Regulation of hepatic P-gp expression and activity by genistein in rats.

P-glycoprotein (P-gp) is an ABC transporter exhibiting high pharmacotoxicological relevance by extruding a wide range of cytotoxic compounds out of the cells. Previously, we demonstrated that the phytoestrogen genistein (GNT) modulates P-gp expression in hepatocellular carcinoma in vitro. Although several beneficial effects (e.g., antioxidant, antimutagenic, anticancer) have been attributed to GNT, the molecular mechanisms have not been totally elucidated. In the present work, we evaluated the effect of GNT on P-gp expression in rat liver, kidney and ileum. We found that GNT (5 mg/kg daily s.c. 3 days) increased hepatic P-gp expression and also Mdr1a (one of the genes encoding P-gp) mRNA levels. Renal and intestinal P-gp remained unchanged after GNT treatment. Hepatic P-gp activity measured with rhodamine-123 and digoxin, both well-known P-gp substrates, was also increased. In vitro experiments using hepatocyte primary cell culture demonstrated that inhibition of ER-α with ICI182/780 did not prevent Mdr1a mRNA up-regulation by GNT (10 µM). In contrast, Mdr1a induction was suppressed after pregnane X receptor (PXR) inhibition by sulforaphane and knockdown of this nuclear receptor. These findings were confirmed in vivo by using the PXR antagonist ketoconazole. In conclusion, we demonstrated the induction of hepatic P-gp expression and activity by GNT in vivo, with PXR being a likely mediator. This suggests that GNT, at concentrations observed in plasma of individuals consuming the phytoestrogen in the diet or through supplements, could affect the clearance of relevant P-gp substrates of therapeutic use as well as toxicity of environmental and food toxicants.

RUNX3 gene polymorphisms and haplotypes in Mexican patients with colorectal cancer.

We analyzed a possible association between RUNX3 gene polymorphisms and haplotypes in Mexican patients with colorectal cancer (CRC). Genomic DNA samples were obtained from the peripheral blood of 176 Mexican patients with CRC at diagnosis and from 195 individuals that formed the control group. The polymorphisms were detected by polymerase chain reaction-restriction fragment length polymorphism. Association was estimated by odds ratio (OR). The haplotypes and linkage disequilibrium were established using the Arlequin v3.5 software. We found that the RUNX3 polymorphisms analyzed were in Hardy-Weinberg equilibrium. The RUNX3 rs2236852 AA genotype and A allele showed association with CRC (OR = 0.39, 95%CI = 0.21-0.73, P < 0.01; OR = 0.65, 95%CI = 0.49-0.87, P < 0.01, respectively), while the rs6672420, rs11249206, and rs760805 polymorphisms did not show significant association with CRC. The TA haplotype (SNPs rs760805 and rs2236852) showed an increased risk for CRC (OR = 2.52, 95%CI = 1.47-4.30, P < 0.001). In conclusion, we found that the AA genotype and A allele of rs2236852 polymorphism confer a decreased CRC risk, while the TA haplotype appears to increase the risk of CRC development in Mexican patients.

Association of MMP7-181A/G and MMP13-77A/G polymorphisms with colorectal cancer in a Mexican population.

Colorectal cancer (CRC) is characterized by enhanced expression and activity of several metalloproteinases (MMPs), including MMP13 and MMP7, which play an important role in tumor invasion and metastasis. The objective of this study was to analyze the association of functional MMP7-181A/G and MMP13-77A/G promoter polymorphisms with susceptibility to CRC in a Mexican population. Genomic DNA samples were obtained from peripheral blood of 102 CRC patients and 125 blood donors who were included as the control group. Identification of polymorphisms was based on polymerase chain reaction-restriction fragment length polymorphism methodology. The association was estimated by the odds ratio (OR) test. The results showed that MMP7-181A/G and MMP13-77A/G variants were associated with CRC. For MMP7-181A/G, the AA (P=0.02, OR=3.38, 95% confidence interval (CI)=1.16-9.84) and AG (P=0.01, OR=3.4, 95%CI=1.17-9.83) genotypes were associated with an increased risk of CRC. For MMP13-77A/G, the AA and AG genotypes were associated with CRC (AA genotype: P=0.04, OR=3.2, 95%CI=1.004-10.2; AG genotype: P=0.01, OR=4.08, 95%CI=1.3-13.07). In conclusion, AA and AG genotype carriers for both polymorphisms are at a higher risk of developing CRC in this Mexican population.

