The Impacts of Climate Change on the Emergence and Reemergence of Mosquito-Borne Diseases in Temperate Zones: An Umbrella Review Protocol.

INTRODUCTION: Mosquito-borne diseases represent a global public health concern and are responsible for over 700 000 deaths globally every year. Additionally, many mosquito species have undergone a dramatic global expansion due to various factors, including climate change, and forecasts indicate that mosquito populations will persist in dispersing beyond their present geographic range, namely in temperate climates. The research literature on this topic has grown in recent years, including some systematic evidence synthesis. However, to provide a comprehensive overview of this growing literature needed for policy action, a summary of this evidence, including existing systematic reviews, is required. This study aims to undertake an umbrella review that explores the impacts of climate change on the emergence and reemergence of diseases transmitted by mosquitoes in temperate zones and the publication of the protocol is a fundamental step to ensure the credibility, transparency and reproducibility of this research. METHODS AND ANALYSIS: Studies published in scientific journals indexed by PubMed, EMBASE, Cochrane Library, Epistemonikos, and Web of Science Core Collection to be included in this umbrella review will meet the following criteria: the topic of study (climate change and mosquito-borne diseases), regions (temperate zones), study designs (systematic reviews and meta-analysis), language (any) and date (since inception until December 31st, 2023). Titles and abstracts from selected articles will be evaluated by two authors independently and any discrepancy will be resolved through consensus or, if not possible, through a third author. The data will be extracted, and the risk of bias will be evaluated. The quality of the methodology of the included reviews will be assessed using AMSTAR 2. A narrative synthesis will examine the included systematic reviews. The quality of evidence for all outcomes will be judged using the Grading of Recommendations Assessment, Development and Evaluation working group methodology.

Region-Specific Decellularization of Porcine Uterine Tube Extracellular Matrix: A New Approach for Reproductive Tissue-Engineering Applications.

The uterine tube extracellular matrix is a key component that regulates tubal tissue physiology, and it has a region-specific structural distribution, which is directly associated to its functions. Considering this, the application of biological matrices in culture systems is an interesting strategy to develop biomimetic tubal microenvironments and enhance their complexity. However, there are no established protocols to produce tubal biological matrices that consider the organ morphophysiology for such applications. Therefore, this study aimed to establish region-specific protocols to obtain decellularized scaffolds derived from porcine infundibulum, ampulla, and isthmus to provide suitable sources of biomaterials for tissue-engineering approaches. Porcine uterine tubes were decellularized in solutions of 0.1% SDS and 0.5% Triton X-100. The decellularization efficiency was evaluated by DAPI staining and DNA quantification. We analyzed the ECM composition and structure by optical and scanning electronic microscopy, FTIR, and Raman spectroscopy. DNA and DAPI assays validated the decellularization, presenting a significative reduction in cellular content. Structural and spectroscopy analyses revealed that the produced scaffolds remained well structured and with the ECM composition preserved. YS and HEK293 cells were used to attest cytocompatibility, allowing high cell viability rates and successful interaction with the scaffolds. These results suggest that such matrices are applicable for future biotechnological approaches in the reproductive field.

Isolation and Identification of Lichen Photobionts Collected from Different Environments in North of Portugal and Evaluation of Bioactivities of Their Extracts.

