Structure of the Bacterial Sex F Pilus Reveals an Assembly of a Stoichiometric Protein-Phospholipid Complex.

Conjugative pili are widespread bacterial appendages that play important roles in horizontal gene transfer, in spread of antibiotic resistance genes, and as sites of phage attachment. Among conjugative pili, the F "sex" pilus encoded by the F plasmid is the best functionally characterized, and it is also historically the most important, as the discovery of F-plasmid-mediated conjugation ushered in the era of molecular biology and genetics. Yet, its structure is unknown. Here, we present atomic models of two F family pili, the F and pED208 pili, generated from cryoelectron microscopy reconstructions at 5.0 and 3.6 Å resolution, respectively. These structures reveal that conjugative pili are assemblies of stoichiometric protein-phospholipid units. We further demonstrate that each pilus type binds preferentially to particular phospholipids. These structures provide the molecular basis for F pilus assembly and also shed light on the remarkable properties of conjugative pili in bacterial secretion and phage infection.

Cryo-EM structure of a type IV secretion system.

Bacterial conjugation is the fundamental process of unidirectional transfer of DNAs, often plasmid DNAs, from a donor cell to a recipient cell1. It is the primary means by which antibiotic resistance genes spread among bacterial populations2,3. In Gram-negative bacteria, conjugation is mediated by a large transport apparatus-the conjugative type IV secretion system (T4SS)-produced by the donor cell and embedded in both its outer and inner membranes. The T4SS also elaborates a long extracellular filament-the conjugative pilus-that is essential for DNA transfer4,5. Here we present a high-resolution cryo-electron microscopy (cryo-EM) structure of a 2.8 megadalton T4SS complex composed of 92 polypeptides representing 8 of the 10 essential T4SS components involved in pilus biogenesis. We added the two remaining components to the structural model using co-evolution analysis of protein interfaces, to enable the reconstitution of the entire system including the pilus. This structure describes the exceptionally large protein-protein interaction network required to assemble the many components that constitute a T4SS and provides insights on the unique mechanism by which they elaborate pili.

The DNA replication initiation protein DnaD recognises a specific strand of the Bacillus subtilis chromosome origin.

Genome replication is a fundamental biological activity shared by all organisms. Chromosomal replication proceeds bidirectionally from origins, requiring the loading of two helicases, one for each replisome. However, the molecular mechanisms underpinning helicase loading at bacterial chromosome origins (oriC) are unclear. Here we investigated the essential DNA replication initiation protein DnaD in the model organism Bacillus subtilis. A set of DnaD residues required for ssDNA binding was identified, and photo-crosslinking revealed that this ssDNA binding region interacts preferentially with one strand of oriC. Biochemical and genetic data support the model that DnaD recognizes a new single-stranded DNA (ssDNA) motif located in oriC, the DnaD Recognition Element (DRE). Considered with single particle cryo-electron microscopy (cryo-EM) imaging of DnaD, we propose that the location of the DRE within oriC orchestrates strand-specific recruitment of helicase during DNA replication initiation. These findings significantly advance our mechanistic understanding of bidirectional replication from a bacterial chromosome origin.

Heparin-Bonded Expanded Polytetrafluoroethylene Is a Solution for Infrapopliteal Revascularization in the Absence of an Adequate Autologous Vein Graft.

