CHEK2 germline variants identified in familial nonmedullary thyroid cancer lead to impaired protein structure and function.

Approximately 5 to 15% of nonmedullary thyroid cancers (NMTC) present in a familial form (familial nonmedullary thyroid cancers [FNMTC]). The genetic basis of FNMTC remains largely unknown, representing a limitation for diagnostic and clinical management. Recently, germline mutations in DNA repair-related genes have been described in cases with thyroid cancer (TC), suggesting a role in FNMTC etiology. Here, two FNMTC families were studied, each with two members affected with TC. Ninety-four hereditary cancer predisposition genes were analyzed through next-generation sequencing, revealing two germline CHEK2 missense variants (c.962A > C, p.E321A and c.470T > C, p.I157T), which segregated with TC in each FNMTC family. p.E321A, located in the CHK2 protein kinase domain, is a rare variant, previously unreported in the literature. Conversely, p.I157T, located in CHK2 forkhead-associated domain, has been extensively described, having conflicting interpretations of pathogenicity. CHK2 proteins (WT and variants) were characterized using biophysical methods, molecular dynamics simulations, and immunohistochemistry. Overall, biophysical characterization of these CHK2 variants showed that they have compromised structural and conformational stability and impaired kinase activity, compared to the WT protein. CHK2 appears to aggregate into amyloid-like fibrils in vitro, which opens future perspectives toward positioning CHK2 in cancer pathophysiology. CHK2 variants exhibited higher propensity for this conformational change, also displaying higher expression in thyroid tumors. The present findings support the utility of complementary biophysical and in silico approaches toward understanding the impact of genetic variants in protein structure and function, improving the current knowledge on CHEK2 variants' role in FNMTC genetic basis, with prospective clinical translation.

Pembrolizumab-induced type 1 diabetes.

INTRODUCTION: Immunotherapy has a crucial role in the current treatment of multiple malignancies. Albeit described as rare, new onset autoimmune diabetes is a potentially life-threatening complication of programmed cell death-1 (PD-1) inhibitors, such as pembrolizumab, and its predisposing factors and pathological mechanism are yet to be clarified. CASE REPORT: We present a case of a 72-year-old man with a high-grade bladder carcinoma undergoing pembrolizumab treatment. He had no personal or family history of diabetes mellitus but was diagnosed with primary hypothyroidism four months after starting pembrolizumab. Two years after starting pembrolizumab, he presented in the emergency department due to abdominal pain, anorexia, polydipsia, polyuria and vomiting over the preceding five days and he met criteria for severe diabetic ketoacidosis (DKA). Three days prior to his admission, he had received prednisolone therapy for suspected hypersensitivity related to a contrast-enhanced imaging that he performed. MANAGEMENT & OUTCOME: Prompt treatment for DKA was started, with transition to insulin basal-bolus therapy after DKA resolution, with progressive glycaemic stabilization. Further investigation revealed low C-peptide levels (0.07 ng/dL, with a fasting blood glucose of 288 mg/dL), HbA1c 9.2% and positive anti-IA2 antibodies, which allowed the diagnosis of new-onset autoimmune diabetes. Pembrolizumab was transiently suspended, and the patient resumed treatment after glycaemic profile optimization under multiple daily insulin administrations two months later. DISCUSSION: This case highlights the importance of clinical suspicion and glycaemic monitoring as an integral part of treatment protocols in patients on pembrolizumab and other immune checkpoint inhibitors. Additional research and investigation into the underlying mechanisms of this condition are necessary to identify potential screening tests for individuals at higher risk of developing DM and to guide the implementation of management and preventive strategies for ketoacidosis complication.

Impact of enhancement filters of a CMOS system on halo artifact expression at the bone-to-implant interface.

