Decreased Gas6 and sAxl Plasma Levels Are Associated with Hair Loss in COVID-19 Survivors.

Post-acute conditions after coronavirus disease 2019 (COVID-19) are quite common, although the underlying pathogenetic mechanisms leading to these conditions are not yet completely understood. In this prospective observational study, we aimed to test the hypothesis that Growth Arrest-Specific 6 (Gas6) and its soluble receptors, Axl (sAxl) and MerTK (sMer), might be implicated. A total of 263 subjects underwent a structured clinical evaluation one year after their hospital discharge for COVID-19, and they consented to donate a blood sample to measure their circulating Gas6, sAxl, and sMer levels. A total of 98 (37.3%) post-COVID-19 subjects complained of at least one residual physical symptom one year after their hospital discharge. Univariate analysis revealed that sAxl was marginally associated with residual symptoms, but at the level of logistic regression analysis, only the diffusing capacity of the lungs for carbon monoxide (DLCO) (OR 0.98, CI 95%: 0.96-0.99; p = 0.007) and the female sex (OR 2.49, CI 95%: 1.45-4.28; p = 0.001) were independently associated with long-lasting symptoms. A total of 69 (26.2%) subjects had hair loss. At the level of univariate analysis, Gas6, sAxl, DLCO, and the female gender were associated with its development. In a logistic regression analysis model, Gas6 (OR 0.96, CI 95%: 0.92-0.99; p = 0.015) and sAxl (OR 0.98, CI 95%; 0.97-1.0; p = 0.014), along with the female sex (OR 6.58, CI 95%: 3.39-12.78; p = 0.0001), were independent predictors of hair loss. Decreased levels of Gas6 and sAxl were associated with a history of hair loss following COVID-19. This was resolved spontaneously in most patients, although 23.7% complained of persistent hair loss one year after hospital discharge.

Virulence factors of Malassezia strains isolated from pityriasis versicolor patients and healthy individuals.

Over the last decade, Malassezia species have emerged as increasingly important pathogens associated with a wide range of dermatological disorders and bloodstream infections. The pathogenesis of Malassezia yeasts is not completely clear, but it seems to be strictly related to Malassezia strains and hosts and needs to be better investigated. This study aimed to assess the enzymatic activities, biofilm formation and in vitro antifungal profiles of Malassezia spp. from pityriasis versicolor (PV) and healthy patients. The potential relationship between virulence attributes, the antifungal profiles and the origin of strains was also assessed. A total of 44 Malassezia strains isolated from patients with (n = 31) and without (n = 13) PV were employed to evaluate phospholipase (Pz), lipase (Lz), and hemolytic (Hz) activities and biofilm formation. In addition, in vitro antifungal susceptibility testing was conducted using the CLSI broth microdilution with some modifications. A high percentage of strains produced Pz, Lz, Hz and biofilm regardless of their clinical origin. The highest number of strains producing high enzymatic activities came from PV patients. A correlation between the intensity of hydrolytic activities (Lz and Pz activities) and the Hz activity was detected. Positive associations between Lz and the low fluconazole susceptibility and Hz and biofilm formation were observed. These results suggest that enzyme patterns and biofilm formation along with antifungal profiles inter-play a role in the pathogenicity of Malassezia spp. and might explain the implication of some Malassezia spp. in invasive fungal infections and in the development of inflammation. LAY SUMMARY: There is still little information on the virulence factors of Malassezia spp., despite their implication in severe diseases. Phospholipase, lipase, and hemolytic activities, biofilm formation and decreased antifungal susceptibility seem to contribute to their virulence in susceptible hosts.

Thioridazine reverts the phenotype in cellular and Drosophila models of amyotrophic lateral sclerosis by enhancing TDP-43 aggregate clearance.

