A phospho-switch constrains BTL2-mediated phytocytokine signaling in plant immunity.

Enabling and constraining immune activation is of fundamental importance in maintaining cellular homeostasis. Depleting BAK1 and SERK4, the co-receptors of multiple pattern recognition receptors (PRRs), abolishes pattern-triggered immunity but triggers intracellular NOD-like receptor (NLR)-mediated autoimmunity with an elusive mechanism. By deploying RNAi-based genetic screens in Arabidopsis, we identified BAK-TO-LIFE 2 (BTL2), an uncharacterized receptor kinase, sensing BAK1/SERK4 integrity. BTL2 induces autoimmunity through activating Ca2+ channel CNGC20 in a kinase-dependent manner when BAK1/SERK4 are perturbed. To compensate for BAK1 deficiency, BTL2 complexes with multiple phytocytokine receptors, leading to potent phytocytokine responses mediated by helper NLR ADR1 family immune receptors, suggesting phytocytokine signaling as a molecular link connecting PRR- and NLR-mediated immunity. Remarkably, BAK1 constrains BTL2 activation via specific phosphorylation to maintain cellular integrity. Thus, BTL2 serves as a surveillance rheostat sensing the perturbation of BAK1/SERK4 immune co-receptors in promoting NLR-mediated phytocytokine signaling to ensure plant immunity.

FMR1 allelic complexity in premutation carriers provides no evidence for a correlation with age at amenorrhea.

BACKGROUND: Premutations in the Fragile X Messenger Ribonucleoprotein 1 (FMR1) gene, defined as between 55 and 200 CGGs, have been implicated in fragile X-associated primary ovarian insufficiency (FXPOI). Only 20% of female premutation carriers develop early ovulatory dysfunction, the reason for this incomplete penetrance is unknown. This study validated the mathematical model in premutation alleles, after assigning each allele a score representing allelic complexity. Subsequently, allelic scores were used to investigate the impact of allele complexity on age at amenorrhea for 58 premutation cases (116 alleles) previously published. METHODS: The allelic score was determined using a formula previously described by our group. The impact of each allelic score on age at amenorrhea was analyzed using Pearson's test and a contour plot generated to visualize the effect. RESULTS: Correlation of allelic score revealed two distinct complexity behaviors in premutation alleles. No significant correlation was observed between the allelic score of premutation alleles and age at amenorrhea. The same lack of significant correlation was observed regarding normal-sized alleles, despite a nearly significant trend. CONCLUSIONS: Our results suggest that the use of allelic scores combination have the potential to explain female infertility, namely the development of FXPOI, or ovarian dysfunction, despite the lack of correlation with age at amenorrhea. Such a finding is of great clinical significance for early identification of females at risk of ovulatory dysfunction, enhancement of fertility preservation techniques, and increasing the probability for a successful pregnancy in females with premutations. Additional investigation is necessary to validate this hypothesis.

Low risk of haematomas with intramuscular vaccines in anticoagulated patients: a systematic review with meta-analysis.

INTRODUCTION: The summary of product characteristics of vaccines administered intramuscularly, including the vaccine for coronavirus SARS-CoV-2 (COVID-19) and Influenza, warned for risks of bleeding in patients treated with oral anticoagulants. We aimed to estimate the incidence of major bleeding events in this setting and to compare these risks against other vaccination routes. METHODS: This systematic review included all prospective and retrospective studies enrolling anticoagulated patients that received intramuscular vaccination, published until December 2020 in CENTRAL, MEDLINE and EMBASE. The outcomes of interest were major bleeding and haematoma related with vaccination. The incidence of the outcomes was estimated through a random-effects meta-analysis using the Freeman-Turkey transformation. The results are expressed in percentages, with 95%-confidence intervals (95%CI), limited between 0 and 100%. When studies compared intramuscular vaccination vs. other route, the data were compared and pooled using random-effects meta-analysis. Risk ratios (RR) with 95%CI were reported. RESULTS: Overall 16 studies with 642 patients were included. No major bleeding event was reported. The pooled incidence of haematomas following vaccination (mostly against Influenza) in patients treated with oral anticoagulants (mostly warfarin; no data with DOACs / NOACs) was 0.46% (95%CI 0-1.53%). Three studies evaluated the intramuscular vs. subcutaneous route of vaccination. Intramuscular vaccines did not increase the risk of haematoma (RR 0.53, 95%CI 0.10-2.82) compared with subcutaneous route. CONCLUSIONS: Intramuscular vaccination in anticoagulated patients is safe with very low incidence of haematomas and the best available evidence suggests that using the intramuscular route does not increase the risk of haematomas compared with the subcutaneous route.

