimilation of data across institutions is acceptable or whether a standard culturing methodology across institutions is necessary. Level of Evidence: V

Mycobacterium massiliense (M. massiliense) is a novel nontuberculous mycobacteria (NTM) and an opportunistic pathogen that lives in the water, soil, food, and air. It is a subspecies of the rapidly growing mycobacteria Mycobacterium abscessus. This atypical pathogen has been reported mainly in patients with lung disease or those undergoing cosmetic or surgical procedures. A 62-year-old woman presented with productive otorrhea for 10 months, no history of surgery, and chronic otitis media. M. massiliense was identified from a tissue specimen using real-time polymerase chain reaction for NTM (Biosewoom, Seoul, Korea), and NTM was identified by acid-fast bacilli culture. Successful treatment consisted of clarithromycin for 4 months. No other case of chronic otitis media related to M. massiliense has been reported. This is the first confirmed case of chronic otitis media caused by M. massiliense in a healthy adult in South Korea. (Korean J Med 2020;95:276-280)

Atherosclerotic plaques can gradually develop in certain arteries. Disruption of fibrous tissue in plaques can result in plaque rupture and thromboembolism, leading to heart attacks and strokes. Collagen fibrils are important tissue building blocks and tissue strength depends on how fibrils are oriented. Fibril orientation in plaque tissue may potentially influence vulnerability to disruption. While X-ray scattering has previously been used to characterize fibril orientations in soft tissues and bones, it has never been used for characterization of human atherosclerotic plaque tissue. This study served to explore fibril orientation in specimens from human plaques using small angle X-ray scattering (SAXS). Plaque tissue was extracted from human femoral and carotid arteries, and each tissue specimen contained a region of calcified material. Three-dimensional (3D) collagen fibril orientation was determined along scan lines that started away from and then extended toward a given calcification. Fibrils were found to be oriented mainly in the circumferential direction of the plaque tissue at the majority of locations away from calcifications. However, in a number of cases, the dominant fibril direction differed near a calcification, changing from circumferential to longitudinal or thickness (radial) directions. Further study is needed to elucidate how these fibril orientations may influence plaque tissue stress–strain behavior and vulnerability to rupture.

The immune tumor microenvironment (iTME) of glioblastoma (GBM) contains microglial, macrophage, other myeloid cell populations and as adaptive immune cells. Recent therapeutic strategies for GBM aim at targeting iTME components to induce antitumoral immunity. A patient-tailored, ex vivo drug testing and response analysis platform would facilitate personalized therapy planning, provide insights into treatment-induced immune mechanisms in the iTME, and enable the discovery of biomarkers of response and resistance. Here, we generated patient-derived, live 3D GBM bioreactors from different tumor regions to assess iTME treatment responses to microglia modulators and immune checkpoint inhibitors. Intact GBM tissue specimens from the tumor center and periphery were cultured for 7 days in the presence or absence of anti-PD1, anti-CD47 antibodies or their combination. Tissues were analyzed by CODEX highly multiplexed microscopy using an immune-centered 54-marker panel, and changes in cytokine and chemokine levels in culture supernatants were investigated. A computational pipeline for integrative therapy response assessment was implemented. Explant cultures from n=8 IDH wt GBM were subjected to this integrative personalized analysis. Tissue integrity after 3D bioreactor cultures was comparable to tissue taken directly after surgery. FFPE CODEX workflow was feasible with adequate staining quality in bioreactor cultures. 850'000 single cells were segmented and clustered. Cellular composition between tumor center and the peripheral invasion zone differed significantly in immune phenotypes, cytokine profile and response to innate, adaptive or combinatorial local immunotherapies. Multiplexed cytokine analysis revealed IFNγ response signatures in a subset of center samples, whereas the peripheral invasion zone displayed a blunted cytokine response. This cytokine signature corresponded to cellular composition shifts within specific cellular neighborhoods. CD4 and CD8 T cells were invigorated and left their vascular niche. Our study demonstrates that local immunotherapies enable an active antitumoral immune response within the tumor center, and provides a multidimensional personalized framework for immunotherapy response assessment.

BACKGROUND The BRAF V600E mutation is present in approximately 50% of patients with melanoma and is an important prerequisite for a response to targeted therapies such as BRAF inhibitors. In the majority of patients, the BRAF mutational status is based on the analysis of tissue samples from the extracranial primary tumor only. Since the extracranial and intracranial BRAF mutational status may be discrepant, the additional information on the BRAF mutational status of melanoma brain metastases would be of clinical value, e.g., for the prediction of response to targeted therapies. Here, we evaluated the potential of MRI radiomics for the determination of the intracranial BRAF mutational status in patients with melanoma brain metastases. PATIENTS AND METHODS Fifty-nine patients with melanoma brain metastases from two university hospitals (group 1, 45 patients; group 2, 14 patients) were operated with subsequent genetic analysis of the intracranial BRAF mutational status. All patients underwent structural MRI preoperatively. Areas of contrast enhancement were manually segmented and analyzed. Group 1 was used for model training and validation, group 2 for model testing. After image preprocessing and radiomics feature extraction, a test-retest analysis was performed to identify robust features prior to feature selection. Finally, the best performing radiomics model was applied to the test data. Diagnostic performances were evaluated using receiver operating characteristic (ROC) analyses. RESULTS Twenty-two patients (49%) in group 1, and 6 patients (43%) in group 2 had an intrametastatic BRAF V600E mutation. Using the best performing six parameter radiomics signature, a linear support vector machine classifier yielded an area under the ROC curve (AUC) of 0.92 (sensitivity, 83%; specificity, 88%) in the test data. CONCLUSION The developed radiomics classifier allows a non-invasive prediction of the intracranial BRAF V600E mutational status in patients with melanoma brain metastases and may be of value for treatment decisions.

