👤 Sónia Barbosa

A novel series of 6,7-dimethoxyquinazoline-hydrazide hybrids 1-4 were designed, synthesized, and tested for their AChE inhibition activity using the qualitative assay, which showed that compounds 3 and 4 exhibited their activity via TLC. The compounds were characterized using HRMS, IR, Show less

Dihydromyricetin (DMY) presents itself as a promising therapeutic candidate due to its inhibitory effects on various receptor tyrosine kinases, prompting an investigation of its structural characteristics, molecular interactions, and biological activity across the FGFR, HER, PDGFR, and VEGFR families. Protein sequences and structures for FGFR1/2, HER2/3, PDGFRA/B, and VEGFR1/2 were retrieved from UniProt/PDB. DMY and reference inhibitors were docked to each kinase using AutoDock Vina. Anti-angiogenic activity was measured by HET-CAM assay with vessel metrics quantified via IKOSA CAM. MTT determined cytotoxicity (IC₅₀) and tumor-selectivity index in 4T1 and L929 cells; data (mean ± SEM) were analyzed by one-way ANOVA with Tukey's test (p < 0.005). DMY exhibited docking scores comparable to established inhibitors, achieved over 45 % inhibition of neovascularization in the HET-CAM assay at nanomolar concentrations, displayed a tumor-selectivity index of less than one in 4T1 versus L929 cells (mirroring many clinical chemotherapeutics), and, notably, coadministration with doxorubicin reduced in vitro cardiotoxicity markers. The high-affinity, multi-kinase binding profile and significant anti-angiogenic efficacy underscore DMY's multifunctional potential, while its tumor-selectivity index aligns with accepted therapeutic risk-benefit balances and its cardioprotective effect suggests a way to mitigate anthracycline toxicity. These findings indicate that DMY is a multifunctional agent exhibiting both antiangiogenic and cytotoxic properties, warranting further preclinical and clinical investigation. Show less

Many membrane proteins, including G protein-coupled receptors (GPCRs), are susceptible to denaturation when extracted from their native membrane by detergents. Therefore, alternative methods have been developed, including amphiphilic copolymers that enable the direct extraction of functional membrane proteins along with their surrounding lipids. Among these amphiphilic copolymers, styrene/maleic acid (SMA) and diisobutylene/maleic acid (DIBMA) polymers have been extensively studied. Despite their many benefits, SMA and DIBMA polymers also have considerable drawbacks limiting their applications. Herein, we describe a series of new amphiphilic copolymers derived from DIBMA via partial amidation of the carboxylate pendant groups with various biocompatible amines. We characterize the new polymer's nanodisc-forming properties and ability to extract the melanocortin 4 receptor (MC Show less

Alzheimer's disease (AD) is a progressive neurodegenerative disorder affecting millions of people worldwide, with its prevalence expected to rise in the coming years. Due to the complexity of AD and the intricate interplay among its pathological mechanisms, the development of multitarget-directed ligands (MTDLs) has emerged as a promising therapeutic strategy. These compounds could simultaneously modulate multiple pathogenic pathways. Specifically, cholinergic and amyloid mechanisms, implicated in the onset of the disease, are regulated by AChE and BACE1, respectively. Therefore, targeting both pathways offers substantial therapeutic potential for AD. Computational tools can be useful in the identification of potential MTDL for these enzymes, reducing both costs and time in the drug discovery process. This review explores the relevance of this approach in the research and development for novel AD therapies, highlighting ongoing efforts focused on the identification and development of MTDLs for AChE and BACE1 inhibition through in silico methods. Virtual screening was the most frequently applied technique for a fast selection of ligands based on their affinity for the enzymes of interest. The in silico ADMET prediction also appears with a technique that allows the screening of compounds with drug-likeness. Moreover, evidence suggests that combining multiple computational methods can effectively identify drug candidates with optimized properties for target modulation and brain bioavailability. Show less