Brain inclusions mainly composed of misfolded and aggregated TAR DNA binding protein 43 (TDP-43), are characteristic hallmarks of amyotrophic lateral sclerosis (ALS). Irrespective of the role played by the inclusions, their reduction represents an important therapeutic pathway that is worth exploring. Their removal can either lead to the recovery of TDP-43 function by removing the self-templating conformers that sequester the protein in the inclusions, and/or eliminate any potential intrinsic toxicity of the aggregates. The search for curative therapies has been hampered by the lack of ALS models for use in high-throughput screening. We adapted, optimised, and extensively characterised our previous ALS cellular model for such use. The model demonstrated efficient aggregation of endogenous TDP-43, and concomitant loss of its splicing regulation function. We provided a proof-of-principle for its eventual use in high-throughput screening using compounds of the tricyclic family and showed that recovery of TDP-43 function can be achieved by the enhanced removal of TDP-43 aggregates by these compounds. We observed that the degradation of the aggregates occurs independent of the autophagy pathway beyond autophagosome-lysosome fusion, but requires a functional proteasome pathway. The in vivo translational effect of the cellular model was tested with two of these compounds in a Drosophila model expressing a construct analogous to the cellular model, where thioridazine significantly improved the locomotive defect. Our findings have important implications as thioridazine cleared TDP-43 aggregates and recovered TDP-43 functionality. This study also highlights the importance of a two-stage, in vitro and in vivo model system to cross-check the search for small molecules that can clear TDP-43 aggregates in TDP-43 proteinopathies.

Point mutations in the N-terminal domain of transactive response DNA-binding protein 43 kDa (TDP-43) compromise its stability, dimerization, and functions.

Transactive response DNA-binding protein 43 (TDP-43) performs multiple tasks in mRNA processing, transport, and translational regulation, but it also forms aggregates implicated in amyotrophic lateral sclerosis. TDP-43's N-terminal domain (NTD) is important for these activities and dysfunctions; however, there is an open debate about whether or not it adopts a specifically folded, stable structure. Here, we studied NTD mutations designed to destabilize its structure utilizing NMR and fluorescence spectroscopies, analytical ultracentrifugation, splicing assays, and cell microscopy. The substitutions V31R and T32R abolished TDP-43 activity in splicing and aggregation processes, and even the rather mild L28A mutation severely destabilized the NTD, drastically reducing TDP-43's in vitro splicing activity and inducing aberrant localization and aggregation in cells. These findings strongly support the idea that a stably folded NTD is essential for correct TDP-43 function. The stably folded NTD also promotes dimerization, which is pertinent to the protein's activities and pathological aggregation, and we present an atomic-level structural model for the TDP-43 dimer based on NMR data. Leu-27 is evolutionarily well conserved even though it is exposed in the monomeric NTD. We found here that Leu-27 is buried in the dimer and that the L27A mutation promotes monomerization. In conclusion, our study sheds light on the structural and biological properties of the TDP-43 NTD, indicating that the NTD must be stably folded for TDP-43's physiological functions, and has implications for understanding the mechanisms promoting the pathological aggregation of this protein.

TDP-43 loss of cellular function through aggregation requires additional structural determinants beyond its C-terminal Q/N prion-like domain.

TDP-43 aggregates are the neurohistological landmark of diseases like amyotrophic lateral sclerosis and frontotemporal dementia. Their role in the pathogenesis of these conditions is not yet clear mainly due to the lack of proper models of aggregation that may allow the study of the mechanism of formation, their interactions with other cellular components and their effect on the cell metabolism. In this work, we have used tandem repeats of the prion like Q/N-rich region of TAR DNA-binding protein (TDP-43) fused to additional TDP-43 protein sequences to trigger aggregate formation in neuronal and non-neuronal cell lines. At the functional level, these aggregates are able to sequester endogenous TDP-43 depleting its nuclear levels and inducing loss of function at the pre-mRNA splicing level. No apparent direct cellular toxicity of the aggregates seems to be present beyond the lack of functional TDP-43. To our knowledge, this is the only system that achieves full functional TDP 43 depletion with effects similar to RNAi depletion or gene deletion. As a result, this model will prove useful to investigate the loss-of-function effects mediated by TDP-43 aggregation within cells without affecting the expression of the endogenous gene. We have identified the N-terminus sequence of TDP-43 as the domain that enhances its interaction with the aggregates and its insolubilization. These data show for the first time that cellular TDP-43 aggregation can lead to total loss of function and to defective splicing of TDP-43-dependent splicing events in endogenous genes.