Evaluation of EGCG Loading Capacity in DMPC Membranes.

Catechins are molecules with potential use in different pathologies such as diabetes and cancer, but their pharmaceutical applications are often hindered by their instability in the bloodstream. This issue can be circumvented using liposomes as their nanocarriers for in vivo delivery. In this work, we studied the molecular details of (-)-epigallocatechin-3-gallate (EGCG) interacting with 1,2-dimyristoyl- sn-glycero-3-phosphocholine (DMPC) monolayer/bilayer systems to understand the catechin loading ability and liposome stability, using experimental and computational techniques. The molecular dynamics simulations show the EGCG molecules deep inside the lipid bilayer, positioned below the lipid ester groups, generating a concentration-dependent lipid condensation. This effect was also inferred from the surface pressure isotherms of DMPC monolayers. In the polarization-modulated infrared reflection absorption spectra assays, the predominant effect at higher concentrations of EGCG (e.g., 20 mol %) was an increase in lipid tail disorder. The steady-state fluorescence data confirmed this disordered state, indicating that the catechin-induced liposome aggregation outweighs the condensation effects. Therefore, by adding more than 10 mol % EGCG to the liposomes, a destabilization of the vesicles occurs with the ensuing release of entrapped catechins. The loading capacity for DMPC seems to be limited by its disordered lipid arrangements, typical of a fluid phase. To further increase the clinical usefulness of liposomes, lipid bilayers with more stable and organized assemblies should be employed to avoid aggregation at large concentrations of catechin.

Classical fragile-X phenotype in a female infant disclosed by comprehensive genomic studies.

BACKGROUND: We describe a female infant with Fragile-X syndrome, with a fully expanded FMR1 allele and preferential inactivation of the homologous X-chromosome carrying a de novo deletion. This unusual and rare case demonstrates the importance of a detailed genomic approach, the absence of which could be misguiding, and calls for reflection on the current clinical and diagnostic workup for developmental disabilities. CASE PRESENTATION: We present a female infant, referred for genetic testing due to psychomotor developmental delay without specific dysmorphic features or relevant family history. FMR1 mutation screening revealed a methylated full mutation and a normal but inactive FMR1 allele, which led to further investigation. Complete skewing of X-chromosome inactivation towards the paternally-inherited normal-sized FMR1 allele was found. No pathogenic variants were identified in the XIST promoter. Microarray analysis revealed a 439 kb deletion at Xq28, in a region known to be associated with extreme skewing of X-chromosome inactivation. CONCLUSIONS: Overall results enable us to conclude that the developmental delay is the cumulative result of a methylated FMR1 full mutation on the active X-chromosome and the inactivation of the other homologue carrying the de novo 439 kb deletion. Our findings should be taken into consideration in future guidelines for the diagnostic workup on the diagnosis of intellectual disabilities, particularly in female infant cases.

Streptococcus mutans in Umbilical Cord Blood, Peripheral Blood, and Saliva from Healthy Mothers.

The aims of this study were to analyze the presence of Streptococcus mutans (SM)-DNA in cord blood (CB), maternal peripheral blood (PB), and maternal saliva (SA) and compare with data collected in health surveys. Sixty-four healthy women with pregnancies to term and without complications attending for elective cesarean section in the Clinical Hospital of Ribeirao Preto, Sao Paulo were included. Samples of PB and unstimulated SA were obtained on the day of hospitalization and samples of CB were collected after the delivery section. Samples were investigated using polymerase chain reaction for the presence of SM-DNA using specific primers. The results show over 50% of the sample of PB and CB showed SM-DNA detectable. There was a positive correlation between the SM detection in PB/CB and SA (P < 0.05). Pregnant women, who reported tooth brushing more than three times a day, often showed detectable SM-DNA in PB and CB (P < 0.05). In conclusion, the majority of children can have contact with SM-DNA during the intrauterine life by the CB. SM probably transferred from salivary habitat to PB and CB. The tooth brushing can be associated to S. mutans detection in blood samples.

