A Prognostic Methylation-Driven Two-Gene Signature in Medulloblastoma.

Medulloblastoma (MB) is one of the most common pediatric brain tumors and it is estimated that one-third of patients will not achieve long-term survival. Conventional prognostic parameters have limited and unreliable correlations with MB outcome, presenting a major challenge for patients' clinical improvement. Acknowledging this issue, our aim was to build a gene signature and evaluate its potential as a new prognostic model for patients with the disease. In this study, we used six datasets totaling 1679 samples including RNA gene expression and DNA methylation data from primary MB as well as control samples from healthy cerebellum. We identified methylation-driven genes (MDGs) in MB, genes whose expression is correlated with their methylation. We employed LASSO regression, incorporating the MDGs as a parameter to develop the prognostic model. Through this approach, we derived a two-gene signature (GS-2) of candidate prognostic biomarkers for MB (CEMIP and NCBP3). Using a risk score model, we confirmed the GS-2 impact on overall survival (OS) with Kaplan-Meier analysis. We evaluated its robustness and accuracy with receiver operating characteristic curves predicting OS at 1, 3, and 5 years in multiple independent datasets. The GS-2 showed highly significant results as an independent prognostic biomarker compared to traditional MB markers. The methylation-regulated GS-2 risk score model can effectively classify patients with MB into high and low-risk, reinforcing the importance of this epigenetic modification in the disease. Such genes stand out as promising prognostic biomarkers with potential application for MB treatment.

Bridging the Gaps in Meta-Omic Analysis: Workflows and Reproducibility.

The past few years have seen significant advances in the study of complex microbial communities associated with the evolution of sequencing technologies and increasing adoption of whole genome shotgun sequencing methods over the once more traditional Amplicon-based methods. Although these advances have broadened the horizon of meta-omic analyses in planetary health, human health, and ecology from simple sample composition studies to comprehensive taxonomic and metabolic profiles, there are still significant challenges in processing these data. First, there is a widespread lack of standardization in data processing, including software choices and the ease of installing and running attendant software. This can lead to several inconsistencies, making comparing results across studies and reproducing original results difficult. We argue that these drawbacks are especially evident in metatranscriptomic analysis, with most analyses relying on ad hoc scripts instead of pipelines implemented in workflow managers. Additional challenges rely on integrating meta-omic data, since methods have to consider the biases in the library preparation and sequencing methods and the technical noise that can arise from it. Here, we critically discuss the current limitations in metagenomics and metatranscriptomics methods with a view to catalyze future innovations in the field of Planetary Health, ecology, and allied fields of life sciences. We highlight possible solutions for these constraints to bring about more standardization, with ease of installation, high performance, and reproducibility as guiding principles.

Revealing metastatic castration-resistant prostate cancer master regulator through lncRNAs-centered regulatory network.

BACKGROUND: Metastatic castration-resistant prostate cancer (mCRPC) is an aggressive form of cancer unresponsive to androgen deprivation therapy (ADT) that spreads quickly to other organs. Despite reduced androgen levels after ADT, mCRPC development and lethality continues to be conducted by the androgen receptor (AR) axis. The maintenance of AR signaling in mCRPC is a result of AR alterations, androgen intratumoral production, and the action of regulatory elements, such as noncoding RNAs (ncRNAs). ncRNAs are key elements in cancer signaling, acting in tumor growth, metabolic reprogramming, and tumor progression. In prostate cancer (PCa), the ncRNAs have been reported to be associated with AR expression, PCa proliferation, and castration resistance. In this study, we aimed to reconstruct the lncRNA-centered regulatory network of mCRPC and identify the lncRNAs which act as master regulators (MRs). METHODS: We used publicly available RNA-sequencing to infer the regulatory network of lncRNAs in mCRPC. Five gene signatures were employed to conduct the master regulator analysis. Inferred MRs were then subjected to functional enrichment and symbolic regression modeling. The latter approach was applied to identify the lncRNAs with greater predictive capacity and potential as a biomarker in mCRPC. RESULTS: We identified 31 lncRNAs involved in cellular proliferation, tumor metabolism, and invasion-metastasis cascade. SNHG18 and HELLPAR were the highlights of our results. SNHG18 was downregulated in mCRPC and enriched to metastasis signatures. It accurately distinguished both mCRPC and primary CRPC from normal tissue and was associated with epithelial-mesenchymal transition (EMT) and cell-matrix adhesion pathways. HELLPAR consistently distinguished mCRPC from primary CRPC and normal tissue using only its expression. CONCLUSION: Our results contribute to understanding the regulatory behavior of lncRNAs in mCRPC and indicate SNHG18 and HELLPAR as master regulators and potential new diagnostic targets in this tumor.