BACKGROUND: To analyze the outcomes of heparin-bonded expanded polytetrafluoroethylene (HePTFE) graft as an alternative conduit in infrapopliteal revascularization of chronic limb-threatening ischemia (CLTI) in the absence of an autologous vein conduit. METHODS: A single-center retrospective analysis of patients with CLTI submitted to infrapopliteal bypasses with autologous vein graft (VEIN group) or HePTFE graft (HePTFE group) was implemented. Primary end points were freedom from CLTI at 12 months and recurrence of CLTI at 3 years. Secondary end points included freedom from major amputation, amputation-free survival (AFS), survival, and primary (PP) and secondary patency (SP) rates at 3 years of follow-up. RESULTS: A total of 348 limbs submitted to infrapopliteal bypasses, 214 with venous graft and 134 with HePTFE graft, were followed-up for a median of 25 months. Most patients of the HePTFE group were male (69%), with a median age of 76 years (interquartile range [IQR] 15). Fifty-nine percent of the limbs of the HePTFE group had Wound grade ≥2, being 46% of them infected. Eighty-eight percent were GLASS stage III. Freedom from CLTI was not significantly different between HePTFE and VEIN groups (75% vs. 84%, adjusted hazard ratio [aHR] 0.88, confidence interval [CI] 0.66-1.18, P = 0.401). Recurrence of CLTI was higher in the HePTFE group (42% vs.18% at 3 years; aHR 2.82, CI 1.59-5.00, P < 0.001). The VEIN group achieved higher rates of freedom from major amputation (87% vs.69% at 3 years; aHR 2.21, CI 1.31-3.75, P = 0.003) and AFS (59% vs. 37% at 3 years; aHR 1.39, CI 1.02-1.88, P = 0.036), but no significance in survival (aHR 1.10, CI 0.72-1.66, P = 0.667). Patency rates were inferior in the HePTFE group, with 2-year PP and SP rates of 52% vs. 74%, and 76% vs. 90%, respectively (PP: aHR 1.70, CI 1.11-2.59, P = 0.014; SP: aHR 2.51, CI 1.42-4.42, P = 0.001). CONCLUSIONS: Infrapopliteal bypass with autologous vein graft is the gold standard to treat CLTI limbs. HePTFE graft should be regarded as an alternative for complex infrapopliteal revascularization when lacking an autologous vein conduct.

Cryoelectron-microscopy structure of the enteropathogenic Escherichia coli type III secretion system EspA filament.

Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic Escherichia coli (EHEC) utilize a macromolecular type III secretion system (T3SS) to inject effector proteins into eukaryotic cells. This apparatus spans the inner and outer bacterial membranes and includes a helical needle protruding into the extracellular space. Thus far observed only in EPEC and EHEC and not found in other pathogenic Gram-negative bacteria that have a T3SS is an additional helical filament made by the EspA protein that forms a long extension to the needle, mediating both attachment to eukaryotic cells and transport of effector proteins through the intestinal mucus layer. Here, we present the structure of the EspA filament from EPEC at 3.4 Å resolution. The structure reveals that the EspA filament is a right-handed 1-start helical assembly with a conserved lumen architecture with respect to the needle to ensure the seamless transport of unfolded cargos en route to the target cell. This functional conservation is despite the fact that there is little apparent overall conservation at the level of sequence or structure with the needle. We also unveil the molecular details of the immunodominant EspA epitope that can now be exploited for the rational design of epitope display systems.

IT-Related Barriers and Facilitators to the Implementation of a New European eHealth Solution, the Digital Survivorship Passport (SurPass Version 2.0): Semistructured Digital Survey.

BACKGROUND: To overcome knowledge gaps and optimize long-term follow-up (LTFU) care for childhood cancer survivors, the concept of the Survivorship Passport (SurPass) has been invented. Within the European PanCareSurPass project, the semiautomated and interoperable SurPass (version 2.0) will be optimized, implemented, and evaluated at 6 LTFU care centers representing 6 European countries and 3 distinct health system scenarios: (1) national electronic health information systems (EHISs) in Austria and Lithuania, (2) regional or local EHISs in Italy and Spain, and (3) cancer registries or hospital-based EHISs in Belgium and Germany. OBJECTIVE: We aimed to identify and describe barriers and facilitators for SurPass (version 2.0) implementation concerning semiautomation of data input, interoperability, data protection, privacy, and cybersecurity. METHODS: IT specialists from the 6 LTFU care centers participated in a semistructured digital survey focusing on IT-related barriers and facilitators to SurPass (version 2.0) implementation. We used the fit-viability model to assess the compatibility and feasibility of integrating SurPass into existing EHISs. RESULTS: In total, 13/20 (65%) invited IT specialists participated. The main barriers and facilitators in all 3 health system scenarios related to semiautomated data input and interoperability included unaligned EHIS infrastructure and the use of interoperability frameworks and international coding systems. The main barriers and facilitators related to data protection or privacy and cybersecurity included pseudonymization of personal health data and data retention. According to the fit-viability model, the first health system scenario provides the best fit for SurPass implementation, followed by the second and third scenarios. CONCLUSIONS: This study provides essential insights into the information and IT-related influencing factors that need to be considered when implementing the SurPass (version 2.0) in clinical practice. We recommend the adoption of Health Level Seven Fast Healthcare Interoperability Resources and data security measures such as encryption, pseudonymization, and multifactor authentication to protect personal health data where applicable. In sum, this study offers practical insights into integrating digital health solutions into existing EHISs.

