Mass spectrometric analysis of digesta does not improve the allergenicity assessment of GM crops.

An investigation of the potential allergenicity of newly expressed proteins in genetically modified (GM) crops comprises part of the assessment of GM crop safety. However, allergenicity is not completely predictable from a definitive assay result or set of protein characteristics, and scientific opinions regarding the data that should be used to assess allergenicity are continuously evolving. Early studies supported a correlation between the stability of a protein exposed to digestive enzymes such as pepsin and the protein's status as a potential allergen, but over time the conclusions of these earlier studies were not confirmed. Nonetheless, many regulatory authorities, including the European Food Safety Authority (EFSA), continue to require digestibility analyses as a component of GM crop risk assessments. Moreover, EFSA has recently investigated the use of mass spectrometry (MS), to make digestion assays more predictive of allergy risk, because it can detect and identify small undigested peptides. However, the utility of MS is questionable in this context, since known allergenic peptides are unlikely to exist in protein candidates intended for commercial development. These protein candidates are pre-screened by the same bioinformatics processes that are normally used to identify MS targets. Therefore, MS is not a standalone allergen identification method and also cannot be used to predict previously unknown allergenic epitopes. Thus, the suggested application of MS for analysis of digesta does not improve the poor predictive power of digestion assays in identifying allergenic risk.

Antimicrobial resistance of three common molecularly identified pathogenic bacteria to Allium aqueous extracts.

The aim of this work was to evaluate the in vitro bacterial inhibition of different types of garlic on Escherichia coli ATCC 25922, Listeria monocytogenes and Staphylococcus aureus. The bacterial strains were molecularly identified using gen 16S rDNA molecular identification. Four different types of garlics were used: 1) white, 2) Japanese, 3) elephant and 3) black, and these were evaluated at two different concentrations (0.25 and 0.125 g/mL) per garlic type. Bioactive compounds present in the garlics were identified using high-performance liquid chromatography coupled to ultraviolet detector (HPLC-UV), and total polyphenols were quantified by the Folin-Ciocalteu technique. The Kirby-Bauber method was used for the bacterial evaluation. Aqueous extract of black garlic had the highest amount of polyphenols 6.26 ± 0.21 mg GAE/mL. The area of inhibition was measured and classified as sensitive, intermediate or resistant. Using the disc diffusion assay, higher concentration (0.25 g/mL) of aqueous extract of white garlic had the highest antibacterial activity area, with 21.46 ± 3.94 mm for L. monocytogenes, 20.61 ± 2.47 mm for S. aureus and 17.83 ± 2.21 mm for E. coli. White garlic had comparable antimicrobial activity as the control (tetracycline at 30 µg) as indicated by the size of the inhibition halos. Based on your results, white garlic can be used as an alternative to synthetic antimicrobials.

Nested Polymerase Chain Reaction and Cutaneous Tuberculosis.

The role of Mycobacterium tuberculosis in the etiology and pathogenesis of cutaneous tuberculosis is controversial because of the difficulties associated with demonstrating the presence of these mycobacteria in tuberculid cutaneous lesions by routinely available microbiological and histological techniques. In this study, we aimed to demonstrate the presence of M. tuberculosis in cutaneous tuberculosis. Multiple polymerase chain reaction (PCR) followed by nested PCR was used to amplify genomic fragments from 3 different mycobacteria species. DNA was isolated from 30 paraffin-embedded skin biopsies. Samples were selected randomly from patients with a clinical and histopathological diagnosis of the most frequent groups of cutaneous tuberculosis in Mexico as follows: 5 cases of scrofuloderma tuberculosis; 2 cases of lupus vulgaris tuberculosis; and 5 cases of tuberculosis verrucosa cutis. The other cases denominated tuberculids in some countries such as Mexico and included the following: 7 cases of rosacea-like tuberculosis; one case of papulonecrotic tuberculosis; and 10 cases of erythema induratum of Bazin. Four normal skin biopsies were included as controls. M. tuberculosis DNA was amplified successfully by nested PCR in 80% of the samples (24 of the 30 samples) assayed. Mycobacterial DNA was not detected in the normal skin biopsies used as controls. Detection of M. tuberculosis DNA in 80% of cutaneous tuberculosis analyzed implicates this mycobacterium in the pathogenesis of multiple clinical forms of cutaneous tuberculosis.