Reverse Engineering of the Pediatric Sepsis Regulatory Network and Identification of Master Regulators.

Sepsis remains a leading cause of death in ICUs all over the world, with pediatric sepsis accounting for a high percentage of mortality in pediatric ICUs. Its complexity makes it difficult to establish a consensus on genetic biomarkers and therapeutic targets. A promising strategy is to investigate the regulatory mechanisms involved in sepsis progression, but there are few studies regarding gene regulation in sepsis. This work aimed to reconstruct the sepsis regulatory network and identify transcription factors (TFs) driving transcriptional states, which we refer to here as master regulators. We used public gene expression datasets to infer the co-expression network associated with sepsis in a retrospective study. We identified a set of 15 TFs as potential master regulators of pediatric sepsis, which were divided into two main clusters. The first cluster corresponded to TFs with decreased activity in pediatric sepsis, and GATA3 and RORA, as well as other TFs previously implicated in the context of inflammatory response. The second cluster corresponded to TFs with increased activity in pediatric sepsis and was composed of TRIM25, RFX2, and MEF2A, genes not previously described as acting in a coordinated way in pediatric sepsis. Altogether, these results show how a subset of master regulators TF can drive pathological transcriptional states, with implications for sepsis biology and treatment.

Ionotropic Receptors as a Driving Force behind Human Synapse Establishment.

The origin of nervous systems is a main theme in biology and its mechanisms are largely underlied by synaptic neurotransmission. One problem to explain synapse establishment is that synaptic orthologs are present in multiple aneural organisms. We questioned how the interactions among these elements evolved and to what extent it relates to our understanding of the nervous systems complexity. We identified the human neurotransmission gene network based on genes present in GABAergic, glutamatergic, serotonergic, dopaminergic, and cholinergic systems. The network comprises 321 human genes, 83 of which act exclusively in the nervous system. We reconstructed the evolutionary scenario of synapse emergence by looking for synaptic orthologs in 476 eukaryotes. The Human-Cnidaria common ancestor displayed a massive emergence of neuroexclusive genes, mainly ionotropic receptors, which might have been crucial to the evolution of synapses. Very few synaptic genes had their origin after the Human-Cnidaria common ancestor. We also identified a higher abundance of synaptic proteins in vertebrates, which suggests an increase in the synaptic network complexity of those organisms.

Anti-inflammatory and antixidant properties of blend formulated with compounds of Malpighia emarginata D.C (acerola) and Camellia sinensis L. (green tea) in lipopolysaccharide-stimulated RAW 264.7 macrophages.

The antioxidant and anti-inflammatory properties of Malpighia emarginata D.C (acerola) and Camellia sinensis L. (green tea) have been studied, particularly as an alternative in medicinal approach for different physio pathological conditions. Here we develop an powder blend formulated with both Malpighia emarginata D.C and Camellia sinensis L. which have in the composition higher content of ascorbic acid and epigallatocathechin-3-gallate respectively. Using different conditions for microencapsulation of biocompounds, we performed the powder production through spray-drying process. After, we evaluate the antioxidant and anti-inflammatory properties of blends formulated with Malpighia emarginata D.C and Camellia sinensis L. in an in vitro model of inflammation, using LPS-stimulated RAW-264.7 macrophage cell line. We observed that co-treatment with blends was able to modulate the redox parameters in cells during the in vitro inflammatory response. Moreover, the co-treatment with blends were able to modulate inflammatory response by altering the secretion of cytokines IL-1ß, IL-6, IL-10, and TNF-α. Taken together, our results demonstrate for the first time the synergistic effects antioxidant and anti-inflammatory of Malpighia emarginata D.C and Camellia sinensis L. These results warrant further use of the blend powder for use in the products to heath beneficial, principally in terms of prevention of chronic diseases.