Lichens are organisms constituted by a symbiotic relationship between a fungus (mycobiont) and a photoautotrophic partner (photobiont). Lichens produce several bioactive compounds; however, the biotechnological exploitation of this organism is hampered by its slow growth. To start studying the possibility of exploiting lichens as alternative sources of bioactive compounds, eighteen lichens were collected in the north of Portugal in order to isolate and study the bioactivity of their photobionts. It was possible to isolate and cultivate only eight photobionts. Three of them, LFR1, LFA2 and LCF3, belong to the Coelastrella genus, the other two (LFA1 and LCF1) belong to the Chlorella genus and for the remaining three photobionts, LFS1, LCA1 and LCR1, it was impossible to isolate their microalgae. These only grow in consortium with bacteria and/or cyanobacteria. All extracts showed antioxidant activity, mainly at a concentration of 10 mg.mL-1. LFS1, a consortium extract, showed the highest antioxidant power, as well as the highest concentration of phenolic compounds (5.16 ± 0.53 mg of gallic acid equivalents (GAE).g-1). The extracts under study did not show significant antibacterial activity against Escherichia coli, Listeria or Salmonella. The Coelastrella sp. and LFA1 extracts showed the highest hyaluronidase inhibition. The LFR1 extract at a concentration of 5 mg.mL-1 showed the highest anti-inflammatory activity (79.77 ± 7.66%). The extracts of Coelastrella sp. and LFA1 also showed greater antidiabetic activity, demonstrating the high inhibitory power of α-amylase and α-glucosidase. LFR1 at a concentration of 5 mg.mL-1, due to its selective cytotoxicity inhibiting the growth of cancer cells (Caco-2 cells), is a promising anticancer agent.

Inflammatory response to bioceramic and epoxy resin-based endodontic sealers implanted in mice subcutaneous tissue: An in vivo study.

To evaluate the inflammatory tissue response to BioRoot™ RCS (BR) and AH Plus Jet (AHPJ) sealers implanted in mice subcutaneous tissue. It was hypothesized that the inflammatory tissue response to BR would be milder than to AHPJ. An in vivo study was carried out using isogenic mice. The sealers were implanted during standardized surgical procedures. The inflammatory response was evaluated by microscopic analysis and von Kossa reaction in the reactionary tissue around the specimens after 7, 21, and 63 days. For comparisons, a zinc oxide and eugenol sealer (ZOE) was used as a positive control, in addition to a negative control without a sealer (n = 10 per group/period). All statistical analyses considered a significance level of 5%. All endodontic sealers triggered an inflammatory tissue response after 7 days. BR had a higher inflammatory cell count and a thicker fibrous capsule when compared with AHPJ, but both were less inflammatory than ZOE (p < .001). After 21 days, BR continued to trigger an intense inflammatory tissue response, higher in both microscopic parameters compared to AHPJ, and a thicker fibrous capsule than ZOE (p < .001). After 63 days, the inflammatory tissue response decreased in BR, matching the fibrous capsule thickness with AHPJ and ZOE. BR promoted intense calcium precipitation in all study periods. After 63 days, AHPJ and BR sealers were more biocompatible to subcutaneous mice tissue, but AHPJ present better early inflammatory response, as well as BR showed potential bioactivity. RESEARCH HIGHLIGHTS: The inflammatory tissue response triggered by a bioceramic endodontic sealer (BR) was not milder than that triggered by an epoxy-resin based endodontic sealer (AHPJ) during the first 3 weeks, considering the microscopic analysis of the reactionary tissue.

Quantifying the relative importance of genetics and environment on the comorbidity between mental and cardiometabolic disorders using 17 million Scandinavians.

Mental disorders are leading causes of disability and premature death worldwide, partly due to high comorbidity with cardiometabolic disorders. Reasons for this comorbidity are still poorly understood. We leverage nation-wide health records and near-complete genealogies of Denmark and Sweden (n = 17 million) to reveal the genetic and environmental contributions underlying the observed comorbidity between six mental disorders and 15 cardiometabolic disorders. Genetic factors contributed about 50% to the comorbidity of schizophrenia, affective disorders, and autism spectrum disorder with cardiometabolic disorders, whereas the comorbidity of attention-deficit/hyperactivity disorder and anorexia with cardiometabolic disorders was mainly or fully driven by environmental factors. In this work we provide causal insight to guide clinical and scientific initiatives directed at achieving mechanistic understanding as well as preventing and alleviating the consequences of these disorders.

Dereplication of 4-Quinolone Alkaloids from Waltheria Indica (Malvaceae) Tissues Using Molecular Network Tools.

