Evaluation of urinary cysteinyl leukotrienes as biomarkers of severity and putative therapeutic targets in COVID-19 patients.

BACKGROUND: Cysteinyl leukotrienes (CysLT) are potent inflammation-promoting mediators, but remain scarcely explored in COVID-19. We evaluated urinary CysLT (U-CysLT) relationship with disease severity and their usefulness for prognostication in hospitalized COVID-19 patients. The impact on U-CysLT of veno-venous extracorporeal membrane oxygenation (VV-ECMO) and of comorbidities such as hypertension and obesity was also assessed. METHODS: Blood and spot urine were collected in "severe" (n = 26), "critically ill" (n = 17) and "critically ill on VV-ECMO" (n = 17) patients with COVID-19 at days 1-2 (admission), 3-4, 5-8 and weekly thereafter, and in controls (n = 23) at a single time point. U-CysLT were measured by ELISA. Routine markers, prognostic scores and outcomes were also evaluated. RESULTS: U-CysLT did not differ between groups at admission, but significantly increased along hospitalization only in critical groups, being markedly higher in VV-ECMO patients, especially in hypertensives. U-CysLT values during the first week were positively associated with ICU and total hospital length of stay in critical groups and showed acceptable area under curve (AUC) for prediction of 30-day mortality (AUC: 0.734, p = 0.001) among all patients. CONCLUSIONS: U-CysLT increase during hospitalization in critical COVID-19 patients, especially in hypertensives on VV-ECMO. U-CysLT association with severe outcomes suggests their usefulness for prognostication and as therapeutic targets.

Diversity, Equity, and Inclusion in the United States Emergency Medical Services Workforce: A Scoping Review.

OBJECTIVE: Emergency medical services (EMS) workforce demographics in the United States do not reflect the diversity of the population served. Despite some efforts by professional organizations to create a more representative workforce, little has changed in the last decade. This scoping review aims to summarize existing literature on the demographic composition, recruitment, retention, and workplace experience of underrepresented groups within EMS. METHODS: Peer-reviewed studies were obtained from a search of PubMed, CINAHL, Web of Science, ProQuest Thesis and Dissertations, and non-peer-reviewed ("gray") literature from 1960 to present. Abstracts and included full-text articles were screened by two independent reviewers trained on inclusion/exclusion criteria. Studies were included if they pertained to the demographics, training, hiring, retention, promotion, compensation, or workplace experience of underrepresented groups in United States EMS by race, ethnicity, sexual orientation, or gender. Studies of non-EMS fire department activities were excluded. Disputes were resolved by two authors. A single reviewer screened the gray literature. Data extraction was performed using a standardized electronic form. Results were summarized qualitatively. RESULTS: We identified 87 relevant full-text articles from the peer-reviewed literature and 250 items of gray literature. Primary themes emerging from peer-reviewed literature included workplace experience (n = 48), demographics (n = 12), workforce entry and exit (n = 8), education and testing (n = 7), compensation and benefits (n = 5), and leadership, mentorship, and promotion (n = 4). Most articles focused on sex/gender comparisons (65/87, 75%), followed by race/ethnicity comparisons (42/87, 48%). Few articles examined sexual orientation (3/87, 3%). One study focused on telecommunicators and three included EMS physicians. Most studies (n = 60, 69%) were published in the last decade. In the gray literature, media articles (216/250, 86%) demonstrated significant industry discourse surrounding these primary themes. CONCLUSIONS: Existing EMS workforce research demonstrates continued underrepresentation of women and nonwhite personnel. Additionally, these studies raise concerns for pervasive negative workplace experiences including sexual harassment and factors that negatively affect recruitment and retention, including bias in candidate testing, a gender pay gap, and unequal promotion opportunities. Additional research is needed to elucidate recruitment and retention program efficacy, the demographic composition of EMS leadership, and the prevalence of racial harassment and discrimination in this workforce.

Compromised Mitotic Fidelity in Human Pluripotent Stem Cells.

