Role of hydroxymethylglutharyl-coenzyme A reductase in the induction of stem-like states in breast cancer.

PURPOSE: De novo synthesis of cholesterol and its rate-limiting enzyme, 3-hydroxy-3-methylglutharyl-coenzyme A reductase (HMGCR), is deregulated in tumors and critical for tumor cell survival and proliferation. However, the role of HMGCR in the induction and maintenance of stem-like states in tumors remains unclear. METHODS: A compiled public database from breast cancer (BC) patients was analyzed with the web application SurvExpress. Cell Miner was used for the analysis of HMGCR expression and statin sensitivity of the NCI-60 cell lines panel. A CRISPRon system was used to induce HMGCR overexpression in the luminal BC cell line MCF-7 and a lentiviral pLM-OSKM system for the reprogramming of MCF-7 cells. Comparisons were performed by two-tailed unpaired t-test for two groups and one- or two-way ANOVA. RESULTS: Data from BC patients showed that high expression of several members of the cholesterol synthesis pathway were associated with lower recurrence-free survival, particularly in hormone-receptor-positive BC. In silico and in vitro analysis showed that HMGCR is expressed in several BC cancer cell lines, which exhibit a subtype-dependent response to statins in silico and in vitro. A stem-like phenotype was demonstrated upon HMGCR expression in MCF-7 cells, characterized by expression of the pluripotency markers NANOG, SOX2, increased CD44 +/CD24low/ -, CD133 + populations, and increased mammosphere formation ability. Pluripotent and cancer stem cell lines showed high expression of HMGCR, whereas cell reprogramming of MCF-7 cells did not increase HMGCR expression. CONCLUSION: HMGCR induces a stem-like phenotype in BC cells of epithelial nature, thus affecting tumor initiation, progression and statin sensitivity.

Antifungal Activity of Fibrate-Based Compounds and Substituted Pyrroles That Inhibit the Enzyme 3-Hydroxy-methyl-glutaryl-CoA Reductase of Candida glabrata (CgHMGR), Thus Decreasing Yeast Viability and Ergosterol Synthesis.

Due to the emergence of multidrug-resistant strains of yeasts belonging to the Candida genus, there is an urgent need to discover antifungal agents directed at alternative molecular targets. The aim of the current study was to evaluate the capacity of three different series of synthetic compounds to inhibit the Candida glabrata enzyme denominated 3-hydroxy-methyl-glutaryl-CoA reductase and thus affect ergosterol synthesis and yeast viability. Compounds 1c (α-asarone-related) and 5b (with a pyrrolic core) were selected as the best antifungal candidates among over 20 synthetic compounds studied. Both inhibited the growth of fluconazole-resistant and fluconazole-susceptible C. glabrata strains. A yeast growth rescue experiment based on the addition of exogenous ergosterol showed that the compounds act by inhibiting the mevalonate synthesis pathway. A greater recovery of yeast growth occurred for the C. glabrata 43 fluconazole-resistant (versus fluconazole-susceptible) strain and after treatment with 1c (versus 5b). Given that the compounds decreased the concentration of ergosterol in the yeast strains, they probably target ergosterol synthesis. According to the docking analysis, the inhibitory effect of 1c and 5b could possibly be mediated by their interaction with the amino acid residues of the catalytic site of the enzyme. Since 1c displayed higher binding energy than α-asarone and 5b, it is the best candidate for further research, which should include structural modifications to increase its specificity and potency. The derivatives could then be examined with in vivo animal models using a therapeutic dose. IMPORTANCE Within the context of the COVID-19 pandemic, there is currently an epidemiological alert in health care services due to outbreaks of Candida auris, Candida glabrata, and other fungal species multiresistant to conventional antifungals. Therefore, it is important to propose alternative molecular targets, as well as new antifungals. The three series of synthetic compounds herein designed and synthesized are inhibitors of ergosterol synthesis in yeasts. Of the more than 20 compounds studied, two were selected as the best antifungal candidates. These compounds were able to inhibit the growth and synthesis of ergosterol in C. glabrata strains, whether susceptible or resistant to fluconazole. The rational design of antifungal compounds derived from clinical drugs (statins, fibrates, etc.) has many advantages. Future studies are needed to modify the structure of the two present test compounds to obtain safer and less toxic antifungals. Moreover, it is important to carry out a more in-depth mechanistic approach.

Mitochondrial dysfunction in anti-3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) immune-mediated necrotising myopathy.