Carotid dysfunction in senescent female mice is mediated by increased α1A-adrenoceptor activity and COX-derived vasoconstrictor prostanoids.

α-Adrenergic receptors are crucial regulators of vascular hemodynamics and essential pharmacological targets for cardiovascular diseases. With aging, there is an increase in sympathetic activation, which could contribute to the progression of aging-associated cardiovascular dysfunction, including stroke. Nevertheless, there is little information directly associating adrenergic receptor dysfunction in the blood vessels of aged females. This study determined the role of a-adrenergic receptors in carotid dysfunction of senescent female mice (accelerated-senescence prone, SAMP8), compared with a nonsenescent (accelerated-senescence prone, SAMR1). Vasoconstriction to phenylephrine (Phe) was markedly increased in common carotid artery of SAMP8 [area under the curve (AUC), 527 ± 53] compared with SAMR1 (AUC, 334 ± 30, P = 0.006). There were no changes in vascular responses to the vasoconstrictor agent U46619 or the vasodilators acetylcholine (ACh) and sodium nitroprusside (NPS). Hyperactivity to Phe in female SAMP8 was reduced by cyclooxygenase-1 and cyclooxygenase-2 inhibition and associated with augmented ratio of TXA2/PGI2 release (SAMR1, 1.1 ± 0.1 vs. SAMP8, 2.1 ± 0.3, P = 0.007). However, no changes in cyclooxygenase expression were seen in SAMP8 carotids. Selective α1A-receptor antagonism markedly reduced maximal contraction, whereas α1D antagonism induced a minor shift in Phe contraction in SAMP8 carotids. Ligand binding analysis revealed a threefold increase of α-adrenergic receptor density in smooth muscle cells (VSMCs) of SAMP8 vs. SAMR1. Phe rapidly increased intracellular calcium (Cai2+) in VSMCs via the α1A-receptor, with a higher peak in VSMCs from SAMP8. In conclusion, senescence intensifies vasoconstriction mediated by α1A-adrenergic signaling in the carotid of female mice by mechanisms involving increased Cai2+ and release of cyclooxygenase-derived prostanoids.NEW & NOTEWORTHY The present study provides evidence that senescence induces hyperreactivity of α1-adrenoceptor-mediated contraction of the common carotid. Impairment of α1-adrenoceptor responses is linked to increased Ca2+ influx and release of COX-derived vasoconstrictor prostanoids, contributing to carotid dysfunction in the murine model of female senescence (SAMP8). Increased reactivity of the common carotid artery during senescence may lead to morphological and functional changes in arteries of the cerebral microcirculation and contribute to cognitive decline in females. Because the elderly population is growing, elucidating the mechanisms of aging- and sex-associated vascular dysfunction is critical to better direct pharmacological and lifestyle interventions to prevent cardiovascular risk in both sexes.

The O-GlcNAc dichotomy: when does adaptation become pathological?