In Situ Cytokine Expression and Morphometric Evaluation of Total Collagen and Collagens Type I and Type III in Keloid Scars.

Keloids are characterized by excessive collagen deposition and growth beyond the edges of the initial injury, and cytokines may be related to their formation. The objective of this study was to evaluate the collagen fibers, analyze in situ expression of cytokines in keloid lesions, and compare to the control group. Results showed that there was a predominance of women and nonwhite and direct black ancestry. Keloid showed a significant increase in total and type III collagen. Significantly, the expression of mRNA for TGF-ß in keloid was increased, the expressions of IFN-γ, IFN-γR1, and IL-10 were lower, and IFN-γR1 and TNF-α had no statistical difference. Correlations between collagen type III and TGF-ß mRNA expression were positive and significant, IFN-γ, IFN-γR1, and IL-10 were negative and significant, and TNF-α showed no statistical difference. We conclude that there was a significant increase of total collagen in keloid and predominance of collagen type III compared to the controls, showing keloid as an immature lesion. There is a significant increase in TGF-ß mRNA in keloid lesions, and a significant decrease in IFN-γ and IL-10, suggesting that these cytokines are related to keloid lesions.

Bioprospection of cytotoxic compounds in fungal strains recovered from sediments of the Brazilian coast.

The cytotoxic activities of extracts (50 µg/ml) from 48 fungal strains, recovered from sediments of Pecém's offshore port terminal (Northeast coast of Brazil), against HCT-116 colon cancer cell lines were investigated. The most promising extract was obtained from strain BRF082, identified as Dichotomomyces cejpii by phylogenetic analyses of partial RPB2 gene sequence. Thus, it was selected for bioassay-guided isolation of the cytotoxic compounds. Large-scale fermentation of BRF082 in potato dextrose broth, followed by chromatographic purification of the bioactive fractions from the liquid medium, yielded gliotoxin (4) and its derivatives acetylgliotoxin G (3), bis(dethio)bis(methylsulfanyl)gliotoxin (1), acetylgliotoxin (5), 6-acetylbis(dethio)bis(methylsulfanyl)gliotoxin (2), besides the quinazolinone alkaloid fiscalin B. All isolated compounds were tested for their cytotoxicities against the tumor cell lines HCT-116, revealing 4 and 3 as the most cytotoxic ones (IC50 0.41 and 1.06 µg/ml, resp.).

Exercise training decreases mitogen-activated protein kinase phosphatase-3 expression and suppresses hepatic gluconeogenesis in obese mice.

Insulin plays an important role in the control of hepatic glucose production. Insulin resistant states are commonly associated with excessive hepatic glucose production, which contributes to both fasting hyperglycaemia and exaggerated postprandial hyperglycaemia. In this regard, increased activity of phosphatases may contribute to the dysregulation of gluconeogenesis. Mitogen-activated protein kinase phosphatase-3 (MKP-3) is a key protein involved in the control of gluconeogenesis. MKP-3-mediated dephosphorylation activates FoxO1 (a member of the forkhead family of transcription factors) and subsequently promotes its nuclear translocation and binding to the promoters of gluconeogenic genes such as phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase). In this study, we investigated the effects of exercise training on the expression of MKP-3 and its interaction with FoxO1 in the livers of obese animals. We found that exercised obese mice had a lower expression of MKP-3 and FoxO1/MKP-3 association in the liver. Further, the exercise training decreased FoxO1 phosphorylation and protein levels of Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and gluconeogenic enzymes (PEPCK and G6Pase). These molecular results were accompanied by physiological changes, including increased insulin sensitivity and reduced hyperglycaemia, which were not caused by reductions in total body mass. Similar results were also observed with oligonucleotide antisense (ASO) treatment. However, our results showed that only exercise training could reduce an obesity-induced increase in HNF-4α protein levels while ASO treatment alone had no effect. These findings could explain, at least in part, why additive effects of exercise training treatment and ASO treatment were not observed. Finally, the suppressive effects of exercise training on MKP-3 protein levels appear to be related, at least in part, to the reduced phosphorylation of Extracellular signal-regulated kinases (ERK) in the livers of obese mice.