Social networks and health status in the elderly: the 'ANZIANI IN-RETE' population-based study.

BACKGROUND: Certain features of the social environment could maintain and even improve not only psychological well-being, but also health and cognition of the elderly. AIMS: We tested the association between social network characteristics and the number of chronic diseases in the elderly. METHODS: A randomized sample of the elderly population of Brescia, Italy, was evaluated (N = 200, age ≥65 years). We performed a comprehensive geriatric assessment, including information on socio-demographic variables (family, friendships, and acquaintance contacts). We measured each person's social network, i.e., degree, efficiency, and variety. RESULTS: The sample included 118 women and 82 men, mean age 77.7 years. The mean number of chronic diseases was 3.5. A higher social network degree, i.e., more social connections, was associated with fewer diseases. We also found that having more contacts with people similar to each other or intense relationships with people who do not know each other were associated with fewer diseases. CONCLUSION: More healthy people tend to share certain characteristics of social networks. Our study indicates that it is important to look at diseases and health as complex phenomena, which requires integrating different levels of analysis.

Cellular model of TAR DNA-binding protein 43 (TDP-43) aggregation based on its C-terminal Gln/Asn-rich region.

TDP-43 is one of the major components of the neuronal and glial inclusions observed in several neurodegenerative diseases such as amyotrophic lateral sclerosis and frontotemporal lobar degeneration. These characteristic aggregates are a "landmark" of the disease, but their role in the pathogenesis is still obscure. In previous works, we have shown that the C-terminal Gln/Asn-rich region (residues 321-366) of TDP-43 is involved in the interaction of this protein with other members of the heterogeneous nuclear ribonucleoprotein protein family. Furthermore, we have shown that the interaction through this region is important for TDP-43 splicing inhibition of cystic fibrosis transmembrane regulator exon 9, and there were indications that it was involved in the aggregation process. Our experiments show that in cell lines and primary rat neuronal cultures, the introduction of tandem repeats carrying the 331-369-residue Gln/Asn region from TDP-43 can trigger the formation of phosphorylated and ubiquitinated aggregates that recapitulate many but not all the characteristics observed in patients. These results establish a much needed cell-based TDP-43 aggregation model useful to investigate the mechanisms involved in the formation of inclusions and the gain- and loss-of-function consequences of TDP-43 aggregation within cells. In addition, it will be a powerful tool to test novel therapeutic strategies/effectors aimed at preventing/reducing this phenomenon.

An oncolytic adenovirus enhanced for toll-like receptor 9 stimulation increases antitumor immune responses and tumor clearance.

Oncolytic viruses represent a multifaceted tool for cancer treatment. In addition to specific killing of cancer cells (oncolysis), these agents also provide danger signals prompting the immune system to stimulate an antitumor immune response. To increase adenovirus adjuvancy, we engineered the genome of Ad5D24 by inserting 18 immunostimulatory islands (Ad5D24-CpG). The toxicity and immunogenicity profile of Ad5D24-CpG showed that the safety of the maternal virus was retained. The efficacy of the CpG-enriched virus was assessed in a xenograft model of lung cancer where a significant increase in antitumor effect was seen in comparison with controls. When the experiment was repeated in animal depleted of natural killer (NK) cells, Ad5D24-CpG lost its advantage. The same was seen when Toll-like receptor (TLR)9 was blocked systemically. In a syngeneic model of melanoma (B16-OVA), we observed a significant increase of OVA-specific T cells and a decrease of activation of myeloid-derived suppressor cells in Ad5D24-CpG-treated mice. In conclusion, we have generated the first genetically modified oncolytic adenovirus backbone able to enhance TLR9-stimulation for increased antitumor activity.

Immunological effects of low-dose cyclophosphamide in cancer patients treated with oncolytic adenovirus.