O-Linked attachment of ß-N-acetylglucosamine (O-GlcNAc) on serine and threonine residues of nuclear, cytoplasmic, and mitochondrial proteins is a highly dynamic and ubiquitous post-translational modification that impacts the function, activity, subcellular localization, and stability of target proteins. Physiologically, acute O-GlcNAcylation serves primarily to modulate cellular signaling and transcription regulatory pathways in response to nutrients and stress. To date, thousands of proteins have been revealed to be O-GlcNAcylated and this number continues to grow as the technology for the detection of O-GlcNAc improves. The attachment of a single O-GlcNAc is catalyzed by the enzyme O-GlcNAc transferase (OGT), and their removal is catalyzed by O-GlcNAcase (OGA). O-GlcNAcylation is regulated by the metabolism of glucose via the hexosamine biosynthesis pathway, and the metabolic abnormalities associated with pathophysiological conditions are all associated with increased flux through this pathway and elevate O-GlcNAc levels. While chronic O-GlcNAcylation is well associated with cardiovascular dysfunction, only until recently, and with genetically modified animals, has O-GlcNAcylation as a contributing mechanism of cardiovascular disease emerged. This review will address and critically evaluate the current literature on the role of O-GlcNAcylation in vascular physiology, with a view that this pathway can offer novel targets for the treatment and prevention of cardiovascular diseases.

Validating a measure for eco-anxiety in Portuguese young adults and exploring its associations with environmental action.

BACKGROUND: Worsening environmental conditions may amplify people's emotional responses to an environmental crisis (eco-anxiety). In Portugal, young people seem to be especially concerned about climate change. However, this phenomenon needs to be interpreted using accurate instruments. Thus, this study aimed to validate the Portuguese version of the Hogg Eco-Anxiety Scale (HEAS) in young adults and examine the associations among eco-anxiety, sociodemographic characteristics, and pro-environmental behaviours. METHODS: A survey was administered to 623 Portuguese university students aged between 18 and 25 years. The survey included our Portuguese translation of the HEAS (obtained through a back-translation and pretesting process), a sociodemographic assessment, and questions related to pro-environmental behaviours. Confirmatory factor analysis was conducted to assess the construct validity of the Portuguese version of the HEAS, and global fit indices were used to assess whether the original four-dimensional structure of the scale was reproduced. The reliability of the Portuguese version of the HEAS was evaluated by Cronbach's alpha and the intraclass correlation coefficient. Measurement invariance examined sex differences in scale interpretation. Linear regressions were used to detect whether sociodemographic variables predict eco-anxiety and whether eco-anxiety predicts pro-environmental behaviours. RESULTS: The factorial structure of the original scale was replicated in the Portuguese version of the HEAS, showing good internal consistency, reliability over time and strict invariance between men and women. A higher paternal education level predicted greater eco-anxiety in children. Two dimensions of eco-anxiety-namely, rumination and anxiety about personal impacts on the environment-predicted higher engagement in pro-environmental behaviours. CONCLUSIONS: The translated scale is an appropriate tool to measure eco-anxiety in the Portuguese context and should be used to collect evidence to drive environmental and health policies. An individual's education level should be considered a determinant of their emotional response to environmental conditions. Importantly, eco-anxiety can act as a protective emotional response to preserving the planet.

Nurses' promotion of Mental Health First Aid Training Programmes for upper secondary students: a modified Delphi approach.

BACKGROUND: Mental Health First Aid Training Programmes can be carried out by nurses in schools. Adolescents have reported the importance of these interventions, the contents to be addressed, and intervention strategies that should be used. Mental health nurses have also discussed the characteristics of these training programmes. This study sought to create a consensus on the features of Mental Health First Aid Training Programmes promoted by nurses aimed at upper secondary students. METHODS: A descriptive mixed method study was developed using the modified e-Delphi technique. Original statements on the topic were drawn from the results section of previous studies with nurses and adolescents. The statements were included in a structured online questionnaire. An expert panel of 78 mental health nurses participated in the two-rounds online survey from October to December 2021. Absolute and relative frequencies of responses were analysed. The experts' comments were also considered. RESULTS: In the first round, experts suggested 4 new ideas about training programmes. Experts took a position on a total of 59 declarations. At least 75% of participants agreed with 58 statements about training programmes, namely about facilitators, intervention foci, evaluation methodology, participants, the implementation context, period and regularity, intervention strategies and content. The experts' opinions differed only in one statement that expressed the impaired sexual behaviour as a mental health problem to be addressed in the training programmes. CONCLUSIONS: Mental health nurses can lead the delivery of training programmes in upper secondary schools, improving adolescent competencies in mental health. Several educational approaches can be used to teach about mental health, related problems and actions underlying these conditions. Valid and appropriate assessment methods must be used. Our findings guide the planning, implementation and evaluation of these interventions in upper secondary schools and encourage nurses to explore these programmes and include them in their educational curricula.