Effects of bacterial direct-fed microbial mixtures offered to beef cattle consuming finishing diets on intake, nutrient digestibility, feeding behavior, and ruminal kinetics/fermentation profile.

Effects of bacterial direct-fed microbial (DFM) mixtures on intake, nutrient digestibility, feeding behavior, ruminal fermentation profile, and ruminal degradation kinetics of beef steers were evaluated. Crossbred Angus ruminally cannulated steers (n = 6; body weight [BW] = 520 ±â€…30 kg) were used in a duplicated 3 × 3 Latin square design and offered a steam-flaked corn-based finisher diet to ad libitum intake for 3, 28-d periods. Treatments were 1) Control (no DFM, lactose carrier only); 2) Treat-A (Lactobacillus animalis, Propionibacterium freudenreichii, Bacillus subtilis, and Bacillus licheniformis), at 1:1:1:3 ratio, respectively; totaling 6 × 109 CFU (50 mg)/animal-daily minimum; and 3) Treat-B, the same DFM combination, but doses at 1:1:3:1 ratio. Bacterial counts were ~30% greater than the minimum expected. Data were analyzed using the GLIMMIX procedure of SAS with the model including the fixed effect of treatment and the random effects of square, period, and animal (square). For repeated measure variables, the fixed effects of treatment, time, and their interaction, and the random effects of square, period, animal (square), and animal (treatment) were used. Preplanned contrasts comparing Control × Treat-A or Treat-B were performed. Intake and major feeding behavior variables were not affected (P ≥ 0.17) by treatments. Steers offered Treat-A had an increased (P = 0.04) ADF digestibility compared with Control. Steers offered Treat-A experienced daily 300 min less (P = 0.04) time under ruminal pH 5.6, a greater (P = 0.04) ruminal pH average and NH3-N concentration (P = 0.05) and tended (P = 0.06) to have a lower ruminal temperature compared to Control. Ruminal VFA was not affected (P ≥ 0.38) by treatments. Steers offered Treat-A increased (P = 0.02) and tended (P = 0.08) to increase the ruminal effective degradable NDF and ADF fractions of the diet-substrate, respectively. When the forage-substrate (low quality) was incubated, steers offered Treat-A tended (P = 0.09) to increase the effective degradable hemicellulose fraction compared to Control. In this experiment, the bacterial combinations did not affect intake and feeding behavior, while the combination with a greater proportion of B. licheniformis (Treat-A) elicited an improved core-fiber digestibility and a healthier ruminal pH pattern, in which the ruminal environment showed to be more prone to induce the effective degradability of fiber fractions, while also releasing more NH3-N.

During the finishing phase, a high-energy diet offers benefits related to beef cattle growth and development. However, it is essential to acknowledge that finisher diets are energy-dense and can pose digestive challenges, such as subacute ruminal acidosis. Digestive disturbances negatively affect animal well-being, growth performance, and economic returns. To address digestive challenges endured by animals on high-energy diets, the current experiment focused on the addition of bacterial direct-fed microbial (DFM) mixtures. A unique combination of bacterial DFM containing Lactobacillus animalis, Propionibacterium freudenreichii, Bacillus subtilis, and Bacillus licheniformis was evaluated. These bacteria have been individually reported to improve cattle nutrient utilization, digestibility, ruminal function, and maintain ruminal pH. The study aimed to investigate the effects of this specific microbial combination and doses when added to beef cattle finisher diets. The DFM mixtures offered seemed to not affect intake and major feeding behavior variables. The DFM combination containing a greater proportion of B. licheniformis (Treat-A) seemed to elicit an improved total tract core-fiber digestibility, and a safer ruminal pH pattern. The ruminal environment was shown to be more prone to improve the ruminal effective degradability of fiber fractions, while also releasing more NH3­N.

Investigating the role of microwave thermal and non-thermal effects on WO3-graphene oxide composite synthesis.