Mitochondrial dysfunction is a plausible cause of muscle fibre damage in a number of myopathies including immune-mediated necrotising myopathy. However, histopathological evidence of mitochondrial dysfunction is not often described in immune-mediated necrotising myopathy and, when present, it is often attributed to patient age. The purpose of this study was to describe features of mitochondrial dysfunction on muscle biopsy in anti-3­hydroxy-3-methylglutaryl-CoA reductase immune-mediated necrotising myopathy and explore whether these features are age-related. In this observational case control study, a statistically significant increase in the number of muscle fibres with increased lipid content (p = 0.004) and cytochrome c oxidase-negative/succinate dehydrogenase-positive fibres (p = 0.037) in anti-3­hydroxy-3-methylglutaryl-coenzyme immune-mediated necrotising myopathy was found compared to age-matched controls. Therefore, histopathological features of mitochondrial dysfunction are more frequent in anti-3­hydroxy-3-methylglutaryl-coenzyme immune-mediated necrotising myopathy than aged-matched controls and therefore, may be contributing to the pathogenesis.

IAF, QGF, and QDF Peptides Exhibit Cholesterol-Lowering Activity through a Statin-like HMG-CoA Reductase Regulation Mechanism: In Silico and In Vitro Approach.

In this study, in silico approaches are employed to investigate the binding mechanism of peptides derived from cowpea ß-vignin and HMG-CoA reductase. With the obtained information, we designed synthetic peptides to evaluate their in vitro enzyme inhibitory activity. In vitro, the total protein extract and <3 kDa fraction, at 5000 µg, support this hypothesis (95% and 90% inhibition of HMG-CoA reductase, respectively). Ile-Ala-Phe, Gln-Gly-Phe, and Gln-Asp-Phe peptides were predicted to bind to the substrate binding site of HMGCR via HMG-CoAR. In silico, it was established that the mechanism of HMG-CoA reductase inhibition largely entailed mimicking the interactions of the decalin ring of simvastatin and via H-bonding; in vitro studies corroborated the predictions, whereby the HMG-CoA reductase activity was decreased by 69%, 77%, and 78%, respectively. Our results suggest that Ile-Ala-Phe, Gln-Gly-Phe, and Gln-Asp-Phe peptides derived from cowpea ß-vignin have the potential to lower cholesterol synthesis through a statin-like regulation mechanism.

Reducción de eventos cardiovasculares de Lomitapida versus Estatinas en pacientes con diagnóstico de hipercolesterolemia fa-miliar. Protocolo de revisión sistemática / Reduction of cardiovascular events of Lomitapida versus Statins in patients with a diagnosis of familial hypercholesterolemia: A systematic review protocol

Introducción: La hipercolesterolemia familiar (HF) un trastorno genético autosómico domi-nante que produce un desarrollo prematuro de enfermedades cardiovasculares. Las estati-nas han sido el medicamento de elección en estos pacientes, sin embargo, un buen por-centaje de pacientes no pueden alcanzar sus objetivos terapéuticas con las dosis máximas por lo que la Lomitapida se podría establecer como una nueva alternativa de tratamiento. Objetivo: El objetivo de esta revisión sistemática es determinar si la Lomitapida reduce los eventos cardiovasculares en pacientes con diagnóstico de Hipercolesterolemia familiar comparado con estatinas. Métodos: Se incluirán ensayos controlados aleatorios (ECA) y cuasialeatorios de pacientes con diagnóstico de HF. Las medidas de resultado los niveles de LDL, HDL pos tratamiento y eventos cardiovasculares. Las búsquedas electrónicas se realizarán en PUBMED, The Coch-rane Central Register of Controlled Trials (CENTRAL), EMBASE y Scientific electronic library (Scielo). En la evaluación del riesgo de sesgo se utilizará la herramienta de Cochrane. Las medidas del efecto del tratamiento serán las diferencias de medias (DM) y los intervalos de confianza (IC) del 95%. La evaluación de heterogeneidad se realizará mediante la inspec-ción visual del diagrama de embudo. La evaluación de la calidad de la evidencia se reali-zará usando la evaluación GRADE.