Type IV secretion systems: Advances in structure, function, and activation.

Bacterial type IV secretion systems (T4SSs) are a functionally diverse translocation superfamily. They consist mainly of two large subfamilies: (i) conjugation systems that mediate interbacterial DNA transfer and (ii) effector translocators that deliver effector macromolecules into prokaryotic or eukaryotic cells. A few other T4SSs export DNA or proteins to the milieu, or import exogenous DNA. The T4SSs are defined by 6 or 12 conserved "core" subunits that respectively elaborate "minimized" systems in Gram-positive or -negative bacteria. However, many "expanded" T4SSs are built from "core" subunits plus numerous others that are system-specific, which presumptively broadens functional capabilities. Recently, there has been exciting progress in defining T4SS assembly pathways and architectures using a combination of fluorescence and cryoelectron microscopy. This review will highlight advances in our knowledge of structure-function relationships for model Gram-negative bacterial T4SSs, including "minimized" systems resembling the Agrobacterium tumefaciens VirB/VirD4 T4SS and "expanded" systems represented by the Helicobacter pylori Cag, Legionella pneumophila Dot/Icm, and F plasmid-encoded Tra T4SSs. Detailed studies of these model systems are generating new insights, some at atomic resolution, to long-standing questions concerning mechanisms of substrate recruitment, T4SS channel architecture, conjugative pilus assembly, and machine adaptations contributing to T4SS functional versatility.

Evaluation of global limb anatomic staging system for patients with chronic limb-threatening ischemia treated with autogenous bypass grafts.

OBJECTIVE: We evaluated the relationship of the global limb anatomic staging system (GLASS) stage with the clinical outcomes for patients with chronic limb-threatening ischemia (CLTI) who had undergone distal bypass with vein grafting. METHODS: We performed a single-center, retrospective analysis of patients with CLTI who had undergone distal bypass with vein grafting from January 2012 to December 2019. The primary end point was freedom from CLTI, including amputation-free survival, complete wound healing, and no ischemic rest pain. The secondary end points included a composite outcome of recurrence (patients who had achieved freedom from CLTI but had developed a new wound or ischemic rest pain), major limb amputation, amputation free-survival, overall survival, major adverse limb events, limb-based patency, and primary and secondary patency rates. RESULTS: A total of 190 patients had undergone 211 distal bypasses with a median follow-up of 30 months. Of these patients, 80% had had diabetes or end-stage renal disease requiring dialysis. Most patients (63%) had experienced major or extensive tissue loss (WIfI [wound, ischemia, foot infection] wound class ≥2), and more than one half of these had had some degree of infection of the foot. A severe anatomic pattern (GLASS stage III) was predominant, with a prevalence of 78%. No significant differences were found between GLASS stage I and II and GLASS stage III groups for all the outcomes analyzed. Approximately 80% of the 211 revascularized limbs were free of CLTI at 12 months after treatment. At 4 years of follow-up, we observed that 25% of the patients had had CLTI recurrence, 83% were free from major amputation, and 61% were free from major adverse limb events. Limb-based patency, primary patency, and secondary patency were 79%, 80%, and 93% at 1 year and 64%, 65%, and 81% at 4 years, respectively. CONCLUSIONS: The GLASS stage was not related to patency or the clinical outcomes after distal bypass with vein grafting. Distal open revascularization resulted in excellent rates of freedom from CLTI with low rates of CLTI recurrence, two key time-integrated outcomes of clinical disease severity for patients with CLTI.

Haematology, physiological parameters, morphometry and parasitological status of rescued bearded capuchin monkeys (Sapajus libidinosus).