Waltheria indica (Malvaceae) is a plant popularly used in folk medicine by traditional African and indigenous communities, and in various countries worldwide, to treat general inflammation. Several biological activities of this plant have been reported, including acetylcholinesterase inhibition and potential anti-human immunodeficiency virus (HIV), antinociceptive, analgesic, antifungal, anticancer, anti-inflammatory, leishmanicidal, trypanocidal, antioxidant, and antibacterial activities. The chemical profile of Waltheria indica was assessed by dereplication analysis using UPLC-MS/MS, and data acquisition was performed using chemoinformatics tools, such as Mass Spectrometry-Data Independent AnaLysis (MS-DIAL) and MS-FINDER softwares. The preprocessed data were sent to the GNPS to build a feature-based molecular network (FBMN). Thirty-three 4-quinolone alkaloids were annotated in the extracts and fractions of stems and roots, whereas 12 were annotated in the extracts and fractions of flowers and leaves. This represents an inaugural chemical investigation study employing UPLC-Q-TOF-MS/MS analysis, along with a molecular network approach, within this species and genus.

Outpatient dental procedures carried out by Pediatric Dentists within Brazil's public healthcare system.

Brazil's public healthcare system (SUS) offers specialized oral health services to Brazilians, but the productivity of specialists, such as Pediatric Dentists, has not been characterized. Therefore, the objective of this study was to characterize the outpatient dental procedures (ODPs) carried out by Pediatric Dentists within the SUS. An epidemiological study with an ecological, longitudinal, retrospective, and quantitative approach was conducted. The ODPs carried out by Pediatric Dentists within the SUS were characterized based on type of procedure, complexity level, and circumstance (urgent or elective). Data were analyzed using a descriptive and analytical approach, considering a significance level of 5%, as well as the impact of the COVID-19 pandemic (the 2020-2022 years were not included in secondary analyses). In the last 15 years, 29,234,972 ODPs were carried out by Pediatric Dentists within the SUS. Clinical procedures were the majority (55.4%), significantly more frequent than all other types of procedures (all p <0.05). Among these, restorative and periodontal procedures were the most common (30.7% and 21.0%, respectively). From 2008 to 2019, excluding COVID-19 pandemic years, the trend over the last 15 years was constant for all types of procedures (all p >0.05). In addition, low complexity ODPs were the majority (90.1%), significantly more frequent than medium (9.7%) and high complexity procedures (0.1%) (both p <0.05). At last, most ODPs were not characterized by circumstance in the outpatient production reports (96.9%). Therefore, it was possible to conclude that Pediatric Dentists carried out diverse ODPs within the SUS over the past 15 years, although there was a dominant pattern of type and complexity.

Territorial Disparities in Dental Care for Disabled Persons and Oral Health-Related Indicators: A Population-Level Approach in Brazil's Public Healthcare System from 2014 to 2023.

This study evaluated territorial disparities in dental care for disabled persons in Brazil's public healthcare system from 2014 to 2023. The person-year incidence of outpatient dental procedures carried out by special care dentistry specialists and hospitalizations for dental procedures for disabled persons were compared across different regions and against the national estimate. In addition, productivity was correlated with oral health-related indicators. The significance level was set at 5%. The northern region exhibited the highest outpatient productivity, while the southern region showed lower productivity compared to the national estimate (both p-value < 0.05). This pattern was reversed in inpatient productivity (both p-value < 0.05), with the northeastern and central-western regions also below average (both p-value < 0.05). There were no significant correlations between the indicators and inpatient productivity, but outpatient productivity was positively correlated with the proportions of inhabitants who self-rated their general and oral health as "poor" or "very poor", who have never visited a dentist, and who visited a dentist for tooth extraction (all p-values < 0.05). Territorial disparities in dental care for disabled persons were observed within Brazil's public healthcare system, and they were correlated with unfavorable oral health-related indicators at the population level.

Reduced sympathetic activity is associated with the development of pain and muscle atrophy in a female rat model of fibromyalgia.