Analysis of Arabidopsis thaliana Redox Gene Network Indicates Evolutionary Expansion of Class III Peroxidase in Plants.

Reactive oxygen species (ROS) are byproducts of aerobic metabolism and may cause oxidative damage to biomolecules. Plants have a complex redox system, involving enzymatic and non-enzymatic compounds. The evolutionary origin of enzymatic antioxidant defense in plants is yet unclear. Here, we describe the redox gene network for A. thaliana and investigate the evolutionary origin of this network. We gathered from public repositories 246 A. thaliana genes directly involved with ROS metabolism and proposed an A. thaliana redox gene network. Using orthology information of 238 Eukaryotes from STRINGdb, we inferred the evolutionary root of each gene to reconstruct the evolutionary history of A. thaliana antioxidant gene network. We found two interconnected clusters: one formed by SOD-related, Thiol-redox, peroxidases, and other oxido-reductase; and the other formed entirely by class III peroxidases. Each cluster emerged in different periods of evolution: the cluster formed by SOD-related, Thiol-redox, peroxidases, and other oxido-reductase emerged before opisthokonta-plant divergence; the cluster composed by class III peroxidases emerged after opisthokonta-plant divergence and therefore contained the most recent network components. According to our results, class III peroxidases are in expansion throughout plant evolution, with new orthologs emerging in each evaluated plant clade divergence.

Sunlight Incidence, Vitamin D Deficiency, and Alzheimer's Disease.

Vitamin D (VD) deficiency is a growing problem, affecting a significant portion of the population in many countries. VD deficiency may be related to several diseases, including Alzheimer's disease (AD). This study aimed to review the relationship between VD deficiency and AD. We describe the proteins involved in AD pathogenesis and how those proteins can be influenced by VD deficiency. We also investigated a relationship between AD death rate and solar radiation and we found an increased AD death rate in countries with low sunlight. It was also observed that amyloid precursor protein, ryanodine receptor, mammalian target of rapamycin complex 1, and receptor for advanced glycation end products are associated with a worse prognosis in AD. While the Klotho protein, phosphatase and tensin homologue, and VD receptor are associated with a better prognosis in the disease. The literature suggests that decline in VD concentrations may be involved in the establishment and progression of AD. According to sunlight data, we can conclude that countries with low average sunlight have high AD death rate.

Lead-interacting proteins and their implication in lead poisoning.

Lead is an important heavy metal used worldwide in several applications, especially in industry. People exposed to lead can develop a wide range of symptoms associated with lead poisoning. Many effects of lead poisoning are reported in the literature, showing a compromising of whole body health, with symptoms related to cardiovascular, immune, bone, reproductive, hematological, renal, gastrointestinal, and nervous system. However, the molecular lead targets as well as the pathways affected by lead poisoning are not completely described. The aim of this study was to construct a map of metabolic pathways impaired in lead poisoning by evaluating which biomolecules are directly affected by lead. Through manual literature curation, we identified proteins which physically interact with lead and subsequently determined the metabolic pathways those proteins are involved with. At total, we identified 23 proteins involved with heme synthesis, calcium metabolism, neurotransmission, among other biological systems, which helps to understand the wide range of lead-poisoning symptoms.

Antioxidant and Anti-Inflammatory Properties of Anacardium occidentale Leaf Extract.

