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Tobacco use is associated with morbidity and mortality. Many individuals who present to treatment facilities with substance use disorders (SUDs) other than tobacco use disorder also smoke cigarettes or have a concomitant tobacco use disorder. Despite high rates of smoking among those with an SUD, and numerous demonstrated benefits of comprehensive SUD treatment for tobacco use in addition to co-occurring SUDs, not all facilities address the treatment of comorbid tobacco use disorder. In addition, facilities vary widely in terms of tobacco use policies on campus. This study examined SUD facility smoking policies in a national sample of N = 16,623 SUD treatment providers in the United States in 2021. Most facilities with outpatient treatment (52.1%) and facilities with residential treatment (67.8%) had a smoking policy that permitted smoking in designated outdoor area(s). A multinomial logistic regression model found that among facilities with outpatient treatment (n = 13,778), those located in a state with laws requiring tobacco free grounds at SUD facilities, those with tobacco screening/education/counseling services, and those with nicotine pharmacotherapy were less likely to have an unrestrictive tobacco smoking policy. Among facilities with residential treatment (n = 3449), those with tobacco screening/education/counseling services were less likely to have an unrestrictive tobacco smoking policy. There is variability in smoking policies and tobacco use treatment options in SUD treatment facilities across the United States. Since tobacco use is associated with negative biomedical outcomes, more should be done to ensure that SUD treatment also focuses on reducing the harms of tobacco use.

This study evaluated the effect of different chemo-mechanical cleaning protocols on the residual presence of epoxy resin-based and bioceramic endodontic sealers, as well as the bond strength of a universal adhesive system for cementing fiber posts to intraradicular dentin. A total of 160 bovine incisors were prepared using the ProTaper system and filled with either an epoxy resin-based sealer (Vioseal) or a bioceramic sealer (BioRoot RCS). Four cleaning protocols were tested: ethanol (ET), amyl acetate (AA), acetone (AC), and an experimental solution (EX) combining amyl acetate, ethanol, and acetone. Residual sealer was analyzed via scanning electron microscopy, and bond strength was assessed through push-out tests. Fracture patterns were examined under a stereomicroscope, and data were analyzed using ANOVA and Kruskal-Wallis tests (α=5%). For epoxy resin-based sealers, AA and AC resulted in greater dentinal tubule opening than ET and EX (p<0.05), with AC leaving the least residue. Conversely, ET and EX showed higher residual presence. For bioceramic sealers, ET and EX achieved the greatest tubule opening (p<0.05). Bond strength was highest with ET in the apical third (epoxy sealer) and in the cervical third (bioceramic sealer) (p<0.05). Ethanol and the experimental solution effectively removed bioceramic sealer residues while preserving bond strength. Although epoxy resin-based sealers left more residue, all tested protocols provided satisfactory adhesion. Este estudio evaluó el efecto de diferentes protocolos de limpieza quimico-mecánica sobre la presencia residual de cementos endodónticos a base de resina epóxica y biocerámico, así como la resistencia de unión de un sistema adhesivo universal para cementar postes de fibra a la dentina intraradicular. Se utilizaron 160 incisivos bovinos que fueron preparados con el sistema ProTaper y obturados con un cemento a base de resina epóxica (Vioseal) o un cemento biocerámico (BioRoot RCS). Se probaron cuatro protocolos de limpieza: etanol (ET), acetato de amilo (AA), acetona (AC) y una solución experimental (EX) que combina acetato de amilo, etanol y acetona. El cemento residual fue analizado mediante microscopía electrónica de barrido, y la resistencia de unión se evaluó a través de pruebas de push-out. Los tipo de falla adhesiva se examinaron bajo un estereomicroscopio, y los datos se analizaron utilizando los tests ANOVA y Kruskal-Wallis (α=5%). Para los cementos a base de resina epóxica, AA y AC resultaron en una mayor apertura de los túbulos dentinarios que ET y EX (p<0.05), siendo AC el que dejó menos residuo. Por el contrario, ET y EX mostraron mayor presencia residual. Para los cementos biocerámicos, ET y EX lograron la mayor apertura de los túbulos (p<0.05). La resistencia de unión fue mayor con ET en el tercio apical (sellador epóxico) y en el tercio cervical (sellador biocerámico) (p<0.05). El etanol y la solución experimental eliminaron eficazmente los residuos del cemento biocerámico mientras preservaban la resistencia de unión. Aunque los selladores a base de resina epóxica dejaron más residuo, todos los protocolos probados proporcionaron una adhesión satisfactoria.