Protocol for a Randomised Controlled Trial Investigating Decompression for Leprous Neuropathy (The DELN Protocol).

Objectives: An expert group of peripheral nerve surgeons, reconstructive surgeons, and immunologists who have extensive experience with Hansen's Disease convened to discuss the status of nerve decompression as a treatment for leprous neuropathy. The expert group recommended an international, multi-center randomised controlled trial (RCT). Subsequently, a study protocol called Decompression for Leprous Neuropathy (DELN) was designed and further refined by multiple investigators worldwide. The DELN Protocol: The DELN RCT seeks to determine the long-term effect of nerve decompression on sensibility, motor function, neuropathic pain, disability, and quality of life. The RCT would enroll patients with clinically diagnosed leprous neuropathy and positive Tinel signs in the upper and lower extremities. Patients would then be randomized to receive nerve decompression or not. Outcomes of interest include sensory function, motor function, pain, disability, and quality of life. The development of ulcers or amputations after surgery and the influence of corticosteroid therapy are also important outcomes that DELN seeks to determine. Conclusions: The study Decompression for Leprous Neuropathy (DELN) is an international, multi-center RCT with the potential to produce high quality data to address whether nerve decompression for leprous neuropathy can conclusively improve patient outcomes. We invite discussion from all those involved in the peripheral nerve and leprosy communities.

Potential of olfactory neuroepithelial cells as a model to study schizophrenia: A focus on GPCRs (Review).

Schizophrenia (SZ) is a multifactorial disorder characterized by volume reduction in gray and white matter, oxidative stress, neuroinflammation, altered neurotransmission, as well as molecular deficiencies such as punctual mutation in Disrupted­in­Schizophrenia 1 protein. In this regard, it is essential to understand the underlying molecular disturbances to determine the pathophysiological mechanisms of the disease. The signaling pathways activated by G protein­coupled receptors (GPCRs) are key molecular signaling pathways altered in SZ. Convenient models need to be designed and validated to study these processes and mechanisms at the cellular level. Cultured olfactory stem cells are used to investigate neural molecular and cellular alterations related to the pathophysiology of SZ. Multipotent human olfactory stem cells are undifferentiated and express GPCRs involved in numerous physiological functions such as proliferation, differentiation and bioenergetics. The use of olfactory stem cells obtained from patients with SZ may identify alterations in GPCR signaling that underlie dysfunctional processes in both undifferentiated and specialized neurons or derived neuroglia. The present review aimed to analyze the role of GPCRs and their signaling in the pathophysiology of SZ. Culture of olfactory epithelial cells constitutes a suitable model to study SZ and other psychiatric disorders at the cellular level.

Nucleoredoxin Redox Interactions Are Sensitized by Aging and Potentiated by Chronic Alcohol Consumption in the Mouse Liver.

Aging is characterized by increased reactive species, leading to redox imbalance, oxidative damage, and senescence. The adverse effects of alcohol consumption potentiate aging-associated alterations, promoting several diseases, including liver diseases. Nucleoredoxin (NXN) is a redox-sensitive enzyme that targets reactive oxygen species and regulates key cellular processes through redox protein-protein interactions. Here, we determine the effect of chronic alcohol consumption on NXN-dependent redox interactions in the liver of aged mice. We found that chronic alcohol consumption preferentially promotes the localization of NXN either into or alongside senescent cells, declines its interacting capability, and worsens the altered interaction ratio of NXN with FLII, MYD88, CAMK2A, and PFK1 proteins induced by aging. In addition, carbonylated protein and cell proliferation increased, and the ratios of collagen I and collagen III were inverted. Thus, we demonstrate an emerging phenomenon associated with altered redox homeostasis during aging, as shown by the declining capability of NXN to interact with partner proteins, which is enhanced by chronic alcohol consumption in the mouse liver. This evidence opens an attractive window to elucidate the consequences of both aging and chronic alcohol consumption on the downstream signaling pathways regulated by NXN-dependent redox-sensitive interactions.

