Transcriptome Analysis of Co-Cultures of THP-1 Human Macrophages with Inactivated Germinated Trichophyton rubrum Conidia.

Although most mycoses are superficial, the dermatophyte Trichophyton rubrum can cause systemic infections in patients with a weakened immune system, resulting in serious and deep lesions. The aim of this study was to analyze the transcriptome of a human monocyte/macrophage cell line (THP-1) co-cultured with inactivated germinated T. rubrum conidia (IGC) in order to characterize deep infection. Analysis of macrophage viability by lactate dehydrogenase quantification showed the activation of the immune system after 24 h of contact with live germinated T. rubrum conidia (LGC). After standardization of the co-culture conditions, the release of the interleukins TNF-α, IL-8, and IL-12 was quantified. The greater release of IL-12 was observed during co-culturing of THP-1 with IGC, while there was no change in the other cytokines. Next-generation sequencing of the response to T. rubrum IGC identified the modulation of 83 genes; of these, 65 were induced and 18 were repressed. The categorization of the modulated genes showed their involvement in signal transduction, cell communication, and immune response pathways. In total, 16 genes were selected for validation and Pearson's correlation coefficient was 0.98, indicating a high correlation between RNA-seq and qPCR. Modulation of the expression of all genes was similar for LGC and IGC co-culture; however, the fold-change values were higher for LGC. Due to the high expression of the IL-32 gene in RNA-seq, we quantified this interleukin and observed an increased release in co-culture with T. rubrum. In conclusion, the macrophages-T. rubrum co-culture model revealed the ability of these cells to modulate the immune response, as demonstrated by the release of proinflammatory cytokines and the RNA-seq gene expression profile. The results obtained permit to identify possible molecular targets that are modulated in macrophages and that could be explored in antifungal therapies involving the activation of the immune system.

Salmonella Typhimurium ST313 isolated in Brazil revealed to be more invasive and inflammatory in murine colon compared to ST19 strains.

Salmonella Typhimurium (ST313) has caused an epidemic of invasive disease in sub-Saharan Africa and has been recently identified in Brazil. As the virulence of this ST is poorly understood, the present study aimed to (i) perform the RNA-seq in vitro of S. Typhimurium STm30 (ST313) grown in Luria-Bertani medium at 37°C; (ii) compare it with the RNA-seq of the S. Typhimurium SL1344 (ST19) and S. Typhimurium STm11 (ST19) strains under the same growing conditions; and (iii) examine the colonization capacity and expression of virulence genes and cytokines in murine colon. The STm30 (ST313) strain exhibited stronger virulence and was associated with a more inflammatory profile than the strains SL1344 (ST19) and STm11 (ST19), as demonstrated by transcriptome and in vivo assay. The expression levels of the hilA, sopD2, pipB, and ssaS virulence genes, other Salmonella pathogenicity islands SPI-1 and SPI-2 genes or effectors, and genes of the cytokines IL-1ß, IFN-γ, TNF-α, IL-6, IL-17, IL-22, and IL-12 were increased during ST313 infection in C57BL/6J mice. In conclusion, S. Typhimurium STm30 (ST313) isolated from human feces in Brazil express higher levels of pathogenesis-related genes at 37°C and has stronger colonization and invasion capacity in murine colon due to its high expression levels of virulence genes, when compared with the S. Typhimurium SL1344 (ST19) and STm11 (ST19) strains. STm30 (ST313) also induces stronger expression of pro-inflammatory cytokines in this organ, suggesting that it causes more extensive tissue damage.

Noninvasive neuromodulation of the prefrontal cortex in young women with obesity: a randomized clinical trial.

