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A 3-month-old, full term female infant, adequate for gestational age, and exclusively breastfed, was admitted with a 10 day history of generalised scaling erythematous dermatitis, affecting the face (perinasal, nasolabial folds and periauricular), acral and intertriginous areas, with irritability and failure to thrive. Her mother had been treated with isoniazid since the third trimester because of family contact with tuberculosis. Based on a diagnosis of suspected impetiginised eczema, the infant was treated with flucloxacillin and prednisolone, and maternal isoniazid was suspended, with no improvement. Investigations found low serum zinc levels in the infant (33 μg/dL; normal range (NR) >60 μg/dL), normal plasma zinc levels in the mother (111.3 μg/dL; NR 68–120 μg/dL) and lower than the normal range of zinc levels in breast milk (270μg/L; NR 1000–2500 μg/L), suggesting acrodermatitis caused by zinc deficiency. Oral zinc supplementation (3 mg/kg/day) was started with a marked improvement in skin lesions, as well as good weight gain. At the age of 6 months, after food diversification, supplementation was suspended, without any recurrence of symptoms.

tRNAs are a driving force of genome evolution in Yeast and Bacteria. Their deregulation is frequently observed in tumors with Serine tRNAs being often overexpressed. This has important functional consequences, such as increased metabolism and tumor growth. In yeast, time and chemical stimulus boost alterations in the genome driven by tRNA deregulation. Therefore, we hypothesized that tRNA deregulation may contribute to the increased genome instability observed in tumors. To study the effect of tRNA deregulation in tumors, we overexpressed tRNA-Ser-AGA-2-1 in a NSCLC cell line, H460. This cell line and a Mock (control) were xenografted in nude mice and collected at 3 timepoints: T1-Naïve; T2-Treated once with cisplatin/vehicle and; T3) treated twice with cisplatin/vehicle. These tumors were characterized by WES, RNAseq and Mass Spectrometry and the data obtained was integrated. The tumor mutation burden was increased in T3 tRNASerOE tumors, regardless of treatment. Although in T1 Mock and tRNASer tumors have a similar number of variants, in T2&3, tRNASerOE tumors display two times more variants than Mock tumors regardless of treatment. Interestingly, tRNASerOE exclusive variants favor proliferation and therapy resistance, which is in line with the phenotypes observed and supported by RNAseq and proteomics data. In conclusion, tRNASerOE increases the tumor mutation burden and the variants detected favor tumor growth, proving tRNA deregulation is enough to induce adaptive mutations in the genome.

The sensitivity of the αβ T-cell receptor (TCR) is enhanced by the coreceptors CD4 and CD8αβ, which are expressed primarily by cells of the helper and cytotoxic T-cell lineages, respectively. The coreceptors bind to major histocompatibility complex (MHC) molecules and associate intracellularly with the Src-family kinase Lck, which catalyzes TCR phosphorylation during receptor triggering. Although coreceptor-kinase occupancy was initially believed to be high, a recent study suggested that most coreceptors exist in an Lck-free state, and that this low occupancy helps to effect TCR antigen discrimination. Here, using the same method, we found instead that the CD4-Lck interaction was stoichiometric (~100%) and that the CD8αβ-Lck interaction was also substantial (~60%). We confirmed our findings in live cells using fluorescence cross-correlation spectroscopy (FCCS) to measure coreceptor-Lck co-diffusionin situ. After introducing structurally guided mutations into the intracellular domain of CD4, we used FCCS to show that stoichiometric Lck coupling required an amphipathic α-helix present in CD4 but not CD8α. In double-positive cells expressing equal numbers of both coreceptors, but limiting amounts of kinase, CD4 out-competed CD8αβ for Lck. In T cells, TCR signaling induced CD4-Lck oligomerization but did not affect the high levels of CD4-Lck occupancy. These findings help settle the question of kinase occupancy and suggest that the binding advantages that CD4 has over CD8 could be important when Lck levels are limiting.Significance statementCD4 and CD8αβ are archetypal coreceptor proteins that potently enhance T-cell antigen sensitivity but how they function is still debated. A fundamental question that remains incompletely resolved is: what fractions of the coreceptors bind the signal-initiating kinase, Lck? Usingin vitroassays and non-invasive fluorescence fluctuation spectroscopy in live cells, we show that most coreceptors are occupied by Lck at the surface of live cells. The structural basis for important differences in the kinase occupancy of CD4 and CD8αβ is also identified. These results provide important context for refining current models of both TCR antigen recognition and cell fate decisions made during thymopoiesis.

Transfer RNA fragments (tRFs) have gene silencing effects similarly to miRNAs, can be sorted into extracellular vesicles (EVs) and are emerging as potential circulating biomarkers for cancer diagnoses. We aimed at analyzing the expression of tRFs in gastric cancer (GC) and understanding their potential as biomarkers. We explored miRNA datasets from gastric tumors and normal adjacent tissues (NATs) from TCGA repository, as well as proprietary 3D-cultured GC cell lines and corresponding EVs, in order to identify differentially represented tRFs using MINTmap and R/Bioconductor packages. Selected tRFs were validated in patient-derived EVs. We found 613 Differentially Expressed (DE)-tRFs in the TCGA dataset, of which 19 were concomitantly upregulated in TCGA gastric tumors and present in 3D cells and EVs, but barely expressed in NATs. Moreover, 20 tRFs were expressed in 3D cells and EVs and downregulated in TCGA gastric tumors. Of these 39 DE-tRFs, 9 tRFs were also detected in patient-derived EVs. Interestingly, the targets of these 9 tRFs affect neutrophil activation and degranulation, cadherin binding, focal adhesion and the cell–substrate junction, highlighting these pathways as major targets of EV-mediated crosstalk with the tumor microenvironment. Furthermore, as they are present in four distinct GC datasets and can be detected even in low quality patient-derived EV samples, they hold promise as GC biomarkers. By repurposing already available NGS data, we could identify and cross-validate a set of tRFs holding potential as GC diagnosis biomarkers.

