Enantioselective Determination of Mexiletine and Its Metabolites p-Hydroxymexiletine and Hydroxymethylmexiletine in Rat Plasma by Normal-Phase Liquid Chromatography-Tandem Mass Spectrometry: Application to Pharmacokinetics

Mexiletine (MEX), hydroxymethylmexiletine (HMM) and P-hydroxy-mexiletine (PHM) were analyzed in rat plasma by LC-MS/MS. The plasma samples were prepared by liquid-liquid extraction using methyl-tert-butyl ether as extracting solvent. MEX, HMM, and PHM enantiomers were resolved on a Chiralpak (R) AD column. Validation of the method showed a relative standard deviation (precision) and relative errors (accuracy) of less than 15% for all analytes studied. Quantification limits were 0.5 ng ml(-1) for the MEX and 0.2 ng ml(-1) for the HMM and PHM enantiomers. The validated method was successfully applied to quantify the enantiomers of MEX and its metabolites in plasma samples of rats (n = 6) treated with a single oral dose of racemic MEX. Chirality 21:648-656, 2009. (C) 2008 Wiley-Liss, Inc.

Respiratory Infection, Exposure to Mouse Allergen and Breastfeeding: Role in Recurrent Wheezing in Early Life

Background: Environmental factors may influence the development of allergen sensitization and asthma. The aim of this study was to evaluate the role of endotoxin and allergen exposure in early life as a risk factor for recurrent wheezing. Methods: One hundred and four infants from low-income families, at high risk of asthma, were enrolled at birth. Dust samples were collected from the bedding and bedroom floor within 6 months after birth. Recurrent wheezing was defined as 3 or more wheezing episodes in the past year. Endotoxin was determined by Limulus amebocyte lysate assay, and major indoor allergens were quantitated by ELISA in dust extracts. IgE antibodies were measured by ImmunoCAP at 30 months of age. Results: At 30 months, 51 of the 99 infants who completed the study (51.5%) had recurrent wheezing. Respiratory infection was strongly associated with recurrent wheezing (OR 6.67, 95% CI 1.96-22.72), whereas exclusive breastfeeding for at least 1 month was a protective factor (OR 0.09, 95% CI 0.01-0.51). Exposure to high levels of mouse allergen was more frequent among non-recurrent wheezers, approaching significance (OR 0.12, 95% CI 0.01-1.13; p=0.064). None of the children were sensitized to mouse. Sensitization to mite was found in 26/90 (28.8%) children, with no association with recurrent wheezing. Conclusion: Respiratory infection was strongly associated with recurrent wheezing in the first 30 months of life, in children at high risk of asthma, living in a socially deprived community in Brazil. Copyright (C) 2009 S. Karger AG, Basel

Obese Patients Have Stronger Peristalsis and Increased Acid Exposure in the Esophagus

Obesity is a risk factor for GERD and a potential modulator of esophageal motility. To assess whether obese patients differ from non-obese patients in terms of esophageal motility and reflux. Patients (n = 332) were categorized in GERD and controls after clinical assessment, esophageal manometry, and pH monitoring. Non-obese (BMI 16-29.9) and obese (BMI 30-68) were compared in regard of distal esophageal amplitude (DEA), LES pressure (LESP), manometric diagnosis, and esophageal acid exposure (EAE). Obese showed higher DEA in both controls (122 +/- A 53 vs. 97 +/- A 36 mmHg, p = 0.041) and GERD patients (109 +/- A 38 vs. 94 +/- A 46 mmHg, p < 0.001), higher LESP in GERD patients (20.5 +/- A 10.6 vs. 18.2 +/- A 10.6 mmHg, p = 0.049), higher frequency of nutcracker esophagus in controls (30 vs. 0%, p = 0.001), lower frequency of ineffective motility in GERD patients (6 vs. 20%, p = 0.001), and higher EAE in both controls [total EAE: 1.6% (0.7-5.1) vs. 0.9% (0.2-2.4), p = 0.027] and GERD patients [upright EAE: 6.5% (3.8-11.1) vs. 5.2% (1.5-10.6), p = 0.048]. Multiple linear regression showed that BMI was associated either with EAE (p < 0.001), DEA (p = 0.006), or LESP (in men, p = 0.007). Obese patients differed from non-obese in terms of esophageal motility and reflux, regardless of the presence of GERD. Obese patients showed stronger peristalsis and increased acid exposure in the esophagus.