Annona neoinsignis H. Rainer (Annonaceae) is a tree native to the Amazon rainforest. Its fruits are also suitable for human consumption in their natural state or are processed to make desserts. In this work, we characterized the chemical composition of the essential oil (EO) from the leaves of A. neoinsignis and evaluated its anti-liver-cancer potential via in vitro and in vivo approaches. Chemical composition analysis revealed β-elemene, (E)-caryophyllene, germacrene D, and germacrene B as the main constituents. The EO had IC50 values ranging from 12.28 to 37.50 μg/mL for B16-F10 cells and MCF-7 cells, whereas an IC50 value of >50 μg/mL was found for noncancerous MRC-5 cells. DNA fragmentation, YO-PRO-1 staining, and loss of mitochondrial transmembrane potential were detected in EO-treated HepG2 cells, indicating the induction of apoptosis. Significant in vivo growth inhibition of 53.7% was observed in mice bearing HepG2 cell xenografts treated with EO at a dosage of 40 mg/kg. These data suggest that EO from A. neoinsignis leaves is a drug source for liver cancer.

Annona amazonica R.E. Fries (synonyms Annona amazonica var. lancifolia R.E. Fries), popularly known in Brazil as “envireira”, is a tropical tree belonging to the Annonaceae family and is traditionally used as a food source. In this work, the in vitro and in vivo anti-liver cancer effects of essential oil (EO) from A. amazonica leaves were investigated for the first time. The chemical composition of the EO was evaluated via GC–MS and GC–FID. The alamar blue assay was used to evaluate the cytotoxicity of EOs against different cancerous and noncancerous cell lines. Cell cycle analyses, YO-PRO-1/PI staining, and rhodamine 123 staining were performed via flow cytometry in HepG2 cells treated with EO. The in vivo antitumor activity of EO was evaluated in NSG mice that were xenografted with HepG2 cells and treated with EO at a dose of 60 mg/kg. The major constituents (>5%) of the EO were (E)-caryophyllene (32.01%), 1,8-cineole (13.93%), α-copaene (7.77%), α-humulene (7.15%), and α-pinene (5.13%). EO increased apoptosis and proportionally decreased the number of viable HepG2 cells. The induction of DNA fragmentation and cell shrinkage together with a significant reduction in the ΔΨm in EO-treated HepG2 cells confirmed that EO can induce apoptosis. A significant 39.2% inhibition of tumor growth in vivo was detected in EO-treated animals. These data indicate the anti-liver cancer potential of EO from A. amazonica leaves.

In the past decades, the production of biopharmaceuticals has gained high interest due to its great sensitivity, specificity, and lower risk of negative effects to patients. Biopharmaceuticals are mostly therapeutic recombinant proteins produced through biotechnological processes. In this context, L-asparaginase (L-asparagine amidohydrolase, L-ASNase (E.C. 3.5.1.1)) is a therapeutic enzyme that has been abundantly studied by researchers due to its antineoplastic properties. As a biopharmaceutical, L-ASNase has been used in the treatment of acute lymphoblastic leukemia (ALL), acute myeloblastic leukemia (AML), and other lymphoid malignancies, in combination with other drugs. Besides its application as a biopharmaceutical, this enzyme is widely used in food processing industries as an acrylamide mitigation agent and as a biosensor for the detection of L-asparagine in physiological fluids at nano-levels. The great demand for L-ASNase is supplied by recombinant enzymes from Escherichia coli and Erwinia chrysanthemi. However, production processes are associated to low yields and proteins associated to immunogenicity problems, which leads to the search for a better enzyme source. Considering the L-ASNase pharmacological and food importance, this review provides an overview of the current biotechnological developments in L-ASNase production and biochemical characterization aiming to improve the knowledge about its production.Key points• Microbial enzyme applications as biopharmaceutical and in food industry• Biosynthesis process: from the microorganism to bioreactor technology• Enzyme activity and kinetic properties: crucial for the final application

