Miracle-dz/UniFAR-FTUs huggingface dataset:
Clinical-Grade Human Pluripotent Stem Cells for Cell Therapy: Characterization Strategy

Human pluripotent stem cells have the potential to change the way in which human diseases are cured. Clinical-grade human embryonic stem cells and human induced pluripotent stem cells have to be created according to current good manufacturing practices and regulations. Quality and safety must be of the highest importance when humans’ lives are at stake. With the rising number of clinical trials, there is a need for a consensus on hPSCs characterization. Here, we summarize mandatory and ′for information only′ characterization methods with release criteria for the establishment of clinical-grade hPSC lines.
From Bivariate to Multivariate Analysis of Cytometric Data: Overview of Computational Methods and Their Application in Vaccination Studies

Flow and mass cytometry are used to quantify the expression of multiple extracellular or intracellular molecules on single cells, allowing the phenotypic and functional characterization of complex cell populations. Multiparametric flow cytometry is particularly suitable for deep analysis of immune responses after vaccination, as it allows to measure the frequency, the phenotype, and the functional features of antigen-specific cells. When many parameters are investigated simultaneously, it is not feasible to analyze all the possible bi-dimensional combinations of marker expression with classical manual analysis and the adoption of advanced automated tools to process and analyze high-dimensional data sets becomes necessary. In recent years, the development of many tools for the automated analysis of multiparametric cytometry data has been reported, with an increasing record of publications starting from 2014. However, the use of these tools has been preferentially restricted to bioinformaticians, while few of them are routinely employed by the biomedical community. Filling the gap between algorithms developers and final users is fundamental for exploiting the advantages of computational tools in the analysis of cytometry data. The potentialities of automated analyses range from the improvement of the data quality in the pre-processing steps up to the unbiased, data-driven examination of complex datasets using a variety of algorithms based on different approaches. In this review, an overview of the automated analysis pipeline is provided, spanning from the pre-processing phase to the automated population analysis. Analysis based on computational tools might overcame both the subjectivity of manual gating and the operator-biased exploration of expected populations. Examples of applications of automated tools that have successfully improved the characterization of different cell populations in vaccination studies are also presented.
A fatal case of COVID-19 pneumonia occurring in a patient with severe acute ulcerative colitis

Since December 2019, a novel coronavirus referred to as SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2)1 was identified as the causative agent of a respiratory syndrome named COVID-19.2 Since February 2020, an outbreak of COVID-19 has developed in Lombardy, in northern Italy, posing serious challenges to healthcare system. As reported, COVID-19 severity directly correlates with patient’s age and several comorbidities,3 but no data are currently available regarding clinical outcome in inflammatory bowel diseases (IBD). Recently, the International Organization for the Study of Inflammatory Bowel Disease (IOIBD) recommended a reduction of corticosteroid therapy in IBD patients, while maintaining thiopurines and biologics,4 even if Authors recognised that specific IBD research on COVID-19 is lacking. On 12 February 2020, days before this outbreak was recognised, an 80-year-old female with a 3-year history of left-sided ulcerative colitis (UC), in maintenance with mesalamine, …
Tailoring translational strength using Kozak sequence variants improves bispecific antibody assembly and reduces product‐related impurities in CHO cells

Optimal production of bispecific antibodies (bsAb) requires efficient and tailored co‐expression and assembly of two distinct heavy and two distinct light chains. Here, we describe a novel technology to modulate the translational strength of antibody chains via Kozak sequence variants to produce bsAb in a single cell line. In this study, we designed and screened a large Kozak sequence library to identify 10 independent variants that can modulate protein expression levels from approximately 0.2 to 1.3‐fold compared with the wild‐type sequence in transient transfection. We used a combination of several of these variants, covering a wide range of translational strength, to develop stable single cell Chinese hamster ovary bispecific cell lines and compared the results with those obtained from the wild‐type sequence. A significant increase in bispecific antibody assembly with a concomitant reduction in the level of product‐related impurities was observed. Our findings suggest that for production of bsAb it can be advantageous to modify translational strength for selected protein chains to improve overall yield and product quality. By extension, tuning of translational strength can also be applied to improving the production of a wide variety of heterologous proteins.
Evaluation of Addition of Intravenous Metronidazole to Oral Vancomycin Therapy in Critically Ill Patients with Non‐Fulminant Severe Clostridioides difficile Infection