Introduction: Familial hypercholesterolemia (FH) is an autosomal dominant genetic disor-der that produces hypercholesterolemia and premature development of cardiovascular diseas-es. Statins are the drug of choice in these patients; however, a high percentage of patients cannot achieve their therapeutic goals with the maximum recommended doses, so Lo-mitapide may prove to be useful as a new treatment alternative to traditional statins. Objective: The objective of this systematic review is to determine if Lomitapide is better than statins at reducing cardiovascular events in patients with a diagnosis of FH. Methods: Randomized controlled trials (RCTs) and quasi-randomized trials of patients di-agnosed with FH will be included. Primary outcome measures included several parameters: 1. Post-treatment low- and high-density lipoprotein (LDL and HDL, respectively) levels and 2. Presence of cardiovascular events. Electronic searches will be conducted in PUBMED, The Cochrane Central Register of Controlled Trials (CENTRAL), EMBASE, and the scientific elec-tronic library (Scielo). The assessment of the risk of bias will be used by the Cochrane tool. The measures of the treatment effect will be considered the mean differences (MD) and the 95% confidence intervals (CI). The evaluation of heterogeneity will be done by visual inspec-tion of the funnel diagram. The evaluation of the quality of the evidence will be done using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) ap-proach.

Reducción de eventos cardiovasculares de Lomitapida versus Estatinas en pacientes con diagnóstico de hipercolesterolemia fa-miliar. Protocolo de revisión sistemática / Reduction of cardiovascular events of Lomitapida versus Statins in patients with a diagnosis of familial hypercholesterolemia: A systematic review protocol

Introducción: La hipercolesterolemia familiar (HF) un trastorno genético autosómico domi-nante que produce un desarrollo prematuro de enfermedades cardiovasculares. Las estati-nas han sido el medicamento de elección en estos pacientes, sin embargo, un buen por-centaje de pacientes no pueden alcanzar sus objetivos terapéuticas con las dosis máximas por lo que la Lomitapida se podría establecer como una nueva alternativa de tratamiento. Objetivo: El objetivo de esta revisión sistemática es determinar si la Lomitapida reduce los eventos cardiovasculares en pacientes con diagnóstico de Hipercolesterolemia familiar comparado con estatinas. Métodos: Se incluirán ensayos controlados aleatorios (ECA) y cuasialeatorios de pacientes con diagnóstico de HF. Las medidas de resultado los niveles de LDL, HDL pos tratamiento y eventos cardiovasculares. Las búsquedas electrónicas se realizarán en PUBMED, The Coch-rane Central Register of Controlled Trials (CENTRAL), EMBASE y Scientific electronic library (Scielo). En la evaluación del riesgo de sesgo se utilizará la herramienta de Cochrane. Las medidas del efecto del tratamiento serán las diferencias de medias (DM) y los intervalos de confianza (IC) del 95%. La evaluación de heterogeneidad se realizará mediante la inspec-ción visual del diagrama de embudo. La evaluación de la calidad de la evidencia se reali-zará usando la evaluación GRADE.

Introduction: Familial hypercholesterolemia (FH) is an autosomal dominant genetic disor-der that produces hypercholesterolemia and premature development of cardiovascular diseas-es. Statins are the drug of choice in these patients; however, a high percentage of patients cannot achieve their therapeutic goals with the maximum recommended doses, so Lo-mitapide may prove to be useful as a new treatment alternative to traditional statins. Objective: The objective of this systematic review is to determine if Lomitapide is better than statins at reducing cardiovascular events in patients with a diagnosis of FH. Methods: Randomized controlled trials (RCTs) and quasi-randomized trials of patients di-agnosed with FH will be included. Primary outcome measures included several parameters: 1. Post-treatment low- and high-density lipoprotein (LDL and HDL, respectively) levels and 2. Presence of cardiovascular events. Electronic searches will be conducted in PUBMED, The Cochrane Central Register of Controlled Trials (CENTRAL), EMBASE, and the scientific elec-tronic library (Scielo). The assessment of the risk of bias will be used by the Cochrane tool. The measures of the treatment effect will be considered the mean differences (MD) and the 95% confidence intervals (CI). The evaluation of heterogeneity will be done by visual inspec-tion of the funnel diagram. The evaluation of the quality of the evidence will be done using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) ap-proach.

