Modelling aptamers with nucleic acid mimics (NAM): From sequence to three-dimensional docking.

Aptamers are single-stranded oligonucleotides, formerly evolved by Systematic Evolution of Ligands by EXponential enrichment (SELEX), that fold into functional three-dimensional structures. Such conformation is crucial for aptamers' ability to bind to a target with high affinity and specificity. Unnatural nucleotides have been used to develop nucleic acid mimic (NAM) aptamers with increased performance, such as biological stability. Prior knowledge of aptamer-target interactions is critical for applying post-SELEX modifications with unnatural nucleotides since it can affect aptamers' structure and performance. Here, we describe an easy-to-apply in silico workflow using free available software / web servers to predict the tertiary conformation of NAM, DNA and RNA aptamers, as well as the docking with the target molecule. Representative 2'-O-methyl (2'OMe), locked nucleic acid (LNA), DNA and RNA aptamers, with experimental data deposited in Protein Data Bank, were selected to validate the workflow. All aptamers' tertiary structure and docking models were successfully predicted with good structural similarity to the experimental data. Thus, this workflow will boost the development of aptamers, particularly NAM aptamers, by assisting in the rational modification of specific nucleotides and avoiding trial-and-error approaches.

Improving aptamer performance with nucleic acid mimics: de novo and post-SELEX approaches.

Aptamers are structural single-stranded oligonucleotides generated in vitro to bind to a specific target molecule. Aptamers' versatility can be enhanced with nucleic acid mimics (NAMs) during or after a selection process, also known as systematic evolution of ligands by exponential enrichment (SELEX). We address advantages and limitations of the technologies used to generate NAM aptamers, especially the applicability of existing engineered polymerases to replicate NAMs and methodologies to improve aptamers after SELEX. We also discuss the limitations of existing methods for sequencing NAM sequences and bioinformatic tools to predict NAM aptamer structures. As a conclusion, we suggest that NAM aptamers might successfully compete with molecular tools based on proteins such as antibodies for future application.

Matrix Metalloproteinase-9 Levels are Associated with Brain Lesion and Persistent Venous Occlusion in Patients with Cerebral Venous Thrombosis.

BACKGROUND: Elucidating mechanisms of brain damage in cerebral venous thrombosis (CVT) would be instrumental to develop targeted therapies and improve prognosis prediction. Matrix metalloproteinase-9 (MMP-9), a gelatinase that degrades major components of the basal lamina, has been associated to blood-brain barrier disruption. We aimed to assess, in patients with CVT, the temporal change in serum concentrations of MMP-9 and its association with key imaging and clinical outcomes. METHODS: Pathophysiology of Venous Infarction-PRediction of InfarctiOn and RecanalIzaTion in CVT (PRIORITy-CVT) was a multicenter prospective cohort study of patients with newly diagnosed CVT. Serial collection of peripheral blood samples performed on day 1, 3, and 8, and standardized magnetic resonance imaging on day 1, 8, and 90. MMP-9 was quantified using enzyme-linked immunosorbent assay in 59 patients and 22 healthy controls. Primary outcomes were parenchymal brain lesion, early evolution of brain lesion, early recanalization, and functional outcome on day 90. RESULTS: CVT patients with parenchymal brain lesion had higher baseline concentrations of MMP-9 compared with controls (adjusted p = 0.001). The area under receiver operating characteristic curve value for MMP-9 for predicting brain lesion was 0.71 (95% confidence interval [CI]: 0.57-0.85, p = 0.009). Patients with venous recanalization showed early decline of circulating MMP-9 and significantly lower levels on day 8 (p = 0.021). Higher MMP-9 on day 8 was associated with persistent venous occlusion (odds ratio: 1.20 [per 20 ng/mL], 95% CI: 1.02-1.43, p = 0.030). CONCLUSION: We report a novel relationship among MMP-9, parenchymal brain damage, and early venous recanalization, suggesting that circulating MMP-9 is a dynamic marker of brain tissue damage in patients with CVT.

Minho Communication Assessment Scale: Development and Validation.

