Effect of Trichomonacide 6-Nitro-1H-benzimidazole Derivative Compounds on Expression Level of Metabolic Genes in Trichomonas vaginalis.

The parasite Trichomonas vaginalis is the etiologic agent of trichomoniasis, the most common non-viral sexually transmitted disease worldwide. This infection often remains asymptomatic and is related to several health complications. The traditional treatment for trichomoniasis is the use of drugs of the 5-nitroimidazole family, such as metronidazole; however, scientific reports indicate an increasing number of drug-resistant strains. Benzimidazole derivatives could offer an alternative in the search for new anti-trichomonas drugs. In this sense, two attractive candidates are the compounds O2N-BZM7 and O2N-BZM9 (1H-benzimidazole derivatives), since, through in vitro tests, they have shown a higher trichomonacide activity. In this study, we determined the effect on the expression level of metabolic genes in T. vaginalis. The results show that genes involved in redox balance (NADHOX, G6PD::6PGL) are overexpressed, as well as the gene that participates in the first reaction of glycolysis (CK); on the other hand, structural genes such as ACT and TUB are decreased in expression in trophozoites treated with the compound O2N-BZM9, which would probably affect its morphology, motility and virulence. These results align with the trichomonacidal activity of the compounds, with benzimidazole O2N-BZM9 being the most potent, with an IC50 value of 4.8 µM. These results are promising for potential future therapeutic applications.

An Overall View of the Functional and Structural Characterization of Glucose-6-Phosphate Dehydrogenase Variants in the Mexican Population.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency, affecting an estimated 500 million people worldwide, is a genetic disorder that causes human enzymopathies. Biochemical and genetic studies have identified several variants that produce different ranges of phenotypes; thus, depending on its severity, this enzymopathy is classified from the mildest (Class IV) to the most severe (Class I). Therefore, understanding the correlation between the mutation sites of G6PD and the resulting phenotype greatly enhances the current knowledge of enzymopathies' phenotypic and genotypic heterogeneity, which will assist both clinical diagnoses and personalized treatments for patients with G6PD deficiency. In this review, we analyzed and compared the structural and functional data from 21 characterized G6PD variants found in the Mexican population that we previously characterized. In order to contribute to the knowledge regarding the function and structure of the variants associated with G6PD deficiency, this review aimed to determine the molecular basis of G6PD and identify how these mutations could impact the structure, stability, and function of the enzyme and its relation with the clinical manifestations of this disease.

Giardia lamblia G6PD::6PGL Fused Protein Inhibitors Decrease Trophozoite Viability: A New Alternative against Giardiasis.

Treatments to combat giardiasis have been reported to have several drawbacks, partly due to the drug resistance and toxicity of current antiparasitic agents. These constraints have prompted many researchers to investigate new drugs that act against protozoan parasites. Enzyme inhibition is an important means of regulating pathogen metabolism and has recently been identified as a significant alternative target in the search for new treatments. Glucose-6-phosphate dehydrogenase and 6-phosphogluconolactonase (G6PD::6PGL) is a bifunctional enzyme involved in the pentose phosphate pathway (PPP) in Giardia lamblia (G. lamblia). The G. lamblia enzyme is unusual since, unlike the human enzyme, it is a fused enzyme. Here, we show, through inhibition assays, that an in-house chemical library of 120 compounds and four target compounds, named CNZ-7, CNZ-8, CMC-1, and FLP-2, are potent inhibitors of the G. lamblia G6PD::6PGL fused enzyme. With a constant (k2) of 2.3, 3.2, and 2.8 M−1 s−1, respectively, they provoke alterations in the secondary and tertiary protein structure and global stability. As a novel approach, target compounds show antigiardial activity, with IC50 values of 8.7, 15.2, 15.3, and 24.1 µM in trophozoites from G. lamblia. Moreover, these compounds show selectivity against G. lamblia, since, through counter-screening in Caco-2 and HT29 human cells, they were found to have low toxicity. This finding positions these compounds as a potential and attractive starting point for new antigiardial drugs.