BACKGROUND: There are few studies on the physiology and haematology of rescued bearded capuchin monkeys. These are necessary to better understand the health and welfare status of the animals, including when performing reintroductions, and to avoid zoonoses. METHODS: We aimed to obtain physiological and haematological values, morphometry and parasitological status of 26 bearded capuchins in two rescue centres in Northeast Brazil. RESULTS: We found sex- and age-related differences in respiratory rate, body weight and body mass index, but not haematology. The haematological values obtained were significantly different from published data but within the reference intervals for the species. Animals infected with Ancylostoma spp. had significantly lower values in the parameters: haemoglobin concentration, mean corpuscular volume and mean corpuscular haemoglobin concentration than non-infected individuals. CONCLUSION: Physiological and haematological values of rescued capuchins were similar to those found in previous studies. Ancylostomiasis appears to cause alterations to haematological values of this species.

Factors Influencing Performance of Cholangioscopy-Guided Lithotripsy Including Available Different Technologies: A Prospective Multicenter Study with 94 Patients.

BACKGROUND: Peroral cholangioscopy (POC)-guided lithotripsy is an effective treatment for difficult biliary stones. A clear definition of factors associated with the efficacy of POC-guided lithotripsy in one session and the performance of electrohydraulic lithotripsy (EHL) and laser lithotripsy (LL) have not clearly emerged. METHODS: This was a non-randomized prospective multicenter study of all consecutive patients who underwent POC lithotripsy (using EHL and/or LL) for difficult biliary stones. The primary endpoint of the study was the number of sessions needed to achieve complete ductal clearance and the factors associated with this outcome. Secondary endpoints included the evaluated efficacies of LL and EHL. RESULTS: Ninety-four patients underwent 113 procedures of EHL or LL. Complete ductal clearance was obtained in 93/94 patients (98.94%). In total, 80/94 patients (85.11%) achieved stone clearance in a single session. In the multivariate analysis, stone size was independently associated with the need for multiple sessions to achieve complete ductal clearance (odds ratio = 1.146, 95% confidence interval: 1.055-1.244; p = 0.001). Using ROC curves and the Youden index, 22 mm was found to be the optimal cutoff for stone size (95% confidence interval: 15.71-28.28; p < 0.001). The majority of the patients (62.8%) underwent LL in the first session. Six patients failed the first session with EHL after using two probes and therefore were crossed over to LL, obtaining ductal clearance in a single additional session with a single LL fiber. EHL was significantly associated with a larger number of probes (2.0 vs. 1.02) to achieve ductal clearance (p < 0.01). The mean procedural time was significantly longer for EHL than for LL [72.1 (SD 16.3 min) versus 51.1 (SD 10.5 min)] (p < 0.01). CONCLUSIONS: POC is highly effective for difficult biliary stones. Most patients achieved complete ductal clearance in one session, which was significantly more likely for stones < 22 mm. EHL was significantly associated with the need for more probes and a longer procedural time to achieve ductal clearance.

CAD-CAM glass fiber compared with conventional prefabricated glass fiber posts: A systematic review.

STATEMENT OF PROBLEM: A prefabricated glass fiber post provides esthetic and biomechanical benefits for severely damaged endodontically treated teeth; however, failures in post retention remain a challenge. PURPOSE: The purpose of this systematic review was to compare the fracture resistance and bond strength of prefabricated glass fiber posts to those made with computer-aided design and computer-aided manufacturing (CAD-CAM). MATERIAL AND METHODS: This review was entered into PROSPERO (CRD42020213668) and was based on the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Electronic systematic searches of PubMed/MEDLINE, Embase, and the Cochrane Library were conducted for published articles until October 2020. Studies that compared prefabricated glass fiber posts with CAD-CAM-fabricated glass fiber posts were selected, and studies that had not been published in English or in the previous 10 years were excluded. The fracture resistance (N) and bond strength (MPa) were the primary outcomes. RESULTS: After reviewing the title, abstract, and the entire text of 227 articles, 8 studies were selected. Four studies were omitted. Four of the 8 studies were retained for qualitative analyses, with 1 having a high risk of bias and 3 having a medium risk of bias. Only 1 study found significantly higher fracture resistance in the CAD-CAM group than in the prefabricated group. The bond strength study reported a significant difference, with the CAD-CAM group having greater bond strength than the prefabricated group. CONCLUSIONS: Despite the high heterogeneity and the few included articles, a tendency was found for the CAD-CAM-fabricated glass fiber post to improve fracture resistance and bond strength. However, the available literature is insufficient to draw definitive conclusions.