Human pluripotent stem cells (PSCs), which include both embryonic and induced pluripotent stem cells, are widely used in fundamental and applied biomedical research. They have been instrumental for better understanding development and cell differentiation processes, disease origin and progression and can aid in the discovery of new drugs. PSCs also hold great potential in regenerative medicine to treat or diminish the effects of certain debilitating diseases, such as degenerative disorders. However, some concerns have recently been raised over their safety for use in regenerative medicine. One of the major concerns is the fact that PSCs are prone to errors in passing the correct number of chromosomes to daughter cells, resulting in aneuploid cells. Aneuploidy, characterised by an imbalance in chromosome number, elicits the upregulation of different stress pathways that are deleterious to cell homeostasis, impair proper embryo development and potentiate cancer development. In this review, we will summarize known molecular mechanisms recently revealed to impair mitotic fidelity in human PSCs and the consequences of the decreased mitotic fidelity of these cells. We will finish with speculative views on how the physiological characteristics of PSCs can affect the mitotic machinery and how their suboptimal mitotic fidelity may be circumvented.

Assessment and comparison of microcirculation and macrocirculation in horses undergoing emergency exploratory celiotomy versus elective surgical procedures.

OBJECTIVE: To assess oral buccal microcirculation by hand-held videomicroscopy in horses during colic surgery, comparing microcirculation values with macrocirculatory parameters and with those of healthy elective surgical horses. STUDY DESIGN: Clinical prospective study. ANIMALS: Client-owned horses (nine in the colic group; 11 in the elective group). METHODS: In the colic group, buccal mucosal side stream dark-field microscopy (DFM) videos, cardiac output (CO), mean arterial pressure (MAP), and lactate were obtained at three timepoints under general anesthesia (30, 90, and 150 min after induction). Video analysis was used to determine total vessel density, proportion of perfused vessels, perfused vessel density, and heterogeneity index. Dark-field microscopy videos, MAP, and lactate were obtained at a single timepoint under general anesthesia (45 min after induction) in the elective group. RESULTS: There were no differences in microcirculatory parameters between colic and elective horses, nor was there a difference across timepoints in the colic group. There was a weak negative correlation between microvascular parameters and CO (rho = -0.23). CONCLUSION: The colic group did not have decreased microcirculation in comparison with the healthy elective group. Dark-field microscopy did not correlate well with macrocirculatory parameters in the colic group. IMPACT: Dark-field microscopy may not be a sensitive enough indicator to detect differences in microcirculation between colic and elective groups. The lack of difference in microcirculation may be due to sample size, probe location, or variation in disease severity.

Do different protocols affect the success rate or bond strength of glass-ceramics repaired with composite resin? A systematic review and meta-analysis.

STATEMENT OF PROBLEM: Intraoral repair techniques prevent unnecessary replacement of ceramic restorations, thereby increasing the survival rate. However, adhesion between ceramics and the composite resin is challenging and how different protocols influence adhesion is unclear. PURPOSE: The purpose of this systematic review and meta-analysis was to analyze the influence of different protocols on repairing glass-ceramic surfaces with composite resins. MATERIAL AND METHODS: PubMed, Scopus, ISI Web of Science, and Embase electronic databases were searched to select studies comparing bond strength values or survival rates of glass-ceramic repaired with composite resins using different surface treatment protocols. No publication year or language restriction was applied. Data sets were extracted from all included studies, and the mean differences calculated. A 95% confidence interval was calculated by using the random effect model (Rev Man 5.4). RESULTS: The search identified 5037 studies, and 165 were assessed for eligibility. Finally, 123 in vitro studies were included in the systematic review and 48 in the meta-analysis. Considering different glass-ceramics, bond strength tests, and aged or not aged specimens, 37 meta-analyses found the effect of repair protocols: only adhesive, silane plus adhesive alone or preceded by hydrofluoric (HF) acid, airborne-particle abrasion (APA) with Al2O3 particles, silica-coated APA (SCAPA), diamond rotary instrument (DRI), and laser irradiation (LI). CONCLUSIONS: For feldspathic porcelain, HF acid, APA, SCAPA, or DRI improved the repair micromechanical retention; applying silane is essential to HF-conditioned surfaces but the use of adhesive is optional when silane is applied. Results for leucite and lithium disilicate were inconclusive in terms of suggesting a treatment other than HF acid plus silane and adhesive applications.