Fibromyalgia (FM) is characterized by chronic widespread musculoskeletal pain accompanied by fatigue and muscle atrophy. Although its etiology is not known, studies have shown that FM patients exhibit altered function of the sympathetic nervous system (SNS), which regulates nociception and muscle plasticity. Nevertheless, the precise SNS-mediated mechanisms governing hyperalgesia and skeletal muscle atrophy in FM remain unclear. Thus, we employed two distinct FM-like pain models, involving intramuscular injections of acidic saline (pH 4.0) or carrageenan in prepubertal female rats, and evaluated the catecholamine content, adrenergic signaling and overall muscle proteolysis. Subsequently, we assessed the contribution of the SNS to the development of hyperalgesia and muscle atrophy in acidic saline-injected rats treated with clenbuterol (a selective ß2-adrenergic receptor agonist) and in animals maintained under baseline conditions and subjected to epinephrine depletion through adrenodemedullation (ADM). Seven days after inducing an FM-like model with acidic saline or carrageenan, we observed widespread mechanical hyperalgesia along with loss of strength and/or muscle mass. These changes were associated with reduced catecholamine content, suggesting a common underlying mechanism. Notably, treatment with a ß2-agonist alleviated hyperalgesia and prevented muscle atrophy in acidic saline-induced FM-like pain, while epinephrine depletion induced mechanical hyperalgesia and increased muscle proteolysis in animals under baseline conditions. Together, the results suggest that reduced sympathetic activity is involved in the development of pain and muscle atrophy in the murine model of FM analyzed.

Effect of preheating on the viscosity of composite resins.

Aim: To determine the effects of time and temperature on the viscosity of preheated composite resins. Materials and Methods: Eleven composite resins were heated to 60°C, and temperature analyses were performed at intervals of 1 min until they had cooled to 25°C. The permanent oscillatory shear test was performed at 25°C, 35°C, 50°C, and 60°C for three composite resins under a shear rate of 1s-1. One- and two-way analysis of variance were used for the analysis (α = 0.05). Results: There was no significant interaction between the composite resin and time (P = 0.9304), and only the main effect time was significantly different (P < 0.0001). A difference was observed between T0 and T6 (P < 0.001), but not after T7. The increase in temperature resulted in a viscosity reduction (P < 0.05). At 25°C, Beautifil II presented higher viscosity. Palfique LX5 showed a significant viscosity reduction with increasing temperature compared with the others (P < 0.05). For Beautifil II and Z100, there was no difference at temperatures of 50°C and 60°C, while for Palfique LX5, no statistical difference was observed at 35°C, 50°C, and 60°C. Conclusions: Ten minutes of preheating were sufficient to reach a temperature of 60°C, reducing viscosity by at least 84%. However, 5 min after removal, the composite resin cooled to room temperature. Clinical Significance: Preheating composite resin has potential benefits. To determine how this approach will work in clinical practice, it is important to define the effects of time and temperature in the protocol of this technique and understand its limitations.

The effects of NLRP3 inflammasome inhibition or knockout in experimental apical periodontitis induced in mice.

OBJECTIVE: To evaluate the effects of NLRP3 inflammasome inhibition or knockout in experimental apical periodontitis (AP) induced in mice. METHODS: The experimental AP was induced by pulpal exposure. To evaluate NLRP3-specific inhibitor medication (MCC950), WT mice received intraperitoneal injections, while the control received PBS (n = 10). In addition, to evaluate NLRP3 knockout, 35 wild-type (WT) and 35 NLRP3-/- mice were divided into a control group (without pulpal exposure, n = 5) and three experimental groups: after 2, 14 and 42 days after pulpal exposure (n = 10). Microscopic and molecular analyzes were carried out using a significance level of 5%. RESULTS: Exposure to MCC950 did not affect the periapical lesion size after 14 days (P = 0.584). However, exposed mice had a lower expression of IL-1ß, IL-18 and caspase-1 (P = 0.010, 0.016 and 0.002, respectively). Moreover, NLRP3-/- mice showed a smaller periapical lesion after 14 and 42 days (P = 0.023 and 0.031, respectively), as well as a lower expression of IL-1ß after 42 days (P < 0.001), of IL-18 and caspase-1 after 14 (P < 0.001 and 0.035, respectively) and 42 days (P = 0.002 and 0.002, respectively). NLRP3-/- mice also showed a lower mRNA for Il-1ß, Il-18 and Casp1 after 2 (P = 0.002, 0.036 and 0.001, respectively) and 14 days (P = 0.002, 0.002 and 0.001, respectively). CONCLUSIONS: NLRP3 inflammasome inhibition or knockout can attenuate the inflammatory events that result in the periapical lesion (AP) formation after pulpal exposure in mice. CLINICAL RELEVANCE: The NLRP3 inflammasome may be a therapeutic target for AP, and new approaches may verify the impact of its inhibition (through intracanal medications or filling materials) on the bone repair process and treatment success.