BACKGROUND/OBJECTIVES: Obesity is associated with reduced neurocognitive performance. Individuals with obesity show decreased activation in the left dorsolateral prefrontal cortex (DLPFC), a key brain region relevant to the regulation of eating behavior. Transcranial direct current stimulation (tDCS) has emerged as a potential technique to correct these abnormalities. However, there is limited information to date, particularly in clinical settings and regarding long-term effects of tDCS. This study aimed to investigate the effects of DLPFC-targeted tDCS in young women with obesity. SUBJECT/METHODS: Randomized, double-blind, sham-controlled parallel-design clinical trial conducted in 38 women, aged 20-40 years, with BMI 30-35 kg/m2. STUDY DESIGN: Phase I: target engagement (immediate effects of tDCS on working memory performance), Phase II: tDCS only (ten sessions, 2 weeks), Phase III: tDCS + hypocaloric diet (six sessions, 30% energy intake reduction, 2 weeks, inpatient), Phase IV: follow-up at 1, 3, and 6 months. PRIMARY OUTCOME: change in body weight. SECONDARY OUTCOMES: change in eating behavior and appetite. Additional analyses: effect of Catechol-O-methyl transferase (COMT) gene variability. Data were analyzed as linear mixed models. RESULTS: There was no group difference in change in body weight during the tDCS intervention. At follow-up, the active group lost less weight than the sham group. In addition, the active group regained weight at 6-month follow-up, compared with sham. Genetic analysis indicated that COMT Met noncarriers were the subgroup that accounted for this paradoxical response in the active group. CONCLUSION: Our results suggest that in young women with class I obesity, tDCS targeted to the DLPFC does not facilitate weight loss. Indeed, we found indications that tDCS could have a paradoxical effect in this population, possibly connected with individual differences in dopamine availability. Future studies are needed to confirm these findings.

Estrous cycle impacts microRNA content in extracellular vesicles that modulate bovine cumulus cell transcripts during in vitro maturation†.

Extracellular vesicles (EVs) are nanoparticles secreted by ovarian follicle cells. Extracellular vesicles are an important form of intercellular communication, since they carry bioactive contents, such as microRNAs (miRNAs), mRNAs, and proteins. MicroRNAs are small noncoding RNA capable of modulating mRNA translation. Thus, EVs can play a role in follicle and oocyte development. However, it is not clear if EV contents vary with the estrous cycle stage. The aim of this study was to investigate the bovine miRNA content in EVs obtained from follicles at different estrous cycle stages, which are associated with different progesterone (P4) levels in the follicular fluid (FF). We collected FF from 3 to 6 mm follicles and evaluated the miRNA profile of the EVs and their effects on cumulus-oocyte complexes during in vitro maturation. We observed that EVs from low P4 group have a higher abundance of miRNAs predicted to modulate pathways, such as MAPK, RNA transport, Hippo, Cell cycle, FoxO, oocyte meiosis, and TGF-beta. Additionally, EVs were taken up by cumulus cells and, thus, affected the RNA global profile 9 h after EV supplementation. Cumulus cells supplemented with EVs from low P4 presented upregulated genes that could modulate biological processes, such as oocyte development, immune responses, and Notch signaling compared with genes of cumulus cells in the EV free media or with EVs from high P4 follicles. In conclusion, our results demonstrate that EV miRNA contents are distinct in follicles exposed to different estrous cycle stage. Supplementation with EVs impacts gene expression and biological processes in cumulus cells.

Appetite effects of prefrontal stimulation depend on COMT Val158Met polymorphism: A randomized clinical trial.

The regulation of appetite is supported by dopamine-modulated brain circuits. Recent studies have shown that transcranial direct current stimulation (tDCS) aimed at increasing the excitability of the dorsolateral prefrontal cortex can reduce appetite, but the underlying mechanisms remain unknown, and response variability is large. The aim of this study was to determine whether individual differences in Catechol-O-methyl transferase (COMT) Val158Met polymorphism can influence tDCS effects on appetite. Thirty-eight adult women with obesity, classified as carriers or non-carriers of the Met allele, underwent a randomized, double-blind, sham-controlled tDCS intervention involving three phases: Phase I, target engagement (immediate effects of tDCS on working memory performance), Phase II, tDCS only (10 sessions, two weeks), and Phase III, tDCS + hypocaloric diet: (6 sessions, two weeks, 30% energy intake reduction, inpatient). Data were analyzed using linear mixed-effects models and mixed ANCOVA. Appetite was evaluated using visual analogue scales. We found that Met-carriers receiving active tDCS were the only participants who experienced a significant reduction of appetite over time. Conversely, Met non-carriers maintained high levels of appetite during the intervention; this effect was driven by a delayed paradoxical rise in appetite after stimulation. Working memory task performance at phase I correlated with subsequent appetite change in a COMT-dependent manner: speed improvements during the task predicted appetite increase in Met carriers and appetite reduction in Met non-carriers. Our findings suggest that genotype differences impacting dopamine levels influence prefrontal tDCS effects on appetite. This source of variability should be considered in the design of future studies.