Alzheimer's Disease (AD) is a progressive neurodegenerative disorder and the leading cause of cognitive decline in older adults. Several biomarkers of AD have been identified, but its pathogenesis has not yet been completely elucidated. One of the most relevant hypotheses proposed to explain the cognitive impairment caused by this disease is the cholinergic hypothesis, which postulates that loss of cholinergic neurons is one of its causes and that the subsequent reduction of acetylcholine levels in the synaptic cleft can be compensated through the inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Another well-known hypothesis is the amyloid-beta hypothesis, which explains the disease as being caused by the formation and accumulation of amyloid plaques in a cascade of enzymatic events starting with the cleavage of an amyloid precursor protein (APP) by beta-secretase 1 (BACE-1). Previous studies have shown that silodosin has the structural requirements for the inhibition of those three enzymes (AChE, BuChE, and BACE-1), which suggests that it can be useful as a multitarget candidate to treat Alzheimer patients. This study aims to assess the effect of silodosin on cellular viability, measure the inhibitory activity against AChE, BuChE, and BACE-1, and evaluate the molecular behavior of all three inhibitor-enzyme systems by molecular dynamics (MD) simulations. Cell viability assays through the MTT method showed that silodosin concentrations of less than 10 μM are safe to be used. Enzymatic assays revealed AChE inhibitory activity at high micromolar levels (IC50 >500.0 μM) but inhibited BuChE at low micromolar levels (IC50 = 3.02 ± 0.05 μM). BACE-1 inhibition assays have shown significant reduction at three micromolar. MD simulations demonstrated that silodosin promotes late stabilization of the AChE complex, but the simulations involving BuChE and BACE-1 revealed that the compound promotes system stabilization at early stages and has the structural requirements to inhibition. Show less

Obicetrapib is a novel cholesteryl ester transfer protein (CETP) inhibitor with promising lipid-lowering effects. While earlier CETP inhibitors have shown inconsistent cardiovascular outcomes and safety concerns, the efficacy and safety of obicetrapib remain under active investigation. We systematically searched PubMed, Embase, and Cochrane Central databases for randomized controlled trials (RCTs) comparing obicetrapib versus placebo in adults with dyslipidemia or at high cardiovascular risk. We pooled mean differences (MDs) with 95 % confidence intervals (CI) with a random effects model. We used R software version 4.4.2 for statistical analysis. We included 7 RCTs comprising 3381 participants, of whom 2151 (63 %) received obicetrapib. The mean age was 64.3 years, and 36 % were women. Compared with placebo, obicetrapib significantly reduced mean LDL-C (MD: -37.21 %; 95 % CI: -41.53 to -32.90; Among patients with dyslipidemia and/or high cardiovascular risk, obicetrapib significantly reduces LDL-C, lipoprotein(a), apolipoprotein B, and non-HDL-C. No significant differences were observed in adverse events, supporting the favorable safety profile of obicetrapib. Show less

Base editors can correct disease-causing genetic variants. After a neonate had received a diagnosis of severe carbamoyl-phosphate synthetase 1 deficiency, a disease with an estimated 50% mortality in early infancy, we immediately began to develop a customized lipid nanoparticle-delivered base-editing therapy. After regulatory approval had been obtained for the therapy, the patient received two infusions at approximately 7 and 8 months of age. In the 7 weeks after the initial infusion, the patient was able to receive an increased amount of dietary protein and a reduced dose of a nitrogen-scavenger medication to half the starting dose, without unacceptable adverse events and despite viral illnesses. No serious adverse events occurred. Longer follow-up is warranted to assess safety and efficacy. (Funded by the National Institutes of Health and others.). Show less

Insufficient dietary fiber intake contributes to gut microbiota dysbiosis, systemic inflammation, and the onset of obesity-related metabolic disorders. Agro-industrial by-products have emerged as sustainable sources to restore microbial and metabolic balance. This study aimed to evaluate the effects of a mango bagasse- and peel-based confectionery (MC) on gut microbiota composition, short-chain fatty acids (SCFAs), and hepatic gene expression in Wistar rats fed either a standard diet (STD) or a high-fat diet (HFD). Twenty-four rats were randomly assigned to four groups (STD, MC-STD, HFD, MC-HFD) and treated for 11 weeks. Eating behavior, body composition, microbiota composition, SCFAs, and hepatic transcriptomics were evaluated. MC supplementation did not significantly alter weight gain or SCFA levels but shifted clustering patterns in principal component analysis, indicating a distinct dietary response. Microbiota analysis revealed a trend toward lower relative abundances of obesogenic species such as MC supplementation may beneficially modulate the gut-liver axis and highlights the nutritional potential of fruit by-products as functional ingredients to promote metabolic health under high-fat dietary conditions. Show less