Reducción de eventos cardiovasculares de Lomitapida versus Estatinas en pacientes con diagnóstico de hipercolesterolemia fa-miliar. Protocolo de revisión sistemática / Reduction of cardiovascular events of Lomitapida versus Statins in patients with a diagnosis of familial hypercholesterolemia: A systematic review protocol

Introducción: La hipercolesterolemia familiar (HF) un trastorno genético autosómico domi-nante que produce un desarrollo prematuro de enfermedades cardiovasculares. Las estati-nas han sido el medicamento de elección en estos pacientes, sin embargo, un buen por-centaje de pacientes no pueden alcanzar sus objetivos terapéuticas con las dosis máximas por lo que la Lomitapida se podría establecer como una nueva alternativa de tratamiento. Objetivo: El objetivo de esta revisión sistemática es determinar si la Lomitapida reduce los eventos cardiovasculares en pacientes con diagnóstico de Hipercolesterolemia familiar comparado con estatinas. Métodos: Se incluirán ensayos controlados aleatorios (ECA) y cuasialeatorios de pacientes con diagnóstico de HF. Las medidas de resultado los niveles de LDL, HDL pos tratamiento y eventos cardiovasculares. Las búsquedas electrónicas se realizarán en PUBMED, The Coch-rane Central Register of Controlled Trials (CENTRAL), EMBASE y Scientific electronic library (Scielo). En la evaluación del riesgo de sesgo se utilizará la herramienta de Cochrane. Las medidas del efecto del tratamiento serán las diferencias de medias (DM) y los intervalos de confianza (IC) del 95%. La evaluación de heterogeneidad se realizará mediante la inspec-ción visual del diagrama de embudo. La evaluación de la calidad de la evidencia se reali-zará usando la evaluación GRADE.

Introduction: Familial hypercholesterolemia (FH) is an autosomal dominant genetic disor-der that produces hypercholesterolemia and premature development of cardiovascular diseas-es. Statins are the drug of choice in these patients; however, a high percentage of patients cannot achieve their therapeutic goals with the maximum recommended doses, so Lo-mitapide may prove to be useful as a new treatment alternative to traditional statins. Objective: The objective of this systematic review is to determine if Lomitapide is better than statins at reducing cardiovascular events in patients with a diagnosis of FH. Methods: Randomized controlled trials (RCTs) and quasi-randomized trials of patients di-agnosed with FH will be included. Primary outcome measures included several parameters: 1. Post-treatment low- and high-density lipoprotein (LDL and HDL, respectively) levels and 2. Presence of cardiovascular events. Electronic searches will be conducted in PUBMED, The Cochrane Central Register of Controlled Trials (CENTRAL), EMBASE, and the scientific elec-tronic library (Scielo). The assessment of the risk of bias will be used by the Cochrane tool. The measures of the treatment effect will be considered the mean differences (MD) and the 95% confidence intervals (CI). The evaluation of heterogeneity will be done by visual inspec-tion of the funnel diagram. The evaluation of the quality of the evidence will be done using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) ap-proach.

Further evidence for the involvement of mitochondrial aquaporin-8 in hepatocyte lipid synthesis.

We recently provided evidence suggesting that mitochondrial aquaporin-8 (mtAQP8), a channel protein able to conduct H2O2, is involved in the modulation of hepatocyte cholesterogenesis. To expand that study, we cultured human hepatocyte-derived Huh-7 cells in medium with lipoprotein-deficient serum (LPDS) to induce the de novo synthesis of cholesterol and fatty acids. We found that LPDS induced mtAQP8 expression and that AQP8 gene silencing significantly down-regulated the LPDS-induced synthesis of cholesterol and fatty acids as well as the expression of the corresponding key biosynthetic enzymes, 3-hydroxy-3-methylglutaryl-CoA reductase and fatty acid synthase. Our data further support a regulatory role of mtAQP8 in hepatocyte lipid homeostasis.

Immune-mediated necrotising myopathy in asymptomatic patients with high creatine kinase.

Subacute symmetrical proximal muscle weakness and persistent elevated creatine kinase levels are typical of immune-mediated necrotising myopathy (IMNM). These conditions are accompanied by copious myofibre necrosis, degeneration and regeneration with minimal to no inflammation on muscle biopsy. We report two cases (case 1 and case 2) of asymptomatic IMNM from different families with hyperCKaemia associated with positive anti-signal recognition particle (SRP) and anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) antibodies, respectively, and we also reviewed the literature. There are only a few previous descriptions of patients with asymptomatic IMNM.The disease onset could be insidious and lead to delayed diagnosis and treatment. We recommend testing for the anti-HMGCR and anti-SRP antibodies in patients with idiopathic hyperCKaemia because they could show no symptoms of this disorder.

Spontaneous symptomatic improvement in a pediatric patient with anti-3-hydroxy-3-methylglutraryl-coenzyme A reductase myopathy.