Kinetic and isotopic characterization of L-proline dehydrogenase from Mycobacterium tuberculosis.

The monofunctional proline dehydrogenase (ProDH) from Mycobacterium tuberculosis performs the flavin-dependent oxidation of l-proline to Δ(1)-pyrroline-5-carboxylate in the proline catabolic pathway. The ProDH gene, prub, was cloned into the pYUB1062 vector, and the C-terminal His-tagged 37 kDa protein was expressed and purified by nickel affinity chromatography. A steady-state kinetic analysis revealed a ping-pong mechanism with an overall kcat of 33 ± 2 s(-1) and Km values of 5.7 ± 0.8 mM and 3.4 ± 0.3 µM for l-proline and 2,6-dichlorophenolindophenol (DCPIP), respectively. The pH dependence of kcat revealed that one enzyme group exhibiting a pK value of 6.8 must be deprotonated for optimal catalytic activity. Site-directed mutagenesis suggests that this group is Lys110. The primary kinetic isotope effects on V/KPro and V of 5.5 and 1.1, respectively, suggest that the transfer of hydride from l-proline to FAD is rate-limiting for the reductive half-reaction, but that FAD reoxidation is the rate-limiting step in the overall reaction. Solvent and multiple kinetic isotope effects suggest that l-proline oxidation occurs in a stepwise rather than concerted mechanism. Pre-steady-state kinetics reveal an overall kred of 88.5 ± 0.7 s(-1), and this rate is subject to a primary kinetic isotope effect of 5.2. These data confirm that the overall reaction is limited by reduced flavin reoxidation in the second half-reaction.

Kinetic, mutational, and structural analysis of malonate semialdehyde decarboxylase from Coryneform bacterium strain FG41: mechanistic implications for the decarboxylase and hydratase activities.

Malonate semialdehyde decarboxylase from Pseudomonas pavonaceae 170 (designated Pp MSAD) is in a bacterial catabolic pathway for the nematicide 1,3-dichloropropene. MSAD has two known activities: it catalyzes the metal ion-independent decarboxylation of malonate semialdehyde to produce acetaldehyde and carbon dioxide and a low-level hydration of 2-oxo-3-pentynoate to yield acetopyruvate. The latter activity is not known to be biologically relevant. Previous studies identified Pro-1, Asp-37, and a pair of arginines (Arg-73 and Arg-75) as critical residues in these activities. In terms of pairwise sequence, MSAD from Coryneform bacterium strain FG41 (designated FG41 MSAD) is 38% identical with the Pseudomonas enzyme, including Pro-1 and Asp-37. However, Gln-73 replaces Arg-73, and the second arginine is shifted to Arg-76 by the insertion of a glycine. To determine how these changes relate to the activities of FG41 MSAD, the gene was cloned and the enzyme expressed and characterized. The enzyme has a comparable decarboxylase activity but a significantly reduced hydratase activity. Mutagenesis along with crystal structures of the native enzyme (2.0 Å resolution) and the enzyme modified by a 3-oxopropanoate moiety (resulting from the incubation of the enzyme and 3-bromopropiolate) (2.2 Å resolution) provided a structural basis. The roles of Pro-1 and Asp-37 are likely the same as those proposed for Pp MSAD. However, the side chains of Thr-72, Gln-73, and Tyr-123 replace those of Arg-73 and Arg-75 in the mechanism and play a role in binding and catalysis. The structures also show that Arg-76 is likely too distant to play a direct role in the mechanism. FG41 MSAD is the second functionally annotated homologue in the MSAD family of the tautomerase superfamily and could represent a new subfamily.