Androgen treatment effects on neurocognition in female-to-male transgender adolescents. / Efectos del tratamiento con andrógenos sobre la neurocognición en adolescentes transgénero de mujer a hombre.

INTRODUCTION: It has been hypothesized that cognitive and memory-related brain function in transgender during cross-sex hormonal treatment might be activated towards that of the subjective gender. However, research on this topic has produced inconsistent results, and to the best of our knowledge no studies have investigated neurocognitive changes in androgen-treated female-to-male (FM) transgender adolescents. SUBJECTS AND METHODS: A total of 15 FM transgender adolescents (14-17 years) underwent neuropsychological testing in order to examine the effects of androgen on visuo-spacial abilities, verbal memory language, processing speed and executive functions. We used a longitudinal design in which 10 participants were tested twice, before and after receiving 12 months of testosterone treatment. This group was also compared with 5 FM transgender adolescents off-androgen treatment. RESULTS: Participants tested before and after 12 months of androgen treatment improved significantly on processing speed in a visuo-spatial (Rey-Osterrieth complex figure test) and in a visuo-oral task (Stroop), their performance on a verbal memory task (TAVEC) and on interference (Stroop) and they exhibited lower impulsivity control (CARAS-R). On-androgen treatment adolescents exhibited worse cognitive impulsivity control than off-androgen treatment adolescents. CONCLUSIONS: The results indicate that androgen has an influence on immediate verbal memory, cognitive interference, impulsivity control and processing speed.

TITLE: Efectos del tratamiento con andrógenos sobre la neurocognición en adolescentes transgénero de mujer a hombre.Introducción. Se ha planteado la hipótesis de que la neurocognición en personas transgénero durante el tratamiento hormonal cruzado podría aproximarse a la del género subjetivo. Sin embargo, la investigación sobre este tema ha producido resultados inconsistentes y, hasta donde sabemos, ningún estudio ha investigado los cambios neurocognitivos en adolescentes transgénero de mujer a hombre (FM) tratados con andrógenos. Sujetos y métodos. Quince adolescentes transgénero FM (14-17 años) se sometieron a pruebas neuropsicológicas para examinar los efectos de los andrógenos en sus habilidades visuoespaciales, memoria verbal, velocidad de procesamiento y funciones ejecutivas. Utilizamos un diseño longitudinal en el que se evaluó a 10 participantes dos veces, antes y después de recibir, durante 12 meses, tratamiento con testosterona. Este grupo también se comparó con cinco adolescentes transgénero FM sin tratamiento con andrógenos. Resultados. Los participantes evaluados antes y después de 12 meses de tratamiento con andrógenos mejoraron significativamente en velocidad de procesamiento en una tarea visuoespacial (prueba de la figura compleja de Rey-Osterrieth) y en una tarea visual (Stroop), en una tarea de memoria verbal (test de aprendizaje verbal España-Complutense) y en interferencia (Stroop), y exhibieron un menor control de la impulsividad (test de percepción de diferencias revisado). Los adolescentes que recibieron tratamiento con andrógenos mostraron un peor control de la impulsividad cognitiva que los adolescentes que no recibieron tratamiento con andrógenos. Conclusiones. Los resultados indican que los andrógenos influyen en la memoria verbal, la interferencia cognitiva, el control de la impulsividad y la velocidad de procesamiento.

MLH1 and XRCC1 polymorphisms in Mexican patients with colorectal cancer.