Risk of anesthesia-related complications in draft horses: a retrospective, single-center analysis.

OBJECTIVE: To report anesthetic-related complications and determine risks associated with anesthesia in draft horses. STUDY DESIGN: Retrospective study. ANIMALS: A total of 401 anesthetic records for draft horse breeds that underwent general anesthesia from January 2010 through December 2020 were reviewed; horses euthanized during general anesthesia were excluded. METHODS: Demographics, perioperative drugs used, procedure type and duration, time to extubation, number of attempts to stand, use of sling in recovery and perioperative morbidity and mortality were investigated. Morbidity and mortality statistical evaluation included univariable logistic regression analysis and ordinal regression analysis. RESULTS: American Society of Anesthesiologists (ASA) status I-II, ASA III-V and total mortality rate for all cases was 0.69% (2/288), 6.19% (7/113) and 2.24% (9/401), respectively, with Belgian horses being overrepresented (6/9). Cardiac arrest occurred in six out of nine horses that died without euthanasia, and five out of six of these horses underwent colic surgery. Factors associated with increased mortality risk included ASA status of III-V, increased body weight, emergency status and horses presenting for colic. Hypotension, hypercarbia and hypoxemia occurred in 56% (224/401), 46% (186/401) and 14% (58/401) of horses, respectively. During recovery from anesthesia, lighter horses and horses undergoing shorter anesthetic procedures were more likely to be successful on the first or second attempt to stand and were less likely to require a sling in recovery. CONCLUSIONS AND CLINICAL RELEVANCE: Draft horses undergoing general anesthesia had a higher mortality rate than previously reported for all types and breeds of horses.

Nuclear phylogenies and genomics of a contact zone establish the species rank of Podarcis lusitanicus (Squamata, Lacertidae).

Unravelling when divergent lineages constitute distinct species can be challenging, particularly in complex scenarios combining cryptic diversity and phylogenetic discordances between different types of molecular markers. Combining a phylogenetic approach with the study of contact zones can help to overcome such difficulties. The Podarcis hispanicus species complex has proven to be prosperous in independent evolutionary units, sometimes associated with cryptic diversity. Previous studies have revealed that one of the species of this complex, P. guadarramae, comprises two deeply divergent yet morphologically indistinguishable evolutionary units, currently regarded as subspecies (P. g. guadarramae and P. g. lusitanicus). In this study we used molecular data to address the systematics of the two lineages of Podarcis guadarramae and the closely related P. bocagei. Firstly, we reconstructed the species tree of these three and two additional taxa based on 30 nuclear loci using the multispecies coalescent with and without gene flow. Secondly, we used SNPs obtained from RADseq data to analyze the population structure across the distribution limits P. g. lusitanicus and P. g. guadarramae, and for comparison, a contact zone between P. bocagei and P. g. lusitanicus. Nuclear phylogenetic relationships between these three taxa are clearly difficult to determine due to the influence of gene flow, but our results give little support to the monophyly of P. guadarramae, potentially due to a nearly simultaneous divergence between them. Genetic structure and geographic cline analysis revealed that the two lineages of P. guadarramae replace each other abruptly across the sampled region and that gene flow is geographically restricted, implying the existence of strong reproductive isolation. Podarcis bocagei and P. g. lusitanicus show a similar degree of genetic differentiation and reproductive isolation, with very low levels of admixture in syntopy. These results support that all three forms are equally differentiated and reproductively isolated. In consequence, we conclude that the two former subspecies of Podarcis guadarramae constitute valid, yet cryptic species, that should be referred to as P. lusitanicus and P. guadarramae.

Evaluation of perfusion index as a noninvasive tool to determine epidural anesthesia effectiveness in dogs.