Immune-mediated necrotizing myopathy with antibodies against 3-hydroxy-3-methylglutaryl-coenzyme A reductase is a subgroup of idiopathic inflammatory myopathies mainly described in adults and requiring long term immunomodulatory therapy for remission. Pediatric patients have been reported as small series or sporadic cases. We report an eight-year-old girl with anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase myopathy, presenting with subacute proximal limb weakness, high creatine kinase and a muscle biopsy displaying necrotizing pattern, initially diagnosed as limb-girdle muscular dystrophy, but subsequently negative genetic testing. A noteworthy spontaneous improvement in her weakness suggested the possibility of an acquired autoimmune myopathy, confirmed by positive testing of anti-HMGCR antibodies titers. After four years of follow-up, she maintains normal strength with high levels of anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase antibody. This patient shows that spontaneous fluctuations and spontaneous long-lasting symptomatic remission can occur in patients with anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase myopathy. Some patients could present a wane and wax clinical course, an important aspect when assessing response to therapy.

The Differential Expression of Mevalonate Pathway Genes in the Gut of the Bark Beetle Dendroctonus rhizophagus (Curculionidae: Scolytinae) Is Unrelated to the de Novo Synthesis of Terpenoid Pheromones.

Bark beetles commonly produce de novo terpenoid pheromones using precursors synthesized through the mevalonate pathway. This process is regulated by Juvenile Hormone III (JH III). In this work, the expression levels of mevalonate pathway genes were quantified after phloem feeding-to induce the endogenous synthesis of JH III-and after the topical application of a JH III solution. The mevalonate pathway genes from D. rhizophagus were cloned, molecularly characterized, and their expression levels were quantified. Also, the terpenoid compounds produced in the gut were identified and quantified by Gas Chromatography Mass Spectrometry (GC-MS). The feeding treatment produced an evident upregulation, mainly in acetoacetyl-CoA thiolase (AACT), 3-hydroxy-3-methylglutaryl-CoA synthase (HMGS), 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), phosphomevalonate kinase (PMK), and isopentenyl diphosphate isomerase (IPPI) genes, and males reached higher expression levels compared to females. In contrast, the JH III treatment did not present a clear pattern of upregulation in any sex or time. Notably, the genes responsible for the synthesis of frontalin and ipsdienol precursors (geranyl diphosphate synthase/farnesyl diphosphate synthase (GPPS/FPPS) and geranylgeranyl diphosphate synthase (GGPPS)) were not clearly upregulated, nor were these compounds further identified. Furthermore, trans-verbenol and myrtenol were the most abundant compounds in the gut, which are derived from an α-pinene transformation rather than de novo synthesis. Hence, the expression of mevalonate pathway genes in D. rhizophagus gut is not directed to the production of terpenoid pheromones, regardless of their frequent occurrence in the genus Dendroctonus.

Inhibition of recombinant enzyme 3-hydroxy-3-methylglutaryl-CoA reductase from Candida glabrata by α-asarone-based synthetic compounds as antifungal agents.

Due to increasing resistance of Candida species to antifungal drugs, especially azoles, new drugs are needed. The proposed compounds 3 and 4 are analogous to α-asarone (2), a naturally occurring potent inhibitor of HMGR with hypolipidemic and antifungal activity. We used the recombinant enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase of Candida glabrata (CgHMGR) as a model to test the effectiveness of the test compounds. Compounds 3 and 4 demonstrated inhibitory kinetics, having lower IC50 values (42.65 µM and 28.77 µM, respectively) than compound 2 (>100 µM). The docking studies showed better binding energies for compounds 3 and 4 (-5.35 and -6.1 kcal/mol, respectively) than for compound 2 (-4.53 kcal/mol). These findings suggest that the tested compounds are better than their natural analogue. Plaque assays were performed on the C. glabrata strain CBS138 by applying ergosterol or cholesterol to evaluate the possible reversal of the inhibition induced by compounds 2, 3 and 4. Inhibition was easily suppressed in all three cases, recovering the viability of C. glabrata. These results reveal that the CgHMGR model is excellent for testing antifungals. Compound 4 produced the best effect and is herein proposed as a new potent antifungal agent.

Mitochondrial aquaporin-8 is involved in SREBP-controlled hepatocyte cholesterol biosynthesis.