Data on the impact of combination therapy (intravenous metronidazole [IV MTZ] plus oral vancomycin [PO VAN]) on clinical outcomes in intensive care unit (ICU) patients with severe non‐fulminant Clostridioides difficile infection (CDI), including NAP1‐positive samples, are lacking.
The Structure, Vibrational Spectra, and Thermal Expansion Study of AVO4 (A=Bi, Fe, Cr) and Co2V2O7

Vanadate is an important functional material. It has been widely studied and applied in luminescence and photocatalysis. Vanadium compounds have been synthesized to investigate the thermal expansion properties and structure. Both BiVO4 and Co2V2O7 are monoclinic at room temperature, FeVO4’s crystal structure is triclinic, and CrVO4 is orthorhombic. The relatively linear, thermal-expansion, and temperature-dependent Raman spectroscopy results showed that the phase transition of BiVO4 occurred at 200 to 300 °C. The coefficient of thermal expansion (CTE) of Co2V2O7 was larger than that of the monoclinic structure BiVO4. The CTE of the tetragonal structure of BiVO4 was 15.27 × 10−6 °C−1 which was the largest CTE in our measurement results, and the CTE of anorthic structure FeVO4 was 2.84 × 10−6 °C−1 and was the smallest.
Salmonella in Raptors and Aquatic Wild Birds in Chile

Abstract: Salmonella enterica is one of the main causes of gastrointestinal disease worldwide. Wild birds are capable of harboring a variety of Salmonella serovars, which could have an important role in the epidemiology of salmonellosis in humans and production animals. We tested 519 fecal samples from raptors and aquatic birds from different regions of central (three rehabilitation centers for wildlife and the coastal area) and southern areas of Chile for Salmonella. All samples were obtained in 2015 and 2017, covering all four seasons. Salmonella was isolated from 12 of the 519 samples (2%) analyzed, from two carnivorous birds, four birds with generalist habits, and six waterfowl. Among the isolates obtained, one showed resistance to gentamicin, and one showed a multidrug-resistance phenotype, with resistance to ampicillin, ceftriaxone, ciprofloxacin, chloramphenicol, streptomycin, gentamicin, kanamycin, trimethoprim-sulfamethoxazole, and tetracycline. These results demonstrated the importance of characterizing Salmonella in wild birds because previous studies have shown genetic and phenotypic evidence suggesting interspecies transmission of Salmonella enterica that is resistant to antimicrobials between humans and wild and domestic birds.
Gout. Diagnosis and its association with cardiovascular diseases

Objectives The gold standard for diagnosing gout is the identification of monosodium urate (MSU) crystals in joint fluid. In secondary care the facilities or expertise to analyze joint fluid are not always available, and gout is diagnosed clinically. To improve the predictive value of the clinical diagnosis of gout in secondary care, a diagnostic rule developed in primary care could be helpful. The aim of this study was to validate this diagnostic rule in a secondary care population with the gold standard as reference test. Methods 390 patients with monoarthritis were included. The variables of the diagnostic rule (male sex, previous arthritis attack, onset <1 day, joint redness, involvement of the first metatarsophalangeal joint, hypertension or one or more cardiovascular disease, and serum uric acid >5.88 mg/dl) were collected and scored. The affected joint was aspirated and joint fluid was analyzed for the presence of MSU crystals. Results In 219 patients (56%) MSU crystals were found. The positive predictive value of a score of ≥8 points was 0.87, the negative predictive value of a score of ≤4 points was 0.95. The area under the receiver operator characteristic curve for the diagnostic rule was 0.86 (95% confidence interval 0.82-0.89). The Hosmer-Lemeshows’ goodness-of-fit test showed that the difference between the expected and the observed probability was non-significant (P=0.64), indicating good agreement. Conclusions An easy-to-use diagnostic rule for gout developed in primary care shows a good performance in secondary care and improves the predictive value of the clinical diagnosis of gout when joint fluid analysis is not available.
Pre‐analytical factors affecting the establishment of a single tube assay for multiparameter liquid biopsy detection in melanoma patients