Case Report A short stature 10 year old boy with pes planus, mild hearing loss, failure of exfoliation of the primary dentition, anterior hyperdontia, malocclusion and open anterior fontanelle was observed in Pediatric Orthopedics consultation, and presenting the following alterations (image). Discussion This child has cleidocranial dysplasia (CCD). Is an autosomal dominant disease caused by an heterozygous loss-of-function mutation in the RUNX2 gene, encoding transcription factor CBFA1, on chromosome 6p21, which plays an important role in osteogenesis and differentiation of osteoblasts. The main clinical features include persistently open skull sutures, hypoplasia or aplasia of the clavicles permitting abnormal facility in opposing the shoulders, short middle phalanx of the fifth fingers, dental anomalies (supernumerary teeth, failure of exfoliation of the primary dentition and malocclusion), pes planus, upper respiratory complications, recurrent otitis media, hearing loss and often vertebral malformation (scoliosis). Conclusion The clinical diagnosis of CCD was made during pregnancy, through the detection of aplasia of the clavicles in fetu, in association with the mother having short stature (1.32 meters), persistence of primary teeth and open anterior fontanelle. With 10 years of age, he underwent mendilioma study which revealed the mutation in the RUNX2 gene (c. 747 del (p.(Arg251 Alafs*7))) classified as a pathogenic variant.

Transitions between epithelial and mesenchymal cellular states (EMT/MET) contribute to cancer progression. We hypothesize that EMT followed by MET promotes cell population heterogeneity favouring tumour growth. We developed an EMT model by on/off exposure of epithelial EpH4 cells (E-cells) to TGFβ1 that mimics phenotypic EMT (M-cells) and MET. We aimed at understanding whether phenotypic MET is accompanied by molecular and functional reversion back to epithelia, by using RNA sequencing, Immunofluorescence (IF), proliferation, wound healing, focus formation and mamosphere formation assays, as well as cell-xenografts in nude mice. Phenotypic reverted-epithelial cells (RE-cells), obtained after MET induction, presented pure epithelial morphology and proliferation rate resembling E-cells. However, RE transcriptomic profile and IF staining of epithelial and mesenchymal markers revealed a unique and heterogeneous mixture of cell-subpopulations, with high self-renewal ability fed by oxidative phosporylation. RE-cells heterogeneity is stably maintained for long periods after TGFβ1 removal, both in vitro and in large derived tumours in nude mice. Overall, we show that phenotypic reverted-epithelial cells (RE-cells) do not return to the molecular and functional epithelial state, present mesenchymal features related with aggressiveness and cellular heterogeneity that favour tumour growth in vivo. This work strengthens epithelial cells reprogramming and cellular heterogeneity fostered by inflammatory cues as a tumour-growth promoting factor in vivo.

Henoch–Schönlein purpura (HSP) is characterized by nonthrombocytopenic palpable purpura, arthritis or arthral­gia, and gastrointestinal and/or renal involvement. Gastrointestinal symptoms are reported in 50%-75% and they are related to a previous group A streptococcal infection in 40%. A healthy 5-year-old girl presented with a three-week history of a recurrent purpuric rash on the lower limbs, ar­thralgia and angioedema, without renal involvement. During the third relapse, she had severe, diffuse and persistent abdominal pain and bloody stools. An abdominal ultrasound revealed transmural edema of the last ileal segment, compatible with ileitis. She received prednisolone for five days, with full clinical recovery. Antistreptolysin O titer was elevated. The remaining laboratory tests were normal (antinuclear, anti-neutrophil and anti-Saccharomyces cerevisiae antibodies; rheumatoid factor; stool cultures, parasitological examination and viral antigen tests). One month later, an abdominal ultrasound revealed no abnormalities. Terminal ileitis is a very rare complication of HSP in children but has a good prognosis.

At red wine pH, malvidin-3-glucoside (mv-3-glc), the major anthocyanin of red wine, is expected to be present mainly in its non-colored hemiketal form. However, due to copigmentation with flavanols (e.g. epicatechin), the stabilization of the colored forms of mv-3-glc occurs. Some flavanols have been linked to astringency, due to their ability to interact/precipitate salivary proteins, namely proline-rich proteins (PRPs). So, a major question is if this copigmentation interaction could affect the ability of flavanols to interact with SP. To answer this, the effect of the interaction between mv-3-glc and epicatechin with basic and acidic PRPs, was investigated by saturation-tranfer difference (STD)-NMR and isothermal titration calorimetry (ITC). The most relevant result was that epicatechin:mv-3-glc mixture presents a synergic effect toward the interaction with both PRPs when compared to individual polyphenols. Furthermore, was observed that epicatechin interaction was driven by hydrophobic and hydrophilic interactions while mv-3-glc interaction was driven by electrostatic interactions.