In tropical America, principally in Northeastern Brazil, the leaf extract of Anacardium occidentale is traditionally used for treatment of different diseases. However, chemical and biological properties and activities of Anacardium occidentale are poorly investigated and known. Here, we evaluated the antioxidant and anti-inflammatory activities "in vitro" of leaf extract from Anacardium occidentale. Our results show that leaf extract exhibits antioxidant activity when used to treat RAW 264.7 macrophage cells. Antioxidant effects were observed by decrease in oxidative damage in macrophage cells treated with 0.5 µg/mL and 5 µg/mL of leaf extract. Moreover, leaf extract reversed oxidative damage and inflammatory parameters induced in LPS-stimulated RAW 264.7 macrophage cells. Leaf extract at 0.5 µg/mL and 5 µg/mL was able to inhibit release of TNF-α and IL-1ß in LPS-stimulated cells. Taken together, our results indicate antioxidant and anti-inflammatory effects of leaf extract from Anacardium occidentale and reveal the positive effects that intake of these products can mediate in biological system.

Analysis of genome instability biomarkers in children with non-syndromic orofacial clefts.

The non-syndromic cleft lip and/or palate (NSCL/P) is a common birth defect caused by a combination of genetic and environmental factors. The possible role of genome instability on NSCL/P patient needs more investigation, since DNA metabolism is an essential cellular function to keep cells with normal genotypes and gene expression patterns according to tissue specificities, which is critical during embryo development because it requires sensitive regulation of cell proliferation, apoptosis and differentiation. Thus, genome stability is ultimately essential to maintain a healthy life. The aim of this study was to assess the frequency of genome instability biomarkers and their relationship with NSCL/P. Cytokinesis-block micronucleus assay was performed to estimate the biomarkers frequency and gene expression was analyzed by the transcriptogram in order to further explore the role of genome instability and other biological processes in this birth defect. The NSCL/P patients had higher baseline frequency of micronucleus, nuclear buds and nucleoplasmic bridges (P < 0.001) than the control group. Moreover, new nuclear morphologies (fused, circular and horseshoe) was detected in the patients' cells analyzed, possibly indicating that chronic folic acid deficiency is interfering in their genome instability. Children with clefts had 2.3 times more risk to have high micronuclei frequency (P = 0.043) according to binary logistic regression. The high genomic instability in children with oral clefts suggests that misrepaired double strand breaks in DNA that create micronuclei representing a significant factor in NSCL/P development. This study was published in 52nd EUROTOX Abstract Book.

Turnera subulata Anti-Inflammatory Properties in Lipopolysaccharide-Stimulated RAW 264.7 Macrophages.

In South America, particularly in the Northeastern regions of Brazil, Turnera subulata leaf extract is used as an alternative traditional medicine approach for several types of chronic diseases, such as diabetes, hypertension, chronic pain, and general inflammation. Despite its widespread use, little is known about the medicinal properties of the plants of this genus. In this study, we evaluate the antioxidant and anti-inflammatory of T. subulata leaf extract in an in vitro model of inflammation, using lipopolysaccharide-stimulated RAW-264.7 macrophage cell line. We observed that cotreatment with T. subulata leaf extract was able to reduce the oxidative stress in cells due to inflammatory response. More importantly, we observed that the leaf extract was able to directly modulate inflammatory response by altering activity of members of the mitogen-activated protein kinase pathways. Our results demonstrate for the first time that T. subulata have antioxidant and anti-inflammatory properties, which warrant further investigation of the medicinal potential of this species.

Gene Expression Profile of NF-κB, Nrf2, Glycolytic, and p53 Pathways During the SH-SY5Y Neuronal Differentiation Mediated by Retinoic Acid.