TFEB; Beyond Its Role as an Autophagy and Lysosomes Regulator.

Transcription factor EB (TFEB) is considered the master transcriptional regulator of autophagy and lysosomal biogenesis, which regulates target gene expression through binding to CLEAR motifs. TFEB dysregulation has been linked to the development of numerous pathological conditions; however, several other lines of evidence show that TFEB might be a point of convergence of diverse signaling pathways and might therefore modulate other important biological processes such as cellular senescence, DNA repair, ER stress, carbohydrates, and lipid metabolism and WNT signaling-related processes. The regulation of TFEB occurs predominantly at the post-translational level, including phosphorylation, acetylation, SUMOylating, PARsylation, and glycosylation. It is noteworthy that TFEB activation is context-dependent; therefore, its regulation is subjected to coordinated mechanisms that respond not only to nutrient fluctuations but also to stress cell programs to ensure proper cell homeostasis and organismal health. In this review, we provide updated insights into novel post-translational modifications that regulate TFEB activity and give an overview of TFEB beyond its widely known role in autophagy and the lysosomal pathway, thus opening the possibility of considering TFEB as a potential therapeutic target.

Biochemical and Kinetic Characterization of the Glucose-6-Phosphate Dehydrogenase from Helicobacter pylori Strain 29CaP.

Helicobacter pylori (H. pylori) has been proposed as the foremost risk factor for the development of gastric cancer. We found that H. pylori express the enzyme glucose-6-phosphate dehydrogenase (HpG6PD), which participates in glucose metabolism via the pentose phosphate pathway. Thus, we hypothesized that if the biochemical and physicochemical characteristics of HpG6PD contrast with the host G6PD (human G6PD, HsG6PD), HpG6PD becomes a potential target for novel drugs against H. pylori. In this work, we characterized the biochemical properties of the HpG6PD from the H.pylori strain 29CaP and expressed the active recombinant protein, to analyze its steady-state kinetics, thermostability, and biophysical aspects. In addition, we analyzed the HpG6PD in silico structural properties to compare them with those of the HsG6PD. The optimal pH for enzyme activity was 7.5, with a T1/2 of 46.6 °C, at an optimum stability temperature of 37 °C. The apparent Km values calculated for G6P and NADP+ were 75.0 and 12.8 µM, respectively. G6P does not protect HpG6PD from trypsin digestion, but NADP+ does protect the enzyme from trypsin and guanidine hydrochloride (Gdn-HCl). The biochemical characterization of HpG6PD contributes to knowledge regarding H. pylori metabolism and opens up the possibility of using this enzyme as a potential target for specific and efficient treatment against this pathogen; structural alignment indicates that the three-dimensional (3D) homodimer model of the G6PD protein from H. pylori is different from the 3D G6PD of Homo sapiens.

Glucose-6-Phosphate Dehydrogenase::6-Phosphogluconolactonase from the Parasite Giardia lamblia. A Molecular and Biochemical Perspective of a Fused Enzyme.

Giardia lamblia is a single-celled eukaryotic parasite with a small genome and is considered an early divergent eukaryote. The pentose phosphate pathway (PPP) plays an essential role in the oxidative stress defense of the parasite and the production of ribose-5-phosphate. In this parasite, the glucose-6-phosphate dehydrogenase (G6PD) is fused with the 6-phosphogluconolactonase (6PGL) enzyme, generating the enzyme named G6PD::6PGL that catalyzes the first two steps of the PPP. Here, we report that the G6PD::6PGL is a bifunctional enzyme with two catalytically active sites. We performed the kinetic characterization of both domains in the fused G6PD::6PGL enzyme, as well as the individual cloned G6PD. The results suggest that the catalytic activity of G6PD and 6PGL domains in the G6PD::6PGL enzyme are more efficient than the individual proteins. Additionally, using enzymatic and mass spectrometry assays, we found that the final metabolites of the catalytic reaction of the G6PD::6PGL are 6-phosphoglucono-δ-lactone and 6-phosphogluconate. Finally, we propose the reaction mechanism in which the G6PD domain performs the catalysis, releasing 6-phosphoglucono-δ-lactone to the reaction medium. Then, this metabolite binds to the 6PGL domain catalyzing the hydrolysis reaction and generating 6-phosphogluconate. The structural difference between the G. lamblia fused enzyme G6PD::6PGL with the human G6PD indicate that the G6PD::6PGL is a potential drug target for the rational synthesis of novels anti-Giardia drugs.