The effects of microwave-assisted synthesis on the morphology and crystalline structure of WO3-graphene oxide (GO) composites have been investigated. Using two different microwave reactors, evidence supports that thermal and non-thermal effects significantly influence the properties of the synthesized materials. The findings reveal that the microwave cavity geometry affects how the microwaves are "delivered" to the reactional cavity as a function of time; it also orientates the growth process of the WO3 particles. Consequently, the crystalline structure and morphology are affected. As a result, the WO3-GO composites produced using a CEM reactor exhibit a rounded shape and hexagonal phase of WO3, besides enhanced reduction of GO. Whereas the composites made using an Anton-Paar reactor are composed of sheets and flowers of WO3 with hexagonal, triclinic and/or WO3 hydrate structures and cause a lower reduction on the GO.

Comparative Analysis of the Tibial Component Migration After Cemented and Uncemented Total Knee Arthroplasty with a Hydroxyapatite Coating in Two, Five, and Ten Years: A Meta-Analysis.

Objective This study evaluated and compared the tibial component migration in cemented and uncemented total knee arthroplasty (TKA) with no hydroxyapatite coating 2, 5, and 10 years after surgery. Methods This meta-analysis followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) based on PubMed and MeSH database queries from June to July 2022. Results The meta-analysis included eight randomized clinical studies evaluating 668 knees undergoing TKA. The maximum total point motion (MTPM) in cemented TKAs was higher in 5 years, with a mean value of 0.67 mm (95% confidence interval [CI], 0.52 to 0.87). Uncemented TKAs also presented higher mean MTPM in 5 years (1 mm; 95% CI, 0.82 to 1.22). Uncemented coated ATKs had a higher mean MTPM in 10 years (1.30 mm; 95% CI, 0.70 to 2.39). MTPM was statistically similar in the short- and long-term for cemented and uncemented techniques, with a standardized mean difference of -0.65 (95% CI, -1.65 to 0.35). Conclusion Tibial component migration in TKA was statistically similar at 2, 5, and 10 years in cemented and uncemented techniques, either with or without coating. However, due to the scarce literature, further studies are required with a longer follow-up time.

Comparative Analysis of TNF-alpha, TNF-R1, and TNF-R2 in Patients with Low-impact Fractures Due to Osteoporosis.

Objective To analyze the serum levels of TNF-alpha and its TNF-R1 and TNF-R2 receptors in the blood of patients with low-impact fractures due to osteoporosis, comparing between genders and with healthy patients. Methods The present study was conducted with a blood sample of 62 patients, divided into patients with osteoporosis and healthy patients. The results were obtained using the ELISA method. Cytokine concentrations were determined based on the absorbance values obtained. Results Serum TNF-alpha levels were undetectable in female patients, while in males they were found only in one patient, with no significant difference. Similar results were found in the analyses of TNF-R1 and TNF-R2 levels, a significant increase in levels of TNF-alpha receptors in the groups of patients with osteoporosis compared with the control group in both sexes. There was no significant difference between the sexes in the dosage of both receptors within the group with osteoporosis. There was also a positive and significant correlation in the levels of TNF-R1 and TNF-R2 only in women. Conclusion The significant increase in TNF-R1 and TNF-R2 levels in women with osteoporosis suggest that the release and expression of these receptors may be contributing differently to the development of osteoporosis in men and women.

Effect of the Case for Carpule as a Visual Passive Distraction Tool on Dental Fear and Anxiety: A Pilot Study.

The present cross-over study aimed to evaluate the effect of a visual passive distraction tool, a case for carpule, in the management of fear and anxiety during invasive dental treatment. Children between the ages of 4 and 8 years that need at least two treatments with inferior alveolar nerve block were selected. All the included children received two treatments at different moments: in one treatment, local anesthesia was performed with a visual passive distraction tool, the crocodile case, for carpule (experimental); in the other treatment, local anesthesia was performed without the device (control). An evaluation of the anxiety levels was performed by measuring the heart rate and salivary cortisol levels. Wilcoxon's nonparametric test was used for a data analysis (p < 0.05). The mean heart rate with the visual passive distraction device fell from 81.8 to 78.1, while the control rose from 91.4 to 100 (p < 0.05). The mean salivary cortisol levels with the visual passive distraction device fell from 2.0 to 1.6, while in the control, they rose from 2.1 to 2.4 (p < 0.05). In conclusion, the crocodile case for carpule as a visual passive distraction device contributed positively to the management of fear and anxiety during inferior alveolar nerve block in children.