Influence of expression of UCP3, PLIN1 and PPARG2 on the oxidation of substrates after hypocaloric dietary intervention.

BACKGROUND & AIMS: In addition to environmental and psychosocial factors, it is known that genetic factors can also influence the regulation of energy metabolism, body composition and determination of excess weight. The objective of this study was to evaluate the influence of UCP3, PLIN1 and PPARG2 genes on the substrates oxidation in women with grade III obesity after hypocaloric dietary intervention. SUBJECTS/METHODS: This is a longitudinal study with 21 women, divided into two groups: Intervention Group (G1): 11 obese women (Body Mass Index (BMI) ≥40 kg/m2), and Control Group (G2): 10 eutrophic women (BMI between 18.5 kg/m2 and 24.9 kg/m2). Weight (kg), height (m), BMI (kg/m2), substrate oxidation (by Indirect Calorimetry) and abdominal subcutaneous adipose tissue were collected before and after the intervention. For the dietary intervention, the patients were hospitalized for 6 weeks receiving 1200 kcal/day. RESULTS: There was a significant weight loss (8.4 ± 4.3 kg - 5.2 ± 1.8%) and reduction of UCP3 expression after hypocaloric dietary intervention. There was a positive correlation between carbohydrate oxidation and UCP3 (r = 0.609; p = 0.04), PLIN1 (r = 0.882; p = 0.00) and PPARG2 (r = 0.791; p = 0.00) expression before dietary intervention and with UCP3 (r = 0.682; p = 0.02) and PLIN1 (r = 0.745; p = 0.00) genes after 6 weeks of intervention. There was a negative correlation between lipid oxidation and PLIN1 (r = -0.755; p = 0.00) and PPARG2 (r = 0.664; p = 0.02) expression before dietary intervention and negative correlation with PLIN1 (r = 0.730; p = 0.02) expression after 6 weeks of hypocaloric diet. CONCLUSION: Hypocaloric diet reduces UCP3 expression in individuals with obesity and the UCP3, PLIN1 and PPARG2 expression correlate positively with carbohydrate oxidation and negatively with lipid oxidation.

Changes in Global Transcriptional Profiling of Women Following Obesity Surgery Bypass.

BACKGROUND: Differential gene expression in peripheral blood mononuclear cells (PBMCs) after Roux-en-Y gastric bypass (RYGB) is poorly characterized. Markers of these processes may provide a deeper understanding of the mechanisms that underlie these events. The main goal of this study was to identify changes in PBMC gene expression in women with obesity before and 6 months after RYGB-induced weight loss. METHODS: The ribonucleic acid (RNA) of PBMCs from 13 obese women was analyzed before and 6 months after RYGB; the RNA of PBMCs from nine healthy women served as control. The gene expression levels were determined by microarray analysis. Significant differences in gene expression were validated by real-time quantitative polymerase chain reaction (RT-qPCR). RESULTS: Microarray analysis for comparison of the pre- and postoperative periods showed that 1366 genes were differentially expressed genes (DEGs). The main pathways were related to gene transcription; lipid, energy, and glycide metabolism; inflammatory and immunological response; cell differentiation; oxidative stress regulation; response to endogenous and exogenous stimuli; substrate oxidation; mTOR signaling pathway; interferon signaling; mitogen-activated protein kinases (MAPK), cAMP response element binding protein (CREB1), heat shock factor 1 (HSF1), and sterol regulatory element binding protein 1c (SREBP-1c) gene expression; adipocyte differentiation; and methylation. CONCLUSIONS: Six months after bariatric surgery and significant weight loss, many molecular pathways involved in obesity and metabolic diseases change. These findings are an important tool to identify potential targets for therapeutic intervention and clinical practice of nutritional genomics in obesity.

Exome sequencing of multiple-sclerosis patients and their unaffected first-degree relatives.