INTRODUCTION: Verbal and non-verbal communication skills are core competencies in medical practice and should be acquired and monitored in medical schools. However, their assessment poses a challenge. The aim of this study is to develop and assess the psychometric properties, validity and reliability of a communication assessment scale. MATERIAL AND METHODS: We developed a communication assessment scale, composed by five dimensions (Structure, Way of Questioning, Behavior and Posture, Clarity of information and Emotional dimension). Two focus groups and a development group, composed by faculty members and standardized patients experienced in assessment, were responsible for creating the scale. RESULTS: The communication assessment scale was tested on 332 students from the 3rd and 6th year of medical school, with a total of 2754 assessments, performed by faculty members and standardized patients. A descriptive analysis, an exploratory factor analysis, a confirmatory factor analysis and a Cronbach's alpha analysis to establish internal reliability were conducted. DISCUSSION: The Minho Communication Assessment Scale can be effectively used by both faculty members and standardized patients, providing correct students assessment and relevant feedback to the students. The final Minho Communication Assessment Scale has a total of 19 items, being simple and intuitive to use. The exploratory factor analysis and confirmatory factor analysis results were satisfactory. Cronbach's alpha value revealed high internal consistency of Minho Communication Assessment Scale. CONCLUSION: The final Minho Communication Assessment Scale proved to be simple to use and to have very good psychometric properties. Our results show that the Minho Communication Assessment Scale is a valid scale to assess communication skills which can be accurately replicated on objective structured clinical examinations focusing on communication.

Introdução: A comunicação verbal e não-verbal são competências essenciais na prática médica e devem ser adquiridas e monitorizadas nas escolas médicas. Contudo, a sua avaliação representa um desafio. O objetivo deste trabalho consistiu em desenvolver e avaliar as propriedades psicométricas, validade e fiabilidade de uma escala de avaliação de comunicação. Material e Métodos: Desenvolvemos uma escala de avaliação de comunicação, composta por cinco dimensões (Estrutura, Modo de Questionar, Comportamento e Postura, Clareza de Informação e a dimensão Emocional). Dois focus groups e um grupo de desenvolvimento, composto por membros do corpo docente e pacientes estandardizados experienciados em avaliação, foram responsáveis por criar a escala. Resultados: A escala de avaliação de comunicação foi testada em 332 alunos de medicina do terceiro e sexto ano curricular, num total de 2754 avaliações, realizada por docentes e pacientes estandardizados. Realizámos uma análise descritiva, uma análise fatorial exploratória, uma análise factorial confirmatória e uma análise de alfa de Cronbach, de modo a estabelecer a fiabilidade interna da escala. Discussão: A Minho Communication Assessment Scale pode ser usada eficazmente por docentes e pacientes estandardizados, fornecendo uma avaliação correta e um feedback relevante aos estudantes. A Minho Communication Assessment Scale final tem um total de 19 itens, sendo simples e intuitiva de usar. Os resultados da análise fatorial exploratória e confirmatória foram satisfatórios. O valor de alfa de Cronbach's revelou valores elevados de consistência interna. Conclusão: A versão final da Minho Communication Assessment Scale, demonstrou ser simples de usar e possuir muito boas propriedades psicométricas. Os resultados demonstram que a Minho Communication Assessment Scale é uma escala válida para avaliar as capacidades de comunicação e pode ser replicada em objective structured clinical examinations que se foquem nessa avaliação.

Early Recanalization in Patients With Cerebral Venous Thrombosis Treated With Anticoagulation.

Background and Purpose- The hypothesis that venous recanalization prevents progression of venous infarction is not established in patients with cerebral venous thrombosis (CVT). Evidence is also scarce on the association between residual symptoms, particularly headache, and the recanalization grade. We aimed to assess, in patients with CVT treated with standard anticoagulation, (1) the rate of early venous recanalization, (2) whether lack of early recanalization was predictor of parenchymal brain lesion progression, and (3) the prevalence and features of persistent headache according to the recanalization grade achieved. Methods- PRIORITy-CVT (Pathophysiology of Venous Infarction - Prediction of Infarction and Recanalization in CVT) was a multicenter, prospective, cohort study including patients with newly diagnosed CVT. Standardized magnetic resonance imaging was performed at inclusion (≤24 hours of therapeutic anticoagulation), days 8 and 90. Potential imaging predictors of recanalization were predefined and analyzed at each anatomical segment. Primary outcomes were rate of early recanalization and brain lesion progression at day 8. Secondary outcomes were headache (days 8 and 90) and functional outcome (modified Rankin Scale at days 8 and 90). Results- Sixty eight patients with CVT were included, of whom 30 (44%) had parenchymal lesions. At the early follow-up (n=63; 8±2 days), 68% (n=43) of patients had partial recanalization and 6% (n=4) full recanalization. Early recanalization was associated both with early regression (P=0.03) and lower risk of enlargement of nonhemorrhagic lesions (P=0.02). Lesions showing diffusion restriction (n=12) were fully reversible in 66% of cases, particularly in patients showing early venous recanalization. Evidence of new or enlarged hemorrhagic lesions, headache at days 8 and 90, and unfavorable functional outcome at days 8 and 90 were not significantly different in patients achieving recanalization. Conclusions- Venous recanalization started within the first 8 days of therapeutic anticoagulation in most patients with CVT and was associated with early regression of nonhemorrhagic lesions, including venous infarction. There was an association between persistent venous occlusion at day 8 and enlargement of nonhemorrhagic lesions.