Childhood obesity and associated comorbidities in adulthood are of great concern worldwide. Evidence highlights the importance of lactation in later disease development. In this sense, obese children are at great risk of developing adult obesity, insulin resistance, type 2 diabetes, and cardiovascular disease at adulthood. PPARα activation during lactation promotes the expression of key enzymes involved in lipid oxidation, and it was associated with reduced adiposity in children. Therefore, we hypothesized that an animal model of childhood obesity, small litter (SL), would lead to the development of obesity and metabolic dysfunction in adulthood, which could be prevented by postnatal PPARα agonism. Wistar dams had their litter reduced, leading to postnatal overfeeding and obesity early in life. SL male pups were treated with fenofibrate, an PPARα agonist, during lactation, from postnatal day (PND) 1 until weaning (PND21), to verify whether PPARα activation prevents the developmental programming at adulthood (PND120). Childhood obesity induced by postnatal overfeeding leads to decreased markers for oxidative metabolism during infancy, leading to increased visceral adiposity and oxidative stress, insulin resistance, hepatic microvesicular steatosis, and increased fibroblast growth factor 21 (Fgf21) expression, followed by decreased brown adipose tissue (BAT) sympathetic nerve activity and decreased Fgfr1 hypothalamic expression in adulthood. Agonist-induced PPARα activation during lactation mitigated the development of aforementioned alterations in adulthood. Postnatal fenofibrate treatment prevents the developmental programming of visceral obesity, liver-associated metabolic dysfunction and BAT autonomic sympathetic hypoactivity in an animal model of childhood obesity. Show less

Coronavirus disease 2019 (COVID-19) has been widely associated with intense systemic inflammation, endothelial injury, and a high incidence of thrombotic complications, which together contribute to disease severity and poor clinical outcomes. While endothelial dysfunction, dysregulated cytokine production, and oxidative stress are recognized features of severe COVID-19, the direct impact of circulating factors from infected individuals on endothelial cell behavior remains insufficiently characterized. Here, we examined how serum from patients with severe COVID-19 and from convalescent individuals modulates endothelial activation, inflammatory responses, and oxidative stress using human umbilical vein endothelial cells as an in vitro model. Venous blood samples were collected from individuals with severe COVID-19 (n = 13), convalescent patients (n = 11), and healthy volunteers (n = 7) during the initial phase of the COVID-19 pandemic. Human umbilical vein endothelial cells (HUVEC) were maintained in culture and exposed to 15% serum from each study group after a period of serum deprivation. The expression of genes associated with endothelial activation, thrombosis, inflammation, and oxidative stress was analyzed by quantitative real-time PCR at defined time points. In addition, the endothelial secretory profile was evaluated in cell culture supernatants using multiplex bead-based immunoassays. Statistical analyses were performed using one-way ANOVA followed by appropriate post hoc tests, receiver operating characteristic (ROC) curve analysis to assess the discriminatory capacity of biomarkers, and multivariate linear regression to identify factors associated with disease severity. We investigated the role of the endothelium in modulating the cytokine storm in severe COVID-19. HUVEC were stimulated with serum from patients with severe COVID-19, convalescent individuals, and healthy volunteers. Stimulation with serum from severe cases induces significant increases in These findings suggest that HUVEC serves as a promising biological sensor for detecting inflammatory responses in COVID-19 patients and shows the crucial role of the endothelium in sustaining the cytokine storm that contributes to patient severity and mortality. Show less