OBJECTIVES: The understanding of complex multifactorial diseases requires the availability of a variety of data for a large-number of affected individuals. In this data note here we provide whole exome sequencing data from a set of non-familiar multiple-sclerosis (MS) patients as well as their unaffected first-degree relatives. This data might help the identification of genomic alterations, including single nucleotide polymorphisms, de novo variations and structural genomic variations, such as copy-number alterations that may impact this disease. DATA DESCRIPTION: This dataset comprises the full exome of 28 Brazilian subjects grouped in eight distinct families, consisting of four complete trios (mother-patient-father) plus another four complete trios with one added unaffected sibling. In total, we present the full exome data of eight patients diagnosed with recurrent remittent multiple sclerosis. Diagnoses were made by experienced neurologists and all enrolled patients had at least 5 years of follow up and specific MS treatment. Exomes were sequenced from leukocyte-derived DNA, after the capture of exons using biotinylated probes, in the Ion Proton platform. For each exome we generated an average of 66.1 million good quality mapped reads with an average length of ~ 160nt. On average, for 90% of the exome a vertical coverage above 20× was reached.

UCP2 and PLIN1 Expression Affects the Resting Metabolic Rate and Weight Loss on Obese Patients.

BACKGROUND: Analysis of the expression of genes related to the energy metabolism can elucidate the understanding of physiological and genetic factors that contribute to obesity. This study aimed to evaluate the expression of genes and its influence on resting metabolic rate and weight loss in obese patients before and after bariatric surgery. METHODS: This study was conducted on 23 women, who were divided into two groups: bariatric surgery (preoperative and 6 months after surgery) and control. Abdominal subcutaneous adipose tissue samples were collected to analyze the gene expression, and the resting metabolic rate (RMR) was measured by indirect calorimetry. RESULTS: Significant differences were observed in weight reduction (22 %, p = 0.01), BMI (22.5 %, p = 0.01), and RMR values (10.5 %, p = 0.01) after the bariatric surgery, while the weight-adjusted RMR increased (15.8 %, p = 0.01). Increased UCP2 expression after 6 months of Roux-en-Y gastric bypass (RYGB) as compared to preoperative period (0.764 to 1.268, p = 0.01) was observed. Analysis with weight-adjusted RMR as dependent variable revealed that UCP2 (r 2 = 0.517, p = 0.01) and PLIN1 (r 2 = 0.420, p = 0.04) expression determine the RMR values in preoperative period. Moreover, UCP2 and PLIN1 expression in preoperative period influenced the percentage of weight loss, even when adjusted for age and BMI. CONCLUSIONS: We have demonstrated that after 6 months of bariatric surgery, there is significant increase in the UCP2 expression. Additionally, the expression of UCP2 and PLIN1 genes influences the resting metabolic rate in obese individuals and could predict the weight loss after bariatric surgery.

Analysis of the presence of the GJB6 mutations in patients heterozygous for GJB2 mutation in Brazil.

Mutations in the GJB2 gene, mainly 35delG, are responsible for most autosomal recessive inherited genetic hearing loss. The audiometric standard of these hearing losses remains inconsistent and other genes, such as GJB6, have been involved in association with GJB2. The objective of the study was to identify the deletions del(GJB6-D13S1830) and del(GJB6-D13S1854) in patients heterozygous for 35delG/GJB2 and analyze the phenotype they present. 101 patients with mild to profound degree of sensorineural hypoacusis were evaluated. The allele-specific PCR technique was used to identify 35delG. The del(GJB6-D13S1830) and del(GJB6-D13S1854) were identified through the PCR multiplex technique. 90% of the subjects presented a normal genotype for the analyzed mutations; 6.93% were shown to be heterozygous for 35delG/GJB2 and 1% presented compound heterozygosis GJB2/GJB6). The data found reinforced the hypothesis of an interaction of more than one gene as the cause of autosomal recessive genetic hearing loss and emphasized the importance of an early diagnosis for appropriate intervention.

Predictive models for mutations in mismatch repair genes: implication for genetic counseling in developing countries.