Yeast Diversity in Honey and Pollen Samples from Stingless Bees in the State of Bahia, Brazil: Use of the MALDI-TOF MS/Genbank Proteomic Technique.

(1) Background: The identification of microorganisms includes traditional biochemical methods, molecular biology methods evaluating the conserved regions of rRNA, and the molecular biology of proteins (proteomics), such as MALDI-TOF MS mass spectrometry. This work aimed to identify the biodiversity of yeasts associated with stingless bee species' honey and pollen, Melipona scutellaris, Nannotrigona testaceicornes, and Tetragonisca angustula, from the region of São Gonçalo dos Campos-Bahia (BA) state, Brazil. (2) Methods: Cellular proteins were extracted from 2837 microbial isolates (pollen and honey) and identified via MALDI-TOF MS. The identified yeast species were also compared to the mass spectra of taxonomically well-characterized reference strains, available from the National Center of Biotechnology Information (NCBI) database. (3) Results: Nine yeast species were identified: Candida maltosa, Candida norvegica, Kazachstania telluris, Schizosaccharomyces pombe, Scheffersomyces insectosus, Meyerozyma guilliermondii, Brettanomyces bruxellensis, Kazachstania exigua, and Starmerella lactis-condensi. Nannotrigona testaceicornes pollen had the highest number of yeast colonies. The yeasts Brettanomyces bruxellensis and Kazachstania telluris showed high populations in the samples of Nannotrigona testaceicornes and Melipona scutellaris, respectively. This work shows that there is some sharing of the same species of yeast between honey and pollen from the same beehive. (4) Conclusions: A total of 71.84% of the identified species present a high level of confidence at the species level. Eight yeast species (Candida maltosa, Candida norvegica, Kazachstania telluris, Schizosaccharomyces pombe, Scheffersomyces insectosus, Meyerozyma guilliermondii, Kazachstania exigua, and Starmerella lactis-condensi) were found for the first time in the samples that the authors inspected. This contributes to the construction of new knowledge about the diversity of yeasts associated with stingless bee products, as well as to the possibility of the biotechnological application of some yeast species.

Root canal conicity determination of primary second molars using nanotomography: An in vitro study.

BACKGROUND: The conicity of the root canals of primary teeth is an important measure for endodontic therapies. However, determining this conicity depends on the methods employed, which requires further investigation. AIM: The aim of this study was to determine the conicity of the root canals of the upper and lower primary second molars using nanotomography (nCT). DESIGN: An in vitro study was performed using nine primary second molars, both upper and lower, subjected to nCT. Comparisons between the diameters of root canals were performed between the thirds (cervical-D0, middle-D5, and apical-D7). The conicity (%) was determined for each root canal from cervical to apical. Data were statistically analyzed with a significance level of 5%. RESULTS: The conicity ranged from 2% to 8% for the upper primary second molars. Significant differences in root canal diameter between the thirds (D0, D5, and D7 points) were observed in the mesio- and distobuccal roots (p < .05), but not in the palatal roots (p > .05). For the lower primary second molars, the conicity ranged from 2% to 17%, as well as significant differences in root canal diameter between the thirds (D0, D5, and D7 points) were observed in all roots (distal, mesiobuccal, and mesiolingual; p < .05). CONCLUSION: The conicity of the upper primary second molars was different from that of the lower ones, which showed a greater variability.

Oral Mucositis Management in Children under Cancer Treatment: A Systematic Review.