The combination of liquid biomarkers from a single blood tube can provide more comprehensive information on tumor development and progression in cancer patients compared to single analysis. Here, we evaluated whether a combined analysis of circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), and circulating cell‐free microRNA (miRNA) in total plasma and extracellular vesicles (EV) from the same blood sample is feasible and how the results are influenced by the choice of different blood tubes. Peripheral blood from 20 stage IV melanoma patients and five healthy donors (HD) was collected in EDTA, Streck, and Transfix tubes. Peripheral blood mononuclear cell fraction was used for CTC analysis, whereas plasma and EV fractions were used for ctDNA mutation and miRNA analysis. Mutations in cell‐free circulating DNA were detected in 67% of patients, with no significant difference between the tubes. CTC was detected in only EDTA blood and only in 15% of patients. miRNA NGS (next‐generation sequencing) results were highly influenced by the collection tubes and could only be performed from EDTA and Streck tubes due to hemolysis in Transfix tubes. No overlap of significantly differentially expressed miRNA (patients versus HD) could be found between the tubes in total plasma, whereas eight miRNA were commonly differentially regulated in the EV fraction. In summary, high‐quality CTCs, ctDNA, and miRNA data from a single blood tube can be obtained. However, the choice of blood collection tubes is a critical pre‐analytical variable.
Complement Component C3 Participates in Early Stages of Niemann–Pick C Mouse Liver Damage

Niemann–Pick type C (NPC), a lysosomal storage disorder, is mainly caused by mutations in the NPC1 gene. Niemann–Pick type C patients and mice show intracellular cholesterol accumulation leading to hepatic failure with increased inflammatory response. The complement cascade, which belongs to the innate immunity response, recognizes danger signals from injured tissues. We aimed to determine whether there is activation of the complement system in the liver of the NPC mouse and to assess the relationship between C3 activation, a final component of the pathway, and NPC liver pathology. Niemann–Pick type C mice showed high levels of C3 staining in the liver which unexpectedly decreased with aging. Using an inducible NPC1 hepatocyte rescue mouse model, we restored NPC1 expression for a short time in young mice. We found C3 positive cells only in non-rescued cells, suggesting that C3 activation in NPC cells is reversible. Then, we studied the effect of C3 ablation on NPC liver damage at two postnatal time points, P56 and P72. Deletion of C3 reduced the presence of hepatic CD68-positive cells at postnatal day 56 and prevented the increase of transaminase levels in the blood of NPC mice. These positive effects were abrogated at P72, indicating that the complement cascade participates only during the early stages of liver damage in NPC mice, and that its inhibition may serve as a new potential therapeutic strategy for the disease.
Experimental Investigations of Timber Beams with Stop-Splayed Scarf Carpentry Joints

The paper presents the results of an experimental investigation of stop-splayed scarf joints, which was carried out as part of a research programme at the Wroclaw University of Science and Technology. A brief description and the characteristics of scarf and splice joints appearing in historical buildings are provided, with special reference to stop-splayed scarf joints (so-called ‘Bolt of lightning’) which were widely used, for example, in Italian renaissance architecture. Analyses and studies of scarf and splice joints in bent elements presented in the literature are reviewed, along with selected examples of analyses and research on tensile joints. It is worth noting that the authors in practically all the cited literature draw attention to the need for further research in this area. Next, the results of the authors’ own research on beams with stop-splayed scarf joints, strengthened using various methods, e.g., by means of drawbolts (metal screws), steel clamps and steel clamps with wooden pegs, which were subjected to four-point bending tests are presented. Load-deflection plots were obtained for load-bearing to bending of each beam in relation to the load-bearing of a continuous reference beam. A comparative analysis of the results obtained for each beam series is presented, along with conclusions and directions for further research.
Cytokines and Growth Factors as Predictors of Response to Medical Treatment in Diabetic Macular Edema