Reaction of cis-3-chloroacrylic acid dehalogenase with an allene substrate, 2,3-butadienoate: hydration via an enamine.

cis-3-Chloroacrylic acid dehalogenase (cis-CaaD) catalyzes the hydrolytic dehalogenation of cis-3-haloacrylates to yield malonate semialdehyde. The enzyme processes other substrates including an allene (2,3-butadienoate) to produce acetoacetate. In the course of a stereochemical analysis of the cis-CaaD-catalyzed reaction using this allene, the enzyme was unexpectedly inactivated in the presence of NaBH(4) by the reduction of a covalent enzyme-substrate bond. Covalent modification was surprising because the accumulated evidence for cis-CaaD dehalogenation favored a mechanism involving direct substrate hydration mediated by Pro-1. However, the results of subsequent mechanistic, pre-steady state and full progress kinetic experiments are consistent with a mechanism in which an enamine forms between Pro-1 and the allene. Hydrolysis of the enamine or an imine tautomer produces acetoacetate. Reduction of the imine species is likely responsible for the observed enzyme inactivation. This is the first reported observation of a tautomerase superfamily member functioning by covalent catalysis. The results may suggest that some fraction of the cis-CaaD-catalyzed dehalogenation of cis-3-haloacrylates also proceeds by covalent catalysis.

Is Nucleoredoxin a Master Regulator of Cellular Redox Homeostasis? Its Implication in Different Pathologies.

Nucleoredoxin (NXN), an oxidoreductase enzyme, contributes to cellular redox homeostasis by regulating different signaling pathways in a redox-dependent manner. By interacting with seven proteins so far, namely disheveled (DVL), protein phosphatase 2A (PP2A), phosphofructokinase-1 (PFK1), translocation protein SEC63 homolog (SEC63), myeloid differentiation primary response gene-88 (MYD88), flightless-I (FLII), and calcium/calmodulin-dependent protein kinase II type alpha (CAMK2A), NXN is involved in the regulation of several key cellular processes, including proliferation, organogenesis, cell cycle progression, glycolysis, innate immunity and inflammation, motility, contraction, protein transport into the endoplasmic reticulum, neuronal plasticity, among others; as a result, NXN has been implicated in different pathologies, such as cancer, alcoholic and polycystic liver disease, liver fibrogenesis, obesity, Robinow syndrome, diabetes mellitus, Alzheimer's disease, and retinitis pigmentosa. Together, this evidence places NXN as a strong candidate to be a master redox regulator of cell physiology and as the hub of different redox-sensitive signaling pathways and associated pathologies. This review summarizes and discusses the current insights on NXN-dependent redox regulation and its implication in different pathologies.

Crystal structures of native and inactivated cis-3-chloroacrylic acid dehalogenase: Implications for the catalytic and inactivation mechanisms.

The isomeric mixture of cis- and trans-1,3-dichloropropene constitutes the active component of a widely used nematocide known as Telone II®. The mixture is processed by various soil bacteria to acetaldehyde through the 1,3-dichloropropene catabolic pathway. The pathway relies on an isomer-specific hydrolytic dehalogenation reaction catalyzed by cis- or trans-3-chloroacrylic acid dehalogenase, known respectively as cis-CaaD and CaaD. Previous sequence analysis and crystallographic studies of the native and covalently modified enzymes identified Pro-1, His-28, Arg-70, Arg-73, Tyr-103, and Glu-114 as key binding and catalytic residues in cis-CaaD. Mutagenesis of these residues confirmed their importance to the dehalogenation reaction. Crystal structures of the native enzyme (2.01Å resolution) and the enzyme covalently modified at the Pro-1 nitrogen by 2-hydroxypropanoate (1.65Å resolution) are reported here. Both structures are at a resolution higher than previously reported (2.75Å and 2.1Å resolution, respectively). The conformation of the covalent adduct is strikingly different from that previously reported due to its interaction with a 7-residue loop (Thr-32 to Leu-38). The participation of another active site residue, Arg-117, in catalysis and inactivation was also examined. The implications of the combined findings for the mechanisms of catalysis and inactivation are discussed.

Phytoestrogen treatment induces testis alterations in dogs. Potential use in population control.