Autophagy has been implicated in various cellular processes, including non-conventional secretion. Our previous findings suggest that ATP is loaded into amphisomes and secreted upon autophagy stimulation at focal adhesion sites in a VAMP7-dependent manner. Here, we demonstrate that the knockout (KO) of VAMP7, along with its partners RAB21 and its guanine nucleotide exchange factor (GEF) VARP, inhibits ATP release, indicating a key role for this pathway in amphisome secretion. Constitutively inactive RAB21 also inhibited ATP secretion. RAB21 overexpression rescued starvation-induced ATP secretion in RAB21 KO, but not in VAMP7 or VARP KO cells. RAB21-LC3-positive vesicles redistributed to the cell periphery upon starvation. KO cells and overexpression experiments showed that RAB21 plays a positive role in autophagosome biogenesis, particularly in controlling the number of LC3-II- and DFCP1-positive structures upon starvation, suggesting a role in the early steps of autophagosome formation. Accordingly, VARP partially colocalized with LC3 upon starvation. Together, these findings identify a novel role for RAB21 in regulating autophagic ATP secretion likely in amphisome biogenesis and their localization in the cell periphery. Show less

A malfunction in the melanocortin-4 receptor (MC4R) is associated with obesity in rare genetic syndromes; setmelanotide is a new drug that activates this receptor and is being used to treat severe obesity. This meta-analysis evaluated the efficacy and safety of setmelanotide for weight loss in severe obesity linked to human MC4R deficiency. We searched PubMed, Embase, and Cochrane for randomized and nonrandomized clinical trials using setmelanotide. We considered a We included 376 patients, of whom 328 (87.2%) received setmelanotide for a mean follow-up of 52 weeks. The mean age was 32.8 (14.67) years. Weight loss was significant (MD -3.52; 95% CI -3.98, -3.05; Our results support the use of setmelanotide in treating severe obesity. Show less

Alzheimer's disease causes chronic neurodegeneration and is the leading cause of dementia in the world. The causes of this disease are not fully understood but seem to involve two essential cerebral pathways: cholinergic and amyloid. The simultaneous inhibition of AChE, BuChE, and BACE-1, essential enzymes involved in those pathways, is a promising therapeutic approach to treat the symptoms and, hopefully, also halt the disease progression. This study sought to identify triple enzymatic inhibitors based on stereo-electronic requirements deduced from molecular modeling of AChE, BuChE, and BACE-1 active sites. A pharmacophore model was built, displaying four hydrophobic centers, three hydrogen bond acceptors, and one positively charged nitrogen, and used to prioritize molecules found in virtual libraries. Compounds showing adequate overlapping rates with the pharmacophore were subjected to molecular docking against the three enzymes and those with an adequate docking score ( Show less

Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive loss of cognitive functions, and it is the most prevalent type of dementia worldwide, accounting for 60 to 70% of cases. The pathogenesis of AD seems to involve three main factors: deficiency in cholinergic transmission, formation of extracellular deposits of β-amyloid peptide, and accumulation of deposits of a phosphorylated form of the TAU protein. The currently available drugs are prescribed for symptomatic treatment and present adverse effects such as hepatotoxicity, hypertension, and weight loss. There is urgency in finding new drugs capable of preventing the progress of the disease, controlling the symptoms, and increasing the survival of patients with AD. This study aims to present new multipurpose compounds capable of simultaneously inhibiting acetylcholinesterase (AChE), butyrylcholinesterase (BChE)-responsible for recycling acetylcholine in the synaptic cleft-and beta-secretase 1 (BACE-1)-responsible for the generation of amyloid-β plaques. AChE, BChE, and BACE-1 are currently considered the best targets for the treatment of patients with AD. Virtual hierarchical screening based on a pharmacophoric model for BACE-1 inhibitors and a dual pharmacophoric model for AChE and BChE inhibitors were used to filter 214,446 molecules by QFIT Show less

To analyze the expression of glucose-dependent insulinotropic polypeptide receptor (GIPR) in somatotropinomas specimens and compare clinical, biochemical, radiological, therapeutic, molecular, and pathological data among those who overexpressed (GIPR +) and those who did not overexpress (GIPR - ) GIPR. Clinical, biochemical, radiological, molecular, and pathological data were collected. GNAS1 sequencing was performed with the Sanger method. Protein expression of somatostatin receptor subtypes 2 and 5 and CAM 5.2 were analyzed by immunohistochemistry. Quantitative real-time PCR was performed to analyze the mRNA expression of GIPR with the TaqMan® method. Positive expression was considered when the fold change (FC) was above 17.2 (GIPR +). A total of 74 patients (54% female) were included. Eighteen tumors (24%) were GIPR + . Gsp mutation was detected in 30 tumors (40%). GIPR + tumors were more frequently densely granulated adenomas (83% vs 47%, p = 0.028). There was no difference in clinical, biochemical, radiological, therapeutic (surgical cure or response to medical therapy), or other pathological features between GIPR + and GIPR -  tumors. Twenty-eight out of 56 (50%) GIPR -  tumors harbored a gsp mutation, whereas two out of 18 (11%) GIPR + tumors harbored a gsp mutation (p = 0.005). We described, for the first time, that GIPR + and gsp mutations are not mutually exclusive, but gsp mutations are less common in GIPR + tumors. GIPR + and GIPR -  tumors have similar clinical, biochemical, radiological, therapeutic, and pathological features, with the exception of a high frequency of densely granulated adenomas among GIPR + tumors. Show less