OBJECTIVE: To assess the impact of enhancement filters on the formation of halo artifacts in radiographs of dental implants obtained with a complementary metal oxide semiconductor (CMOS) system. METHODS: Digital radiographs of dental implants placed in dry human mandibles were processed with the Noise Reduction smoothing filter, as well as the Sharpen 1, Sharpen 4, and Sharpen UM high-pass filters available in the CLINIVIEW™ software (Instrumentarium Dental, Tuusula, Finland). Subjective analysis involved evaluating the left, right, and apical surfaces of each implant for the presence of much, few, or no halo. The objective analysis involved measurement of the halo area using the Trainable Weka Segmentation plugin (ImageJ, National Institutes of Health, Bethesda, MD, USA). Data were analyzed using Friedman's test (subjective analysis) and ANOVA (objective analysis) (α = 5%). RESULTS: In the subjective evaluation, the Sharpen 4 filter produced more radiographs with much halo present, and in the objective evaluation, a bigger halo area when compared to the original images and the Noise Reduction filter for all surfaces (p < 0.05). CONCLUSIONS: When evaluating dental implants, priority should be given to original images and those enhanced with smoothing filters since they exhibit fewer halo artifacts. CLINICAL RELEVANCE: Post-processing tools, such as enhancement filters, may improve the image quality and assist some diagnostic tasks. However, little is known regarding the impact of enhancement filters in halo formation on CMOS systems, which have been increasingly used in dental offices.

Efficacy of a radiographic film holder adapter in generating radiographs of dental implants with improved geometric accuracy and sharpness.

STATEMENT OF PROBLEM: Some radiographic film holders produce radiographs with geometric distortion that may interfere with diagnosis. However, whether the distortion can be corrected by adjusting the design of the radiographic film holder is unclear. PURPOSE: The purpose of the study was to develop an adapter for a radiographic film holder model aiming to generate radiographs with greater sharpness and a more accurate geometric representation of dental implants. MATERIAL AND METHODS: The 2-piece adapter was designed using the SketchUp software program and was 3-dimensionally (3D) printed. Implants with internal conical connections were installed in 19 maxillary prototypes in the central incisor region. Five dentists obtained 285 digital periapical radiographs with 3 different radiographic film holders: standard Cone Indicator, Rinn XCP, and adapted Cone Indicator. They then evaluated the radiographic sharpness of the implants threads and their dimensions using the ImageJ software program. The data were analyzed using the Friedman test with the Durbin-Conover post hoc test and MANOVA with the Tukey post hoc test (α=.05). RESULTS: On the mesial surface of the implants, the threads were sharper for the adapted than for the standard Cone Indicator radiographic film holder (P<.05). The adapted Cone Indicator showed a smaller difference between the radiographic and actual implant diameters compared with the Rinn XCP and standard Cone Indicator radiographic film holders (P<.05). CONCLUSIONS: The developed adapter provided radiographs of dental implants with improved sharpness and geometric accuracy.

Managing Allergic Nickel Dermatitis in Occupational Settings: A Case Report.

Contact dermatitis is a common cutaneous inflammatory condition, triggered by exposure to irritant substances or allergens. Nickel is the most prevalent allergen, a metal widely used in accessories, furniture, office materials, food and in industry, with multiple exposure pathways, making it difficult to assess which exposure is causing allergic dermatitis. Here, we report a case of an administrative worker with chronic hand eczema, limited to the radial metacarpophalangeal region of the left hand, caused by occupational exposure to nickel, confirmed by nickel deposition test on the hand and a positive test with a metallic stapler used at her workplace.

Biometric Recognition: A Systematic Review on Electrocardiogram Data Acquisition Methods.

In the last decades, researchers have shown the potential of using Electrocardiogram (ECG) as a biometric trait due to its uniqueness and hidden nature. However, despite the great number of approaches found in the literature, no agreement exists on the most appropriate methodology. This paper presents a systematic review of data acquisition methods, aiming to understand the impact of some variables from the data acquisition protocol of an ECG signal in the biometric identification process. We searched for papers on the subject using Scopus, defining several keywords and restrictions, and found a total of 121 papers. Data acquisition hardware and methods vary widely throughout the literature. We reviewed the intrusiveness of acquisitions, the number of leads used, and the duration of acquisitions. Moreover, by analyzing the literature, we can conclude that the preferable solutions include: (1) the use of off-the-person acquisitions as they bring ECG biometrics closer to viable, unconstrained applications; (2) the use of a one-lead setup; and (3) short-term acquisitions as they required fewer numbers of contact points, making the data acquisition of benefit to user acceptance and allow faster acquisitions, resulting in a user-friendly biometric system. Thus, this paper reviews data acquisition methods, summarizes multiple perspectives, and highlights existing challenges and problems. In contrast, most reviews on ECG-based biometrics focus on feature extraction and classification methods.

Effect of Dietary Incorporation of Yellow Mealworm as a Partial Fishmeal Replacer on Growth, Metabolism, and Intestinal Histomorphology in Juvenile Meagre (Argyrosomus regius).