Polymorphisms -455G/A and -148C/T and Fibrinogen Plasmatic Level as Risk Markers of Coronary Disease and Major Adverse Cardiovascular Events.

Some polymorphisms in genes codifying for fibrinogen have been correlated with plasma levels of this protein, and several studies reported their associations with acute cardiovascular events. In the present study, 118 subjects with unstable and stable coronary diseases were enrolled to determinate the associations among fibrinogen gene polymorphisms, plasma fibrinogen levels, and major cardiovascular adverse events in a sample of southwestern Mexico. The groups, including 81 control subjects, were matched for age, sex, body mass index, and sedentarism. Plasma fibrinogen levels and the polymorphisms 455G/A, -148C/T, +1689T/G, and Bcl I of the gene of fibrinogen were compared in all groups. Plasma fibrinogen levels (>465 mg/dl) were significant in patients with coronary disease. Fibrinogen plasma values > 450 mg/dl were associated with cardiovascular mortality during the follow-up analysis of the unstable coronary disease group (p = 0.04). The allelic loads of -455A and -148T were associated with plasma fibrinogen levels > 450 mg/dl (p < 0.003 and p = 0.03, respectively) and with coronary disease (p = 0.016 and p < 0.006, respectively). The follow-up of posterior events after an acute coronary event showed that the genetic load of the -148T allele was associated with major adverse cardiovascular events (RR = 1.8, 95%CI = 1.01-3.35, p = 0.04). Fibrinogen plasmatic levels > 450 mg/dl and the fibrinogen polymorphisms -455G/A and 148C/T had association with MACE and coronary disease. This study suggests that these gene polymorphisms are associated with cardiovascular risk.

Clinical characterization of acute and convalescent illness of confirmed chikungunya cases from Chiapas, S. Mexico: A cross sectional study.

BACKGROUND: The emerging chikungunya virus (CHIKV), is an arbovirus causing intense outbreaks in North America. The situation in Mexico is alarming, and CHIKV threatens to spread further throughout North America. Clinical and biological features of CHIKF outbreaks in Mexico have not been well described; thus, we conducted a cross sectional study of a CHIKV outbreak in Chiapas, Southern Mexico to further characterize these features. METHODOLOGY/PRINCIPAL FINDINGS: We collected blood samples from patients suspected of having chikungunya fever (CHIKF) who presented to Clinical Hospital ISSSTE Dr. Roberto Nettel in Tapachula, Chiapas, Mexico. In addition to the clinical examination, real-time polymerase chain reaction (PCR) standardized for the Asian Chikungunya lineage and/or enzyme-linked immunosorbent assay for immunoglobulin M (IgM) were used to confirm CHIKV diagnosis. Of a total of 850 patients who presented with probably CHIKV at Hospital "Dr. Roberto Nettel", 112 probable CHIKF cases were enrolled in this study from November 2014- June 2015, of which 95 patients (84.8%) were CHIKV positive and 17 were negative (15.2%). Of these 95 CHIKV positive patients, 62 were positive by real-time reverse transcriptase PCR (+qRT-PCR); and 33 were seropositive to +IgM with a negative qRT-PCR. The most frequent symptoms reported were fever (100%), headache (82.3%), polyarthralgia (72.1%), and exanthem (82.3%). Biological abnormalities observed during CHIKV infection were lymphopenia (41.1%), leukopenia (51.6%), elevated transaminases (30.5%-46.3%) and high LDH (46.3%) and CRP (60.0%). CONCLUSION: Clinical and biological data obtained from this study is providing more useful information for benchmarking purposes with outbreaks from different parts of the world and would be helpful for better patient care and treatment.