Aldosterone Negatively Regulates Nrf2 Activity: An Additional Mechanism Contributing to Oxidative Stress and Vascular Dysfunction by Aldosterone.

High levels of aldosterone (Aldo) trigger oxidative stress and vascular dysfunction independent of effects on blood pressure. We sought to determine whether Aldo disrupts Nrf2 signaling, the main transcriptional factor involved in antioxidant responses that aggravate cell injury. Thoracic aorta from male C57Bl/6J mice and cultured human endothelial cells (EA.hy926) were stimulated with Aldo (100 nM) in the presence of tiron [reactive oxygen species (ROS) scavenger, eplerenone [mineralocorticoid receptor (MR) antagonist], and L-sulforaphane (SFN; Nrf2 activator). Thoracic aortas were also isolated from mice infused with Aldo (600 µg/kg per day) for 14 days. Aldo decreased endothelium-dependent vasorelaxation and increased ROS generation, effects prevented by tiron and MR blockade. Pharmacological activation of Nrf2 with SFN abrogated Aldo-induced vascular dysfunction and ROS generation. In EA.hy926 cells, Aldo increased ROS generation, which was prevented by eplerenone, tiron, and SFN. At short times, Aldo-induced ROS generation was linked to increased Nrf2 activation. However, after three hours, Aldo decreased the nuclear accumulation of Nrf2. Increased Keap1 protein expression, but not activation of p38 MAPK, was linked to Aldo-induced reduced Nrf2 activity. Arteries from Aldo-infused mice also exhibited decreased nuclear Nrf2 and increased Keap1 expression. Our findings suggest that Aldo reduces vascular Nrf2 transcriptional activity by Keap1-dependent mechanisms, contributing to mineralocorticoid-induced vascular dysfunction.

Ablation of piezoelectric polyvinylidene fluoride with a 193 nm excimer laser.

The unique flexible and piezoelectric properties of polyvinylidene fluoride (PVDF) films would allow for new applications for integrated bioelectronic devices. The use of these films has been precluded by the difficulty in machining them into small, discrete features without damaging the properties of the material. The etching of piezoelectric PVDF by means of a 193 nm excimer laser is explored and characterized. Etch rates are shown for common laser fluence values, along with images of the quality of the cuts to provide the reader with an understanding of the compromise between etch rate and edge roughness. The authors describe a novel method for the etching of piezoelectric, ß -phase PVDF. While PVDF is flexible, acoustically matched to biological tissue, and has a wide resonance bandwidth, it is often overlooked as a piezoelectric material for micro-electrical-mechanical-system devices because of the difficulty in fabrication. In this paper, the authors characterize the etch rate and quality while using a 193 nm argon fluoride excimer laser for patterning.

Mitochondrial DNA: A new driver for sex differences in spontaneous hypertension.