Introduction: Due to lack of data for the early diagnosis and treatment of multi drug resistant spinal tuberculosis, the aim of the present study was to estimate the prevalence of multi drug resistant spinal tuberculosis and study the clinical outcomes. Materials & methods: A total of 80 patients were selected for the study. On the basis of the clinico-radiological diagnosis the anti-tubercular treatment was started in all the included patients diagnosed with drug resistant spinal tuberculosis. All the tissue specimen from the biopsy and samples obtained were subjected to histopathological examination, CBNAAT and MGIT culture.Results: Functional and clinical outcome of the patients were evaluated by VAS and ASIA grading. The baseline score of the VAS score was found to be 6 – 7, this score was used to compare with the follow up score. When VAS and ASIA score was compared with previous follow up visit VAS score, significant improvement was noted up to 9 months. Conclusion: In conclusion, early detection of MDR Spinal Tuberculosis would be helpful to avoid consequences arising due to in appropriate chemotherapy and, hence, would also result in favourable outcomes. However, further similar studies with larger samples are needed before we can make definitive statement about the usage of CBNAAT Test as routine diagnostic tool in suspected MDR Spinal Tuberculosis cases.

Biomechanical failure of the artery wall can lead to rupture, a catastrophic event with a high rate of mortality. Thus, there is a pressing need to understand failure behavior of the arterial wall. Uniaxial testing remains the most common experimental technique to assess tissue failure properties. However, the relationship between intrinsic failure parameters of the tissue and measured uniaxial failure properties is not fully established. Furthermore, the effect of the experimental variables, such as specimen shape and boundary conditions, on the measured failure properties is not well understood. We developed a finite element model capable of recapitulating pre-failure and post-failure uniaxial biomechanical response of the arterial tissue specimen. Intrinsic stiffness, strength and fracture toughness of the vessel wall tissue were used as the input material parameters to the model. Two uniaxial testing protocols were considered: a conventional setup with a rectangular specimen held at the grips by cardboard inserts, and the other used a dogbone specimen with soft foam inserts at the grips. Our computational study indicated negligible differences in the peak stress and post-peak mechanical behavior between these two testing protocols. It was also found that the tissue experienced only modest localized failure until higher levels of applied stretch beyond the peak stress. A robust cohesive model was capable of modeling the post-peak biomechanical response, which was primarily governed by tissue fracture toughness. Our results suggest that the post-peak region, in conjunction with the peak stress, must be considered to evaluate the complete biomechanical failure behavior of the soft tissue.

Analysis of the chemical makeup of the brain enables a deeper understanding of several neurological processes. Molecular imaging that deciphers the spatial distribution of neurochemicals with high specificity and sensitivity is an exciting avenue in this aspect. The past two decades have witnessed a significant surge of mass spectrometry imaging (MSI) that can simultaneously map the distribution of hundreds to thousands of biomolecules in the tissue specimen at a fairly high resolution, which is otherwise beyond the scope of other molecular imaging techniques. In this review, we have documented the evolution of MSI technologies in imaging the anatomical distribution of neurochemicals in the human brain in the context of several neuro diseases. This review also addresses the potential of MSI to be a next-generation molecular imaging technique with its promising applications in neuropathology.

The occurrence of neurotrophic tyrosine receptor kinase (NTRK) gene fusions in a wide range of tumor types presents an attractive opportunity for using a tropomyosin receptor kinase (TRK) inhibitor as cancer therapy. Recent clinical studies have demonstrated highly efficacious outcomes associated with the use of TRK inhibitors, such as larotrectinib and entrectinib in NTRK fusion-bearing cancers, in both adult and pediatric populations. While NTRK gene fusions are commonly found in some uncommon adult and pediatric malignancies, they are also found, albeit rarely, in a wide range of more common malignancies. The potential value of testing for NTRK gene fusions in practically all advanced malignancies is underpinned by the remarkable therapeutic outcomes that TRK inhibitors offer. This requirement presents practical and financial challenges in real-world oncological practice. Furthermore, different testing platforms exist to detect NTRK gene fusions, each with its advantages and disadvantages. It is, therefore, imperative to develop strategies for NTRK gene fusion testing in an attempt to optimize the use of limited tissue specimen and financial resources, and to minimize the turnaround time. A multidisciplinary task force of Singapore medical experts in both public and private sectors was convened in late 2020 to propose testing algorithms for adult colorectal tumors, sarcomas, non-small cell lung cancer, and pediatric cancers, with particular adaptation to the Singapore oncological practice. The recommendations presented here highlight the heterogeneity of NTRK-fusion positive cancers, and emphasize the need to customize the testing methods to each tumor type to optimize the workflow.