Diabetic macular edema (DME) is the most common cause of visual loss in patients with diabetes. Antivascular endothelial growth factors (anti-VEGF) agents are first-line therapy for DME. Nevertheless, up to 60% of patients (depending on the anti-VEGF drug used) have an inadequate response to anti-VEGF treatment. Several cytokines are increased in aqueous humor of patients with DME. Differences in response to treatment may be related to baseline cytokine levels. Intravitreal corticosteroids may be used as an alternative to anti-VEGF agents. Steroids have a different pharmacological profile and act on different pathophysiologic mechanisms. Their effect on aqueous humor cytokines is different from the effect of anti-VEGF therapy. This review highlights the major cytokines involved in DME and evaluates whether baseline cytokine levels could be predictors of response to treatment in DME. SIGNIFICANCE STATEMENT Antivascular endothelial growth factor (anti-VEGF) agents are efficient as diabetic macular edema (DME) treatment. However, in some cases, DME fails to respond to anti-VEGF intravitreal injections. Changes in cytokine levels after treatment supported the idea that other cytokines than VEGF are implicated in DME pathogenesis and could be predictors of response to anti-VEGF treatment or corticosteroids allowing targeted and individualized therapy guided by cytokine levels.
In Vitro and In Vivo Characterization of Dibenzothiophene Derivatives [125I]Iodo-ASEM and [18F]ASEM as Radiotracers of Homo- and Heteromeric α7 Nicotinic Acetylcholine Receptors

The α7 nicotinic acetylcholine receptor (α7 nAChR) is involved in several cognitive and physiologic processes; its expression levels and patterns change in neurologic and psychiatric diseases, such as schizophrenia and Alzheimer’s disease, which makes it a relevant drug target. Development of selective radioligands is important for defining binding properties and occupancy of novel molecules targeting the receptor. We tested the in vitro binding properties of [125I]Iodo-ASEM [(3-(1,4-diazabycyclo[3.2.2]nonan-4-yl)-6-(125I-iododibenzo[b,d]thiopentene 5,5-dioxide)] in the mouse, rat and pig brain using autoradiography. The in vivo binding properties of [18F]ASEM were investigated using positron emission tomography (PET) in the pig brain. [125I]Iodo-ASEM showed specific and displaceable high affinity (~1 nM) binding in mouse, rat, and pig brain. Binding pattern overlapped with [125I]α-bungarotoxin, specific binding was absent in α7 nAChR gene-deficient mice and binding was blocked by a range of α7 nAChR orthosteric modulators in an affinity-dependent order in the pig brain. Interestingly, relative to the wild-type, binding in β2 nAChR gene-deficient mice was lower for [125I]Iodo-ASEM (58% ± 2.7%) than [125I]α-bungarotoxin (23% ± 0.2%), potentially indicating different binding properties to heteromeric α7β2 nAChR. [18F]ASEM PET in the pig showed high brain uptake and reversible tracer kinetics with a similar spatial distribution as previously reported for α7 nAChR. Blocking with SSR-180,711 resulted in a significant decrease in [18F]ASEM binding. Our findings indicate that [125I]Iodo-ASEM allows sensitive and selective imaging of α7 nAChR in vitro, with better signal-to-noise ratio than previous tracers. Preliminary data of [18F]ASEM in the pig brain demonstrated principal suitable kinetic properties for in vivo quantification of α7 nAChR, comparable to previously published data.
Cognition in Children with Arachnoid Cysts