DNA repair proteins maintain DNA integrity; polymorphisms in genes coding for these proteins can increase susceptibility to colorectal cancer (CRC) development. We analyzed a possible association of MLH1 -93G>A and 655A>G and XRCC1 Arg194Trp and Arg399Gln polymorphisms with CRC in Mexican patients. Genomic DNA samples were obtained from peripheral blood of 108 individuals with CRC (study group) at diagnosis and 120 blood donors (control group) from Western Mexico; both groups were mestizos. The polymorphisms were detected by PCR-RFLP. Association was estimated by calculating the odds ratio (OR). We found that the MLH1 and XRCC1 polymorphisms were in Hardy- Weinberg equilibrium. The MLH1 655A>G polymorphism in the 655G allele was associated with a 2-fold increase risk for CRC (OR = 2.04 and 95% confidence interval (95%CI) = 1.12-3.69; P < 0.01), while the MLH1 -93G>A polymorphism allele was associated with a protective effect (OR = 0.60, 95%CI = 0.40-0.89; P = 0.01 in the -93A allele and OR = 0.32, 95%CI = 0.13-0.79; P = 0.01 in the AA genotype). The XRCC1 Arg194Trp and Arg399Gln polymorphisms did not show any significant associations. In conclusion, we found that MLH1 -93G>A and 655A>G polymorphisms are associated with CRC in Mexican patients.

Enzymatic modification of chitosan with quercetin and its application as antioxidant edible films.

Quercetin, rutin, naringin, hesperidin and chrysin were tested as substrates for chloroperoxidase to produce reactive quinones to graft onto chitosan. Quercetin and rutin quinones were successfully chemically attached to low molecular weight chitosan. The quercetin-modified chitosan showed an enhancement of plastic, antioxidant and antimicrobial properties as well as of thermal degradability. Finally, chitosan-quercetin films visibly decreased enzymatic oxidation when applied to Opuntia ficus indica cladodes.

Molecular epidemiology of Mycobacterium tuberculosis in adult Filipino TB-HIV co-infected patients.

BACKGROUND: TB is the leading cause of death from a single infectious disease, particularly among people living with HIV (PLHIV). Molecular epidemiology provides information on prevalent genotypes of Mycobacterium tuberculosis and disease transmission dynamics, which aid in TB control. Identification of mutations that confer drug resistance is essential for the rapid diagnosis of drug-resistant TB, especially in high TB burden settings, like the Philippines.METHODS: This study aimed to determine mutations in M. tuberculosis drug resistance-conferring genes and circulating genotypes in PLHIV. MIRU-VNTR (mycobacterial interspersed repetitive unit-variable number of tandem repeats) typing using a set of 24-loci and sequencing of drug resistance-conferring genes were performed in 22 M. tuberculosis isolates from TB-HIV co-infected patients.RESULTS: The prevalence of resistance to any drug was 31.8%, 18.2% for isoniazid monoresistance, 4.5% for streptomycin monoresistance and 9.1% for multidrug resistance. The identified mutations in the katG, rpoB, pncA, rpsL and gyrA genes have been reported in the literature; none was found in the inhA and embB genes. All isolates belonged to the EAI2-Manila family and were grouped into four clusters based on their phenotypic drug resistance and mutation profiles.CONCLUSION: The use of 24-loci set may be used as a more discriminatory MIRU-VNTR typing in settings where the East African-Indian lineage is predominant, like the Philippines.

[Caring for unaccompanied minors: an experience from Lausanne]. / Soins aux mineurs non accompagnés (MNA): l'expérience lausannoise.

Medical treatment of an unaccompanied minor is made more complicated firstly by its connections with the politics of immigration and secondly by the difficulty in gaining recognition of the priority of the minor's interests. Enabling healthcare teams to travel and meet these particularly vulnerable youths makes medical care more accessible to them and facilitates an optimal bio-psycho-social treatment. For most of these adolescents it is their lack of plans for the future which remains the major obstacle to their development and mental and physical health.

Evaluation of novel particles as an inhalation system for GLP-1.