BACKGROUND: Lynch syndrome (LS) is the most common form of inherited predisposition to colorectal cancer (CRC), accounting for 2-5% of all CRC. LS is an autosomal dominant disease characterized by mutations in the mismatch repair genes mutL homolog 1 (MLH1), mutS homolog 2 (MSH2), postmeiotic segregation increased 1 (PMS1), post-meiotic segregation increased 2 (PMS2) and mutS homolog 6 (MSH6). Mutation risk prediction models can be incorporated into clinical practice, facilitating the decision-making process and identifying individuals for molecular investigation. This is extremely important in countries with limited economic resources. This study aims to evaluate sensitivity and specificity of five predictive models for germline mutations in repair genes in a sample of individuals with suspected Lynch syndrome. METHODS: Blood samples from 88 patients were analyzed through sequencing MLH1, MSH2 and MSH6 genes. The probability of detecting a mutation was calculated using the PREMM, Barnetson, MMRpro, Wijnen and Myriad models. To evaluate the sensitivity and specificity of the models, receiver operating characteristic curves were constructed. RESULTS: Of the 88 patients included in this analysis, 31 mutations were identified: 16 were found in the MSH2 gene, 15 in the MLH1 gene and no pathogenic mutations were identified in the MSH6 gene. It was observed that the AUC for the PREMM (0.846), Barnetson (0.850), MMRpro (0.821) and Wijnen (0.807) models did not present significant statistical difference. The Myriad model presented lower AUC (0.704) than the four other models evaluated. Considering thresholds of ≥ 5%, the models sensitivity varied between 1 (Myriad) and 0.87 (Wijnen) and specificity ranged from 0 (Myriad) to 0.38 (Barnetson). CONCLUSIONS: The Barnetson, PREMM, MMRpro and Wijnen models present similar AUC. The AUC of the Myriad model is statistically inferior to the four other models.

In vitro and in vivo studies on CCR10 regulation by Annexin A1.

The mode of action of annexin A1 (ANXA1) is poorly understood. By using rapid subtraction hybridization we studied the effects of human recombinant ANXA1 and the N-terminal ANXA1 peptide on gene expression in a human larynx cell line. Three genes showed strong downregulation after treatment with ANXA1. In contrast, expression of CCR10, a seven transmembrane G-protein coupled receptor for chemokine CCL27 involved in mucosal immunity, was increased. Moreover the reduction in CCR10 expression induced by ANXA1 gene deletion was rescued by intravenous treatment with low doses of ANXA1. These findings provide new evidence that ANXA1 modulates gene expression.

Heterogeneity of the genome ancestry of individuals classified as White in the state of Rio Grande do Sul, Brazil.

One hundred nineteen individuals classified as White, living in different localities of the Brazilian state of Rio Grande do Sul, were studied in relation to the HVS-I region of the mitochondrial DNA (mtDNA). The male fraction of the sample (N = 74) was also tested for seven Y-chromosome polymorphisms. In a specific population (Veranópolis), a city characterized by a large influence of the Italian immigration of the 19th century, the results from the maternal and paternal sides indicated almost complete European ancestry. However, another sample identified as White, from different localities of Rio Grande do Sul, presented significant fractions of Native American (36%) and African (16%) mtDNA haplogroups. These results indicate that Brazilian populations are remarkably heterogeneous; while some present an overwhelming majority of transplanted European genomes, with a complete correspondence between physical appearance and ancestry, others reflect a history of extensive admixture with dissociation between physical appearance and ancestry.

Ethnicity and type 2 diabetes in Rio de Janeiro, Brazil, with a review of the prevalence of the disease in Amerindians.

To what extent can ethnic factors contribute to the prevalence of type 2 diabetes and impaired glucose tolerance (IGT) in an urban Brazilian population? Conversely, how can environmental factors such as diet change these prevalences in a given ethnic group, in this case Brazilian Indians? To answer these questions estimates of ethnic admixture in Afro- and Euro-Brazilians from Rio de Janeiro, Brazil, were established using eight genetic systems and compared with the prevalences of these conditions obtained previously. This information was integrated with results obtained inside and outside of Brazil. The similarity of prevalences for type 2 diabetes and IGT in Afro- and Euro-Brazilians may be related to the extensive gene flow that occurred between them and to similar socioeconomic levels in the samples investigated. On the other hand, changes in the traditional diet are probably conditioning the appearance of diabetes among Brazilian and other South American Indians.