Study question Does endoplasmic reticulum (ER) stress evaluated by X-box binding protein 1 (XBP–1) among patients aged 18–30 years play a role in recurrent pregnancy loss (RPL)? Summary answer High levels of XBP–1 observed in patients with known RPL indicate that ER stress plays an important role in RPL. What is known already Female reproductive tract undergoes dynamic changes during oogenesis which require protein synthesis, folding, maturation, which take place in the ER. ER is also responsible for destruction of unfolded/misfolded proteins. Excess accumulation of these faulty proteins leads to ER stress, which actives unfolded protein response (UPR). XBP–1 is a transcription factor involved in UPR and regulates ER stress-mediated apoptosis. Regulation of ER homeostasis is important in folliculogenesis, oocyte maturation and embryogenesis. It is also known that ER stress has a positive correlation with age and it is associated with age-related diseases. Study design, size, duration This prospective case-controlled study was conducted at University of Health Sciences Turkey, Istanbul Kanuni Sultan Suleyman Training and Research Hospital Department of Obstetrics and Gynecology between March 2020 – September 2020. A total of 70 subjects were included in the study. All patients gave their written informed consent before their enrollment in the study. Participants/materials, setting, methods 38 patients aged 18–30 years with a history of RPL were included in the study. Patients who had miscarriages due to fetal abnormalities, patients with infections, endocrine or genetic disorders, smokers, alcohol and/or drug abusers, with acute/chronic inflammatory diseases, patients using steroids, anti-inflammatory and antioxidant medications were excluded from the study. Age-matched 32 healthy subjects without RPL were included in the control group. XBP–1 levels were determined using Human XBP–1 ELISA Kit (Elabscience Co., USA). Main results and the role of chance The mean age in the control group and in the study group were 25.21±3.3 and 25.26±2.6, respectively and they were statistically similar (p = 0.324). When groups were compared according to thyroid stimulating hormone (TSH) levels and body mass index (BMI), which are additional risk factors of RPL both groups were statistically similar (p = 0.642 and 0.942, respectively). As expected gravidity and abortus numbers were significantly higher in the study group (p Limitations, reasons for caution Small sample size is an important limitation of this study. In addition, evaluating XBP–1 only in serum samples does not let us drive any conclusions on the local changes of ER stress. Studies with larger samples sizes and studying XBP–1 levels in tissue samples of endometrial material is needed. Wider implications of the findings: The significantly high levels of XBP–1 in RPL patients younger than 30 years, indicate higher ER stress in this group even when age dependent increase in ER stress is calculated out of the equation. XBP–1 can be a promising marker in evaluating patients with a fertility wish for RPL risk. Trial registration number NCT04455256

Though transcriptomics technologies evolve rapidly in the past decades, integrative analysis of mixed data between microarray and RNA-seq remains challenging due to the inherent variability difference between them. Here, Rank-In was proposed to correct the nonbiological effects across the two technologies, enabling freely blended data for consolidated analysis. Rank-In was rigorously validated via the public cell and tissue samples tested by both technologies. On the two reference samples of the SEQC project, Rank-In not only perfectly classified the 44 profiles but also achieved the best accuracy of 0.9 on predicting TaqMan-validated DEGs. More importantly, on 327 Glioblastoma (GBM) profiles and 248, 523 heterogeneous colon cancer profiles respectively, only Rank-In can successfully discriminate every single cancer profile from normal controls, while the others cannot. Further on different sizes of mixed seq-array GBM profiles, Rank-In can robustly reproduce a median range of DEG overlapping from 0.74 to 0.83 among top genes, whereas the others never exceed 0.72. Being the first effective method enabling mixed data of cross-technology analysis, Rank-In welcomes hybrid of array and seq profiles for integrative study on large/small, paired/unpaired and balanced/imbalanced samples, opening possibility to reduce sampling space of clinical cancer patients. Rank-In can be accessed at http://www.badd-cao.net/rank-in/index.html.