Antibody-targeted horseradish peroxidase associated with indole-3-acetic acid induces apoptosis in vitro in hematological malignancies

Indole-3-acetic acid (IAA), when oxidized by horseradish peroxidase (HRP), is transformed into cytotoxic molecules capable of inducing cell injury. The aim of this study was to test if, by targeting hematopoietic tumors with HRP-conjugated antibodies in association with IAA treatment, there is induction of apoptosis. We used two lineages of hematologic tumors: NB4, derived from acute promyelocytic leukemia (APL) and Granta-519 from mantle cell lymphoma (MCL). We also tested cells from 12 patients with acute myeloid leukemia (AML) and from 10 patients with chronic lymphocytic leukemia (CLL). HRP targeting was performed with anti-CD33 or anti-CD19 antibodies (depending on the origin of the cell), followed by incubation with goat anti-mouse antibody conjugated with HRP. Eight experimental groups were analyzed: control, HRP targeted, HRP targeted and incubated with 1, 5 and 10 mM IAA, and cells not HRP targeted but incubated with 1, 5 and 10 mM IAA. Apoptosis was analyzed by flow cytometry using annexin V-FITC and propidium iodide labeling. Results showed that apoptosis was dependent on the dose of IAA utilized, the duration of exposure to the prodrug and the origin of the neoplasia. Targeting HRP with antibodies was efficient in activating IAA and inducing apoptosis. (C) 2010 Elsevier Ltd. All rights reserved.

Sensitisation to aeroallergens in Brazilian adolescents living at the periphery of large subtropical urban centres

Objectives: To evaluate the sensitization to aeroallergens determined by skin prick test (SPT) in Brazilian adolescents, and to correlate its positivity with the diagnosis of asthma and/or rhinitis based on the written questionnaire (WQ) of ISAAC phase III study. Patients and Methods: A total of 996 adolescents (387 boys) were selected by systematic samples. A standard allergen extracts panel (positive/negative control, D pteronyssinus [Dpt], P americana [Pal, B germanica [Bg], dog, cat, fungal and grass mix) was used and its positivity compared with positive responses to asthma, rhinitis or both. Results: Positive SPT to at least one allergen was observed in 466 adolescents (46.8 %), with sensitisation to Dpt in 79.1 %. Positivity to more than one allergen occurred in 232 students (49.8 %). The frequency of positive SPTs was significantly higher among adolescents with asthma (OR = 2.16), rhinitis (OR = 1.69), and asthma and rhinitis (OR = 2.03). Positive SPT to four or more allergens were higher among asthmatics (OR = 2.6) and among adolescents with asthma and rhinitis (OR = 3). Conclusions: A high sensitisation rate to aeroallergens was observed, significantly higher among those with asthma, rhinitis or a combination of both, especially in multiple sensitisations.

Salicylates dilate blood vessels through inhibiting PYK2-mediated RhoA/Rho-kinase activation

Aims Compared with other non-steroid anti-inflammatory drugs (NSAIDs), aspirin is not correlated to hypertension. It has been shown that aspirin has unique vasodilator action in vivo, offering an explanation for the unique blood pressure effect of aspirin. In the present study, we investigate the mechanism whereby salicylates (aspirin and sodium salicylate) dilate blood vessels. Methods and results Rat aortic or mesenteric arterial rings were used to test the vascular effect of salicylates and other NSAIDs. RhoA translocation and the phosphorylation of MYPT1, the regulatory subunit of myosin light chain phosphatase, were measured by western blot, as evidenced for RhoA/Rho-kinase activation. Salicylates, but not other NSAIDs, relaxed contraction induced by most tested constrictors except for calyculin A, indicating that RhoA/Rho-kinase-mediated calcium sensitization is involved. The involvement of RhoA/Rho kinase in vasodilation by salicylates was confirmed by measurements of RhoA translocation and MYPT1 phosphorylation. The calculated half maximal inhibitory concentration (IC(50)) of vasodilation was apparently higher than that of cyclooxygenase inhibition, but comparable to that of proline-rich tyrosine kinase 2 (PYK2) inhibition. Over-expression of PYK2 induced RhoA translocation and MYPT1 phosphorylation, and these effects were markedly inhibited by sodium salicylate treatment. Consistent with the ex vitro vascular effects, sodium salicylate acutely decreased blood pressure in spontaneous hypertensive rats but not in Wistar Kyoto rats. Conclusion Salicylates dilate blood vessels through inhibiting PYK2-mediated RhoA/Rho-kinase activation and thus lower blood pressure.

Blockade of NMDA or NO in the dorsal premammillary nucleus attenuates defensive behaviors

The dorsal premammillary nucleus (PMd) is a hypothalamic structure that plays a pivotal role in the processing of predatory threats. Lesions of this nucleus virtually eliminate the expression of defensive responses to predator exposure. However, little is known about the neurotransmitters responsible for these behavioral responses. Since PMd neurons express ionotropic glutamate receptors and exposure to predators have been shown to activate nitric oxide (NO) producing cells in this region, the aim of this study was to verify the involvement of glutamate and NO-mediated neurotransmission in defensive reactions modulated by the PMd. We tested in male Wistar rats the hypothesis that intra-PMd injection of the NMDA receptor antagonist, AP7, or the NO synthase inhibitor, N-propyl-L-arginine (NP), would attenuate behavioral responses induced by cat exposure. Our results showed that both AP7 and NP significantly attenuated the behavioral responses induced by the live cat. These results suggest that the NMDA/NO pathway plays an important role in the behavioral responses mediated by the PMd. (C) 2011 Elsevier Inc. All rights reserved.