Patients with advanced solid tumors refractory to and progressing after conventional therapies were treated with three different regimens of low-dose cyclophosphamide (CP) in combination with oncolytic adenovirus. CP was given with oral metronomic dosing (50 mg/day, N = 21), intravenously (single 1,000 mg dose, N = 7) or both (N = 7). Virus was injected intratumorally. Controls (N = 8) received virus without CP. Treatments were well tolerated and safe regardless of schedule. Antibody formation and virus replication were not affected by CP. Metronomic CP (oral and oral + intravenous schedules) decreased regulatory T cells (T(regs)) without compromising induction of antitumor or antiviral T-cell responses. Oncolytic adenovirus given together with metronomic CP increased cytotoxic T cells and induced Th1 type immunity on a systemic level in most patients. All CP regimens resulted in higher rates of disease control than virus only (all P < 0.0001) and the best progression-free (PFS) and overall survival (OS) was seen in the oral + intravenous group. One year PFS and OS were 53 and 42% (P = 0.0016 and P < 0.02 versus virus only), respectively, both which are unusually high for chemotherapy refractory patients. We conclude that low-dose CP results in immunological effects appealing for oncolytic virotherapy. While these first-in-human data suggest good safety, intriguing efficacy and extended survival, the results should be confirmed in a randomized trial.

Malassezia: Zoonotic Implications, Parallels and Differences in Colonization and Disease in Humans and Animals.

Malassezia spp. are commensals of the skin, oral/sinonasal cavity, lower respiratory and gastrointestinal tract. Eighteen species have been recovered from humans, other mammals and birds. They can also be isolated from diverse environments, suggesting an evolutionary trajectory of adaption from an ecological niche in plants and soil to the mucocutaneous ecosystem of warm-blooded vertebrates. In humans, dogs and cats, Malassezia-associated dermatological conditions share some commonalities. Otomycosis is common in companion animals but is rare in humans. Systemic infections, which are increasingly reported in humans, have yet to be recognized in animals. Malassezia species have also been identified as pathogenetic contributors to some chronic human diseases. While Malassezia species are host-adapted, some species are zoophilic and can cause fungemia, with outbreaks in neonatal intensive care wards associated with temporary colonization of healthcare worker's hands from contact with their pets. Although standardization is lacking, susceptibility testing is usually performed using a modified broth microdilution method. Antifungal susceptibility can vary depending on Malassezia species, body location, infection type, disease duration, presence of co-morbidities and immunosuppression. Antifungal resistance mechanisms include biofilm formation, mutations or overexpression of ERG11, overexpression of efflux pumps and gene rearrangements or overexpression in chromosome 4.

Delivery and effectiveness of entomopathogenic fungi for mosquito and tick control: Current knowledge and research challenges.

Insects, ticks, and mites represent a threat to animal health globally, mainly due to their role as vectors of pathogens. Among the most important diseases, those transmitted by mosquitoes (e.g., malaria and arboviral infections) and ticks (e.g., Lyme borreliosis, babesiosis, and viral haemorrhagic fever) have a huge impact on human health. The principal methods available for reducing the public health burden of most vector-borne diseases are vector-based intervention relying to insecticides and acaricides. However, the use of these products is challenged by the introduction of invasive species, the quick development of physiological insecticide and acaricide resistance, and their non-target effects on human health and environment. In this scenario, insecticide/acaricide-free control approaches based on the employment of entomopathogenic fungi (EPFs) are currently considered a promising tool in Integrated Pest/Vector Management, even if their large-scale use is still limited. In this article, we provide an overview on current knowledge about the role of EPFs for mosquito and tick management to assess solutions improving the delivery and efficacy of EPFs in the field. Laboratory research provided solid evidence that EPFs represent a next-generation control tool to manage mosquito and tick populations. However, the viability, infectivity, and persistence of fungal spores under field conditions are still inadequate. Herein we also discuss the development and optimization of EPF-based lure and kill approaches through biopolymers to improve cost-competitive, safety and eco-friendly pest and vector control tools.

Remodelling is inversely proportional to left ventricular dimensions in a real-life population of patients with chronic heart failure after therapy with sacubitril/valsartan.