The T cell receptor (TCR) expressed by T lymphocytes initiates protective immune responses to pathogens and tumors. To explore the structural basis of how TCR signaling is initiated when the receptor binds to peptide-loaded major histocompatibility complex (pMHC) molecules, we used cryogenic electron microscopy to determine the structure of a tumor-reactive TCRαβ/CD3δγε2ζ2 complex bound to a melanoma-specific human class I pMHC at 3.08 Å resolution. The antigen-bound complex comprises 11 subunits stabilized by multivalent interactions across three structural layers, with clustered membrane-proximal cystines stabilizing the CD3-εδ and CD3-εγ heterodimers. Extra density sandwiched between transmembrane helices reveals the involvement of sterol lipids in TCR assembly. The geometry of the pMHC/TCR complex suggests that efficient TCR scanning of pMHC requires accurate pre-positioning of T cell and antigen-presenting cell membranes. Comparisons of the ligand-bound and unliganded receptors, along with molecular dynamics simulations, indicate that TCRs can be triggered in the absence of spontaneous structural rearrangements.

Funding Acknowledgements Type of funding sources: Other. Main funding source(s): MG and CS were supported by a PhD FCT grant (SFRH/BD/128893/2017) and by an individual grant from CAPES [BEX 0554/14-6], respectively. This work was financially supported by the project POCI-01-0145-FEDER-030011, funded by FEDER, through COMPETE2020-POCI, and by national funds, through FCT/MCTES (PTDC/MEC-CAR/30011/2017). iBiMED is a research unit supported by the Portuguese Foundation for Science and Technology (REF: UID/BIM/04501/2020) and FEDER/Compete2020 funds). Introduction Amyloid-like protein aggregates play a decisive role in the pathology of heart failure. Alterations in protein homeostasis, in particular, the clearance of toxic amyloid-like aggregates are emerging therapeutic targets in cardiovascular medicine. The clinical benefits of cardiac rehabilitation and exercise training are widely accepted in heart failure; however, little is known about the potential benefit of exercise training in amyloid-like protein aggregates. Purpose To assess the effects of a moderate-intensity exercise training program on amyloid-like protein aggregates levels among patients with heart failure with reduced ejection fraction. Methods Eighteen subjects participated in the study; eight patients (age: 66.6 ± 5.9 years; FEVE: 38.4 ± 8.9%) with heart failure with reduced ejection fraction participated in a 3-month exercise training program (2 x 60 min sessions per week of moderate-intensity aerobic and resistance exercise). Ten healthy subjects (age: 68. 4 ± 3.1 years) were recruited to an age-matched reference group. Amyloid-like protein aggregates were assessed before and after 3 months of exercise training. Clinical data, medication, anthropometrics, and cardiorespiratory fitness were also assessed. Thioflavin T (ThT) dye fluorescence was used to quantify the plasma levels of amyloid-like aggregates and the Fourier transform infrared spectroscopy (FTIR) was applied to evaluate the conformation of cross-β-sheet structures characteristic of amyloid protein aggregates. Results Exercise program improved cardiorespiratory fitness by 14.0 ± 17.1% (17.4 ± 3.2 to 19.7 ± 2.9 ml/kg/min) and reduced NT-proBNP levels by 16.5% (34.2) (median concentration of 632 pg/mL (720.8) to 517.5 pg/mL (707.0)) in the heart failure patients. A slight decrease of amyloid-like aggregates levels was observed in post-exercise training samples (a reduction of 3.1%); interestingly, after the exercise training program, the heart failure patients showed levels of amyloid-like aggregates similar to the reference group (1132.0 ± 114.2 vs. 1094.8 ± 132.9 a.u.). Additionally, the PLS-R multivariate analysis of the amide I region of the FTIR spectra revealed enrichment of antiparallel β-sheets (1693 cm-1) assigned to amyloid-like oligomers in the samples of heart failure patients before, but not after, the exercise program. Of note, oligomeric species, as intermediates of amyloid assembly, can contribute to the increase of amyloid burden, but also, some have been reported to be highly reactive and toxic to cells, being key elements of amyloid pathogenesis. Conclusions Our preliminary results indicate that 3 months of exercise training may have significant effects on amyloid-like oligomers, and start hindering the formation of the larger ThT-positive aggregates among patients with heart failure. Abstract Figure.

Introduction With the widespread introduction of conjugate meningococcal and pneumococcal vaccines, the prevalence and etiology of invasive bacterial infections have changed. We aimed to review all cases of bacteremia in a level II pediatric department over a ten-year period in the post-pneumococcal conjugate vaccine era. Methods We reviewed all positive blood cultures (BC) obtained in our department between 2007 and 2016. Results were classified as contaminants, potential pathogens or confirmed pathogens, based on species, number of positive BC in the episode and the patients’ medical history. Demographic and clinical data were collected for patients with identified pathogens. Results A total of 638 positive BC were identified (6.6% of total BC); 120 (1.2%) were considered to represent true bacteremia. The most frequently identified microorganism was Streptococcus pneumoniae (29.2%), with a decrease in the number of cases between 2008 and 2015. Staphylococcus aureus was the second most common organism (19.2%) being 21.7% of these methicillin-resistant. Escherichia coli was the most common isolate in children aged less than three months. Conclusion We found a rate of true bacteremia in children similar to recent studies. Although Streptococcus pneumoniae remains the most common microorganism, its prevalence may be declining. Monitoring microbiological data in children has implications in practice, particularly in local antibiotic prescription.