[Maternal-fetal transmission of Trypanosoma cruzi, a health problem slightly studied in Mexico: case Chiapas]. / Transmisión materno-fetal de Trypanosoma cruzi, un problema de salud poco estudiado en México: caso Chiapas.

OBJECTIVE: To determine the Trypanosoma cruzi infection prevalence in 1125 pregnant women and the transmission frequency to their children from Tapachula and Palenque, Chiapas. MATERIALS AND METHODS: We determined the prevalence by serology tests and the transmission frequency by polymerase chain reaction (PCR) and T. cruzi reactivity capacity after 12 months. RESULTS: Total maternal infection prevalence were 23/1 125 (2.04%), 9/600 (1.5%) were from Tapachula and 14/525 (2.6%) from Palenque. The seropositive women were between 20 and 35 years old, 31.8% have Premature Rapture of Membrane and 9.1% have history of perinatal death. The total percentage of positive newborns by PCR was 9/23 (39.13%), out of those 2/9 (22.2%) are from Tapachula and 7/14 (50%) from Palenque. The Maternal Fetal transmission frequency was. 2/9 (22.2%) in Tapachula and 1/14 (7.14%) in Palenque, all positive infants were asynthomatic. CONCLUSION: The maternal-fetal transmission rate in Chiapas State is variable; the reason could be the maternal immunological status and T. cruzi strain.

Transmisión materno-fetal de Trypanosoma cruzi, un problema de salud poco estudiado en México: caso Chiapas / Maternal-fetal transmission of Trypanosoma cruzi, a health problem slightly studied in Mexico: case Chiapas

Resumen: Objetivo: Determinar la prevalencia de infección de Trypanosoma cruzi en 1 125 mujeres embarazadas y la frecuencia con que éstas la transmiten a sus hijos en Tapachula y Palenque, Chiapas. Material y métodos: Se determinó la prevalencia materna por serología, la frecuencia de transmisión por reacción en cadena de la polimerasa (PCR) y la capacidad reactiva a T. cruzi a los 12 meses de edad del niño. Resultados: La prevalencia materna total fue 23/1 125 (2.04%); 9/600 (1.5%) en Tapachula y 14/525 (2.6%) en Palenque. Las mujeres seropositivas tenían entre 20 y 35 años; 31.8% tuvo ruptura prematura de membrana (RPM) y 9.1% tuvo antecedentes de muerte perinatal. La frecuencia de transmisión materno-fetal en Tapachula fue de 2/9(22.2%) y en Palenque de 1/14(7.14%); todos los niños positivos eran asintomáticos. Conclusión: La tasa de transmisión materno-fetal en el estado de Chiapas es variable; la razón podría ser el estado inmunológico de la madre o la cepa de T. cruzi.

Abstract: Objective: To determine the Trypanosoma cruzi infection prevalence in 1125 pregnant women and the transmission frequency to their children from Tapachula and Palenque, Chiapas. Materials and methods: We determined the prevalence by serology tests and the transmission frequency by polymerase chain reaction (PCR) and T. cruzi reactivity capacity after 12 months. Results: Total maternal infection prevalence were 23/1 125 (2.04%), 9/600 (1.5%) were from Tapachula and 14/525 (2.6%) from Palenque. The seropositive women were between 20 and 35 years old, 31.8% have Premature Rapture of Membrane and 9.1% have history of perinatal death. The total percentage of positive newborns by PCR was 9/23 (39.13%), out of those 2/9 (22.2%) are from Tapachula and 7/14 (50%) from Palenque. The Maternal Fetal transmission frequency was. 2/9 (22.2%) in Tapachula and 1/14 (7.14%) in Palenque, all positive infants were asynthomatic. Conclusion: The maternal-fetal transmission rate in Chiapas State is variable; the reason could be the maternal immunological status and T. cruzi strain.