Efforts have been made to find alternatives to fish meal (FM), as the sustainability of aquaculture depends on it. Insect meal (IM) is a potential candidate to partially replace FM, being more sustainable and economically viable. In this experimental trial, three diets were tested with different yellow mealworm incorporation: a control diet with no IM, a diet with an inclusion of 10% IM (Ins10), and a diet with an incorporation of 20% IM (Ins20). The diets were tested on 10.5 g meagre for 47 days. The results showed that an IM inclusion higher than 10% affected both growth (2.6 vs. 2.2) and FCR (1.5 vs. 1.9) of meagre juveniles. However, this reduction in growth did not result from lower protein retention or changes in muscle fibre area or density. Little differences were observed in the activity of pancreatic and intestinal enzymes except for aminopeptidase total activity which was higher in the control and Ins10 compared to Ins20 (3847 vs. 3540 mU/mg protein), suggesting no limitations in protein synthesis. Also, the alkaline phosphatase intestinal maturation index was higher in the control group compared to the IM groups (437 vs. 296). On the contrary, several differences were also found in the proteolytic activity in the hepatic and muscle tissues of meagre juveniles fed the Ins10 diet. The inclusion of IM had no impact on intestine histomorphology but changes were detected in the enterocytes of fish from control and Ins10 which showed hypervacuolization and nucleus misplacement compared to the Ins20 treatment. Nevertheless, a higher percentage of Vibrionaceae was recorded for meagre fed on the Ins20 diet. Since no signs of inflammation were observed in the distal intestine, this suggests IM incorporation could have had an important impact on intestinal health due to its antimicrobial properties. This is supported by an increase in the haematocrit in the treatments where IM was added (20 to 25%). In conclusion, incorporations of IM at percentages up to 10% do not seem to have a negative impact on meagre performance at this age but can enhance the fish immune system and protection against intestinal inflammation.

Initial Study Using Electrocardiogram for Authentication and Identification.

Recently, several studies have demonstrated the potential of electrocardiogram (ECG) to be used as a physiological signature for biometric systems (BS). We investigated the potential of ECG as a biometric trait for the identification and authentication of individuals. We used data from a public database, CYBHi, containing two off-the-person records from 63 subjects, separated by 3 months. For the BS, two templates were generated: (1) cardiac cycles (CC) and (2) scalograms. The identification with CC was performed with LDA, kNN, DT, and SVM, whereas a convolutional neural network (CNN) and a distance-based algorithm were used for scalograms. The authentication was performed with a distance-based algorithm, with a leave-one-out cross validation, for impostors evaluation. The identification system yielded accuracies of 79.37% and 69.84% for CC with LDA and scalograms with CNN, respectively. The authentication yielded an accuracy of 90.48% and an impostor score of 13.06% for CC, and it had an accuracy of 98.42% and an impostor score of 14.34% for scalograms. The obtained results support the claim that ECG can be successfully used for personal recognition. To the best of our knowledge, our study is the first to thoroughly compare templates and methodologies to optimize the performance of an ECG-based biometric system.

Plant-based diets fed to juvenile meagre Argyrosomus regius with low methionine and taurine supplementation led to an overall reduction in fish performance and to an increase in muscle fibre recruitment.

Methionine and taurine are amino acids (AA) that are usually deficient when fish meal is replaced by plant proteins. In this study, three diets were tested in juvenile meagre (initial weight: 13.4 g) for 8 weeks. The D1 diet had 0.2% methionine and 1% taurine supplementation; the D2 and D3 diets had 0.6% methionine and 1% and 2% taurine supplementation, respectively. The results showed that meagre fed the D1 diet had lower specific growth rate (2.2 to 2.5), lower feed efficiency (0.9 to 1.2) and higher food conversion rate (FCR, 1.1 to 0.8) as well as a lower activity of the alanine aminotransferase (ALAT) enzyme. Furthermore, a higher recruitment of muscle fibres (46% compared to 36%) as well as a higher fibre density was observed (1019 compared to 870 fibres mm-2 ). This study shows that meagre requires a sufficient quantity of methionine in plant-based diets to avoid a reduction in fish performance. Furthermore, taurine supplementation in the D1 diet was not able to mitigate the effects of methionine deficiency. A higher taurine supplementation did not improve meagre performance.

Dehydropeptide Supramolecular Hydrogels and Nanostructures as Potential Peptidomimetic Biomedical Materials.