OBJECTIVE: To evaluate perfusion index (PI) as a noninvasive tool to determine effectiveness and onset of epidural anesthesia in dogs. STUDY DESIGN: Prospective clinical trial. ANIMALS: A total of 21 adult dogs, aged 6.5 ± 3 years and weighing 34.9 ± 6.4 kg, undergoing a tibial plateau leveling osteotomy. METHODS: Dogs were premedicated intramuscularly with acepromazine (0.03 mg kg-1) and hydromorphone (0.1 mg kg-1) and anesthetized with intravenous propofol (to effect) and isoflurane in oxygen. A surface transflectance probe was secured to the tail base to monitor PI and a dorsal pedal artery catheter was placed for invasive blood pressure monitoring. A lumbosacral epidural was performed with the dog in sternal recumbency. Dogs were randomly assigned for inclusion of epidural morphine (0.1 mg kg-1) or morphine (0.1 mg kg-1) and lidocaine (4 mg kg-1). PI was recorded following instrumentation of each dog just prior to the epidural (baseline), at 10 minute intervals for 30 minutes, before and after the surgical skin incision and before and after completion of the osteotomy. Physiological variables and end-tidal isoflurane were recorded at the same time points. RESULTS: There was no significant difference in PI between the groups at any time point. There was a significant change in end-tidal isoflurane before and after the skin incision in the epidural morphine and epidural morphine-lidocaine groups (p = 0.04, p = 0.05, respectively) and before and after the osteotomy in each group for heart rate (p = 0.001, p = 0.04), diastolic (p = 0.01, p = 0.01) and mean arterial blood pressure (p = 0.03, p = 0.05). CONCLUSIONS AND CLINICAL RELEVANCE: PI did not provide an objective means for determining the onset or effectiveness of epidural anesthesia in anesthetized dogs and alternate methods of noninvasive assessment should be investigated.

Effects of perioperative saphenous and sciatic nerve blocks, lumbosacral epidural or morphine-lidocaine-ketamine infusion on postoperative pain and sedation in dogs undergoing tibial plateau leveling osteotomy.

OBJECTIVE: To compare the quality of postoperative analgesia and sedation after preoperative saphenous and sciatic nerve blockade, preoperative lumbosacral epidural injection and perioperative intravenous (IV) morphine, lidocaine and ketamine infusions in dogs undergoing stifle arthroscopy and tibial plateau leveling osteotomy (TPLO) under general anesthesia. STUDY DESIGN: Prospective, blinded, randomized, clinical comparison study. ANIMALS: A total of 45 dogs weighing 33.9 (15.9-56.7) kg and aged 5.2 (1.0-12.0) years, mean (range), undergoing elective unilateral TPLO for spontaneous cranial cruciate ligament rupture. METHODS: Client-owned dogs were enrolled. Dogs were randomly assigned to one of three groups: group MLK, perioperative IV morphine, lidocaine and ketamine infusion; group EPID, lumbosacral epidural with ropivacaine and morphine; or group SSNB, saphenous and sciatic nerve blockade with ropivacaine. Routine stifle arthroscopy followed by TPLO surgery was performed. Sedation and pain scores were assessed at 0, 2, 4, 8 and 24 hours following extubation. Rescue analgesia was administered as prescribed by Glasgow composite pain score-short form score >5. RESULTS: Sedation scores for MLK were higher than EPID and SSNB. Pain scores for SSNB were lower than those for EPID and MLK. No significant differences were found in anesthesia duration or surgery duration among groups. No dogs required rescue analgesia. CONCLUSIONS AND CLINICAL RELEVANCE: Although analgesia was adequate in all groups, the best combination of analgesia without increased sedation was recorded for SSNB.

Evaluation of intramuscular anesthetic protocols in healthy domestic horses.