Determinación molecular de marcadores genéticos en síndromes coronarios agudos y su relación con eventos cardiovasculares adversos / Determination of molecular genetic markers in acute coronary syndromes and their relationship to cardiovascular adverse events

Introducción: Algunos genes relacionados con proteínas que participan en algunas rutas metabólicas podrían tener un papel en el desarrollo de los síndromes coronarios agudos. Objetivo: Correlación entre polimorfismos y su relación con eventos adversos en síndromes coronarios agudos. Método: Prospectivo, seguimiento hospitalario y a un año. Inclusión: síndromes coronarios agudos con desnivel o elevación del ST secundario a aterotrombosis, estabilidad clínica. En todos, reacción de cadena de polimerasa y polimorfismos de la longitud de los fragmentos de restricción. Al estandarizar reacciones de cadena y genotipificación se realizó análisis preliminar de distribución de genotipos para cada polimorfismo y en ninguno se observaron desviaciones en la ley de equilibrio de Hardy-Weinberg (p > 0,05). Resultados: De 2003 a 2005 se ingresaron 150 sujetos. Se analizaron 14 polimorfismos en 9 genes (fibrinógeno, factor V, VII, II, XIII, activador e inhibidor del plasminógeno-1 y proteína C reactiva). En síndromes coronarios agudos, un fibrinógeno > 450mg/dL y leucocitos > 8, 500 cél/mm³ fueron marcadores de mal pronóstico a un año. Los análisis de regresión identificaron al -148 CT/TT del fibrinógeno y al -717 AG/GG de la proteína C reactiva como marcadores de isquemia recurrente y al 1691GA+AA para reinfarto. Conclusión: En pacientes con síndromes coronarios agudos se demostró una relación entre polimorfismos relacionados con hemostasia e inflamación con eventos adversos, por lo que podrían considerarse marcadores de enfermedad coronaria. Se requiere una muestra mayor para confirmar estos resultados.

Introduction: The genes coding for proteins due to their activity in several metabolic pathways could be related with the onset of acute coronary syndromes. Objective: Relationship among polymorphisms and adverse events in. Methods: Prospective. In - hospital, one - year follow-up. Inclusion Acute coronary syndromes with ST elevation or depression secondary to atherothrombosis, clinical stability. In all, polymerase chain reaction and length polymorphism of restriction fragments. By standardizing chain reactions and genotyping,a preliminary analysis of distribution of genotypes was performed for each polymorphism and no deviations were observed in the law of Hardy-Weinberg equilibrium (P> .05). Results: From 2003 to 2005, 150 subjects were enrolled. We analyzed 14 polymorphisms in 9 genes (fibrinogen, factor V, VII, II, XIII, plasminogen activator and inhibitor-1, C-reactive protein). In acute coronary syndromes, fibrinogen > 450 mg/dL and white blood count 8500 cells/mm³ were markers of poor prognosis to one year. Regression analysis identified the -148 CT/TT and fibrinogen -717 AG/GG of C-reactive protein as a marker of recurrent ischemia and reinfarction 1691GA +AA. Conclusion: We are showing a relationship among polymorphisms involved in inflammation and hemostasis with adverse events in the acute phase and follow-up in acute coronary syndromes patients that could be considered as markers of ischemic heart disease. Larger sample is needed to confirm these results.

[Determination of molecular genetic markers in acute coronary syndromes and their relationship to cardiovascular adverse events]. / Determinación molecular de marcadores genéticos en síndromes coronarios agudos y su relación con eventos cardiovasculares adversos.