We determined the relationships between DNA sequence variation and DNA methylation using blood samples from 3,799 Europeans and 3,195 South Asians. We identify 11,165,559 SNP-CpG associations (methylation quantitative trait loci (meQTL), P < 10 Show less

Arterial hypertension (AH) is implicated in vascular health and contributes significantly to cardiovascular morbidity and mortality. In addition to the contribution of usual risk factors for AH, elucidating the influence of genetic factors is a promising area of investigation. Therefore, we evaluated the association between AH and cardiovascular risk factors (CVRFs) and genetic polymorphisms in communities in Southeast Brazil. A total of 515 adults aged 18-91 years, who were cross-sectionally assessed between 2015-2016, were included. Demographic, clinical, behavioral, anthropometric characteristics, and laboratory parameters and 12 single nucleotide polymorphisms in seven candidate genes involved in cardiovascular risk ( There was a significant association between age >60 years (odds ratio [OR] =6.74), alcohol dependence (OR=3.84), smoking (OR=1.74), overweight (OR=1.74), high plasma triglyceride (TG) levels (OR=1.98) and low high-density lipoprotein (HDL-c) (OR=6.22), diabetes (OR=3.68), and insulin resistance (OR=2.40) and AH. A significant association was observed between rs4721 in The interaction of the T allele of the rs4721 polymorphism in Show less

Bipolar disorder (BD) diagnosis currently relies on assessment of clinical symptoms, mainly retrospective and subject to memory bias. BD is often misdiagnosed as Major Depressive Disorder (MDD) resulting in ineffective treatment and worsened clinical outcome. The primary purpose of this study was to identify blood biomarkers that discriminate MDD from BD patients when in a depressed state. We have used clinical data and serum samples from two independent naturalistic cohorts of patients with a Major Depressive Episode (MDE) who fulfilled the criteria of either BD or MDD at inclusion. The discovery and replication cohorts consisted of 462 and 133 patients respectively. Patients were clinically assessed using standard diagnostic interviews, and clinical variables including current treatments were recorded. Blood was collected and serum assessed for levels of 31 cytokines using a sensitive multiplex assay. A penalized logistic regression model combined with nonparametric bootstrap was subsequently used to identify cytokines associated with BD. Interleukin (IL)-6, IL-10, IL-15, IL-27 and C-X-C ligand chemokine (CXCL)-10 were positively associated with BD in the discovery cohort. Of the five cytokines identified as discriminant features in the discovery cohort, IL-10, IL-15 and IL-27 were also positively associated with BD in the replication cohort therefore providing an external validation to our finding. Should our results be validated in a prospective cohort, they could provide new insights into the pathophysiological mechanisms of mood disorders. Show less

Leukemias bearing fusions of the AF10/MLLT10 gene are associated with poor prognosis, and therapies targeting these fusion proteins (FPs) are lacking. To understand mechanisms underlying AF10 fusion-mediated leukemogenesis, we generated inducible mouse models of acute myeloid leukemia (AML) driven by the most common AF10 FPs, PICALM/CALM-AF10 and KMT2A/MLL-AF10, and performed comprehensive characterization of the disease using transcriptomic, epigenomic, proteomic, and functional genomic approaches. Our studies provide a detailed map of gene networks and protein interactors associated with key AF10 fusions involved in leukemia. Specifically, we report that AF10 fusions activate a cascade of JAK/STAT-mediated inflammatory signaling through direct recruitment of JAK1 kinase. Inhibition of the JAK/STAT signaling by genetic Jak1 deletion or through pharmacological JAK/STAT inhibition elicited potent antioncogenic effects in mouse and human models of AF10 fusion AML. Collectively, our study identifies JAK1 as a tractable therapeutic target in AF10-rearranged leukemias. Show less