Background and Aims Perinatal asphyxia (PA) is associated with more than half a million mature newborn deaths yearly. It may lead to severe complications including hypoxic encephalopathy, renal- hepatic- and cardiovascular injury, as well as respiratory distress, Basic research and clinical trials mainly focus on mitigating central nervous system damage by selective head or whole body cooling, which is currently the only routinely used treatment in clinical practice. However, the extent of PA-associated multi-organ damage is not clarified yet and effective therapies are lacking. Our aim was to investigate the acute renal, hepatic and cardiac impairment following PA and to identify pathways involved in the pathomechanism. In addition, we aimed to explore long-term effects of PA on permanent organ damage and susceptibility to ischemia/-reperfusion injury in adulthood. Method Postnatal 7 day-old male Wistar rat pups (n=5-10/group) were randomly grouped as follows: (i) Baseline; (ii) Control; (iii) PA. The PA group was separated from the dam and received asphyxic gas mixture (4% O2; 20% CO2 in N2) for 15 minutes, while Control animals received normal air following separation. Serum and tissue samples were collected after 4 (T4) or 24 (T24) hours. In a second experiment 35 min bilateral renal ischemic insult was performed on control and PA rats aged 6 months (n=6-7/group). Serum and tissue samples were collected 24 (T24 IR) hours after reperfusion (Figure 1). Serum levels of electrolytes, kidney and liver functional parameters, and myocardial ischemic protein Troponin I were determined. Highly selective and sensitive tubular injury markers (Kim1, Ngal) were measured. Expressions of hypoxic (Hif1a, Hif2a) inflammatory (Il1α, Il1β, Il6, Tnfα, Mcp1, Tlr2), apoptotic (Bax, Bcl-2) and angiogenic genes (Vegf, Epo) and heat shock proteins (Hsp27, Hsp72) were investigated. Periodic-Acid Schiff stained kidney sections and Hematoxylin & Eosin stained liver sections were evaluated for structural injury. Results Blood urea nitrogen (BUN) and serum GPT were elevated at T4 following PA. Kim1, Ngal and heat shock protein expressions were increased, inflammatory and angiogenic pathways were activated in the kidney after PA. In the liver hypoxic and apoptotic pathways were activated at T24 in controls and after asphyxia, but not in the Baseline group. Vacuolisation, cytoplasmic degradation, and the onset of necrosis were observed in the liver following PA. Serum Troponin I was elevated indicating myocardial damage, moreover inflammatory cytokines and heat shock proteins increased in the heart. In adult PA rats BUN levels were elevated, suggesting a long-term detrimental effect of PA on renal function. In addition, adult PA rats were more susceptible to renal ischemic insult, confirmed by higher serum creatinine and GPT levels, as well as increased expression of tubular injury, hypoxic and inflammatory markers compared to Control rats subjected to ischemia. Conclusion Acute renal, hepatic and myocardial impairment was observed after PA. These results may justify the need for clinical follow-up and novel treatment strategies for possible multi-organ damage. The molecular pathways described here are potential targets for therapeutic intervention. In addition, birth asphyxia may increase sensitivity to renal injury in adulthood, which may be worth considering in clinical situations with potential renal impairment.

This study was designed to examine osteoconductive effects of Mg in rats tibia. The animals were sacrificed after 1, 2, and 8 weeks. The elemental analysis was performed using SEM/EDX at week 1. Following X-ray micrography at weeks 2 and 8, samples were embedded in paraffin. The expression of osteocalcin was observed by immunohistochemical staining. The element concentrations of fibrous capsules around the specimens were also measured by ICP-MS. The concentrations of Ca and P on the surface of the Mg specimen increased in SEM/EDX. The tissue specimen showed new bone formation on the bone surface near the implanted area. The concentrations of Mg, Ca, and P were high in the fibrous capsules surrounding Mg. Implantation induced differentiation of osteoblasts, and this process was considered to be associated with new bone formation. Induction of cell differentiation may be influenced by corrosion products in addition to corroding magnesium.

Formalin-fixed paraffin-embedded tissue, the most common tissue specimen stored in clinical practice, presents challenges in the analysis due to formalin-induced artifacts. Here, we present Strand Orientation Bias Detector (SOBDetector), a flexible computational platform compatible with all the common somatic SNV-calling pipelines, designed to assess the probability whether a given detected mutation is an artifact. The underlying predictor mechanism is based on the posterior distribution of a Bayesian logistic regression model trained on The Cancer Genome Atlas whole exomes. SOBDetector is a freely available cross-platform program, implemented in Java 1.8.

Objectives This study aimed to develop and validate a deep learning algorithm to screen digitized acid fast–stained (AFS) slides for mycobacteria within tissue sections. Methods A total of 441 whole-slide images (WSIs) of AFS tissue material were used to develop a deep learning algorithm. Regions of interest with possible acid-fast bacilli (AFBs) were displayed in a web-based gallery format alongside corresponding WSIs for pathologist review. Artificial intelligence (AI)–assisted analysis of another 138 AFS slides was compared to manual light microscopy and WSI evaluation without AI support. Results Algorithm performance showed an area under the curve of 0.960 at the image patch level. More AI-assisted reviews identified AFBs than manual microscopy or WSI examination (P < .001). Sensitivity, negative predictive value, and accuracy were highest for AI-assisted reviews. AI-assisted reviews also had the highest rate of matching the original sign-out diagnosis, were less time-consuming, and were much easier for pathologists to perform (P < .001). Conclusions This study reports the successful development and clinical validation of an AI-based digital pathology system to screen for AFBs in anatomic pathology material. AI assistance proved to be more sensitive and accurate, took pathologists less time to screen cases, and was easier to use than either manual microscopy or viewing WSIs.