Dog overpopulation is considered a human health risk; they are the terrestrial vector of rabies and reservoirs for other human diseases. Surgical neutering and intratesticular injections have been used in male dogs. Physiological and morphological alterations in reproductive organs can be induced by phytoestrogens. Our goal was to evaluate the effect of oral coumestrol on dog ejaculates and testis histology. Two groups of 5 healthy adult dogs were used. One coumestrolcontaining biscuit was given once a week for a 4 week period to the experimental group. Ejaculates were obtained and evaluated. After treatment, testis were obtained and processed for histology. Compared to controls, treated dogs have reduced tubules (462 +/- 1.4 vs 336 +/- 2 micron(2)), spermatogenic epithelium (49.1 +/- 0.01 vs 13.3 +/- 0.01 micron(2)), and lumen opening (891 +/- 1.4 vs 530 +/- 26.9 micron). Ejaculates from treated animals have increased numbers of abnormal spermatozoa and reduced sperm concentration.

Characterization of Cg10062 from Corynebacterium glutamicum: implications for the evolution of cis-3-chloroacrylic acid dehalogenase activity in the tautomerase superfamily.

A 149-amino acid protein designated Cg10062 is encoded by a gene from Corynebacterium glutamicum. The physiological function of Cg10062 is unknown, and the gene encoding this protein has no obvious genomic context. Sequence analysis links Cg10062 to the cis-3-chloroacrylic acid dehalogenase ( cis-CaaD) family, one of the five known families of the tautomerase superfamily. The characterized tautomerase superfamily members have two distinctive characteristics: a beta-alpha-beta structure motif and a catalytic amino-terminal proline. Pro-1 is present in the Cg10062 amino acid sequence along with His-28, Arg-70, Arg-73, Tyr-103, and Glu-114, all of which have been implicated as critical residues for cis-CaaD activity. The gene for Cg10062 has been cloned and the protein overproduced, purified, and subjected to kinetic and mechanistic characterization. Like cis-CaaD, Cg10062 functions as a hydratase: it converts 2-oxo-3-pentynoate to acetopyruvate and processes 3-bromopropiolate to a species that inactivates the enzyme by acylation of Pro-1. Kinetic and (1)H NMR spectroscopic studies also show that Cg10062 processes both isomers of 3-chloroacrylic acid at low levels with a clear preference for the cis isomer. Pro-1 is critical for the dehalogenase and hydratase activities because the P1A mutant no longer catalyzes either reaction. The presence of the six key catalytic residues and the hydratase activity coupled with the absence of an efficient cis-CaaD activity and the lack of isomer specificity implicate factors beyond this core set of residues in cis-CaaD catalysis and specificity. This work sets the stage for in-depth mechanistic and structural studies of Cg10062, which could identify the additional features necessary for a fully active and highly specific cis-CaaD. Such results will also shed light on how cis-CaaD emerged in the tautomerase superfamily because Cg10062 could be characteristic of an intermediate along the evolutionary pathway for this dehalogenase.

Structural and mechanistic analysis of trans-3-chloroacrylic acid dehalogenase activity.

Trans-3-chloroacrylic acid dehalogenase (CaaD) is a critical enzyme in the trans-1,3-dichloropropene (DCP) degradation pathway in Pseudomonas pavonaceae 170. This enzyme allows bacteria to use trans-DCP, a common component in commercially produced fumigants, as a carbon source. CaaD specifically catalyzes the fourth step of the pathway by cofactor-independent dehalogenation of a vinyl carbon-halogen bond. Previous studies have reported an X-ray structure of CaaD under acidic conditions with a covalent modification of the catalytic betaPro1 residue. Here, the 1.7 A resolution X-ray structure of CaaD under neutral (pH 6.5) conditions is reported without the presence of the covalent adduct. In this new structure, a substrate-like acetate molecule is bound within the active site in a position analogous to the putative substrate-binding site. Additionally, a catalytically important water molecule was identified, consistent with previously proposed reaction schemes. Finally, flexibility of the catalytically relevant side chain alphaGlu52 is observed in the structure, supporting its role in the catalytic mechanism.

Fluorescence and inmunohistological detection of estrogen receptors in dog testis and epidydimis after oral coumestrol administration.