SH-SY5Y cells, a neuroblastoma cell line that is a well-established model system to study the initial phases of neuronal differentiation, have been used in studies to elucidate the mechanisms of neuronal differentiation. In the present study, we investigated alterations of gene expression in SH-SY5Y cells during neuronal differentiation mediated by retinoic acid (RA) treatment. We evaluated important pathways involving nuclear factor kappa B (NF-κB), nuclear E2-related factor 2 (Nrf2), glycolytic, and p53 during neuronal differentiation. We also investigated the involvement of reactive oxygen species (ROS) in modulating the gene expression profile of those pathways by antioxidant co-treatment with Trolox®, a hydrophilic analogue of α-tocopherol. We found that RA treatment increases levels of gene expression of NF-κB, glycolytic, and antioxidant pathway genes during neuronal differentiation of SH-SY5Y cells. We also found that ROS production induced by RA treatment in SH-SY5Y cells is involved in gene expression profile alterations, chiefly in NF-κB, and glycolytic pathways. Antioxidant co-treatment with Trolox® reversed the effects mediated by RA NF-κB, and glycolytic pathways gene expression. Interestingly, co-treatment with Trolox® did not reverse the effects in antioxidant gene expression mediated by RA in SH-SY5Y. To confirm neuronal differentiation, we quantified endogenous levels of tyrosine hydroxylase, a recognized marker of neuronal differentiation. Our data suggest that during neuronal differentiation mediated by RA, changes in profile gene expression of important pathways occur. These alterations are in part mediated by ROS production. Therefore, our results reinforce the importance in understanding the mechanism by which RA induces neuronal differentiation in SH-SY5Y cells, principally due this model being commonly used as a neuronal cell model in studies of neuronal pathologies.

Transcriptomic analysis reveals pH-responsive antioxidant gene networks.

Reactive oxygen species (ROS) are produced in different physiological conditions. In response to ROS imbalance cells activate oxidative stress defenses, which include more than 60 antioxidant genes. It has been suggested that gene products associated with ROS detoxification can work coordinately, acting as an antioxidant-defense network. However, the functional overlap among oxidative stress defenses and other related cell functions makes difficult the characterization of this network. We previously described a network-based model to characterize the interactions existing among different antioxidant gene products and their substrates. Here, we test whether this network-based model of human antioxidant genes can respond to different physiological conditions. We used a systems biology approach applied to the analysis of two independent gene expression datasets: transcriptomes from HeLa cells and primary astrocytes maintained under hypoxic conditions and transcriptomes from SKGT4 cells exposed to low pH environment. We found that the proposed gene network model responds selectively to both hypoxia and acidosis. We anticipate that this antioxidant gene network model can be helpful to describe stress-responsive expression profiles in different cell types.

Can electrons travel through actin microfilaments and generate oxidative stress in retinol treated Sertoli cell?

In early reports our research group has demonstrated that 7 microM retinol (vitamin A) treatment leads to many changes in Sertoli cell metabolism, such as up-regulation of antioxidant enzyme activities, increase in damage to biomolecules, abnormal cellular division, pre-neoplasic transformation, and cytoskeleton conformational changes. These effects were observed to be dependent on the production of reactive oxygen species (ROS), suggesting extra-nuclear (non-genomic) effects of retinol metabolism. Besides 7 microM retinol treatment causing oxidative stress, we have demonstrated that changes observed in cytoskeleton of Sertoli cells under these conditions were protective, and seem to be an adaptive phenomenon against a pro-oxidant environment resulting from retinol treatment. We have hypothesized that the cytoskeleton can conduct electrons through actin microfilaments, which would be a natural process necessary for cell homeostasis. In the present study we demonstrate results correlating retinol metabolism, actin architecture, mitochondria physiology and ROS, in order to demonstrate that the electron conduction through actin microfilaments might explain our results. We believe that electrons produced by retinol metabolism are dislocated through actin microfilaments to mitochondria, and are transferred to electron transport chain to produce water. When mitochondria capacity to receive electrons is overloaded, superoxide radical production is increased and the oxidative stress process starts. Our results suggested that actin cytoskeleton is essential to oxidative stress production induced by retinol treatment, and electrons conduction through actin microfilaments can be the key of this correlation.