BACKGROUND: Left ventricular (LV) remodelling is a major mechanism underlying disease progression in patients with heart failure (HF) with reduced ejection fraction (EF). Previous studies that LVEF improvement and reverse remodelling can be achieved after therapy with Sacubitril/Valsartan in real-world settings. Therefore, we sought to investigate possible predictors of LV remodelling, in particular echocardiographic parameters derived by Tissue Doppler Imaging. METHODS: Patients with chronic HF, LV dysfunction (EF < 35%), NYHA class II-III were followed up between September 2016 and January 2019. All patients underwent clinical and echocardiography follow up at baseline and after 12 months of therapy with sacubitril/valsartan. RESULTS: Fifty-four consecutive outpatients were enrolled in the study. At follow-up visit LVEF (38 ± 9 vs. 30 ± 5%, p < 0.0001), LVEDD (61 ± 8 vs. 62 ± 8 mm, p = 0.0085), LVESV (114 ± 57 vs. 130 ± 56 mm3, p = 0.0001), mitral regurgitation severity (1 ± 1 vs. 2 ± 1, p < 0.0001), and left atrial area (23 ± 6 vs. 24 ± 6 mm2, p = 0.0121) changed compared to the baseline value. Changes in LVEF (follow up vs baseline) correlated with baseline levels of heart rate (r = 0.24, p = 0.048), LVEDD (r= -0.33, p = 0.004), LVEDV (r= -0.39, p = 0.001), LVESV (r = 0.37, p = 0.002), and changes in LVESV (r=-0.34, p = 0.006). Correlations remained significant even after correction at multivariate analysis including age and gender. CONCLUSIONS: Treatment with sacubitril/valsartan in patients with systolic dysfunction is associated with an improvement in LVEF in a real world scenario. Smaller LV volumes are associated with better reverse LV remodelling.

Rapid Point-Of-Care Serology and Clinical History Assessment Increase Protection Provided by RT-PCR Screening: A Pilot Study Involving Three Nursing Homes in Brescia, a Hotspot of Lombardy.

Background: COVID-19 outbursts have been registered worldwide within care homes with asymptomatic transmission combined with shortage/inaccuracy of diagnostic tests undermining the efforts at containment of the disease. Nursing facilities in Lombardy (Italy) were left with no, or limited, access to testing for 8 weeks after the outbreak of COVID-19. Methods: This study includes 246 residents and 286 workers of three different nursing homes in Brescia-Lombardy. Clinical questionnaires and rapid serology tests were devised to integrate the data of the first available RT-PCR screening. Follow-up serology after 60-days was performed on 67 of 86 workers with positive serology or clinically suspicious. Findings: Thirty-seven residents and 18 workers had previous positive RT-PCR. Thorough screening disclosed two additional RT-PCR-positive workers. Serology screening revealed antibodies in 59 residents and 48 workers, including 32/37 residents and all workers previously positive at RT-PCR. Follow up serology disclosed antibodies in two additional workers with recent symptoms at the time of screening. The professionals in close contact with residents had more infections (47/226-20.79% vs. 1/60-1.66%; p = 0.00013 Fisher exact-test). A suspicious clinical score was present in 44/64 residents and in 41/50 workers who tested positive with either method with totally asymptomatic disease more frequent among residents 28.1 vs. 10.0% (p = 0.019 Fisher exact-test). Interpretation: Based on the available RT-PCR ± results at the time of symptoms/contacts, our integrated clinical and serological screening demonstrated sensitivity 89% and specificity 87%. This multimodal assessment proved extremely useful in understanding the viral spread in nursing homes, in defining its stage and in implementing protective measures. Rapid serology tests demonstrated efficient and particularly suited for older people less able to move/cooperate.

The Role in Teledermoscopy of an Inexpensive and Easy-to-Use Smartphone Device for the Classification of Three Types of Skin Lesions Using Convolutional Neural Networks.