Recent genomic studies on the glioblastoma (GBM) subtypes (e.g., mesenchymal, proneural, and classical) pave a way for effective clinical treatments of the recurrent brain tumor. However, identification of the GBM subtype is complicated by the intratumoral heterogeneity that results in coexistence of multiple subtypes within the tissue specimen. Here, we present the use of hyperspectral stimulated Raman scattering (SRS) microscopy for rapid, label-free molecular assessment of GBM intratumoral heterogeneity with submicron resolution. We develop a unique label-free Raman imaging diagnostic platform consisting of the spectral focusing hyperspectral SRS imaging of the large-area GBM tissue specimens, SRS images, and spectrum retrieval using the multivariate curve resolution algorithm and subtype classification based on the quadratic support vector machine model for rapid molecular subtyping of GBMs. Both the stain-free SRS histological images and 2D subtype maps can be obtained within 20-30 min which is superior to the days of the conventional single-cell RNA sequencing. While the SRS histology assesses the demyelination status as a new diagnostic feature, the SRS mapping provides a new insight into intratumoral heterogeneity across GBM tissue specimens. We find that the major proportions of the GBM tissues agree with the diagnostic results of the genomic analysis, but nontrivial portions of the remaining SRS image tiles in the specimens are found to belong to other molecular subtypes, implying the substantial degree of GBM heterogeneity. The rapid SRS imaging diagnostic platform developed has shown the ability of unveiling tumor heterogeneity in GBM tissues accurately, which would promote the improvement of the GBM-targeted therapy in near future.

Purpose Although the fresh frozen (FF) cadaver is preferred for surgical applications, it is limited due to short usage time, unsuitable for reuse and the risk of infection. Due to its limited use, FF cadavers, which are covered by import in countries with insufficient body donation cause low-cost effectiveness. With the increase of real human tissue specimen necessities for surgical training, long-term preservation of the cadavers is crucial due to changes in mechanical properties. Therefore, studies on embalming solutions have increased in recent years. Methods We quantify the biomechanical properties of human parietal bones preserved via modified larssen solution (MLS) and compare the results with the specimens preserved as FF and fixed with 10% formalin-based solution (F10). The rectangular samples of 24 parietal bones of male individuals were resected from MLS-embalmed, F10-embalmed and FF cadavers to form three groups each containing eight samples. These specimens were tested longitudinally to identify mechanical properties. Results The tensile test results showed that there is not a significant difference between the groups in terms of stiffness, elastic modulus, strain at ultimate stress, failure strain and effective plastic strain. However, the yield stress, ultimate stress, yield strain, failure stress and total energy and post-yield properties are significantly lower in F10 than MLS and FF groups. Conclusion It is observed that the mechanical properties of MLS preserved and FF parietal bones have almost similar properties. Thus, it can be concluded that MLS is a suitable fixative solution for bone studies and bone-related surgical anatomy training applications.

AimsTo determine if a simple prewash step added to the processing workflow of tissue procurement by a core needle biopsy device will recover enough cells to expand the laboratory testing armamentarium.MethodsTissue was obtained from unfixed resection specimens using a core needle device and washed in a buffered solution before fixation. This creates a liquid aliquot from which dislodged cells can be kept and separated from the tissue specimen, the latter of which can then undergo traditional formalin-fixed, paraffin-embedded processing.ResultsCells dislodged from the tissue during the biopsy procedure are recoverable, are representative of the tissue section and of sufficient quantities for additional laboratory testing.ConclusionsThe core needle biopsy wash is an under-recognised and underutilised approach to extending the diagnostic capabilities of the limited amount of targeted material obtained during this common procedure. The ability to recover supplemental amounts of diagnostic material yields great potential as a substrate for a multitude of current and developing laboratory assays.

This study used large format autopsy tissue samples to compare radio-pathomic maps of brain cancer to a current tumor segmentation algorithm. We hypothesized that an MRI-based machine learning model trained with actual histology rather than radiologist annotations cellularity would 1) improve delineation between tumor and treatment effect, and 2) detect abnormal pathology beyond the contrast-enhancing tumor region. Seventeen patients with pathologically confirmed glioma were included in this study. At autopsy, 43 tissue samples were collected from 17 subjects from whole brain slices sectioned to align with the last axial MRI prior to death. Cellularity was calculated using a color deconvolution on 40X digitized H&E stained slides from the tissue samples. In-house custom software was used to align tissue samples and cellularity information to the FLAIR image using manually defined control points. The DeepMedic algorithm was trained to segment tumors using the BraTs 2017 dataset, and then applied to our patients in order to create automated tumor probability maps. An MRI-based ensemble algorithm using a 5x5 voxel searchlight (input: T1, T1C, FLAIR, ADC) was used to predict cellularity at each voxel, using tissue samples from 14 subjects as ground truth. Both models were applied to 3 withheld test subjects in order to compare tumor probability and cellularity predictions to the pathological ground truth. The mutual information between tumor probability and actual cellularity was 1X10-15, relatively low compared to the rad-path predicted cellularity (=0.16), despite the tumor prediction model accurately highlighting regions of contrast enhancement. Additionally, the radio-pathomic ensemble model correctly identify regions of hypercellularity beyond the tumor segmentation model as well as regions of within the segmented tumor area. This study demonstrates the utility of training machine learning models with pathological ground truth rather than radiologist annotations for predicting localized tumor information, particularly in the post-treatment stage.