Background Pre-infarct angina (PIA) has been shown to reduce reperfusion injury and infarct size in patients with ST-elevation myocardial infarction (STEMI) and currently represents the most efficient form of myocardial conditioning yet discovered. The role of diabetes on ischemic preconditioning remains controversial – while some pre-clinical studies suggest that diabetes blunts ischemic conditioning, clinical studies are lacking. Methods We retrospectively evaluated consecutive patients with STEMI admitted in our hospital from January 2008 to August 2018 who underwent primary angioplasty (PCI). PIA was defined as chest, arm or jaw pain during the preceding 48h before STEMI diagnosis. Peak creatine kinase and peak Troponin T levels were used as a surrogate of infarct size. Ischemic time (IT) was defined as the time between the onset of symptoms to the restoration of flow after either guidewire passage, thrombus aspiration or first balloon inflation. Results Of the 1143 included patients, 74% (n=845) were male and mean age was 62.6±13.1 years. A quarter of STEMI-patients had diabetes (25%, n=285). Almost a third of the patients (32%, n=359) had a history of angina in the preceding 48h before STEMI (PIA). The proportion of PIA was similar between diabetic and non-diabetic patients. In patients with diabetes, PIA was associated with lower creatine kinase (CK) (1144 [500–2212] vs 1715 [908–3309] U/L, p=0.0029) and Troponin T (TnT 3.30 [1.90–6.58] vs 4.88 [2.50–9.58] ng/mL, p=0.0022) despite similar IT as compared to those without PIA (328 [200–554] vs. 258 [180–530] minutes, p=0.1365). In non-diabetic patients, PIA was not significantly associated with infarct size (TnT 3.74 [2.23–7.11] vs 4.56 [2.44–7.77] ng/mL, p=0.1945; CK 1549 [910 - 2909] vs 1793 [996 - 3078] U/L, p=0.0653) even after adjustment for the increased ischemic time (240 [150–550] vs. 210 [140–405] minutes, p=0.0128) (β=−0.12, p=0.085 for CK and β=−0.11, p=0.183 for TnT). A significant interaction was observed between the existence of PIA and diabetes on peak TnT (p=0.026 for interaction) and CK (p=0.047 for interaction), which was independent of the culprit vessel and IT. During a median follow-up period of 18.0 [12.1–25.5] months, 268 (24.0%) MACE events have occurred (165 deaths, 27 strokes, 46 myocardial infarctions and 26 target vessel revascularization). PIA was associated with a significant reduction in the incidence of MACE (HR 0.66 (95% CI: 0.48–0.89)) driven by a reduction on mortality (HR 0.44 (95% CI: 0.28–0.70)). Diabetes was associated with an increased incidence of MACE (HR 1.42 (95% CI: 1.07–1.89)). No interaction was found between diabetes and PIA on their effect on MACCE events. Conclusion PIA is a strong predictor of favourable outcomes in the setting of STEMI. The effect of PIA on myocardial protection in patients with STEMI undergoing primary PCI seems to be modulated by the presence of diabetes. Distribution of Peak CK and Peak TnT Funding Acknowledgement Type of funding source: None

Introduction and Objectives: Acute appendicitis (AA) is one of the most common causes of urgent surgery in children and is still associated with diagnostic difficulties. As the red blood width (RDW) and mean platelet volume (MVP) may be related to inflammatory processes, this study aims to evaluate the diagnostic value of RDW in AA in pediatric population. Methods: A retrospective and observational study was conducted. Data was collected from medical records of all patients with clinical suspicion of AA who underwent appendectomy at a secundary hospital during a period of 36 months. Preoperative analytical values were analyzed and compared with a control group. Results: Medical records of 305 patients with a mean age of 12.1 years were analyzed. Children with simple or perforated appendicitis had significantly higher RDW than those with a normal appendix. Compared with the controls mean RDW was significantly higher in the acute appendicitis group (14.16 ± 1.2% vs.13.09 ± 0.9%, p

Polyphenols may contribute directly to plant-based foodstuffs flavor, in particular to astringency and bitterness. In this work, the bitterness of a small library of polyphenols from different classes [procyanidin dimers type B, ellagitannins (punicalagin, castalagin, and vescalagin) and phenolic acid ethyl esters (protocatechuic, ferulic, and vanillic acid ethyl esters] was studied by a cell-based assay. The bitter taste receptors (TAS2Rs) activated by these polyphenols and the half-maximum effective concentrations (EC50) of each agonist-TAS2Rs pair were determined. Computational methodologies were used to understand the polyphenol molecular region responsible for receptor activation and to get insights into the type of bonds established in the agonist-TAS2Rs pairs. The results show the combinatorial pattern of TAS2Rs activation. TAS2R5 seems to be the only receptor exhibiting a bias toward the activation by condensed tannins, while TAS2R7 seems more tuned for hydrolyzable (ellagi)tannins. Additionally, at the concentrations usually found for these compounds in foodstuffs, they can actively contribute to bitter taste, especially ellagitannins.