The Brief Measure of Emotional Preoperative Stress (B-MEPS) was developed to evaluate the preoperative individual vulnerability to emotional stress. To obtain a refined version of B-MEPS suitable for an app approach, this study aimed: (i) to identify items with more discriminant properties; (ii) to classify the level of preoperative emotional stress based on cut-off points; (iii) to assess concurrent validity through correlation with the Central Sensitization Inventory (CSI) score; (iv) to confirm whether the refined version of B-MEPS is an adequate predictive measure for identification of patients prone to intense postoperative pain. We include 1016 patients who had undergone surgical procedures in a teaching hospital. The generalized partial credit model of item response theory and latent class model were employed, respectively, to reduce the number of items and to create cut-off points. We applied the CSI and assessed pain by Visual Analog Scale (0-10) and by the amount of postoperative morphine consumption. The refined B-MEPS shows satisfactory reliability (Cronbach's alpha 0.79). Preoperative emotional stress, according to the cut-off points, is classified into categories: low, intermediate or high stress. The refined B-MEPS exhibited a linear association with the CSI scores (r2 = 0.53, p < 0.01). Patients with higher levels of emotional stress displayed a positive association with moderate to severe pain and greater morphine consumption. The refined version of B-MEPS, along with an interface of easy applicability, assess emotional vulnerability at the bedside before surgery. This app may support studies focused on intervening with perioperative stress levels.

Sphingolipids (SLs) are essential components of all eukaryotic cellular membranes. In fungi, plants and many protozoa, the primary SL is inositol-phosphorylceramide (IPC). Trypanosoma cruzi is a protozoan parasite that causes Chagas disease (CD), a chronic illness for which no vaccines or effective treatments are available. IPC synthase (IPCS) has been considered an ideal target enzyme for drug development because phosphoinositol-containing SL is absent in mammalian cells and the enzyme activity has been described in all parasite forms of T . cruzi . Furthermore, IPCS is an integral membrane protein conserved amongst other kinetoplastids, including Leishmania major , for which specific inhibitors have been identified. Using a CRISPR-Cas9 protocol, we generated T . cruzi knockout (KO) mutants in which both alleles of the IPCS gene were disrupted. We demonstrated that the lack of IPCS activity does not affect epimastigote proliferation or its susceptibility to compounds that have been identified as inhibitors of the L . major IPCS. However, disruption of the T . cruzi IPCS gene negatively affected epimastigote differentiation into metacyclic trypomastigotes as well as proliferation of intracellular amastigotes and differentiation of amastigotes into tissue culture-derived trypomastigotes. In accordance with previous studies suggesting that IPC is a membrane component essential for parasite survival in the mammalian host, we showed that T . cruzi IPCS null mutants are unable to establish an infection in vivo , even in immune deficient mice.