The prevalence of arterial hypertension (AH) is higher in men than in premenopausal women of the same age. AH has been characterized as a chronic inflammatory disease and activation of Toll-like receptors (TLR) by damage-associated molecular patterns (DAMPs) is involved. Mitochondrial DNA (mtDNA) may be released by end-organ damage, which is recognized and activates TLR9. The serum level of mtDNA is increased in AH. The aim of this study was to compare the serum mtDNA levels between male and female spontaneously hypertensive rats (SHR) and to evaluate the sex differences in the effect of mtDNA on the function, inflammation and signaling pathway related to TLR9 in the vasculature. Male and female 15-week-old SHR and Wistar rats were used to evaluate the arterial blood pressure, serum mtDNA, contractile response, inflammatory markers and signaling pathway related to TLR9. Male SHR had higher arterial blood pressure values and serum mtDNA compared to female SHR and to male and female normotensive Wistar rats. In male SHR aorta, mtDNA incubation increased the contractile response to phenylephrine, which was blunted by inhibition of TLR9, and also increased pro-inflammatory molecules IL-6 and TNF-α. However, in female SHR aorta, mtDNA incubation did not change the contractile response, reduced pro-inflammatory molecules and prevented oxidative stress. mtDNA incubation did not change the expression of TLR9, MyD88 and eNOS neither in male nor in female SHR aorta, but it increased the phosphorylation of ERK1/2 in male and reduced in female SHR aorta. The mtDNA differential modulation of vascular response in male and female SHR might contribute to sex differences in AH. This study contributes to the understanding of a need for more personalized therapeutic strategies for men and women with hypertension. Keywords: Sex differences, Arterial hypertension, Mitochondrial DNA, Toll-Like receptor 9.

Electrocardiographic parameters of Sapajus libidinosus (SPIX, 1823) after chemical immobilization with tiletamine-zolazepam.

BACKGROUND: Tiletamine-zolazepam is a widely used as an alternative chemical immobilization method in non-human primates, with a safe application pathway and necessary relaxation. In order to determine the electrocardiographic parameters of Sapajus libidinosus after intramuscular tiletamine-zolazepam administration at the dose of 5 mg/kg, nine animals were submitted to anesthesia. METHODS: The interpretation of the electrocardiogram determined: heart rate in bpm and heart rhythm, P wave, P-R interval, R wave, QRS complex, T wave, Q-T interval, corrected QT interval and electrical axis. The mean HR was 206 ± 32 bpm. RESULTS: The majority of the monkeys showed normal sinus rhythm, but one animal showed sinus tachycardia. The most commonly observed electrical axis was between +30° and +90°. Two animals presented bigger alterations in the tracing such as low amplitude QRS and T wave bigger than 1/4 of the R wave. CONCLUSIONS: The administration of tiletamine-zolazepam was safe and efficacious, with minimal cardiovascular effects.

Translation, cross-cultural adaptation and validation of the Osteoarthritis Quality of Life (OAQoL) questionnaire for use in Portugal.

Osteoarthritis (OA) is the most prevalent rheumatic disease and is a leading cause of decreased quality of life (QoL). The OA Quality of Life questionnaire (OAQoL) is an OA-specific patient-reported outcome measures. The aim of this study was to translate and validate the original UK English version of the Osteoarthritis Quality of Life (OAQoL) questionnaire into European Portuguese. The translation of the questionnaire was carried out according to a dual panel methodology (bilingual panel followed by lay panel). This was followed by cognitive debriefing interviews (CDIs) with OA patients to assess comprehension and relevance of the translated questionnaire. Finally, a validation survey was conducted to assess its psychometric properties. The Portuguese OAQoL, a comparator scale (the Nottingham Health Profile-NHP) as well as questions relating to demographic and disease information were administered to OA patients. A sub-sample of patients also completed the Portuguese OAQoL two weeks later, to assess test-retest reliability. The internal consistency, construct validity and known group validity (according to perceived OA severity) of the scale was also assessed. Both the bilingual and lay panels consisted of five individuals and no major difficulties relating to the translation process were identified. A total of ten patients with OA participated in the CDIs. The mean time to complete the questionnaire was 5 min. These interviews revealed that the Portuguese version of the OAQoL was clear, relevant and easy to complete. Finally, 53 OA patients (44 females; mean age of 67.6 years) completed the validation survey. Cronbach's alpha coefficient was 0.87, demonstrating high internal consistency. Test-retest reliability, assessed by Spearman's rank correlation coefficient, was 0.86. Moderate correlations were found with the majority of the NHP sections, providing evidence of construct validity. Significant differences in OAQoL scores were found between patients who differed according to their perceived OA severity, providing evidence of known group validity. The Portuguese version of the OAQoL is a valid and reliable questionnaire that can be used to assess QoL in OA, both in clinical practice and for research purposes.