Las fracturas por estrés afectan, con mayor frecuencia, a personas físicamente activas con hueso normal y son infrecuentes en los niños con placa de crecimiento abierta. Aun más infrecuentes son las fracturas por estrés del cuello femoral en la población pediátrica. Sin embargo, constituyen entidades muy importantes debido al riesgo de complicaciones graves, como la necrosis avascular. Se describe el caso de una niña de 7 años medicada con metilfenidato que sufrió una fractura por estrés del cuello del fémur atípica. La paciente consulta por dolor inguinal derecho sin limitaciones en las actividades cotidianas. La radiografía muestra una fractura por estrés del cuello del fémur, que se confirma con tomografía. Se instaura un tratamiento conservador, y la paciente está asintomática a las cuatro semanas. Este caso representa una alerta sobre esta infrecuente entidad en la que podrían presentarse errores diagnósticos. Investigaciones recientes también sugieren la posible participación de fármacos, como el metilfenidato, en la desmineralización ósea, que podría constituir un posible factor de riesgo de fractura. AbstractStress fractures most commonly affect physically active individuals with normal bone, and they are rare in children with open growth plates. Even rarer are femoral neck stress fractures in pediatric age. Nevertheless, they constitute a very important entity due to the risk of severe complications, such as avascular necrosis. A seven-year-old girl, treated with methylphenidate, who suffered an atypical femoral neck stress fracture is presented. Patient complained of right inguinal pain without daily life activity limitations. The radiograph showed a stress femoral neck fracture, confirmed by CT imaging. Conservative treatment was instituted and the patient became asymptomatic after 4 weeks.This case report alerts the clinicians about this rare entity that might be misdiagnosed. Recent research also suggests a possible involvement of drugs, such as methylphenidate, in bone demineralization, which might constitute a possible risk factor for fractures.

Deregulation of tRNAs, aminoacyl-tRNA synthetases (aaRS) or tRNA modifying enzymes, increase the level of protein synthesis errors (PSE) and are associated with several diseases, but the cause-effect mechanisms of these pathologies remain elusive. To clarify the role of PSE in human biology, we have engineered a HEK293 cell line to overexpress a wild type (Wt) tRNASer and two tRNASer mutants that misincorporate serine at non-cognate codon sites. Then, we followed long-term adaptation to PSE of such recombinant cells by analysing cell viability, protein synthesis rate and activation of protein quality control mechanisms (PQC). Engineered cells showed higher level of misfolded and aggregated proteins; activated the ubiquitin-proteasome system (UPS) and the unfolded protein response (UPR), indicative of proteotoxic stress. Adaptation to PSE involved increased protein turnover, UPR up-regulation and altered protein synthesis rate. Gene expression analysis showed that engineered cells presented recurrent alterations in the endoplasmic reticulum, cell adhesion and calcium homeostasis. Herein, we unveil new phenotypic consequences of protein synthesis errors in human cells and identify the protein quality control processes that are necessary for long-term adaptation to PSE and proteotoxic stress. Our data provide important insight on how chronic proteotoxic stress may cause disease and highlight potential biological pathways that support the association of PSE with disease. published

Vaginal bleeding can occur shortly after delivery in 3%–5% of newborns as a consequence of placental hormone withdrawal . Although usually benign, its differential diagnosis includes central precocious puberty, tumours and other pathological conditions. A girl born at 26 weeks of gestation presented with five episodes of vaginal bleeding, each lasting less than a week, initiated at 4 months of age. Luteinising hormone and oestradiol levels were in the pubertal range. Later, she exhibited breast development, with no other pubertal signs. An ultrasonography test revealed an impregnated endometrium and a right ovarian cyst with 43 mm of diameter. A cranioencephalic MRI was unremarkable. Clinicians adopted expectant management and there was clinical, hormonal and radiological resolution in 3 months. The spontaneous resolution suggested mini-puberty of infancy. This is usually an asymptomatic condition, but to date, four cases of an exacerbated form in extremepremature infants have been reported. Long-term follow-up data are missing.A girl born at 26 weeks of gestation presented with five episodes of vaginal bleeding, each lasting less than a week, initiated at 4 months of age. Luteinising hormone and oestradiol levels were in the pubertal range. Later, she exhibited breast development, with no other pubertal signs. An ultrasonography test revealed an impregnated endometrium and a right ovarian cyst with 43 mm of diameter. A cranioencephalic MRI was unremarkable. Clinicians adopted expectant management and there was clinical, hormonal and radiological resolution in 3 months. The spontaneous resolution suggested mini-puberty of infancy. This is usually an asymptomatic condition, but to date, four cases of an exacerbated form in extremepremature infants have been reported. Long-term follow-up data are missing.

Context: i* is one of the most influential languages in the Requirements Engineering research community. Perhaps due to its complexity and low adoption in industry, it became a natural candidate for studies aiming at improving its concrete syntax and the stakeholders' ability to correctly interpret i* models. Objectives: We evaluate the impact of semantic transparency on understanding and reviewing i* models, in the presence of a language key. Methods: We performed a quasi-experiment comparing the standard i* concrete syntax with an alternative that has an increased semantic transparency. We asked 57 novice participants to perform understanding and reviewing tasks on i* models, and measured their accuracy, speed and ease, using metrics of task success, time and effort, collected with eye-tracking and participants' feedback. Results: We found no evidence of improved accuracy or speed attributable to the alternative concrete syntax. Although participants' perceived ease was similar, they devoted significantly less visual effort to the model and the provided language key, when using the alternative concrete syntax. Conclusions: The context provided by the model and language key may mitigate the i* symbol recognition deficit reported in previous works. However, the alternative concrete syntax required a significantly lower visual effort.