Human T-lymphotropic virus type 1 (HTLV-1) has worldwide distribution and is considered endemic in many world regions, including southwestern Japan and Brazil. Japanese immigrants and their descendants have a high risk of acquiring this infection due to intense population exchange between Brazil and Japan. This cross-sectional study aimed to estimate the prevalence of HTLV, analyze the main risk factors associated with this infection, identify the main circulating types and subtypes of HTLV in Japanese immigrants and descendants living in Campo Grande-MS (Middle-West Brazil), as well as analyze the phylogenetic relationship among isolates of HTLV. A total of 219 individuals were interviewed and submitted to blood collection. All collected blood samples were submitted for detection of anti-HTLV-1/2 using the immunoassay ELISA and confirmed by immunoblot method. The proviral DNA of the 14 samples HTLV- 1 positive were genotyped by nucleotide sequencing. The overall prevalence of HTLV-1 was 6.8% (IC 95%: 3,5-10,2). Descriptive analysis of behavioral risk factors showed statistical association between HTLV-1 and age greater than or equal to 45 years. The proviral DNA of HTLV-1 was detected in all HTLV-1 positive samples. Of these, 14 were sequenced and classified as Cosmopolitan subtype, and 50% (7/14) belonged to subgroup A (transcontinental) and 50% (7/14) to the subgroup B (Japanese). The high prevalence of HTLV-1 found evidence of the importance of early diagnosis and counseling of individuals infected with HTLV-1 for the control and prevention of the spread of this infection among Japanese immigrants and their descendants in Central Brazil.

Dog training methods range broadly from those using mostly positive punishment and negative reinforcement (aversive-based) to those using primarily positive reinforcement (reward-based). Although aversive-based training has been strongly criticized for negatively affecting dog welfare, there is no comprehensive research focusing on companion dogs and mainstream techniques, and most studies rely on owner-reported assessment of training methods and dog behavior. The aim of the present study was to evaluate the effects of aversive- and reward-based training methods on companion dog welfare within and outside the training context. Ninety-two companion dogs were recruited from three reward-based schools (Group Reward, n = 42), and from four aversive-based schools, two using low proportions of aversive-based methods (Group Mixed, n = 22) and two using high proportions of aversive-based methods (Group Aversive, n = 28). For evaluating welfare during training, dogs were video recorded for three sessions and six saliva samples were collected, three at home (baseline levels) and three after training (post-training levels). Video recordings were used to examine the frequency of stress-related behaviors (e.g., lip lick, yawn) and the overall behavioral state of the dog (e.g., tense, relaxed), and saliva samples were analyzed for cortisol concentration. For evaluating welfare outside the training context, dogs participated in a cognitive bias task. Results showed that dogs from Group Aversive displayed more stress-related behaviors, were more frequently in tense and low behavioral states and panted more during training, and exhibited higher post-training increases in cortisol levels than dogs from Group Reward. Additionally, dogs from Group Aversive were more ‘pessimistic’ in the cognitive bias task than dogs from Group Reward. Dogs from Group Mixed displayed more stress-related behaviors, were more frequently in tense states and panted more during training than dogs from Group Reward. Finally, although Groups Mixed and Aversive did not differ in their performance in the cognitive bias task nor in cortisol levels, the former displayed more stress-related behaviors and was more frequently in tense and low behavioral states. These findings indicate that aversive-based training methods, especially if used in high proportions, compromise the welfare of companion dogs both within and outside the training context.

Shrinking natural habitats exposes some non-human primates to the risk of accidents associated with electrical transmission lines. We examined dead marmosets (Callithrix penicillata) collected in the region from January 2015 to April 2018 to determine the animals’ cause of death and for electrocuted animals we examined the locations the animals had died as well as the configuration of the power lines at these sites. We also recorded the sex of the animal, the body region affected, and characteristics of the injuries. We diagnosed electrocutions in 11% (n = 34) of the marmosets studied. Most of the affected animals were male (n = 22) with single or double sites of injury on the limbs. Animals were injured in urban (n = 26) and peri-urban (n = 8) areas on lower-voltage alternate current lines, and we detected no seasonality or hotspots of electrocution. Our findings suggest that movement along transmission lines composed of bundled conductors is a major factor in electrocutions of marmosets in the Federal District and surrounding areas. The planning of electrical power grid infrastructure should consider arboreal primates to prevent electrocutions.