Supramolecular peptide hydrogels are gaining increased attention, owing to their potential in a variety of biomedical applications. Their physical properties are similar to those of the extracellular matrix (ECM), which is key to their applications in the cell culture of specialized cells, tissue engineering, skin regeneration, and wound healing. The structure of these hydrogels usually consists of a di- or tripeptide capped on the N-terminus with a hydrophobic aromatic group, such as Fmoc or naphthalene. Although these peptide conjugates can offer advantages over other types of gelators such as cross-linked polymers, they usually possess the limitation of being particularly sensitive to proteolysis by endogenous proteases. One of the strategies reported that can overcome this barrier is to use a peptidomimetic strategy, in which natural amino acids are switched for non-proteinogenic analogues, such as D-amino acids, ß-amino acids, or dehydroamino acids. Such peptides usually possess much greater resistance to enzymatic hydrolysis. Peptides containing dehydroamino acids, i.e., dehydropeptides, are particularly interesting, as the presence of the double bond also introduces a conformational restraint to the peptide backbone, resulting in (often predictable) changes to the secondary structure of the peptide. This review focuses on peptide hydrogels and related nanostructures, where α,ß-didehydro-α-amino acids have been successfully incorporated into the structure of peptide hydrogelators, and the resulting properties are discussed in terms of their potential biomedical applications. Where appropriate, their properties are compared with those of the corresponding peptide hydrogelator composed of canonical amino acids. In a wider context, we consider the presence of dehydroamino acids in natural compounds and medicinally important compounds as well as their limitations, and we consider some of the synthetic strategies for obtaining dehydropeptides. Finally, we consider the future direction for this research area.

Exploring the properties and potential biomedical applications of NSAID-capped peptide hydrogels.

The development of strategies to minimise the adverse side-effects of non-steroidal anti-inflammatory drugs (NSAIDs) remains a challenge for medicinal chemists. One such strategy is the development of NSAID-peptide prodrug conjugates and this conjugation to a peptide often confers the additional property of hydrogelation. This review summarises the work published by our research group, alongside other research groups, on supramolecular hydrogels consisting of short peptides conjugated to NSAIDs. Generally, supramolecular low molecular weight hydrogels (LMWHs) are composed of amphiteric molecules, usually consisting of short peptides attached to an aromatic capping group. When the aromatic capping group is switched for an NSAID to afford hybrid gelators, some conjugates exhibit retained or improved anti-inflammatory properties of the parent drug, and sometimes new and unexpected biological activities are observed. Conjugation to peptides often provides selective COX-2 inhibition over COX-1 inhibtion, which is key to retaining the anti-inflammatory benefits of NSAIDs whilst minimising gastric side-effects. Naproxen is the most commonly employed NSAID capping group, partly due to its similarity in structure to commonly employed naphthalene capping groups. Biomimetic approaches, where canonical amino acids are switched for non-natural amino acids such as d-amino acids or dehydroamino acids, are often employed, to tune the stability. The future direction for this area of research is discussed.

A Notch-independent mechanism contributes to the induction of Hes1 gene expression in response to hypoxia in P19 cells.

Hes1 is a Notch target gene that plays a major role during embryonic development. Previous studies have shown that HIF-1α can interact with the Notch intracellular domain and enhance Notch target gene expression. In this study, we have identified a Notch-independent mechanism that regulates the responsiveness of the Hes1 gene to hypoxia. Using P19 cells we show that silencing the Notch DNA binding partner CSL does not prevent hypoxia-dependent upregulation of Hes1 expression. In contrast to CSL, knockdown of HIF-1α or Arnt expression prevents Hes1 induction in hypoxia. Deletion analysis of the Hes1 promoter identified a minimal region near the transcription start site that is still responsive to hypoxia. In addition, we show that mutating the GA-binding protein (GABP) motif significantly reduced Hes1 promoter-responsiveness to hypoxia or to HIF-1 overexpression whereas mutation of the hypoxia-responsive element (HRE) present in this region had no effect. Chromatin immunoprecipitation assays demonstrated that HIF-1α binds to the proximal region of the Hes1 promoter in a Notch-independent manner. Using the same experimental approach, the presence of GABPα and GABPß1 was also observed in the same region of the promoter. Loss- and gain-of-function studies demonstrated that Hes1 gene expression is upregulated by hypoxia in a GABP-dependent manner. Finally, co-immunoprecipitation assays demonstrated that HIF-1α but not HIF-2α is able to interact with either GABPα or GABPß1. These results suggest a Notch-independent mechanism where HIF-1 and GABP contribute to the upregulation of Hes1 gene expression in response to hypoxia.