FCT-MCTES SFRH/BD/108492/2015 Stakeholders without formal training in requirements modelling languages, such as KAOS, struggle to understand requirements specifications. The lack of semantic transparency of the KAOS goal model concrete syntax is perceived as a communication barrier between stakeholders and requirements engineers. We report on a series of related empirical experiments that include the proposal of alternative concrete syntaxes for KAOS by leveraging design contributions from novices and their evaluation with respect to semantic transparency, in contrast with the standard KAOS goal model concrete syntax. We propose an alternative concrete syntax for KAOS that increases its semantic transparency (mean difference of.23, in [−1.00.1.00]) leading to a significantly higher correct symbol identification (mean difference of 19%) by novices. These results may be a stepping stone for reducing the communication gap between stakeholders and requirements engineers. preprint published

Polyphenols interaction with salivary proteins (SP) has been related with organoleptic features such as astringency. The aim of this work was to study the interaction between some human SP and tannins through two spectroscopic techniques, fluorescence quenching, and saturation transfer difference-nuclear magnetic resonance (STD-NMR). Generally, the results showed a significant interaction between SP and both condensed tannins and ellagitannins. Herein, STD-NMR proved to be a useful tool to map tannins’ epitopes of binding, while fluorescence quenching allowed one to discriminate binding affinities. Ellagitannins showed the greatest binding constants values (KSV from 20.1 to 94.1 mM–1; KA from 0.7 to 8.3 mM–1) in comparison with procyanidins (KSV from 5.4 to 40.0 mM–1; KA from 1.1 to 2.7 mM–1). In fact, punicalagin was the tannin that demonstrated the highest affinity for all three SP. Regarding SP, P-B peptide was the one with higher affinity for ellagitannins. On the other hand, cystatins showed in gener...

Innate and adaptive cellular immunity is dependent on interactions of cell surface receptors that initiate signaling, resulting in the formation of the immunological synapse and targeted delivery of effector functions. There has been considerable interest over the past 30 years in methods for isolating the extracellular regions of these receptors and components of the cytoplasmic signaling networks. This chapter describes our current protein expression toolkit used for structural studies of signaling proteins and the functional reconstitution of model cell surfaces, which comprises both bacterial and mammalian cell-based protein expression methodologies.

Congenital aniridia is a rare genetic disorder characterized by varying degrees of iris hypoplasia that are associated with additional ocular abnormalities. More than 90% of the causal mutations identified are found in the PAX6 gene, a transcription factor of critical importance in the process of neurogenesis and ocular development. Here, we investigate clinical, molecular, and craniofacial features of a large Brazilian family with congenital aniridia. Among the 56 eyes evaluated, phenotype variation encompassed bilateral total aniridia to mild iris defects with extensive variation between eyes of the same individual. PAX6 molecular screening indicated a heterozygous splice mutation (c.141 + 1G>A). Thus, we hypothesize that this splicing event may cause variation in the expression of the wild-type transcript, which may lead to the observed variation in phenotype. Affected individuals were more brachycephalic, even though their face height and cephalic circumference were not significantly different when compared to those of non-affected relatives. From this, we infer that the head shape of affected subjects may also be a result of the PAX6 splice-site mutation. Our data summarize the clinical variability associated with the ocular phenotype in a large family with aniridia, and help shed light on the role of PAX6 in neurocranial development. Fil: Fernandes Lima, Z.S.. Universidade Federal do Rio Grande do Sul; Brasil Fil: Paixão-Côrtes, V.R.. Universidade Federal do Rio Grande do Sul; Brasil Fil: de Andrade, A.K.M.. Faculdade de Medicina; Brasil Fil: Fernandes, A.S.. Universidade Federal de Alagoas; Brasil Fil: Coronado, B.N.L.. Universidade Federal de Alagoas; Brasil Fil: Monte Filho, H.P.. Universidade Federal de Alagoas; Brasil Fil: Santos, M.J.. Universidade Federal de Alagoas; Brasil Fil: Omena Filho, R.L.. Universidade Federal de Alagoas; Brasil Fil: Biondi, Facundo Carmelo. Universidade Federal do Rio Grande do Sul; Brasil Fil: Ruiz Linares, A.. University College London; Estados Unidos Fil: Ramallo, Virginia. Universidade Federal do Rio Grande do Sul; Brasil. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Hunemeier, T.. Universidade Federal do Rio Grande do Sul; Brasil Fil: Schuler Faccini, L.. Universidade Federal do Rio Grande do Sul; Brasil Fil: Monlleo, I.L.. Universidade Federal de Alagoas; Brasil

Invasive pneumococcal disease predominantly affects younger children, elderly, and immunocompromised patients. Pneumococcal meningitis is a particularly important form of presentation, considering its high rate of morbimortality. We present the case of a previously healthy 12-year-old adolescent male who was hospitalized due to suspicion of osteoarticular infection in his left foot. A few hours later, he developed meningeal signs, exhibiting slight pleocytosis andStreptococcus pneumoniaeisolates in both cerebrospinal fluid and blood. Imaging studies were inconclusive regarding the nature of the foot disorder. We considered the hypothesis of osteomyelitis of the navicular bone as the most likely, for which he completed six weeks of antibiotic therapy. There was a favorable clinical evolution, along with complete absence of osteoarticular or neurological sequelae. The relevance of this clinical case resides in the unusual presentation of invasive pneumococcal disease in this age group, as well as in the rare form of orthopedic involvement.