Background In the era of personalized therapy liquid biopsy is considered an important diagnostic tool in the clinical management of cancer patients. Tissue specimen represents “gold standard” for molecular evaluation of specific gene targets alterations that lead cancer patients to benefit of a “tailed therapy” based on molecular features of the tumor. This innovative source of nucleic acids was introduced in clinical setting only for NSCLC patients to test Epidermal Grow Factor Receptor (EGFR) mutations when tissue is not available or to monitor acquired resistance mutation after a first line of treatment. The study aimed at assessing the diagnostic potential of liquid biopsy in balanced tertiary screening modeling. Methods The cases relating to 5 years of activity regarding to molecular diagnostics performed on liquid biopsy specimens in the Predictive Diagnostic laboratory of AOU Federico II were reviewed. Laboratory data were collected in SPSS. Non parametric analysis were performed in order to test the differences between patients WILD TYPE or not. A multivariate logistic model was performed in order to assess the effect of mutation, age and sex, on the tumor progression. The results of the revision concern 515 total cases (almost of all plasma or peripheral blood) allowed to evaluate the liquid biopsies for women and men. The average age of the Patients is 66.3 years, and the 25 percentile is 59 years. Results The cases are 221 basal and 294 by progression. The cases with mutation, as expected, have an OR 4,15 compared to the basal to have a tumor progression (95% IC: 2,7 - 6,3) regardless of sex and age. The mutations detected were 131 from different types of pulmonary carcinomas. Conclusions Working on case data, specifying the characteristics of the Patients with mutations will drive a further estimate in tertiary prevention screening designs. Key messages In the last five years, the liquid biopsy has been used for different purposes in oncology. To specify the characteristics of the Patients with mutations will drive a further estimate in tertiary prevention screening designs.

Fourier transform infrared spectroscopy has emerged as a powerful tool for tissue specimen investigation. Its nondestructive and label-free character enables direct determination of biochemical composition of samples. Furthermore, the introduction of polarization enriches this technique by the possibility of molecular orientation study apart from purely quantitative analysis. Most of the molecular orientation studies focused on polymer samples with a well-defined molecular axis. Here, a four-polarization approach for Herman’s in-plane orientation function and azimuthal angle determination was applied to a human tissue sample investigation for the first time. Attention was focused on fibrous tissues rich in collagen because of their cylindrical shape and established amide bond vibrations. Despite the fact that the tissue specimen contains a variety of molecules, the presented results of molecular ordering and orientation agree with the theoretical prediction based on sample composition and vibration directions.

At the heart of the phenome-to-genome approach is high throughput assays, which are liable to produce false results. This risk can be mitigated by minimizing the sample bias, specifically, recycling the same tissue specimen for both phenotypic and genotypic investigations. Therefore, our aim is to suggest a methodology of obtaining robust results from frozen specimens of compromised quality, particularly if the sample is produced in conditions with limited resources. For example, generating samples at the International Space Station (ISS) is challenging because the time and laboratory footprint allotted to a project can get expensive. In an effort to be economical with available resources, snap-frozen euthanized mice are the straightforward solution; however, this method increases the risk of temperature abuse during the thawing process at the beginning of the tissue collection. We found that prolonged immersion of snap frozen mouse carcass in 10% neutral buffered formalin at 4°C yielded minimal microscopic signs of ice crystallization and delivered tissues with histomorphology that is optimal for hematoxylin and eosin (H&E) staining and fixation on glass slides. We further optimized a method to sequester the tissue specimen from the H&E slides using an incubator shaker. Using this method, we were able to recover an optimal amount of RNA that could be used for downstream transcriptomics assays. Overall, we demonstrated a protocol that enables us to maximize scientific values from tissues collected in austere condition. Furthermore, our protocol can suggest an improvement in the spatial resolution of transcriptomic assays.