Cholesterol, via sterol regulatory element-binding protein (SREBP) transcription factors, activates or represses genes involved in its hepatic biosynthetic pathway, and also modulates the expression of hepatocyte mitochondrial aquaporin-8 (mtAQP8), a channel that can function as peroxiporin by facilitating the transmembrane diffusion of H2O2. Here we tested the hypothesis that mtAQP8 is involved in the SREBP-mediated regulation of hepatocyte cholesterol biosynthesis. Using human hepatocyte-derived Huh-7 cells and primary rat hepatocytes, we found that mtAQP8 knockdown significantly downregulated de novo cholesterol synthesis as well as protein expressions of SREBP-2 and its target gene, a rate-limiting enzyme in cholesterol synthesis 3-Hydroxy-3-Methylglutaryl-CoA Reductase (HMGCR). In contrast, adenovirus-mediated human AQP8 mitochondrial expression significantly increased de novo cholesterol synthesis and protein expressions of SREBP-2 and HMGCR. In mtAQP8-overexpressed hepatocytes, mitochondrial H2O2 release was found to be increased; and a mitochondria-targeted antioxidant prevented the upregulation of mitochondrial H2O2 release and that of cholesterol synthesis. Our results suggest that peroxiporin mtAQP8 plays a role in the SREBP-controlled hepatocyte cholesterogenesis, a finding that might be relevant to cholesterol-related metabolic disorders.

Proposed cut-off for reactivity of anti-HMGCR and anti-SRP antibodies in patients statin-exposed and statin-unexposed.

The therapeutic approach with statins is widely used in the control of dyslipidemias. However, there is no laboratory evaluation to elect patients to make use of this class of therapeutic drugs.We analyzed the prevalence of anti-signal recognition particle (anti-SRP) and anti-3-hydroxy-3-methylglutaryl-coenzyme A reductase (anti-HMGCR) antibodies in a heterogeneous cohort of 85 patients in order to determine cutoff reference values for these antibodies.Serum samples from 85 patients were screened for the presence of anti-HMGCR and anti-SRP autoantibodies by enzyme-linked immunosorbent assay. The demographic, clinical, and morphological features were also correlated with anti-HMGCR and anti-SRP antibodies. The patients were divided in 2 groups: A, statin-exposed, and B, statin-unexposed.There was no significant association (P > .05) among anti-HMGCR and anti-SRP titers in relation to age, sex, statin exposure, and CK level. The concentrations of both antibodies were not correlated with symptoms, CK level, or statin exposure. Eleven (12.9%) patients had anti-HMGCR antibodies. We found a tendency (P = .051) toward greater anti-HMGCR positivity in women with no symptoms. Twelve (14.1%) patients had anti-SRP antibodies. There was no sex predominance, and only 1 patient had muscle complaints. Muscular symptoms were present in 31 (36.5%) patients, 4 (12.9%) were positive for anti-HMGCR antibodies, and 1 (3.2%) was positive for anti-SRP antibodies. A total of 54 (63.5%) patients had no muscle symptoms, 7 (13%) were anti-HMGCR positive, and 11 (20.4%) were anti-SRP positive. We found statistical significance for patients with anti-SRP antibodies when asymptomatic and symptomatic patients were compared (P = .029). In contrast, there was no statistically significant difference between symptoms and positivity for anti-HMG antibodies.One of the main aims of this study was to define a cutoff point in a heterogeneous population with different diagnoses. We also demonstrated that anti-HMGCR and anti-SRP antibodies are not 100% specific to immune-mediated necrotizing myopathy. We believe that these antibodies must be tested and interpreted within the specific context.

Potential Molecular Targets of Statins in the Prevention of Hepatocarcinogenesis.

Hepatocellular carcinoma (HCC) represents 90% of liver tumors. Statins, may reduce the incidence of various tumors, including HCC. Antitumoral activities may be mediated by changes in transforming growth factor-beta (TGF-ß1) and thyroid hormones (TH) regulation. INTRODUCTION AND AIM: Hepatocellular carcinoma (HCC) represents 90% of liver tumors. Statins, may reduce the incidence of various tumors, including HCC. Antitumoral activities may be mediated by changes in transforming growth factor-beta (TGF-ß1) and thyroid hormones (TH) regulation. Aim. The aim of our study is to establish the statins mechanism of action and the potential key molecules involved in an in vivo and in vitro HCC model. MATERIALS AND METHODS: We used two models: in vivo (in rats) using diethylnitrosamine (DEN) and hexachlorobenzene (HCB) to develop HCC. We analyzed cell proliferation parameters (proliferating cel nuclear antigen, PCNA) and cholesterol metabolism (hydroxy-methylglutaryl-CoA reductase, HMGCoAR). In vitro (Hep-G2 cells) we evaluated the effects of different doses of Atorvastatin (AT) and Simvastatin (SM) on HCB induced proliferation and analyzed proliferative parameters, cholesterol metabolism, TGF-ß1 mRNA, c-Src and TH levels. RESULTS: In vivo, we observed that cell proliferation significantly increased as well as cholesterol serum levels in rats treated with HCB. In vitro, we observed the same results on PCNA as in vivo. The statins prevented the increase in HMG-CoAR mRNA levels induced by HCB, reaching levels similar to controls at maximum doses: AT (30 µM), and SM (20 µM). Increases in PCNA, TGF-ß1, and pc-Src, and decreases in deiodinase I mRNA levels induced by HCB were not observed when cells were pre-treated with AT and SM at maximum doses. CONCLUSION: Statins can prevent the proliferative HCB effects on Hep-G2 cells. TGF-ß1, c-Src and TH may be the statins molecular targets in hepatocarcinogenesis.