BACKGROUND: The use of teledermatology has spread over the last years, especially during the recent SARS-Cov-2 pandemic. Teledermoscopy, an extension of teledermatology, consists of consulting dermoscopic images, also transmitted through smartphones, to remotely diagnose skin tumors or other dermatological diseases. The purpose of this work was to verify the diagnostic validity of images acquired with an inexpensive smartphone microscope (NurugoTM), employing convolutional neural networks (CNN) to classify malignant melanoma (MM), melanocytic nevus (MN), and seborrheic keratosis (SK). METHODS: The CNN, trained with 600 dermatoscopic images from the ISIC (International Skin Imaging Collaboration) archive, was tested on three test sets: ISIC images, images acquired with the NurugoTM, and images acquired with a conventional dermatoscope. RESULTS: The results obtained, although with some limitations due to the smartphone device and small data set, were encouraging, showing comparable results to the clinical dermatoscope and up to 80% accuracy (out of 10 images, two were misclassified) using the NurugoTM demonstrating how an amateur device can be used with reasonable levels of diagnostic accuracy. CONCLUSION: Considering the low cost and the ease of use, the NurugoTM device could be a useful tool for general practitioners (GPs) to perform the first triage of skin lesions, aiding the selection of lesions that require a face-to-face consultation with dermatologists.

Variability of semiautomated lung nodule volumetry on ultralow-dose CT: comparison with nodule volumetry on standard-dose CT.

The study investigates the effect of a substantial dose reduction on the variability of lung nodule volume measurements by assessing and comparing nodule volumes using a dedicated semiautomated segmentation software on ultralow-dose computed tomography (ULD-CT) and standard-dose computed tomography (SD-CT) data. In 20 patients, thin-slice chest CT datasets (1 mm slice thickness; 20% reconstruction overlap) were acquired at ultralow-dose (120 kV, 5 mAs) and at standard-dose (120 kV, 75 mAs), respectively, and analyzed using the segmentation software OncoTREAT (MeVis, Bremen, Germany; version 1.3). Interobserver variability of volume measurements of 202 solid pulmonary nodules (mean diameter 11 mm, range 3.2-44.5 mm) was calculated for SD-CT and ULD-CT. With respect to interobserver variability, the 95% confidence interval for the relative differences in nodule volume in the intrascan analysis was measured with -9.7% to 8.3% (mean difference -0.7%) for SD-CT and with -12.6% to 12.4% (mean difference -0.2%) for ULD-CT. In the interscan analysis, the 95% confidence intervals for the differences in nodule volume ranged with -25.1% to -23.4% and 26.2% to 28.9% (mean difference 1.4% to 2.1%) dependent on the combination of readers and scans. Intrascan interobserver variability of volume measurements was comparable for ULD-CT and SD-CT data. The calculated variability of volume measurements in the interscan analysis was similar to the data reported in the literature for CT data acquired with equal radiation dose. Thus, the evaluated segmentation software provides nodule volumetry that appears to be independent of the dose level with which the CT source dataset is acquired.

Sacubitril/valsartan improves right ventricular function in a real-life population of patients with chronic heart failure: The Daunia Heart Failure Registry.

BACKGROUND: Previous studies and case-series showed improvement in left ventricular (LV) function and reverse remodeling after sacubitril/valsartan therapy in real-world studies. We therefore aimed to evaluate whether also right ventricular (RV) function may improve after sacubitril/valsartan therapy. METHODS: Sixty consecutive patients with chronic heart failure and NYHA class II-III were followed up for 12 months after therapy with sacubitril/valsartan. Left and (RV) function was assessed at baseline and after 12 months of therapy. RESULTS: At 12-month control, therapy with sacubitril/valsartan was associated with a significant improvement in a series of echo parameters: LVEF (p < 0.05), LV end-systolic volume (p < 0.01), left atrium area (p < 0.05).Right ventricular echo parameters were also improved after sacubitril/valsartan therapy: PAsP (31.0 ± 12.8 vs 34.7 ± 12.5 mmHg, p < 0.05), TAPSE (17.8 ± 3.9 vs 16.5 ± 4.0 mm, p < 0.001); mean PAsP reduction was 3.7 ± 11.4 mmHg (-6.3 ± 37.7%), mean TAPSE increase 1.3 ± 2.5 mm (+9.5 ± 15.7%).Indexed (%) improvement in PAsP (r 0.33, p < 0.01) and TAPSE (r -0.42, p < 0.01) values were proportional to baseline levels. Improvement in PAsP and TAPSE were independent of left ventricular improvements except for PAsP and end-systolic volumes (r 0.44, p < 0.01). CONCLUSIONS: In a real world scenario, sacubitril/valsartan was associated with an improved RV function.