Children undergoing antineoplastic treatment often present severe side effects due to the dosage and duration of treatments, with oral mucositis emerging as one of the most prevalent and painful inflammatory conditions. There is a growing body of evidence on therapeutic interventions such as cryotherapy, low-level laser therapy, and natural compounds for this condition. The aim of this systematic review was to identify and compare therapies for the management of cancer treatment-induced oral mucositis in pediatric patients. From 2655 articles obtained in initial searches, 39 articles were considered in this systematic review, after applying inclusion/exclusion criteria. Low-level laser therapy, palifermin, honey, and zinc demonstrated reductions in oral mucositis incidence, duration, severity, and pain reported by the patient. Although there are several therapies in place for the prevention and treatment of oral mucositis in children, evidence of their efficacy is still inconclusive to establish accurate clinical protocols.

Leukotriene receptor antagonists as adjuvant therapy of antihistamines in chronic urticaria: a systematic review and meta-analysis.

Certain guidelines recommend a second-generation H1-antihistamine (AH) as first-line treatment for patients with chronic urticaria (CU). However, some patients show insufficient response to a standard dose of this therapy and might benefit from adding leukotriene receptor antagonists (LTA). Therefore, we aimed to perform a systematic review and meta-analysis comparing LTA plus antihistamines with antihistamines alone. We performed a systematic review and meta-analysis, searching PubMed, EMBASE, and Cochrane Central for randomized clinical trial (RCT) data comparing LTA plus AH treatment to AH alone in patients with CU. Statistical analysis was performed using R Studio 4.3.2. Heterogeneity was assessed with I2 statistics. Three studies comprising 234 patients with urticaria were included. The mean age was 37.23 years in the leukotriene antagonist + antihistamines (LTA + AH) group and 39.14 years in the antihistamines (AH) group. Follow-up ranged from 2 to 18 months between studies. There was no statistically significant difference between groups in terms of TSS level (SMD: -74.82; 95% CI: -222.66 to 73.02; P = 0.32; I2 = 98%), neither in terms of pruritus (MD: -0.07; 95% CI: -0.42 to 0.28; P = 0.70; I2 = 74%). After sensitivity analysis, with the systematic exclusion of each study from the grouped estimates, the result for TSS level did not change. These findings suggest that leukotriene receptor antagonists with antihistamines do not have better outcomes than antihistamines alone regarding TSS and pruritus in patients with CU.

Suprachoroidal gene transfer with nonviral nanoparticles in large animal eyes.

Suprachoroidal nonviral gene therapy with biodegradable poly(ß-amino ester) nanoparticles (NPs) provides widespread expression in photoreceptors and retinal pigmented epithelial (RPE) cells and therapeutic benefits in rodents. Here, we show in a human-sized minipig eye that suprachoroidal injection of 50 µl of NPs containing 19.2 µg of GFP expression plasmid caused GFP expression in photoreceptors and RPE throughout the entire eye with no toxicity. Two weeks after injection of 50, 100, or 200 µl, there was considerable within-eye and between-eye variability in expression that was reduced 3 months after injection of 200 µl and markedly reduced after three suprachoroidal injections at different locations around the eye. Reduction of bacterial CpG sequences in the expression plasmid resulted in a trend toward higher expression. These data indicate that nonviral suprachoroidal gene therapy with optimized polymer, expression plasmid, and injection approach has potential for treating photoreceptors throughout the entire retina of a human-sized eye.

Quantifying the Relative Importance of Genetics and Environment on the Comorbidity between Mental- and Cardiometabolic Disorders: A Comprehensive Analysis of National Register Data from 17 million Scandinavians.

Mental disorders (MDs) are leading causes of disability and premature death worldwide, partly due to high comorbidity with cardiometabolic disorders (CMDs). Reasons for this comorbidity are still poorly understood. We leverage nation-wide health records and complete genealogies of Denmark and Sweden (n=17 million) to reveal the genetic and environmental contributions underlying the observed comorbidity between six MDs and 14 CMDs. Genetic factors contributed about 50% to the comorbidity of schizophrenia, affective disorders, and autism spectrum disorder with CMDs, whereas the comorbidity of attention-deficit/hyperactivity disorder and anorexia with CMDs was mainly or fully driven by environmental factors. These findings provide causal insight to guide clinical and scientific initiatives directed at achieving mechanistic understanding as well as preventing and alleviating the consequences of these disorders.