Astringency is closely related to the interaction between procyanidins and salivary proteins (SP). The aim of this work was to study the interaction between mucin, a SP responsible for saliva lubricating properties, with different procyanidins (B4 dimer, tetramer (TT) and fractions of oligomeric procyanidins), and the influence of several conditions [pH, ionic strength, procyanidins' mean degree of polymerization (mDP) and different solvents (ethanol or dimethylsulfoxide)] on this interaction by fluorescence quenching and Saturation Transfer Difference (STD)-NMR. For fractions of oligomeric procyanidins, the mucin-procyanidin interaction increased with mDP; however, for pure compounds, procyanidin TT has lower affinity than dimer B4 which could be due to a lower structural flexibility imposed by its complex structure. Furthermore, EtOH and DMSO can disrupt the main driving forces of these interactions, hydrophobic interactions and hydrogen bonds, respectively, lowering significantly the binding constants.

The i* community has published guidelines, including model layout guidelines, for the construction of models. Our goal is to evaluate the effect of the layout guidelines on the i* novice stakeholders' ability to understand and review i* models. We performed a quasi-experiment where participants were given two understanding and two reviewing tasks. Both tasks involved a model with a bad layout and another model following the i* layout guidelines. We evaluated the impact of layouts by combining the success level in those tasks and the required effort to accomplish them. Effort was assessed using time, perceived complexity (with NASA TLX), and eye-tracking data. Participants were more successful in understanding than in reviewing tasks. However, we found no statistically significant difference in the success, time taken, or perceived complexity, between tasks conducted with models with a bad layout and models with a good layout. Most participants had little to no prior knowledge in i*, making them more representative of stakeholders with no requirements engineering expertise. They were able to understand the models fairly well after a short tutorial, but struggled when reviewing models. In the end, adherence to the existing i* layout guidelines did not significantly impact i* model understanding and reviewing performance.

// Ilaria Guerriero 1 , Daniela D’Angelo 2 , Pierlorenzo Pallante 2 , Mafalda Santos 1 , Marianna Scrima 1 , Donatella Malanga 3 , Carmela De Marco 3 ,Maria Ravo 4 , Alessandro Weisz 4 , Carmelo Laudanna 3 , Michele Ceccarelli 1, 5 , Geppino Falco 1, 6 , Antonia Rizzuto 7 , Giuseppe Viglietto 1, 3 1 Biogems.c.ar.l., Istituto di Ricerche Genetiche “Gaetano Salvatore”, Ariano Irpino, Avellino, Italy 2 Istituto per l’Endocrinologia e l’Oncologia Sperimentale, IEOS-CNR, c/o Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Universita “Federico II” of Napoli, Napoli, Italy 3 Dipartimento di Medicina Sperimentale e Clinica, Universita Magna Graecia of Catanzaro, Italy 4 Laboratorio di Medicina Molecolare e Genomica, Dipartimentodi Medicina e Chirurgia, Universita di Salerno, Baronissi, Italy 5 Department of Biological and Environmental Studies, Universita del Sannio, Benevento, Italy 6 Dipartimento di Biologia, Universita degli Studi di Napoli Federico II, Complesso Universitario Monte S.Angelo, Napoli, Italy 7 Dipartimento di Scienze Mediche e Chirurgiche, Universita Magna Graecia of Catanzaro, Italy Correspondence to: Giuseppe Viglietto, email: viglietto@unicz.it Keywords: NSCLC, PI3K/AKT, microRNA, miR-196a Received: January 11, 2016 Accepted: November 02, 2016 Published: November 17, 2016 ABSTRACT Hyperactivation of the PI3K/AKT pathway is observed in most human cancer including lung carcinomas. Here we have investigated the role of miRNAs as downstream targets of activated PI3K/AKT signaling in Non Small Cell Lung Cancer (NSCLC). To this aim, miRNA profiling was performed in human lung epithelial cells (BEAS-2B) expressing active AKT1 (BEAS-AKT1-E17K), active PI3KCA (BEAS-PIK3CA-E545K) or with silenced PTEN (BEAS-shPTEN). Twenty-four differentially expressed miRNAs common to BEAS-AKT1-E17K, BEAS-PIK3CA-E545K and BEAS-shPTEN cells were identified through this analysis, with miR-196a being the most consistently up-regulated miRNA. Interestingly, miR-196a was significantly overexpressed also in human NSCLC-derived cell lines (n=11) and primary lung cancer samples (n=28). By manipulating the expression of miR-196a in BEAS-2B and NCI-H460 cells, we obtained compelling evidence that this miRNA acts downstream the PI3K/AKT pathway, mediating some of the proliferative, pro-migratory and tumorigenic activity that this pathway exerts in lung epithelial cells, possibly through the regulation of FoxO1, CDKN1B (hereafter p27) and HOXA9.

Sternoclavicular arthritis is an unusual osteoarticular infection and can be associated with severe complications. Cases in a paediatric population are infrequently reported, making this approach challenging. Kingella kingae is an agent of increasing recognition in paediatric invasive infections, principally below 2 years of age. A case of K. kingae osteoarthritis in a 17-month-old child is described with a review of the literature.