The authors report the clinical and genetic investigation of a family with hypertrophic cardiomyopathy (HCM). The individuals described are three affected first-degree relatives (father, daughter and son), one affected niece and unaffected nephew and niece. Those affected all share a very similar phenotype consisting of asymmetric HCM, with hypertrophy particularly affecting the septum and the anterior wall, and similar electrocardiographic features, including a short PR interval. Case 1 (proband) presented with obstructive HCM and had undergone myectomy and mitral valve replacement. Case 2 (oldest offspring of Case 1) had non-obstructive HCM with exertional angina and NYHA II heart failure (HF) symptoms; she developed non-sustained ventricular tachycardia during follow-up and received a single-chamber ICD for primary prevention of sudden cardiac death. Case 3 (son of case 1) presented with asymptomatic non-obstructive HCM and developed NYHA II HF symptoms during follow-up. Case 4 had non-obstructive HCM, mainly with NYHA II HF symptoms. Testing of the proband for sarcomeric mutations and phenocopies was initially negative. After eight years of clinical follow-up, the suspicion of an undiscovered pathogenic gene mutation shared among the members of this family led us to enroll the proband in a whole-genome sequencing research project, which revealed a heterozygous pathogenic intronic MYBPC3 variant (c.1227-13G>A [rs397515893]), cosegregating with the phenotype. Show less

Patients with coronary artery disease (CAD) and previous ischemic cerebrovascular events (ICVE, ischemic stroke, or transitory ischemic attack) constitute a high-risk subgroup for cardiovascular outcomes. High-density lipoprotein cholesterol (HDL-C) levels are correlated with cardiovascular events. Lipid transfer to HDL affects structure size and HDL subclass profile. Impairment of this transfer could influence ischemic risk seen in patients with CAD + ICVE. The objective was to evaluate the HDL ability to receive the lipids in patients with CAD with or without ICVE. Patients with CAD + ICVE (n = 60) and patients with CAD only (n = 60) were matched by age, sex, acute coronary syndromes (ACS) event type, and time elapsed between the ACS event and inclusion in the study. Lipid transfer to HDL was evaluated by incubating donor lipid nanoparticles labeled with radioactive unesterified cholesterol (UC) and esterified cholesterol (EC), phospholipid (PL), and triglyceride (TG) with whole plasma. After the chemical precipitation of non-HDL fractions and nanoparticles, the supernatant was counted for HDL radioactivity. CAD + ICVE group presented with impaired lipid transfer to HDL for PL (CAD + ICVE: 21.14 ± 2.7% vs CAD: 21.67 ± 3.1%, P = .03), TG (CAD + ICVE: 4.88 ± 0.97% vs CAD: 5.63 ± 0.92%, P = .002), and UC (CAD + ICVE: 5.55 ± 1.19% vs CAD: 6.16 ± 1.14%, P = .009). Lipid transfer to HDL was similar in both groups for EC. Adjusted models showed similar results. Patients with CAD and ICVE have reduced lipid transfer to HDL compared to those with CAD only. Dysfunctional HDL may account for the higher incidence of ischemic outcomes observed in this population. Show less

CREB-H, an ER-anchored transcription factor, plays a key role in regulating secretion in metabolic pathways, particularly triglyceride homeostasis. It controls the production both of secretory pathway components and cargoes, including apolipoproteins ApoA-IV and ApoC-II, contributing to VLDL/HDL distribution and lipolysis. The key mechanism controlling CREB-H activity involves its ER retention and forward transport to the Golgi, where it is cleaved by Golgi-resident proteases, releasing the N-terminal product, which traffics to the nucleus to effect transcriptional responses. Here we show that a serine-rich motif termed the P-motif, located in the N-terminus between serines 73 and 90, controls release of the precursor transmembrane form from the ER and its forward transport to the Golgi. This motif is subject to GSK-3 phosphorylation, promoting ER retention, while mutation of target serines and drug inhibition of GSK-3 activity coordinately induce both forward transport of the precursor and cleavage, resulting in nuclear import. We previously showed that for the nuclear product, the P-motif is subject to multiple phosphorylations, which regulate stability by targeting the protein to the SCF Show less