A tool to assess risk of bias in non-randomized follow-up studies of exposure effects (ROBINS-E).

BACKGROUND: Observational epidemiologic studies provide critical data for the evaluation of the potential effects of environmental, occupational and behavioural exposures on human health. Systematic reviews of these studies play a key role in informing policy and practice. Systematic reviews should incorporate assessments of the risk of bias in results of the included studies. OBJECTIVE: To develop a new tool, Risk Of Bias In Non-randomized Studies - of Exposures (ROBINS-E) to assess risk of bias in estimates from cohort studies of the causal effect of an exposure on an outcome. METHODS AND RESULTS: ROBINS-E was developed by a large group of researchers from diverse research and public health disciplines through a series of working groups, in-person meetings and pilot testing phases. The tool aims to assess the risk of bias in a specific result (exposure effect estimate) from an individual observational study that examines the effect of an exposure on an outcome. A series of preliminary considerations informs the core ROBINS-E assessment, including details of the result being assessed and the causal effect being estimated. The assessment addresses bias within seven domains, through a series of 'signalling questions'. Domain-level judgements about risk of bias are derived from the answers to these questions, then combined to produce an overall risk of bias judgement for the result, together with judgements about the direction of bias. CONCLUSION: ROBINS-E provides a standardized framework for examining potential biases in results from cohort studies. Future work will produce variants of the tool for other epidemiologic study designs (e.g. case-control studies). We believe that ROBINS-E represents an important development in the integration of exposure assessment, evidence synthesis and causal inference.

µ-Opioid Receptor Activation at the Dorsal Reticular Nucleus Shifts Diffuse Noxious Inhibitory Controls to Hyperalgesia in Chronic Joint Pain in Male Rats.

BACKGROUND: The dorsal reticular nucleus is a pain facilitatory area involved in diffuse noxious inhibitory control (DNIC) through opioidergic mechanisms that are poorly understood. The hypothesis was that signaling of µ-opioid receptors is altered in this area with prolonged chronic inflammatory pain and that this accounts for the loss of DNICs occurring in this condition. METHODS: Monoarthritis was induced in male Wistar rats (n = 5 to 9/group) by tibiotarsal injection of complete Freund's adjuvant. The immunolabeling of µ-opioid receptors and the phosphorylated forms of µ-opioid receptors and cAMP response element binding protein was quantified. Pharmacologic manipulation of µ-opioid receptors at the dorsal reticular nucleus was assessed in DNIC using the Randall-Selitto test. RESULTS: At 42 days of monoarthritis, µ-opioid receptor labeling decreased at the dorsal reticular nucleus, while its phosphorylated form and the phosphorylated cAMP response element binding protein increased. [d-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin acetate (DAMGO) enhanced DNIC analgesia in normal animals (means ± SD: pre-DNIC: 126.9 ± 7.0 g; DNIC - DAMGO: 147.5 ± 8.0 g vs. DNIC + DAMGO: 198.1 ± 19.3 g; P < 0.001), whereas it produced hyperalgesia in monoarthritis (pre-DNIC: 67.8 ± 7.5 g; DNIC - DAMGO: 70.6 ± 7.7 g vs. DNIC + DAMGO: 32.2 ± 2.6 g; P < 0.001). An ultra-low dose of naloxone, which prevents the excitatory signaling of the µ-opioid receptor, restored DNIC analgesia in monoarthritis (DNIC - naloxone: 60.0 ± 6.1 g vs. DNIC + naloxone: 98.0 ± 13.5 g; P < 0.001), compared to saline (DNIC - saline: 62.5 ± 5.2 g vs. DNIC + saline: 64.2 ± 3.8 g). When injected before DAMGO, it restored DNIC analgesia and decreased the phosphorylated cAMP response element binding protein in monoarthritis. CONCLUSIONS: The dorsal reticular nucleus is likely involved in a facilitatory pathway responsible for DNIC hyperalgesia. The shift of µ-opioid receptor signaling to excitatory in this pathway likely accounts for the loss of DNIC analgesia in monoarthritis.