Zaluzania montagnifolia: essential oil composition and biological properties / Zaluzania montagnifolia: composición del aceite esencial y actividades biológicas

The chemical composition of the seasonal essential oils (2015-2016) from the leaves and flowers of Zaluzania montagnifolia is presented. The chemical content of those oils showed quantitative and qualitative differences. Germacrene D (19.9-29.8%), camphor (12.4- 19.4%) and ß-caryophyllene (13.7-18.5%) were the most abundant volatiles in the leaves. The essential oils from the flowers contained high amounts of camphor (32.7-37.2%) limonene (19.8-24.9%) and germacrene D (3.2-7.3%). All the seasonal essential oils showed a potent in vitro inhibition against HMG-CoA reductase. The essential oils from flowers (IC50, 40.5-55.1 µg mL-1) showed better inhibition properties than those of leaves (IC50, 84.4-123.5 µg mL-1). Camphor (IC50, 72.5 µg mL-1) and borneol (IC50, 84.4 µg mL-1) exerted a non-competitive inhibition on the enzyme. Additionally, the hydrodistillates exhibited antibacterial activity against the phytopathogenic Pseudomonas syringae pv. tabaci TBR2004 (MIC, 62.7-76.5 µg mL-1) P. syringae pv. tomato DC3000 (MIC, 45.4-50.4 µg mL-1) and P. syringae pv. phaseolicola NPS3121 (MIC, 26.7-31.9 µg mL-1). Germacrene D (MIC, 35.4-66.2 µg mL-1) and ß-caryophyllene (MIC, 36.5-54.2 µg mL-1) were the strongest anti-Pseudomonas syringae agents.

Se presenta la composición química de los aceites esenciales estacionales (2015-2016) provenientes de hojas y flores de Zaluzania montagnifolia. El contenido químico de los aceites esenciales mostró diferencias cualitativas y cuantitativas. El germacreno D (19.9-29.8%), alcanfor (12.4-19.4%) y ß-cariofileno (13.7-18.5%) fueron los volátiles más abundantes en las hojas. Los aceites esenciales de las flores contuvieron altas concentraciones de alcanfor (32.7-37.2%), limoneno (19.8-24.9%) y germacreno D (3.2-7.3%). Todos los aceites esenciales estacionales mostraron una potente inhibición in vitro contra la HMG-CoA reductasa. Los aceites esenciales de las flores (IC50, 40.5-55.1 µg mL-1) mostraron mejores propiedades inhibitorias que aquellos de las hojas (IC50, 84.4-123.5 µg mL-1). El alcanfor (IC50, 72.5 µg mL-1) y el borneol (IC50, 84.4 µg mL-1) ejercieron una inhibición no competitiva sobre la enzima. Adicionalmente, los hidrodestilados exhibieron una actividad antibacterial contra los fitopatógenos Pseudomonas syringae pv. tabaci TBR2004 (MIC, 62.7-76.5 µg mL-1) P. syringae pv. tomato DC3000 (MIC, 45.4-50.4 µg mL-1) y P. syringae pv. phaseolicola NPS3121 (MIC, 26.7-31.9 µg mL-1). El germacreno D (MIC, 35.4-66.2 µg mL-1) y ß-cariofileno (MIC, 36.5-54.2 µg mL-1) fueron los agentes más fuertes contra los patovares de Pseudomonas syringae.

In vitro and in silico studies of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase inhibitory activity of the cowpea Gln-Asp-Phe peptide.