OBJECTIVE: To assess anesthetic induction, recovery quality and cardiopulmonary variables after intramuscular (IM) injection of three drug combinations for immobilization of horses. STUDY DESIGN: Randomized, blinded, three-way crossover prospective design. ANIMALS: A total of eight healthy adult horses weighing 470-575 kg. METHODS: Horses were administered three treatments IM separated by ≥1 week. Combinations were tiletamine-zolazepam (1.2 mg kg-1), ketamine (1 mg kg-1) and detomidine (0.04 mg kg-1) (treatment TKD); ketamine (3 mg kg-1) and detomidine (0.04 mg kg-1) (treatment KD); and tiletamine-zolazepam (2.4 mg kg-1) and detomidine (0.04 mg kg-1) (treatment TD). Parametric data were analyzed using mixed model linear regression. Nonparametric data were compared using Skillings-Mack test. A p value <0.05 was considered statistically significant. RESULTS: All horses in treatment TD became recumbent. In treatments KD and TKD, one horse remained standing. PaO2 15 minutes after recumbency was significantly lower in treatments TD (p < 0.0005) and TKD (p = 0.001) than in treatment KD. Times to first movement (25 ± 15 minutes) and sternal recumbency (55 ± 11 minutes) in treatment KD were faster than in treatments TD (57 ± 17 and 76 ± 19 minutes; p < 0.0005, p = 0.001) and TKD (45 ± 18 and 73 ± 31 minutes; p = 0.005, p = 0.021). There were no differences in induction quality, muscle relaxation score, number of attempts to stand or recovery quality. CONCLUSIONS AND CLINICAL RELEVANCE: In domestic horses, IM injections of tiletamine-zolazepam-detomidine resulted in more reliable recumbency with a longer duration when compared with ketamine-detomidine and tiletamine-zolazepam-ketamine-detomidine. Recoveries were comparable among protocols.

Aberrantly expressed microRNAs and their implications in childhood central nervous system tumors.

Even though the treatment of childhood cancer has evolved significantly in recent decades, aggressive central nervous system (CNS) tumors are still a leading cause of morbidity and mortality in this population. Consequently, the identification of molecular targets that can be incorporated into diagnostic practice, effectively predict prognosis, follow treatment response, and materialize into potential targeted therapeutic approaches are still warranted. Since the first evidence of the participation of miRNAs in cancer development and progression 20 years ago, notable progress has been made in the basic understanding of the contribution of their dysregulation as epigenetic driver of tumorigenesis. Nevertheless, among the plethora of articles in the literature, microRNA profiling of pediatric tumors are scarce. This article gives an overview of the recent advances in the diagnostic/prognostic potential of miRNAs in a selection of pediatric CNS tumors: medulloblastoma, ependymoma, pilocytic astrocytoma, glioblastoma, diffuse intrinsic pontine glioma, atypical teratoid/rhabdoid tumors, and choroid plexus tumors.

P-glycoprotein activation by 1-(propan-2-ylamino)-4-propoxy-9H-thioxanthen-9-one (TX5) in rat distal ileum: ex vivo and in vivo studies.

In vitro studies showed that 1-(propan-2-ylamino)-4-propoxy-9H-thioxanthen-9-one (TX5) increases P-glycoprotein (P-gp) expression and activity in Caco-2 cells, preventing xenobiotic toxicity. The present study aimed at investigating TX5 effects on P-gp expression/activity using Wistar Han rats: a) in vivo, evaluating intestinal P-gp activity; b) ex vivo, evaluating P-gp expression in ileum brush border membranes (BBM) and P-gp activity in everted intestinal sacs; c) ex vivo, evaluating P-gp activity in everted intestinal sacs of the distal and proximal ileum. TX5 (30 mg/kg, b.w.), gavage, activated P-gp in vivo, given the significant decrease in the AUC of digoxin (0.25 mg/kg, b.w.). The efflux of rhodamine 123 (300 µM), a P-gp fluorescent substrate, significantly increased in TX5-treated everted sacs from the distal portion of the rat ileum, when P-gp activity was evaluated in the presence of TX5 (20 µM), an effect abolished by the P-gp inhibitor verapamil (100 µM). No increases on P-gp expression or activity were found in TX5-treated BBM of the distal ileum and everted distal sacs, respectively, 24 h after TX5 (10 mg/kg, b.w.) administration. In vivo, no differences were found on digoxin portal concentration between control (digoxin 0.025 mg/kg, b.w., intraduodenal) and TX5-treated (digoxin+TX5 20 µM, intraduodenal) rats. The observed discrepancies in digoxin results can be related to differences in TX5 dose administered and used methodologies. Thus, the results show that TX5 activates P-gp at the distal portion of the rat ileum, and, at the higher dose tested (30 mg/kg, b.w.), seems to modulate in vivo the AUC of P-gp substrates.