Effects of a novel direct-fed microbial on growth performance, carcass characteristics, nutrient digestibility, and ruminal morphology of beef feedlot steers.

The effects of a novel direct-fed microbial (DFM) on feedlot performance, carcass characteristics, digestibility, ruminal morphology, and volatile fatty acid (VFA) profile of finishing steers were evaluated. Single-source Angus-crossbred yearling steers (n = 144; initial body weight (BW) = 371 ±â€…19 kg) were used in a randomized complete block design. Steers were blocked by initial BW and randomly assigned to treatments (12 pens/treatment; 4 steers/pen). Treatments included (A) CONTROL (no DFM, tylosin, or monensin, (B) MONTY (monensin sodium [330 mg/animal-daily] and tylosin phosphate [90 mg/animal-daily]), and (C) MONPRO (monensin sodium [same as previous] and Lactobacillus salivarius L28 [1 × 106 CFU/animal-daily]). Treatments were included in a steam-flaked corn-based finisher diet offered once daily using a clean-bunk management for ~149 d. The digestibility assessment was performed from days 70 to 74. Ruminal fluid and rumen tissue samples were collected at the slaughter for VFA profile and papillae morphology analyses, respectively. Data were analyzed using the GLIMMIX procedure of SAS with pen serving as the experimental unit, treatment as fixed effect, and BW block as random effect. Steers offered MONPRO had on average 5.3% less (P < 0.01) dry matter intake (9.56 kg/d) compared with either CONTROL (10.16 kg/d) or MONTY (9.96 kg/d). The carcass-adjusted final BW (613 kg; P = 0.23), overall average daily gain (1.64 kg/d; P = 0.23), and gain-efficiency (0.165; P = 0.61) were not affected by treatments. Steers offered CONTROL had greater (P < 0.01) marbling score and tended (P = 0.06) to have less carcasses grading Select and tended (P = 0.10) to have more carcasses grading Upper-Choice, while other carcass characteristics and liver-abscesses were not affected (P ≥ 0.23) by treatments. The digestibility of nutrients (P ≥ 0.13) and the ruminal VFA profile (P ≥ 0.12) were not affected by treatments. Steers offered MONPRO tended (P = 0.09) to have 16% greater average papillae number compared to other treatments. Yearlings offered finishing diets containing L. salivarius L28 plus monensin did not affect growth performance, digestibility, or ruminal VFA, but reduced feed intake. Carcass quality was negatively affected by treatments, while animals consuming L. salivarius L28 and monensin tended to improve ruminal morphology. Current findings in ruminal morphology and feed intake may warrant further assessment of diets containing L. salivarius L28 on beef cattle food safety aspects.

Antimicrobial resistance is a growing concern to public health and medically important antibiotics have been listed in the Veterinary Feed Directive. Nutritional technologies, such as direct-fed microbials, are being increasingly studied for the development of an effective use on beef cattle production systems. The newly isolated strain of Lactobacillus salivarius L28 has demonstrated pathogenic inhibition of Escherichia coli, Salmonella, and Listeria monocytogenes on in vitro assessments. The potential benefits have warranted the exploration of L. salivarius L28 in a feedlot setting. Single-source Angus-crossbred yearling steers were offered steam-flaked corn-based finishing diets containing no feed additive, or either a combination of tylosin plus monensin or L. salivarius L28 plus monensin. Steers offered L. salivarius L28 plus monensin consumed 5.3% less feed compared with other treatments, while other growth performance variables and the digestibility of nutrients were not affected. Carcasses from cattle supplemented with monensin had slightly lower carcass quality grades than those not supplemented with monensin. Lactobacillus salivarius L28 plus monensin tended to improve steers ruminal morphology. Current findings may warrant further food safety assessments when cattle are offered diets containing L. salivarius L28.

Impact of the Health Gym Program on hospital admissions for stroke in the state of Pernambuco, Brazil.