Initial experience with a chest pain protocol using 320-slice volume MDCT.

We sought to determine the feasibility and image quality of 320-slice volume computed tomography (CT) angiography for the evaluation of patients with acute chest pain. Thirty consecutive patients (11 female, 19 male, mean age 63.2 +/- 14.2 years) with noncritical, acute chest pain underwent 320-slice CT using a protocol consisting of a nonspiral, nongated CT of the entire chest, followed by a nonspiral, electrocardiography-gated CT study of the heart. Data were acquired following a biphasic intravenous injection of 90 ml iodinated contrast agent. Vessel attenuation values of different thoracic vascular territories were recorded, and image quality scored on a five-point scale by two readers. Mean attenuation was 467 +/- 69 HU in the ascending aorta, 334 +/- 52 HU in the aortic arch, 455 +/- 71 HU in the descending aorta, 492 +/- 94 HU in the pulmonary trunk, and 416 +/- 63 HU and 436 +/- 62 HU in the right and left coronary artery, respectively. Radiation exposure estimates ranged between 7 and 14 mSv. The CT protocol investigated enabled imaging of the thoracic aorta, coronary and pulmonary arteries with an excellent diagnostic quality for chest pain triage in all patients. This result was achieved with less contrast material and reduced radiation exposure compared with previously investigated imaging protocols.

Piomiositis en pediatría: experiencia de 10 años en un hospital pediátrico de alta complejidad en Argentina / Pyomyositis in pediatrics: 10-year experience in a high-complexity pediatric hospital in Argentina

INTRODUCCIÓN: La piomiositis es una infección bacteriana agudasubaguda del músculo esquelético. OBJETIVO: Estimar la incidencia de piomiositis en pacientes internados, describir e identificar factores de riesgo para bacteriemia y hospitalización, y evaluar diferencias entre Staphylococccus aureus sensible y resistente a meticilina (SASM y SARM). PACIENTES Y MÉTODOS: Estudio descriptivo, retrospectivo, observacional, con pacientes de 1 mes a 18 años de edad, internados entre el 1 de enero de 2008 y 31 de diciembre de 2018. Variables: sexo, edad, hacinamiento en el hogar, existencia de lesión previa, estacionalidad, localización anatómica e imágenes, antibioterapia previa, estadio clínico, parámetros de laboratorio, cultivos y antibiograma, días de tratamiento intravenoso (IV), de internación, de fiebre y bacteriemia. RESULTADOS: Se incluyeron 188 pacientes. Incidencia: 38,9 casos / 10.000 admisiones (IC95 % 33,7 - 44,9). Días de internación y tratamiento IV: 11 (RQ 8-15 y RQ 8-14, respectivamente). El desarrollo de bacteriemia se asoció a PCR elevada (p = 0,03) y fiebre prolongada (p < 0,001). No hubo diferencias en la evolución y parámetros de laboratorio entre SASM y SARM. La leucocitosis (p = 0,004), neutrofilia (p = 0,005) y bacteriemia (p = 0,001) se asociaron a mayor estadía hospitalaria. CONCLUSIONES: Este estudio recaba la experiencia de más de 10 años de niños internados con diagnóstico de piomiositis y proporciona información sobre sus características. Se describen parámetros asociados a bacteriemia y estadía hospitalaria.