Immune response of Brazilian farmers exposed to multiple pesticides.

Occupational exposure to pesticides has been identified as a factor that predisposes to disorders of the immune system. Immunosuppression, autoimmunity, cancer of various organs and other diseases in people who apply these products have been reported by the studies. This study aimed to investigate the relationship between occupational exposure to pesticides and the immunological profile in 43 farmers exposed to mixtures of pesticides for at least 15 years. A control group composed of 30 individuals without a history of occupational exposure to pesticides was also evaluated. Peripheral blood samples were processed by flow cytometry and cells were labelled with an 8-color monoclonal antibody panel. Plasma cytokines were also measured. Significant increase in classical monocytes (p < 0.001) and dendritic cells (p < 0.001) in the exposed group was observed as well in total T cells (p = 0.04), central memory CD8 T cells (p = 0.02) and effector memory CD8 T cells (p = 0.01). On the other hand, the activation markers of T cells as the expression of CD57, HLA-DR, CD25 and CD28 were evaluated and no difference was found between groups. When the B cells were analyzed, a significant decrease in total B cells (p = 0.01), regulatory B cells (p < 0.001) and plasmablasts (p < 0.001) in the exposed group, compared to healthy controls, was observed. Pro-inflammatory IL-6 was significantly elevated (p = 0.04) in the plasma of farmers compared to that of controls. The constant antigenic stimulus that occurs during exposure to pesticides can favor the recruitment of dendritic cells and macrophages (APCs) presents in the skin and respiratory tract. In the secondary lymphoid organs, the CD4 T and B cells that process such antigens are possibly undergoing proliferative exhaustion, with the consequent depletion of all mature B subpopulations. The resulting drop in humoral immunity may be offset by an increase in the number of circulating CD8 T lymphocytes due to their cytotoxic action.

DNA damage, oxidative stress, and inflammation in children with celiac disease.

The objective of this study was to evaluate the level of genomic instability in patients with celiac disease and to establish a relationship between inflammation, oxidative stress, and DNA damage in these patients. Myeloperoxidase (MPO) activity, adenosine deaminase, nitric oxide (NOx), thiobarbituric acid, catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and DNA damage were evaluated in peripheral blood samples from 47 celiac disease patients and 31 controls. Patients with celiac disease presented higher levels of DNA damage in comparison to controls (p=0.023). This difference was also observed for markers of oxidative stress, such as CAT (p=0.011) and SOD (p=0.013), and inflammatory markers such as MPO (p < 0.001) and NOx (p=0.009). Positive correlations were found between DNA damage levels and the values of CAT (r=0.405; p=0.009) and SOD (r=0.516; p < 0.001). Positive correlations were also found between GPx and NOx (r=0.349; p=0.030) and MPO and NOx (r=0.239; p=0.039). CAT and NOx showed a negative correlation (r= -0.315; p=0.042). In conclusion, intestinal inflammation can have systemic effects, causing an imbalance between oxidant and antioxidant markers, which may promote increased levels of DNA damage.

Newly Synthesized Oxygenated Xanthones as Potential P-Glycoprotein Activators: In Vitro, Ex Vivo, and In Silico Studies.

P-glycoprotein (P-gp) plays a crucial role in the protection of susceptible organs, by significantly decreasing the absorption/distribution of harmful xenobiotics and, consequently, their toxicity. Therefore, P-gp has been proposed as a potential antidotal pathway, when activated and/or induced. Knowing that xanthones are known to interact with P-gp, the main goal was to study P-gp induction or/and activation by six new oxygenated xanthones (OX 1-6). Furthermore, the potential protection of Caco-2 cells against paraquat cytotoxicity was also assessed. The most promising compound was further tested for its ability to increase P-gp activity ex vivo, using everted intestinal sacs from adult Wistar-Han rats. The oxygenated xanthones interacted with P-gp in vitro, increasing P-gp expression and/or activity 24 h after exposure. Additionally, after a short-incubation period, several xanthones were identified as P-gp activators, as they immediately increased P-gp activity. Moreover, some xanthones decreased PQ cytotoxicity towards Caco-2 cells, an effect prevented under P-gp inhibition. Ex vivo, a significant increase in P-gp activity was observed in the presence of OX6, which was selectively blocked by a model P-gp inhibitor, zosuquidar, confirming the in vitro results. Docking simulations between a validated P-gp model and the tested xanthones predicted these interactions, and these compounds also fitted onto previously described P-gp induction and activation pharmacophores. In conclusion, the in vitro, ex vivo, and in silico results suggest the potential of some of the oxygenated xanthones in the modulation of P-gp, disclosing new perspectives in the therapeutics of intoxications by P-gp substrates.