Introducao: O termo «crise hipertensiva» corresponde a uma variedade de situacoes clinicas que diferem entre si, pela gravidade dos valores da pressao arterial e pela necessidade de reducao mais ou menos rapida dos mesmos. Esta entidade e arbitrariamente definida como qualquer elevacao da pressao arterial diastolica acima de 120 mmHg e sintomas acompanhantes relacionados. Dependendo da existencia de lesao de orgao-alvo aguda ou rapidamente progressiva, a «crise hipertensiva» pode manifestar-se como urgencia ou emergencia hipertensiva. Objectivos: Pretende-se apresentar uma breve revisao de conceitos basicos sobre o que e, e como diferenciar, uma verdadeira crise hipertensiva (urgencia ou emergencia hipertensiva) de uma «pseudocrise», qual a marcha diagnostica desta patologia e qual a terapeutica mais adequada a implementar. Metodos: Revisao bibliografica incluindo tratados de referencia e artigos recentemente publicados na area de hipertensao arterial, nomeadamente referentes a urgencias e emergencias hipertensivas. Conclusao: As urgencias hipertensivas e as emergencias hipertensivas sao igualmente denominadas «crises hipertensivas» e ambas se caracterizam por uma elevacao subita e sintomatica da pressao arterial que pode ou nao associar-se a uma lesao de orgao-alvo e/ou envolver risco de morte imediato ou potencial. O doente que se encontra numa situacao de emergencia hipertensiva, se considerado em estado grave e/ou agudo, deve ser internado numa unidade de cuidados intensivos, com monitorizacao da pressao arterial de forma continua e cuidadosa, sendo necessaria uma reducao imediata da pressao arterial em minutos ou horas. A reducao nao necessita de atingir valores «normais» mas sim valores que possam afastar o risco de morte e impedir o agravamento ou deterioracao de orgao-alvo. As urgencias hipertensivas definem-se como elevacoes da pressao arterial sistolica e/ou diastolica sem evidencia de lesao de orgao-alvo a curto prazo, sendo necessaria uma reducao gradual da pressao arterial em 24-48 horas para que ocorra a recuperacao e a readaptacao do doente.

The regioselective addition of deprotonated alkynes (phenyl-1-propyne, propargyl ether or tetrahydropyran-protected 3-butyn-1-ol) to the imine group was identified as a competitive process to the nucleophilic addition to the keto group of (1S)-3-oxo-camphorsulfonylimine. The selectivity of the process depends on the characteristics of the nucleophile and the reaction conditions. In the case of propargyl ether it was possible to render the imine addition the main process. The structural characterization of compounds 1, 2, 6 by X-ray diffraction analysis show that the C2 - C3 bond becomes longer upon nucleophilic addition to the imine group of (1S)-3-oxo-camphorsulfonylimine. This trend is believed to favour the ring opening process that undergoes the formation of the spiro type compound 7.

The urinary tract infections associated with catheterization are very common in hospital and home care contexts. Currently there are several recommendations for its prevention, however, when approaching the kind of solute used in the urinary meatus prior to catheterization doubts continue to persist. Thus this study aimed at determining the effectiveness of cleaning the urinary meatus with water or saline comparing to its sterilization through a systematic review and meta-analysis. In order to do so, the principles proposed by the Cochrane Handbook were followed, a critical analysis was conducted by two researchers and the statistical analysis was performed with the use of STATA 11.1. We concluded that the cleaning or disinfection of the urinary canal prior to bladder catheterization is not statistically significant (OR=1.07, CI 95%=0.68-1.68, p=0.779) and that there is some evidence that the use of water/saline reduces rates of UTI (urinary tract infection).

Systemic sclerosis (SS) is a rare severe autoimmune disease involving the connective tissue. The pathophysiology is not clearly understood. It is characterized by a remarkable clinical heterogeneity, and virtually all organs can be affected. Concerning diagnosis, the presence of antinuclear antibodies (ANA) can be found in more than 90% of patients, but the diagnosis is made gathering clinical manifestations, autoimmune panel, nailfold capillaroscopy and in some cases biopsy of the organ involved. The disease course is also weakly understood, although some serological patterns can be distinguished. Current therapeutic options target few aspects of pathologic mechanism and clinical management remains a challenge.The authors presented a rare case of a SS ANA negative, which demonstrates the diagnostic challenge of this disease.

Epidermal Nevus Syndrome (ENS) is characterized by epidermal nevi associated with abnormalities involving the nervous, skeletal, and other systems. Rarely, hypophosphatemic rickets has been observed in association with epidermal nevi. A patient with ENS with right-sided serpiginous skin lesions, generalized weakness, and diffuse osteopenia associated with hypophosphatemic rickets is described. Medical management was enough to correct the clinical picture. The pathogenic mechanism involved in the onset of hypophosphatemic rickets in ENS is not fully clarified. Different studies suggest that phosphaturia, caused by circulating factor(s), called "phosphatonin(s)," may be secreted by an epidermal nevus. The nature of the phosphaturic factor(s) is not well understood, but elevated levels of circulating FGF-23 were recently reported in one patient with hypophosphatemic rickets. The authors suggest that serum FGF-23 measurement be included in the workup of this kind of rickets because there is growing evidence that in these situations the epidermal nevi produce a phosphaturic factor.

Focal fibrocartilaginous dysplasia is a rare benign entity, previously described as a factor responsible for causing tibia vara at the walking age. We report here the case of a child with tibia valga due to focal fibrocartilaginous dysplasia of the lateral part of the proximal tibia. To our knowledge, this is the first observation of a lesion located in the lateral proximal tibia. Clinical, radiographic and magnetic resonance imaging surveillance was performed. Spontaneous resolution of the lesion with correction of the angular deformity did occur.