INTRODUCTION: Estrogens are well recognized as important hormones in male reproduction and act as ligands to alpha and beta estrogen receptors. Both estrogen receptors could interact with estrogen-mimicking compounds such as the fluorescent phytoestrogen coumestrol, which acts both in an agonist or antagonist fashion. OBJECTIVE: To investigate the presence of Coumestrol-Estrogen Receptor complexes by fluorescence in testis and epididymis, its effect in the ER expression by immunostain in the same tissues and the effect of this binding in the testis histological characteristics. DESIGN: Adult healthy and sexually active dogs were assigned to either the experimental or control group .Coumestrol impregnated dog biscuits were given to each animal from the experimental group once a week for a 4 week period. The control group received a biscuit with no Coumestrol, also once a week and for the same period. Testis morphology, ER immunodetection, and coumestrol-receptor binding were evaluated. SETTING: The experiment was done in the facilities of the Mexico City canine shelter. Animals were caged individually with food and water ad libitum and having at least two daily hours for exercise. RESULTS: Morphological alterations in testis after oral administration of coumestrol were detected. The main alterations include decreased germinal epithelium in tubule, and the loss of a continuous proliferation and differentiation gamete layer. Fluorescence signals in testis interstitial Leydig cells and epididymus indicating ER-coumestrol complexes were detected at the same points to those Immunohystochemically detected ER. CONCLUSIONS: Coumestrol administration induces testis alterations and coumestrol-ER complexes can be co-localized by binding-enhanced fluorescence and immunoprecipitation.

[Genetic study of the vasovagal syncope associated to the Arg389Gly polymorphism of the beta1 adrenergic receptor]. / Estudio genético del síncope vasovagal asociado al polimorfismo Arg389Gly del receptor adrenérgico beta1.

The purpose of this study was to evaluate the correlation between the vasovagal syncope (VVS) and the beta1 adrenergic receptor polymorphism at the 389 position. Seventy individuals with VVS were selected. DNA was extracted from peripheral blood by salting out and subjected to the amplification-restriction test. Genotype identification was made by polyacrylamide gel electrophoresis. A higher frequency in genotype and allele frequencies were found in individuals with positive tilted table test respect individuals with negative test, as well as a marked preference of the GlyGly phenotype in women. Genotype Arg389Gly was the most frequent between individuals with positive response in passive phase with respect to those in the induced phase. When the genotype was analyzed based on the hemodynamic response (VASIS) a gradient is observed in the frequency of Arg389Gly with the highest major frequency in the cardio-inhibitory response followed by the mixed response, and finally the vasodepressor response. These results suggest that the SVV has a genetic component associated with the Arg389Gly polymorphism of the adrenergic receptor. The Gly allele has a high risk association and it is maintained in the population through heterozygosis.

Protease resistance of food proteins: a mixed picture for predicting allergenicity but a useful tool for assessing exposure.

BACKGROUND: Susceptibility to pepsin digestion of candidate transgene products is regarded an important parameter in the weight-of-evidence approach for allergenicity risk assessment of genetically modified crops. It has been argued that protocols used for this assessment should better reflect physiological conditions encountered in representative food consumption scenarios. AIM: To evaluate whether inclusion of more physiological conditions, such as sub-optimal and lower pepsin concentrations, in combination with pancreatin digestion, improved the performance of digestibility protocols used in characterization of protein stability. METHODS: Four pairs of established allergens and their related non/weakly-allergenic counterparts (seed albumins, muscle tropomyosins, plant lipid transfer proteins [LTP] and collagens) plus fish parvalbumin, were subjected to nine combinations of pH (1.2-2.5-4.0) and pepsin-to-protein ratio (PPR: 10-1-0.1 U/µg) for pepsin digestion, followed by pancreatin digestion in the presence of bile salts. Digestion was monitored by SDS-PAGE in conjunction with Coomassie staining and immunoblotting using rabbit antisera and human IgE. RESULTS: At pH 4.0 and at PPR 0.1 most proteins, both allergen and non-allergen, were highly resistant to pepsin. Under conditions known to favor pepsin proteolysis, the established major allergens Ara h 2, Pru p 3 and Pen a 1 were highly resistant to proteolysis, while the allergen Cyp c 1 was not. However, this resistance to pepsin digestion only made Ara h 2 and to a lesser extent Pen a 1 and Pru p 3 stand out compared to their non-allergenic counterparts. Largely irrespective of preceding pepsin digestion conditions, pancreatin digestion was very effective for all tested proteins, allergens and non-allergens, except for Cyp c 1 and bovine collagen. CONCLUSIONS: Sub-optimal pH, low pepsin-to protein ratio, and sequential pepsin and pancreatin digestion protocols do not improve the predictive value in distinguish allergens from non-allergens. Digestion conditions facilitating such distinction differ per protein pair.