INTRODUCTION: The genes coding for proteins due to their activity in several metabolic pathways could be related with the onset of acute coronary syndromes. OBJECTIVE: Relationship among polymorphisms and adverse events in. METHODS: Prospective. In--hospital, one--year follow-up. Inclusion Acute coronary syndromes with ST elevation or depression secondary to atherothrombosis, clinical stability. In all, polymerase chain reaction and length polymorphism of restriction fragments. By standardizing chain reactions and genotyping,a preliminary analysis of distribution of genotypes was performed for each polymorphism and no deviations were observed in the law of Hardy-Weinberg equilibrium (P>.05). RESULTS: From 2003 to 2005, 150 subjects were enrolled. We analyzed 14 polymorphisms in 9 genes (fibrinogen, factor v, vii, ii, xiii, plasminogen activator and inhibitor-1, C-reactive protein). In acute coronary syndromes, fibrinogen>450 mg/dL and white blood count 8500 cells/mm(3) were markers of poor prognosis to one year. Regression analysis identified the -148 CT/TT and fibrinogen -717 AG/GG of C-reactive protein as a marker of recurrent ischemia and reinfarction 1691GA + AA. CONCLUSION: We are showing a relationship among polymorphisms involved in inflammation and hemostasis with adverse events in the acute phase and follow-up in acute coronary syndromes patients that could be considered as markers of ischemic heart disease. Larger sample is needed to confirm these results.

Análisis preliminar de la actividad antimicrobiana de la planta medicinal Chik chawa (Tagetes nelsonii Greenm) / Preliminary analysis of antimicrobial activity of medicinal plant Chik chawa (Tagetes nelsonii Greenm)

INTRODUCCIÓN: Tagetes nelsonii Greenm., comúnmente llamada Chik chawa, es una planta utilizada en San Cristóbal de las Casas, Chiapas, México, porque los indígenas de esa región la han utilizado de manera tradicional para el tratamiento empírico de las diarreas, dolor de cabeza y fiebre; por lo cual los pobladores la han adoptado como una planta medicinal. OBJETIVO: determinar la actividad antimicrobiana in vitro de extractos de T. nelsonii. MÉTODOS: se prepararon 3 diferentes extractos, hexánico, metanólico y diclorometánico, de follaje de T. nelsonii, así como también infusión, los cuales fueron probados frente a 7 diferentes cepas microbianas por el método de dilución en placa de agar. A los extractos activos se les determinó la concentración mínima inhibitoria frente a las cepas microbianas que fueron susceptibles y las concentraciones mínimas bactericidas se determinaron mediante el método de dilución en caldo.RESULTADOS: 4 de las 7 cepas estudiadas fueron sensibles, estas fueron P. mirabilis al extracto metanólico, S. aureus y P. aeruginosa (ATCC 27863) a los extractos metanólico, hexánico y diclorometánico y C. albicans al extracto diclorometánico. La infusión de chik chawa no mostró actividad antimicrobiana. El extracto metanólico frente a P. mirabilis y S. aureus mostró efecto bactericida. CONCLUSIONES: los extractos orgánicos de Chik chawa mostraron actividad antimicrobiana sobre cepas bacterianas asociadas comúnmente con enfermedades humanas. Ninguno de los extractos causó efecto antifúngico. Se necesitan otros estudios para dilucidar los principios activos responsables de la actividad antibacteriana en los extractos probados.

INTRODUCTION: Tagetes nelsonii Greenm, usually called Chik chawa, is a plant used in San Cristobal de las Casas, Chiapas, México, because of natives from that region used it in a traditional way for empirical treatment of diarrheas, headache and fever, which has been adopted as a medicinal plant.OBJECTIVE: to determine the in vitro antimicrobial activity of T. nelsonii extracts. METHODS: three extracts were prepared: hexanic, methanolic and dichloromethanic from the T. nelsonii foliage, as well as a infusion, which were assayed versus 7 different microbial strains by agar plate diffusion method. In active extracts the inhibitory minimal concentration were determined versus susceptible microbial strains and the bactericidal minimal concentrations were determined by broth dilution method. RESULTS: four of the 7 study strains were sensible including P. mirabilis to methanolic extract, S. aureus and P. aeruginosa (ATCC 27863) to methanolic, hexanic and dichloromethanic extracts, and C. albicans to dichloromethanic extract. Chik chawa infussion has not antimicrobial activity. CONCLUSIONS: methanolic extract versus P. mirabilis y S. aureus showed this kind of activity on bacterial strains commonly associated with human diseases. Other studies are necessary to elucidate the active principles accounting for the antibacterial activity in the assayed extracts.