Background and Aims Vascular calcification (VC) is an important contributor to the high rate of cardiovascular mortality associated to chronic kidney disease. The inability to eliminate phosphorus (P) and the subsequent P retention promotes CV. P metabolism and uremic VC are influenced by obesity and by the caloric content of the diet. Caloric restriction (CR) has been shown to have multiple beneficial effects on health, for example, CR has been reported to improve vascular health and retard vascular ageing. However, to our knowledge the effect of CR on the development of uremic VC has not been explored. We hypothesize that CR may be beneficial to prevent the development of uremic VCs. Thus, the objective of the present study was to determine if rats subjected to CR were protected against VC. Method 48 Wistar rats were divided in four groups. The control diet provided Metabolizable Energy = 3.528 kcal/g and contained 0.6% Calcium (Ca) and 0.6% P. Additional diets of identical composition to the control diet but containing varying levels of Ca and P: 0.9% Ca, 0.9% P; 0.6% Ca, 1.2% P; and 0.9% Ca, 1.8% P, were also used in the experiments. Rats in Group 1 and 3 were fed 15 g/day of the control diet. Rats in Group 2 and 4 were calorie restricted and fed 10 g/day of diet with Ca/P = 0.9%/0.9%. Thus the daily P intake should be identical in the four groups. Uremia was induced by 5/6 nephrectomy (Nx). After Nx rats in Group 1 and 2 were fed ad libitum a diet with 0.6% Ca and 1.2% P. While rats in Group 3 and 4 were fed ad libitum a diet with 0.9% Ca and 1.8% P. Rats were supplemented with calcitriol. At the end of the experiment, rats were sacrificed to obtain blood samples and tissue samples (thoracic and abdominal aortas). After blood collection, plasma was separated by centrifugation and stored at –20° C until assayed. Plasma creatinine, Ca and P were measured by spectrophotometry. Energy intake was calculated based on food intake. VC was studied by histology and by measuring the tissue Ca content. Values are expressed as mean ± standard error (SE), the difference between groups was assessed by ANOVA. Fisher LSD test was used as a post-hoc procedure. p Results Before Nx, caloric intake was significantly lower in calorie restricted rats (35.4 ± 0.1 and 35.8 ± 0.1 kcal/day) than in rats eating normal calories (52.7 ± 0.1 and 52.8 ± 0.2 kcal/day); however, P intake was almost identical in the four groups and ranged between 89.8 and 91.6 mg/day. After Nx, rats in all groups reduced food intake and, consequently, caloric intake. Thus, although the P content of the diet was increased after Nx, daily P intake was not increased in Groups 1 and 2; however, P intake was significantly increased in Groups 3 and 4 (120.9 ± 4.6 and 122.2 ± 6.2 mg/day, respectively). In all groups, rats had high plasma concentrations of creatinine and P, and low plasma concentrations of Ca. Also, all rats had elevated Ca content in the aorta. No significant differences between the study groups were found in any of these parameters (Table 1). Von Kossa staining of the aortas showed abundant mineral deposition in the four groups. Conclusion This study shows that, contrary to what was expected, CR did not prevent or ameliorate uremic calcifications.

Background and Aims Percutaneous renal biopsy is the mainstay of renal pathological diagnosis. Not only does it secure an accurate diagnosis, but also lends itself to guiding the prognosis of a myriad of renal disorders. An important diagnostic modality for nephrologists, they have contributed greatly to the field of nephrology, both in terms of the classification of diseases and assisting in understanding their pathogenesis. Adequacy of the tissue samples of at least ten to fifteen glomeruli is generally preferred. The aim of our retrospective study was to review the renal biopsies done in one large UK renal centre, namely Salford Royal Foundation Trust, over the last twenty years. This Trust covers a catchment area that serves approximately 1.5 million people. Method A list of all renal biopsies performed at Salford Royal Foundation Trust between January 2000 to January 2020 was generated by our pathology department. Following this, each individual biopsy report was reviewed. In addition to this, certain information was collated from individual patient records; namely patient demographics, the indication for biopsy, tissue type (native or transplant), histological diagnosis and outcomes. Our centre has an excellent electronic patient record (EPR). This enabled us to analyse full clinical data retrospectively for each case. Results Between January 2000 and January 2020 a total of 3555 renal biopsies were performed at Salford Royal Foundation Trust. These biopsies were performed solely by nephrology specialists and supervised trainees, under ultrasound guidance. 177.5 biopsies were performed on average, per year, ranging between 140 – 203. 90% of biopsies were performed on native kidneys, with 10% being performed on transplanted kidneys. 10% of biopsies had normal histology morphologically when reviewed by light microscopy. There diagnoses of these included entities minimal change disease and thin basement membrane disease most commonly. IgA nephropathy was the most common glomerular pathology identified, in keeping with the wealth of published literature. There were over 525 biopsies done confirming IgAN as the primary pathology. 7% of biopsies had a crescentic glomerular pathology when examined histologically. The main causes included crescentic IgA nephropathy and anca-associated vasculitis. Conclusion Kidney biopsies have long been regarded as safe, minimally invasive procedures are of paramount importance to our diagnostic armamentarium. Our data echoes that of already published pathology series, IgAN being the most common glomerular pathology in our series. We further plan to analyse the various conditions and their outcomes in further detail.

Background: In India, musculoskeletal tuberculosis (TB) accounts for 10%–25% of extrapulmonary TB. Data on drug-resistant skeletal TB are lacking. At present, the diagnosis is based mainly on radiological techniques. Laboratory confirmation of skeletal TB is delayed as 6–8 weeks are required for culture results. Xpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) assay is a fully automated test which simultaneously detects MTB and RIF resistance within 3 h. Hence, this study was done to compare the yield of case detection using Xpert assay in comparison with culture in specimens received from clinically suspected skeletal TB cases. Methods: Retrospective analysis of microscopy, culture and Xpert assay results was carried out on specimens received in laboratory from skeletal TB cases from January 2016 to December 2017. Results: Of the 201 patients analysed, majority of the specimens were obtained from the spine (55.72%). MTB was detected in 48.68% of tissue and 24% of pus specimens. Xpert assay was detected MTB in 67 (33.33%) specimens of which 53 (47.32%) were from the spine. Culture was detected MTB in 66 (32.83%) specimens. Xpert assay was detected two specimens more than culture. One specimen was positive by only culture. RIF-resistant MTB was detected in 10 (14.92%) specimens by Xpert assay. Conclusion: The spine is the most common site involved. Tissue specimen is better for early diagnosis. High RIF resistance in skeletal TB is an alarming situation. Ability of Xpert MTB/RIF assay for rapid and simultaneous detection of MTB and RIF resistance in comparison with culture makes it a useful diagnostic tool in skeletal TB.