Vampire bat reproductive control by a naturally occurring phytooestrogen.

Rabies transmission by wild animals has not being controlled satisfactorily. One major rabies vector to humans and cattle is the hematophagous vampire bat Desmodus rotundus whose distribution is still increasing in the Americas. Of all of the strategies currently in place to control this vector, none of them are really specific and some have ecological impacts. In the present study we used a naturally occurring phytoestrogen on a small vampire bat colony. After collection, bats were fed bovine blood containing 200 microg coumestrol for a 30-day period. After treatment, gonads were excised and processed for histological evaluation. Data indicate that coumestrol adversely affects gonad histology and has a possible impact on the fertility of both male and female vampire bats.

Influence of genetic variants of CYP2D6, CYP2C9, CYP2C19 and CYP3A4 on antiepileptic drug metabolism in pediatric patients with refractory epilepsy.

BACKGROUND: Identified the polymorphisms of CYP2D6, CYP2C9, CYP2C19 and CYP3A4, within a rigorously selected population of pediatric patients with drug-resistant epilepsy. METHOD: The genomic DNA of 23 drug-resistant epilepsy patients and 7 patients with good responses were analyzed. Ten exons in these four genes were genotyped, and the drug concentrations in saliva and plasma were determined. RESULTS: The relevant SNPs with pharmacogenomics relations were CYP2D6*2 (rs16947) decreased your activity and CYP2D6*4 (rs1065852), CYP2C19*2 (rs4244285) and CYP3A4*1B (rs2740574) by association with poor metabolizer. The strongest risk factors were found in the AA genotype and allele of SNP rs3892097 from the CYP2D6 gene, followed by the alleles A and T of SNPs rs2740574 and rs2687116, respectively from CYP3A4. The most important concomitance was between homozygous genotype AA of rs3892097 and genotype AA of rs2740574 with 78.3% in drug-resistant epilepsy patients as compared to 14.3% in control patients. CONCLUSION: The results demonstrated the important role of the CYP 3A4*1B allelic variant as risk factor for developing drug resistance and CYP2D6, CYP2C19 SNPs and haplotypes may affect the response to antiepileptic drugs.

Arg347Cys polymorphism of α1a-adrenergic receptor in vasovagal syncope. Case-control study in a Mexican population.

BACKGROUND: Vasovagal syncope is a common clinical condition, consequential to reduced cerebral blood flow resulting from a failure in cardiovascular homeostasis during orthostasis. Blood pressure regulation is the basis for syncope development. In this regulation, the α1a-adrenergic receptor plays a major role. Some studies have found a positive correlation between the Arg347Cys polymorphism of the α1a-adrenergic receptor to hypertension and heart autonomic control. The goal of this study is to evaluate the possible association between the Arg347Cys α1a-adrenergic receptor polymorphism and vasovagal syncope in a Mexican population. METHODS/MAJOR FINDINGS: A sample of 89 vasovagal syncope patients and 40 healthy controls were studied. Arg347Cys α1a-adrenergic receptor polymorphism was determined by the PCR-RFLP method. We found an increased frequency of genotype ArgArg in vasovagal syncope patients. In a logistic regression model significant associations were found in two genetic models, in codominant model (OR=13.21: CI 95% 3.69-54.99, p<0.001) and in additive model (OR=12.68: CI 95% 3.5-53.07, p<0.001) for ArgArg genotype with CysCys as reference. CONCLUSIONS: Our data suggests an important participation of Arg347Cys polymorphism as susceptibility factor in patients with vasovagal syncope. ArgArg genotype could be a marker for vasovagal syncope susceptibility in the Mexican population.