Prevalence of occult hepatitis B in a population from the Brazilian Amazon region.

In the present study, we evaluated the prevalence of occult hepatitis B (OBI) in a population from the Brazilian Amazon region, identify circulating genotypes, and mutations in the S gene. One hundred eighty-one patients with negative serology for HBsAg and anti-HBs and positive serology for anti-HBc participated in the study. Detection of viral DNA, genotyping by sequencing, and analysis of nucleotide sequences to detect possible mutations were performed. HBV DNA was detected in 14.36% of the patients. Genotyping revealed genotype A in 88.46% of HBV DNA-positive subjects, with subgenotype A1 being the most prevalent (78.26%) followed by subgenotype A2 (21.74%). Genotype F was detected in 11.54% (all of them subgenotype F2). Amino acid substitutions were observed in the amplified S gene in individuals with OBI compared to HBsAg-positive individuals (evident infection). In conclusion, the results show a high prevalence of OBI in the population studied, with a pattern of genotypes A and F that circulate in the Brazilian Amazon region. Amino acid substitutions were detected in part of the S gene in patients with OBI. Further studies on the molecular epidemiology of HBV in this region are important to identify patients considered healthy but who are potential transmitters of the disease.

Polymethoxylated Flavones from Orange Peels Inhibit Cell Proliferation in a 3D Cell Model of Human Colorectal Cancer.

Polymethoxylated flavones (PMFs) have been recognized to inhibit colorectal cancer proliferation through various mechanisms, however most of these studies have been performed on cells grown as monolayers that present limitations in mimicking the 3D tumor architecture and microenvironment. The main aim of this study was to investigate the anticancer potential of an orange peel extract (OPE) enriched in PMFs in a 3D cell model of colorectal cancer. The OPE was developed by supercritical fluid extraction and the anticancer effect was evaluated in HT29 spheroids cultures in a stirred-tank based system. Results showed that OPE inhibited cell proliferation, induced cell cycle arrest (G2/M phase), promoted apoptosis, and reduced ALDH+ population on HT29 spheroids. The antiproliferative activity was significantly lower than that obtained for 2D model (EC50 value of 0.43 ± 0.02 mg/mL) and this effect was dependent on diameter and cell composition/phenotype of spheroids derived from different culture days (day 3 - 0.53 ± 0.05 mg/mL; day 5 - 0.55 ± 0.03 mg/mL; day 7 - 1.24 ± 0.15 mg/mL). HT29 spheroids collected at day 7 presented typical characteristics of in vivo solid tumors including a necrotic/apoptotic core, hypoxia regions, presence of cancer stem cells, and a less differentiated invasive front. Nobiletin, sinesentin, and tangeretin were identified as the main compounds responsible for the anticancer activity.

Markers of genotoxicity and oxidative stress in farmers exposed to pesticides.