Acetone precipitation was evaluated as a rapid, simple, low-cost, and efficient method for the selective purification of O-glycopeptides from enzymatic digests of glycoproteins. Ovalbumin (OVA), human and bovine α Show less

CREB‑H, an endoplasmic reticulum-anchored transcription factor, plays a key role in regulating secretion and in metabolic and inflammatory pathways, but how its activity is modulated remains unclear. We examined processing of the nuclear active form and identified a motif around S87-S90 with homology to DSG-type phosphodegrons. We show that this region is subject to multiple phosphorylations, which regulate CREB-H stability by targeting it to the SCF(Fbw1a) E3 ubiquitin ligase. Data from phosphatase treatment, use of phosophospecific antibody, and substitution of serine residues demonstrate phosphorylation of candidate serines in the region, with the core S87/S90 motif representing a critical determinant promoting proteasome-mediated degradation. Candidate kinases CKII and GSK-3b phosphorylate CREB-H in vitro with specificities for different serines. Prior phosphorylation with GSK-3 at one or more of the adjacent serines substantially increases S87/S90-dependent phosphorylation by CKII. In vivo expression of a dominant-negative Cul1 enhances steady-state levels of CREB‑H, an effect augmented by Fbw1a. CREB-H directly interacts with Fbw1a in a phosphorylation-dependent manner. Finally, mutations within the phosphodegron, when incorporated into the full-length protein, result in increased levels of constitutively cleaved nuclear protein and increased transcription and secretion of a key endogenous target gene, apolipoprotein A IV. Show less

Glucose-dependent insulinotropic peptide receptor (GIPR) and LHCGR are G-protein-coupled receptors with a wide tissue expression pattern. Aberrant expression of these receptors has rarely been demonstrated in adult sporadic adrenocortical tumors with a lack of data on pediatric tumors. We quantified the GIPR and LHCGR expression in a large cohort of 55 patients (25 children and 30 adults) with functioning and non-functioning sporadic adrenocortical tumors. Thirty-eight tumors were classified as adenomas whereas 17 were carcinomas. GIPR and LHCGR expression were analyzed by real-time PCR and normal human pancreatic and testicular tissue samples were used as positive controls. Mean expression values were determined by fold increase in comparison with a normal adrenal pool. GIPR mRNA levels were significantly higher in adrenocortical carcinomas than in adenomas from both pediatric and adult groups. LHCGR expression was similar in both carcinomas and adenomas from the pediatric group but significantly lower in carcinomas than in adenomas from the adult group (median 0.06 and 2.3 respectively, P<0.001). GIPR was detected by immunohistochemistry in both pediatric and adult tumors. Staining and real-time PCR results correlated positively only when GIPR mRNA levels were increased at least two-fold in comparison with normal adrenal expression levels. In conclusion, GIPR overexpression was observed in pediatric and adult adrenocortical tumors and very low levels of LHCGR expression were found in all adult adrenocortical carcinomas. Show less

Alzheimer's disease (AD) is the most common neurodegenerative disorder in the elderly and is also considered a progeroid genetic syndrome. The etiology of AD is complex and the mechanisms underlying its pathophysiology remain to be clarified. It has been suggested that a high serum cholesterol level is a risk factor for (AD), and that some polymorphisms of genes encoding proteins regulating cholesterol metabolism are associated with AD development. APOA5 is a recently discovered apolipoprotein involved primarily with triglyceride (TG) metabolism disorder. This study investigates the association of AD with the APOA5 gene -1131T>C polymorphisms in samples of 106 patients with Alzheimer's disease (AD), 76 elderly healthy controls and 93 young healthy controls. DNA samples were isolated from blood cells, amplified by PCR and digested with Tru1l. We observed that the genotype distributions of APOA5 variants were within Hardy-Weinberg equilibrium in all subject samples. Furthermore, chi-square test comparison for genotype distributions and allele frequencies did not reveal any significant difference among the three groups of subjects P>0.05). These results support the idea that these variants are not involved as a risk factor for developing AD. Show less