Background: This study aims to evaluate if children with temporal arachnoid cysts (AC) have cognitive symptoms and if neurosurgery improves these. Methods: A prospective case series study including consecutive pediatric patients with temporal AC. The children underwent neuroradiology, neuroopthalmologic evaluation, and a standard electroencephalography (EEG). Additionally, a neuropsychologist performed a standardized set of evaluations, with a one-year follow-up consisting of Weschler Intelligence Scale for Children version IV (WISC-IV), FAS (for verbal fluency), Boston Naming Test (BNT, for visual naming ability) and NEPSY-II (Developmental NEuroPSYchological Assessment) for verbal memory. Results: Fifteen children, 9 boys and 6 girls, were evaluated and 11 underwent surgery. The Full Scale IQ subscore (FSIQ) improved from M = 84.8 to M = 93.0 (p = 0.005). The preoperative Verbal Comprehension Index (VCI) was in the low average range (M = 86.7), improving to a level within the average range (M = 94.7, p = 0.001). Preoperative Perceptual Speed Index (PSI) was in the below average range (M = 81.5), improving to a level within the average range (M = 92.5, p = 0.004). Conclusion: ACs are a common finding in a pediatric neurosurgical setting. Our data suggest that some temporal AC have a negative effect on general cognitive ability and that this impairment can be improved by surgery. We suggest a standardized evaluation, including comprehensive and validated neuropsychological assessment tools, to thoroughly assess symptoms as well as the postoperative outcome.
Disability among patients with opioid use disorders and its relationship with stigma toward substance use

Background: From a medical model perspective of substance use disorders (SUDs), opioid use disorders are associated with some degree of disability. This study aimed to assess the disability among patients with opioid use disorders (OUDs) and its relationship with internalized stigma. Methodology: This cross-sectional study assessed patients with SUDs at a tertiary care center. Disability among patients with OUDs was assessed using Indian Disability Evaluation and Assessment Scale (IDEAS) while stigma was measured using Internalized Stigma of Mental Illness Scale (ISMIS). Results: Among 168 patients with OUD, the disability was greatest in the domain of work followed by interpersonal activities. About 48.2% and 3.0% of the participants had moderate and severe disability according to IDEAS. Disability (IDEAS) scores had a robust correlation with the stigma (ISMIS) scores (r = 0.453, P < 0.01). Multiple regression analysis found that internalized stigma (ISMIS score) was an independent predictor of disability among patients with OUDs (β =0.42, P < 0.01). Conclusions: From a medical perspective, OUDs are associated with considerable disability which has significant correlation with internalized stigma. Designing interventions which can target internalized stigma among patients with OUD may help in reducing the disability associated with it.
Analysis of Predisposing Factors for Rapid Dental Calculus Formation

Background: Calculus accumulation varies widely between individuals. Dental calculus has been associated with the principal periodontal diseases. The aim of this study was to analyze individual characteristics, and salivary and microbiological parameters among patients considered to be rapid calculus formers and patients who form calculus slowly. Methods: Individual characteristics were recorded in a sample of 74 patients (age, sex, smoking, periodontal diagnosis, and dental crowding), as well as salivary parameters (unstimulated saliva flow, pH, and biochemical analysis of saliva) and microbiological parameters (by means of semi-quantitative polymerase chain reaction (PCR) analysis). Results: A statistically significant association (p = 0.002) was found between the rate of calculus formation and the diagnosis of periodontal disease. A greater presence of dental crowding was observed among the group of rapid calculus formers. Urea and phosphorus levels were higher among rapid calculus formers. Regarding microbiological parameters, differences were found in Streptococcus mutans, this being higher in the group of slow formers. Conclusions: Rapid calculus formation appears to be linked to patients diagnosed with more severe periodontal diseases. Rapid calculus-forming patients present more dental crowding and a lower proportion of S. mutans.