The effects of chronic exposure to pesticides can lead to the development of several diseases, including different types of cancer, since the genotoxic and mutagenic capacity of these substances can be observed. The objective of this study is to investigate the relation between the occupational exposure to various pesticides and the presence of DNA damage and oxidative stress. Blood samples from 50 rural workers (41 men and 9 women) exposed to pesticides, 46 controls (20 men and 26 women) from the same city (Antônio Carlos, Santa Catarina state, Brazil) and 29 controls (15 men and 14 women) from another city (Florianópolis, Santa Catarina state, Brazil), were evaluated using the comet assay and the cytokinesis-block micronucleus (CBMN) technique for genetic damage, and the test of thiobarbituric acid reactive substances (TBARS) and catalase (CAT) activity for the oxidative stress. Cholinesterase activities were also determined, but there was no statistical difference among exposed workers and controls. Significant differences were found in DNA damage among groups. The comet assay performed on peripheral blood lymphocytes of these individuals had a significantly higher DNA damage index in the exposed group comparing to controls (p < 0.0001). MNi (p < 0.001), NBUDs (p < 0.005) and NPBs (p < 0.0001) were also found to be significantly higher in the exposed group. The TBARS values were significantly higher comparing to the Florianopolis control group (p < 0.0001). Even though CAT values were higher than controls, there was no statistical difference. Thus, it is concluded that the exposed individuals, participants of this study, are more subject to suffer genetic damage and, consequently, more susceptible to diseases resulting from such damages.

Effect of oral trazodone on the minimum alveolar concentration of isoflurane in dogs.

OBJECTIVE: To determine the effect of oral trazodone on the minimum alveolar concentration (MAC) of isoflurane in dogs. STUDY DESIGN: Prospective blinded, single-observer, randomized crossover experimental study. ANIMALS: Six adult (age 6.8 ± 1.6 months) healthy dogs (three males and three females), weighing 24.8 ± 3.4 kg (mean ± standard deviation). METHODS: Each dog was anesthetized twice with a minimum of 7 days between anesthetic episodes. Dogs were randomly assigned to be administered two treatments in a crossover design: premedication with trazodone (8 mg kg-1; TRAZ-ISO) orally 2 hours prior to an anesthetic episode or no (ISO). Dogs were anesthetized with intravenous propofol (6 mg kg-1) and isoflurane in >95% oxygen. Isoflurane MAC was determined using an iterative bracketing technique with electrodes placed in the buccal mucosa. Hemodynamic variables were compared at the lowest end-tidal isoflurane concentration at which each dog did not respond. A paired t test was used to assess the effect of treatment on outcome variables with significance set to a value of p < 0.05. RESULTS: The MAC concentration (mean ± standard deviation) in dogs administered TRAZ-ISO was 0.85 ± 0.17% compared with 1.02 ± 0.11% in those administered ISO (p = 0.01, 95% confidence interval -0.25 to -0.05), resulting in a mean MAC reduction of 17 ± 12%. There were no differences in hemodynamic variables between treatments. CONCLUSIONS AND CLINICAL RELEVANCE: Premedication of dogs with oral trazodone (8 mg kg-1) 2 hours prior to anesthetic induction has a significant isoflurane MAC sparing effect with no significant observed hemodynamic benefit.

Quantification of 1-(propan-2-ylamino)-4-propoxy-9H-thioxanthen-9-one (TX5), a newly synthetized P-glycoprotein inducer/activator, in biological samples: method development and validation.

A simple, rapid and economical method was developed and validated for the analysis and quantification of 1-(propan-2-ylamino)-4-propoxy-9H-thioxanthen-9-one (TX5), a P-glycoprotein inducer/activator, in biological samples, using reverse-phase high-performance liquid chromatography (HPLC). A C18 column and a mobile phase composed of methanol-water (90/10, v/v) with 1% (v/v) triethylamine, at a flow rate of 1 mL/min, were used for chromatographic separation. TX5 standards (0.5-150 µm) were prepared in human serum. Methanol was used for TX5 extraction and serum protein precipitation. After filtration, samples were injected into the HPLC apparatus and TX5 was quantified by a conventional UV detector at 255 nm. The TX5 retention time was 13 min in this isocratic system. The method was validated according to ICH guidelines for specificity/selectivity, linearity, accuracy, precision, limits of detection and quantification (LOD and LOQ) and recovery. The method was proved to be selective, as there were no interferences of endogenous compounds with the same retention time of TX5. Also, the developed method was linear (r2 ≥ 0.99) for TX5 concentrations between 0.5 and 150 µm and the LOD and LOQ were 0.08 and 0.23 µm, respectively. The results indicated that the reported method could meet the requirements for TX5 analysis in the trace amounts expected to be present in biological samples.