BACKGROUND: Pyomyositis is an acute-subacute bacterial infection of skeletal muscle. AIM: To estimate the incidence of pyomyositis in hospitalized patients, describe and identify risk factors for bacteremia and hospitalization, and evaluate differences between MSSA and MRSA. METHODS: Descriptive, retrospective, observational study with patients aged 1 month to 18 years hospitalized between January, 1, 2008 and December 1, 2018. Variables: sex, age, home overcrowding, previous injury, seasonality, anatomical location and images, previous antibiotherapy, clinical stage, laboratory, cultures and antibiogram, days of intravenous (IV) treatment, hospitalization, fever and bacteremia. RESULTS: 188 patients were included. Incidence: 38.9 cases/10,000 admissions (95% CI 33.7 - 44.9). Days of hospitalization and IV treatment: 11 (RQ 8-15 and RQ 8-14, respectively). The development of bacteremia was associated with elevated CRP (p = 0.03) and prolonged fever (p < 0.001). There were no differences in the evolution and laboratory parameters between MSSA and MRSA. Leukocytosis (p = 0.004), neutrophilia (p = 0.005), and bacteremia (p = 0.001) were associated with a longer hospital stay. CONCLUSIONS: This study collects the experience of more than 10 years of hospitalized children diagnosed with pyomyositis and provides information on its characteristics. Parameters associated with bacteremia and hospital stay are described.

A computational protocol to evaluate the effects of protein mutants in the kinase gatekeeper position on the binding of ATP substrate analogues.

BACKGROUND: The determination of specific kinase substrates in vivo is challenging due to the large number of protein kinases in cells, their substrate specificity overlap, and the lack of highly specific inhibitors. In the late 90s, Shokat and coworkers developed a protein engineering-based method addressing the question of identification of substrates of protein kinases. The approach was based on the mutagenesis of the gatekeeper residue within the binding site of a protein kinase to change the co-substrate specificity from ATP to ATP analogues. One of the challenges in applying this method to other kinase systems is to identify the optimal combination of mutation in the enzyme and chemical derivative such that the ATP analogue acts as substrate for the engineered, but not the native kinase enzyme. In this study, we developed a computational protocol for estimating the effect of mutations at the gatekeeper position on the accessibility of ATP analogues within the binding site of engineered kinases. RESULTS: We tested the protocol on a dataset of tyrosine and serine/threonine protein kinases from the scientific literature where Shokat's method was applied and experimental data were available. Our protocol correctly identified gatekeeper residues as the positions to mutate within the binding site of the studied kinase enzymes. Furthermore, the approach well reproduced the experimental data available in literature. CONCLUSIONS: We have presented a computational protocol that scores how different mutations at the gatekeeper position influence the accommodation of various ATP analogues within the binding site of protein kinases. We have assessed our approach on protein kinases from the scientific literature and have verified the ability of the approach to well reproduce the available experimental data and identify suitable combinations of engineered kinases and ATP analogues.

A novel Drosophila model of TDP-43 proteinopathies: N-terminal sequences combined with the Q/N domain induce protein functional loss and locomotion defects.

Transactive response DNA-binding protein 43 kDa (TDP-43, also known as TBPH in Drosophila melanogaster and TARDBP in mammals) is the main protein component of the pathological inclusions observed in neurons of patients affected by different neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration (FTLD). The number of studies investigating the molecular mechanisms underlying neurodegeneration is constantly growing; however, the role played by TDP-43 in disease onset and progression is still unclear. A fundamental shortcoming that hampers progress is the lack of animal models showing aggregation of TDP-43 without overexpression. In this manuscript, we have extended our cellular model of aggregation to a transgenic Drosophila line. Our fly model is not based on the overexpression of a wild-type TDP-43 transgene. By contrast, we engineered a construct that includes only the specific TDP-43 amino acid sequences necessary to trigger aggregate formation and capable of trapping endogenous Drosophila TDP-43 into a non-functional insoluble form. Importantly, the resulting recombinant product lacks functional RNA recognition motifs (RRMs) and, thus, does not have specific TDP-43-physiological functions (i.e. splicing regulation ability) that might affect the animal phenotype per se. This novel Drosophila model exhibits an evident degenerative phenotype with reduced lifespan and early locomotion defects. Additionally, we show that important proteins involved in neuromuscular junction function, such as syntaxin (SYX), decrease their levels as a consequence of TDP-43 loss of function implying that the degenerative phenotype is a consequence of TDP-43 sequestration into the aggregates. Our data lend further support to the role of TDP-43 loss-of-function in the pathogenesis of neurodegenerative disorders. The novel transgenic Drosophila model presented in this study will help to gain further insight into the molecular mechanisms underlying neurodegeneration and will provide a valuable system to test potential therapeutic agents to counteract disease.