High Bone Mineral Density of the Lumbar Spine Is Positively Associated with Breast Cancer.

OBJECTIVE: The objective of this study was to verify possible associations between bone mineral density (BMD) and breast cancer in recently diagnosed women in the Brazilian Mid-west region, considering the menopausal status of patients. METHODS: A case-control study was conducted with 142 cases of breast cancer and 234 controls matched by for age, body mass index (BMI), and menopausal status (pre- and postmenopause), performed in a university hospital in the Brazilian Mid-west. Lumbar spine (L1-L4), femoral neck, and total femur BMD were measured by the dual-energy X-ray absorptiometry (DXA) method. For association, a logistic regression analysis was used. RESULTS: Women in the highest lumbar spine BMD quartile presented had a higher chance of developing breast cancer (OR = 2.31; 1.02-5.25; p = 0.045), after adjusting for the confounding variables. Nonetheless, there were no statistically significant differences in the association between pre- and postmenopause in that quartile and breast cancer. CONCLUSIONS: High lumbar spine BMD was positively associated with breast cancer in the total sample. In evaluating the BMD of the femoral neck and total femur, such an association was not observed.

Pneumocephalus Following Unidentified Dural Puncture: A Case Report with an Unusual Neurological Presentation.

Pneumocephalus is a rare consequence of epidural anesthesia, which may occur following inadvertent or unidentified dural puncture when the loss of resistance to air technique is applied to identify the epidural space. Headache is the most common symptom presented in this condition, usually with sudden onset. This case report describes an unusual presentation of diffuse pneumocephalus after an unidentified dural puncture. The patient (male, 67 years old) was submitted to epidural catheter placement for the treatment of acute exacerbation of ischemic chronic pain using loss of resistance to air technique. No cerebrospinal fluid or blood flashback was observed after needle withdrawal. Shortly after the intervention, the patient presented symptoms of lethargy, apathy, and hypophonia, which are not commonly associated with pneumocephalus. No motor or sensory deficits were detected. Cranial computed tomography showed air in the frontal horn of the left ventricle, subarachnoid space at interhemispheric fissure and basal cisterns, confirming the diagnosis of diffuse pneumocephalus. The patient remained under vigilance with oxygen therapy and the epidural catheter left in place. After 24 hours, cranial computed tomography showed air in the temporal and frontal horns of the left ventricle, with no air in the subarachnoid space. The patient presented no neurological signs or symptoms at this time. Although headache is the most common symptom presented in reported cases of pneumocephalus, this case shows the need for the clinician to be aware of other signs and symptoms that may be indicative of this condition, in order to properly diagnose and treat these patients.Key words: Pneumocephalus, continuous epidural analgesia, ischemic chronic pain, loss-of-resistance to air technique, dural puncture, headache, unusual presentation.

Does the Interplay Between Aging and Neuroinflammation Modulate Alzheimer's Disease Clinical Phenotypes? A Clinico-Pathological Perspective.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder and is the most common cause of dementia worldwide. Cumulative data suggests that neuroinflammation plays a prominent and early role in AD, and there is compelling data from different research groups of age-associated dysregulation of the neuroimmune system. From the clinical point of view, despite clinical resemblance and neuropathological findings, there are important differences between the group of patients with sporadic early-onset (<65 years old) and late-onset AD (>65 years old). Thus, it seems important to understand the age-dependent relationship between neuroinflammation and the underlying biology of AD in order to identify potential explanations for clinical heterogeneity, interpret biomarkers, and promote the best treatment to different clinical AD phenotypes. The study of the delicate balance between pro-inflammatory or anti-inflammatory sides of immune players in the different ages of onset of AD would be important to understand treatment efficacy in clinical trials and eventually, not only direct treatment to early disease stages, but also the possibility of establishing different treatment approaches depending on the age of the patient. In this review, we would like to summarize what is currently known about the interplay between "normal" age associated inflammatory changes and AD pathological mechanisms, and also the potential differences between early-onset and late-onset AD taking into account the age-related neuroimmune background at disease onset.