Targeted alpha therapy has the properties of direct ionization, oxygen independency, high linear energy transfer, a short half-life, and minimal depth of penetration, which are advantageous in cancer treatment. This modified, inexpensive, high-resolution, autoradiographic study of 232Th decay in a tissue specimen, to both locate and quantify individual alpha-particle origins, track lengths, and dose rates, utilized conventional Ilford Pan F Plus, 135–36, Black and White, 35 mm, ISO 50 film. Dose rates measured 0.14 cGy d−1 in the liver, 0.03 cGy d−1 in the right kidney, and 0.003 cGy d−1 in the heart, leading to a dosimetric concern of where the final alpha-particle deposition occurs.

Context.—Specimen misidentification is the most significant error in laboratory medicine, potentially accounting for hundreds of millions of dollars in extra health care expenses and significant morbidity in patient populations in the United States alone. New technology allows the unequivocal documentation of specimen misidentification or contamination; however, the value of this technology currently depends on suspicion of the specimen integrity by a pathologist or other health care worker.Objective.—To test the hypothesis that there is a detectable incidence of unsuspected tissue specimen misidentification among cases submitted for routine surgical pathology examination.Design.—To test this hypothesis, we selected specimen pairs that were obtained at different times and/or different hospitals from the same patient, and compared their genotypes using standardized microsatellite markers used commonly for forensic human DNA comparison in order to identify unsuspected mismatches between the specimen pairs as a trial of “molecular auditing.” We preferentially selected gastrointestinal, prostate, and skin biopsies because we estimated that these types of specimens had the greatest potential for misidentification.Results.—Of 972 specimen pairs, 1 showed an unexpected discordant genotype profile, indicating that 1 of the 2 specimens was misidentified. To date, we are unable to identify the etiology of the discordance.Conclusions.—These results demonstrate that, indeed, there is a low level of unsuspected tissue specimen misidentification, even in an environment with careful adherence to stringent quality assurance practices. This study demonstrates that molecular auditing of random, routine biopsy specimens can identify occult misidentified specimens, and may function as a useful quality indicator.

AimsIn the diagnostic or research field, there is a need that the histopathological evaluation provides as much information as possible, with the shortest time and lowest cost. The development of the tissue microarray (TMA) technology has significantly facilitated and accelerated studies with tissue analyses using in situ technologies. Nevertheless, one of the most recognised limitations of TMA is that the small cores used to construct a TMA may not accurately represent characteristics of the whole tissue specimen. The aim is to present the technical aspects of a simple system to construct tissue macroarrays (TMaAs), its advantages and limitations, and some results of its use in for research purposes.MethodsThe study presents three possibilities of preparing the specimens with up to 6 mm diameter to be included into the TMaA paraffin block using our method.ResultsWith this technique, it was possible to obtain glass slides with multiple whole fragments, which were used in five different studies.ConclusionsWe presented a method for construction of multi-TMaA paraffin blocks, which is very simple for optimising laboratory techniques requiring paraffin-embedded tissue sections and can be easily implemented in any institution with a histology laboratory.

Background Intestinal Behçet’s disease (BD) and Crohn’s disease (CD) present similar manifestations, but there are no specific pathognomonic clinical, laboratory, or histological diagnostic tests to differentiate intestinal BD from CD. We used a proteomic approach to discover a novel diagnostic biomarker specific to intestinal BD. Methods The colon mucosa tissue samples were obtained using colonoscopy-guided biopsy of the affected bowel from patients with intestinal BD or CD at the Inflammatory Bowel Disease Clinic of Severance Hospital, Seoul, Korea. Peptides from seven intestinal BD and seven CD patients were extracted and labeled using tandem mass tag (TMT) reagents. The labeled peptides were identified and quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The differentially expressed proteins were further validated using immunohistochemical (IHC) analysis with tissue samples and an ELISA test with serum samples from 20 intestinal BD and 20 CD patients. Results A total of 3,266 proteins were identified using TMT/LC-MS/MS-based proteomic quantification, including 39 candidate proteins differentially expressed between intestinal BD and CD. Beta-2 glycoprotein 1 (APOH) and maltase-glucoamylase (MGAM) showed significantly higher intensity in the IHC staining of the intestinal BD tissues than that of CD tissues. Furthermore, the serum MGAM level was significantly higher in patients with intestinal BD than in patients with CD. Conclusion Our proteomic analysis revealed that some proteins were differentially expressed in intestinal BD patients compared to CD patients. Differential MGAM expression in intestinal BD suggests its role as a potential novel diagnostic biomarker in the differentiation of intestinal BD from CD.