This study aimed to evaluate the impact of the Health Gym Program (HGP) on hospital admissions for stroke in the state of Pernambuco, Brazil. This policy impact evaluation used a quasi-experimental approach consisting of a difference-in-differences estimator, weighted by propensity score matching to deal with potential confounding variables. The study comprised socioeconomic, demographic, and epidemiological data from official Brazilian databases from 2010 to 2019. The treatment group was composed of the 134 municipalities that implemented the HGP since 2011. The 51 municipalities that did not were allocated to the comparison group. The nearest neighbor algorithm (N5) was used to pair treatment and comparison group municipalities and create the weights to evaluate the average treatment effect on the treated (ATT) in the difference-in-differences estimator. In 2010, 2,771 people were hospitalized for stroke (0.51% of all hospitalizations) and in 2019, 11,542 (2%). Municipalities that implemented the HGP had 18.37% fewer hospitalizations than their counterparts in the comparison group. The program's impact in reducing hospitalization rates was incrementally greater among men (ATT: -0.1932) and those aged 71 to 80 years (ATT: -0.1911). All results were statistically significant at the 5% level. The HGP reduced hospitalization for stroke in several population groups, but primarily in those whose underlying prevalence of stroke is highest, reinforcing the importance of public investments in health promotion policies designed to encourage lifestyle changes.

Atlantic flower-invertebrate interactions: A data set of occurrence and frequency of floral visits.

Encounters between flowers and invertebrates are key events for the functioning of tropical forests. Assessing the structure of networks composed of the interactions between those partners leads to a better understanding of ecosystem functioning and the effects of environmental factors on ecological processes. Gathering such data is, however, costly and time-consuming, especially in the highly diverse tropics. We aimed to provide a comprehensive repository of available flower-invertebrate interaction information for the Atlantic Forest, a South American tropical forest domain. Data were obtained from published works and "gray literature," such as theses and dissertations, as well as self-reports by co-authors. The data set has ~18,000 interaction records forming 482 networks, each containing between one and 1061 interaction links. Each network was sampled for about 200 h or less, with few exceptions. A total of 641 plant genera within 136 different families and 39 orders were reported, with the most abundant and rich families being Asteraceae, Fabaceae, and Rubiaceae. Invertebrates interacting with these plants were all arthropods from 10 orders, 129 families, and 581 genera, comprising 2419 morphotypes (including 988 named species). Hymenoptera was the most abundant and diverse order, with at least six times more records than the second-ranked order (Lepidoptera). The complete data set shows Hymenoptera interacting with all plant orders and also shows Diptera, Lepidoptera, Coleoptera, and Hemiptera to be important nodes. Among plants, Asterales and Fabales had the highest number of interactions. The best sampled environment was forest (~8000 records), followed by pastures and crops. Savanna, grasslands, and urban environments (among others) were also reported, indicating a wide range of approaches dedicated to collecting flower-invertebrate interaction data in the Atlantic Forest domain. Nevertheless, most reported data were from forest understory or lower strata, indicating a knowledge gap about flower-invertebrate interactions at the canopy. Also, access to remote regions remains a limitation, generating sampling bias across the geographical range of the Atlantic Forest. Future studies in these continuous and hard-to-access forested areas will yield important new information regarding the interactions between flowers and invertebrates in the Atlantic Forest. There are no copyright restrictions on the data set. Please cite this data paper if the data are used in publications and teaching events.

Use of the FMR1 Gene Methylation Status to Assess the X-Chromosome Inactivation Pattern: A Stepwise Analysis.

X-chromosome inactivation (XCI) is a developmental process to compensate the imbalance in the dosage of X-chromosomal genes in females. A skewing of the XCI pattern may suggest a carrier status for an X-linked disease or explain the presence of a severe phenotype. In these cases, it is important to determine the XCI pattern, conventionally using the gold standard Human Androgen-Receptor Assay (HUMARA), based on the analysis of the methylation status at a polymorphic CAG region in the first exon of the human androgen receptor gene (AR). The aim of this study was to evaluate whether the methylation status of the fragile mental retardation protein translational regulator gene (FMR1) can provide an XCI pattern similar to that obtained by HUMARA. A set of 48 female carriers of FMR1 gene normal-sized alleles was examined using two assays: HUMARA and a FMR1 methylation PCR (mPCR). Ranges were defined to establish the XCI pattern using the methylation pattern of the FMR1 gene by mPCR. Overall, a 77% concordance of the XCI patterns was obtained between the two assays, which led us to propose a set of key points and a stepwise analysis towards obtaining an accurate result for the XCI pattern and to minimize the underlying pitfalls.