Previous studies have shown that cowpea protein positively interferes with cholesterol metabolism. In this study, we evaluated the ability of the fraction containing peptides of <3 kDa, as well as that of the Gln-Asp-Phe (QDF) peptide, derived from cowpea ß-vignin protein, to inhibit HMG-CoA reductase activity. We established isolation and chromatography procedures to effectively obtain the protein with a purity above 95%. In silico predictions were performed to identify peptide sequences capable of interacting with HMG-CoA reductase. In vitro experiments showed that the fraction containing peptides of <3 kDa displayed inhibition of HMG-CoA reductase activity. The tripeptide QDF inhibits HMG-CoA reductase (IC50 = 12.8 µM) in a dose-dependent manner. Furthermore, in silico studies revealed the binding profile of the QDF peptide and hinted at the molecular interactions that are responsible for its activity. Therefore, this study shows, for the first time, a peptide from cowpea ß-vignin protein that inhibits HMG-CoA reductase and the chemical modifications that should be investigated to evaluate its binding profile.

Down-regulation of single-stranded DNA-binding protein 1 expression induced by HCMV infection promotes lipid accumulation in cells.

The objective of this study was to observe the infection of human cytomegalovirus (HCMV) to human umbilical vein endothelial cells, and its effect on the expression of single-stranded DNA-binding protein (SSBP1) and on lipid metabolism in endothelial cells. We screened the differential expression of mRNAs after HCMV infection by suppression subtractive hybridization and the expression levels of SSBP1 mRNA and protein after HCMV infection by real-time PCR and western blot. After verification of successful infection by indirect immunofluorescent staining and RT-PCR, we found a differential expression of lipid metabolism-related genes including LDLR, SCARB, CETP, HMGCR, ApoB and LPL induced by HCMV infection. The expression levels of SSBP1 mRNA and protein after HCMV infection were significantly down-regulated. Furthermore, we found that upregulation of SSBP1 inhibited the expression of atherosclerosis-associated LDLR, SCARB, HMGCR, CETP as well as the accumulation of lipids in the cells. The results showed that the inhibition of SSBP1 by HCMV infection promotes lipid accumulation in the cells.

Cholesterol can modulate mitochondrial aquaporin-8 expression in human hepatic cells.

Hepatocyte mitochondrial aquaporin-8 (mtAQP8) works as a multifunctional membrane channel protein that facilitates the uptake of ammonia for its detoxification to urea as well as the mitochondrial release of hydrogen peroxide. Since early oligonucleotide microarray studies in liver of cholesterol-fed mice showed an AQP8 downregulation, we tested whether alterations of cholesterol content per se modulate mtAQP8 expression in human hepatocyte-derived Huh-7 cells. Cholesterol loading with methyl-ß-cyclodextrin (mßCD):cholesterol complexes downregulated the proteolytic activation of cholesterol-responsive sterol regulatory element-binding protein (SREBP) transcriptions factors 1 and 2, and the expression of the target gene 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR). Under such conditions, mtAQP8 mRNA and protein expressions were significantly reduced. In contrast, cholesterol depletion using mßCD alone increased SREBP-1 and 2 activation and upregulated HMGCR and mtAQP8 mRNA and protein expressions. The results suggest that cholesterol can regulate transcriptionally human hepatocyte mtAQP8 expression likely via SREBPs. The functional implications of our findings are discussed. © 2017 IUBMB Life, 69(5):341-346, 2017.

Recombinant 3-Hydroxy 3-Methyl Glutaryl-CoA Reductase from Candida glabrata (Rec-CgHMGR) Obtained by Heterologous Expression, as a Novel Therapeutic Target Model for Testing Synthetic Drugs.

The enzyme 3-hydroxy-3-methyl-glutaryl CoA reductase (HMGR) is a glycoprotein of the endoplasmic reticulum that participates in the mevalonate pathway, the precursor of cholesterol in human and ergosterol in fungi. This enzyme has three domains: transmembrane, binding, and soluble. In this study, we expressed and purified the soluble fraction of the HMGR enzyme from Candida glabrata (CgHMGR) in an Escherichia coli heterologous system and used it as a model for studying its inhibitory activity. The soluble fraction of CgHMGR was fused to the maltose binding protein (MBP), purified, and characterized. Optimal pH was 8.0, and its optimal temperature activity was 37 °C. The k m and V max for the HMG-CoA were 6.5 µM and 2.26 × 10-3 µM min-1, respectively. Recombinant CgHMGR was inhibited by simvastatin presenting an IC50 at 14.5 µM. In conclusion, our findings suggest that the recombinant HMGR version from C. glabrata may be used as a study model system for HMGR inhibitors such as statins and newly synthesized inhibitor compounds that might be used in the treatment of hypercholesterolemia or mycosis.