Allergic contact dermatitis caused by (meth)acrylates in nail cosmetic products in users and nail technicians - a 5-year study.

BACKGROUND: The increasing use of long-lasting nail aesthetic products has led to a growing number of cases of allergic contact dermatitis (ACD) caused by (meth)acrylates in recent years. OBJECTIVES: To provide information on ACD caused by (meth)acrylates related to nail cosmetic products. METHODS: We retrospectively reviewed files of patients with ACD caused by (meth)acrylates related to nail cosmetic products, who were patch tested between January 2011 and December 2015 in 13 departments of dermatology in Portugal. RESULTS: Two-hundred and thirty cases of ACD caused by (meth)acrylates (55 technicians, 56 consumers, and 119 with mixed exposure) had been documented, mostly as chronic hand eczema (93%). The most common sensitizers were: 2-hydroxyethyl methacrylate (HEMA), which was positive in 90% of the tested patients, 2-hydroxypropyl methacrylate (HPMA), which was positive in 64.1%, and ethyleneglycol dimethacrylate, which was positive in 54.5%. CONCLUSION: HEMA and HPMA were the most frequent positive allergens. HEMA, which identified 90% of cases, can be considered to be a good screening allergen. The high number of cases of ACD caused by (meth)acrylates in nail cosmetic products certainly warrants better preventive measures at the occupational level, and specific regulation in the field of consumer safety.

Hypoxia-induced and calpain-dependent cleavage of filamin A regulates the hypoxic response.

The cellular response to hypoxia is regulated by hypoxia-inducible factor-1α and -2α (HIF-1α and -2α). We have discovered that filamin A (FLNA), a large cytoskeletal actin-binding protein, physically interacts with HIF-1α and promotes tumor growth and angiogenesis. Hypoxia induces a calpain-dependent cleavage of FLNA to generate a naturally occurring C-terminal fragment that accumulates in the cell nucleus. This fragment interacts with the N-terminal portion of HIF-1α spanning amino acid residues 1-390 but not with HIF-2α. In hypoxia this fragment facilitates the nuclear localization of HIF-1α, is recruited to HIF-1α target gene promoters, and enhances HIF-1α function, resulting in up-regulation of HIF-1α target gene expression in a hypoxia-dependent fashion. These results unravel an important mechanism that selectively regulates the nuclear accumulation and function of HIF-1α and potentiates angiogenesis and tumor progression.

HIF-1α and HIF-2α induce angiogenesis and improve muscle energy recovery.

BACKGROUND: Cardiovascular patients suffer from reduced blood flow leading to ischaemia and impaired tissue metabolism. Unfortunately, an increasing group of elderly patients cannot be treated with current revascularization methods. Thus, new treatment strategies are urgently needed. Hypoxia-inducible factors (HIFs) upregulate the expression of angiogenic mediators together with genes involved in energy metabolism and recovery of ischaemic tissues. Especially, HIF-2α is a novel factor, and only limited information is available about its therapeutic potential. METHODS: Gene transfers with adenoviral HIF-1α and HIF-2α were performed into the mouse heart and rabbit ischaemic hindlimbs. Angiogenesis was evaluated by histology. Left ventricle function was analysed with echocardiography. Perfusion in rabbit skeletal muscles and energy recovery after electrical stimulation-induced exercise were measured with ultrasound and (31)P-magnetic resonance spectroscopy ((31)P-MRS), respectively. RESULTS: HIF-1α and HIF-2α gene transfers increased capillary size up to fivefold in myocardium and ischaemic skeletal muscles. Perfusion in skeletal muscles was increased by fourfold without oedema. Especially, AdHIF-1α enhanced the recovery of ischaemic muscles from electrical stimulation-induced energy depletion. Special characteristic of HIF-2α gene transfer was a strong capillary growth in muscle connective tissue and that HIF-2α gene transfer maintained left ventricle function. CONCLUSIONS: We conclude that both AdHIF-1α and AdHIF-2α gene transfers induced beneficial angiogenesis in vivo. Transient moderate increases in angiogenesis improved energy recovery after exercise in ischaemic muscles. This study shows for the first time that a moderate increase in angiogenesis is enough to improve tissue energy metabolism, which is potentially a very useful feature for cardiovascular gene therapy.