AIM: The feasibility of administering native glucagon-like peptide 1 (GLP-1) as GLP-1 Technosphere Inhalation Powder for diabetes therapy has been demonstrated in a rat model. METHODS: GLP-1 Technosphere Inhalation Powders containing 5, 10 and 15% GLP-1 were prepared and administered to healthy female Sprague-Dawley rats and to male Zucker diabetic obese rats. Rats received a single dose of GLP-1 Technosphere Powder by pulmonary insufflation. GLP-1 pharmacokinetic and pharmacodynamic responses were measured. RESULTS: Maximum circulating GLP-1 concentrations were achieved at approximately 10 min after dosing with detectable levels at 40 min. In a food consumption study, Sprague-Dawley rats receiving GLP-1 Technosphere Powder once-daily consumed less food than control rats for up to 24 h after dosing. Cumulative food consumption was decreased approximately 10% after 78 h. In an intraperitoneal glucose tolerance test, Zucker diabetic fatty rats receiving 2 mg GLP-1 Technosphere Powder (0.3 mg GLP-1) by pulmonary insufflation exhibited lower glucose concentrations and higher insulin concentrations than control rats. Pancreatic evaluations showed no differences in apoptotic index or cell proliferation of beta-cells. In addition, a dose-related increase in insulin expression within the pancreas was observed. CONCLUSIONS: These data demonstrate the feasibility of administering native GLP-1 as GLP-1 Technosphere Inhalation Powder for diabetes therapy.

[Implementation of the guidelines for the introduction of new drugs (GINF) in Andalusian hospitals]. / Implantación de la guía para la incorporación de nuevos fármacos (GINF) en los hospitales andaluces.

OBJECTIVE: To measure the level of implementation of the GINF (guidelines for the introduction of new drugs) in Andalusian hospitals, describe the characteristics of this implementation and analyse if any of the hospital s dependent variables could influence these characteristics. METHOD: A telephone survey was carried out in the hospitals included in the Department of Health list. The survey consisted of 11 closed questions on different variables in the hospital and the GINF use profile, and an open question about the improvements carried out and proposals for improvement. The results were analysed according to the type of hospital (category, training, geographical location) in order to detect possible differences. RESULTS: A target population of 31 hospitals was identified. The survey was carried out in 29 of these; the level of implementation was 96.5% in the responding hospitals. 23 hospitals used the GINF for 100% of drugs, 6 had carried out local modifications and 80% made proposals for improvement. Significant differences were found in the implementation of the GINF according to resident/intern pharmacist training (p = 0.049), the geographical location (p = 0.004) and the hospital category (p < 0.001). CONCLUSIONS: The GINF have been implemented in almost all public Andalusian hospitals as the guidelines for requesting new drugs. Very few local modifications have been carried out to the guidelines, although numerous proposals for improvement have been made. Differences in use have been identified (No. of drugs, different versions) according to the hospital characteristics (location, training and complexity classification). They are considered a useful tool and influence the drug selection process, in particular in training hospitals with a higher classification.

Nest use and patterns of egg laying and damage by 4 strains of laying hens in an aviary system.

Laying hens are strongly motivated to use nests for egg laying, and alternative production systems (e.g., aviaries) provide artificial sites to meet this need and ensure efficient collection of clean, undamaged eggs. However, nests are typically not provided to allow simultaneous use by all hens; therefore, competition or mislaid eggs can result. To understand the influence of strain on laying eggs outside nests and damage to eggs, we compared daily patterns of nests use and egg laying among 4 laying hen strains (Hy-Line Brown (HB), Bovans Brown (BB), DeKalb White (DW), and Hy-Line W36 (W36)). Hens were observed over 3 consecutive days in aviaries with colony nests in the enclosure's top tier (2 nests/unit, 4 aviary units/strain, 144 hens/unit). The number and location of hens in nests and the number, location and condition of eggs throughout aviaries were recorded. Most eggs (90 to 95%) were laid in nests; however, brown hens consistently laid more non-nest eggs and damaged more eggs than white hens (P ≤ 0.05). Higher nest occupancy by brown hens was correlated with more non-nest and damaged eggs (P ≤ 0.05). In the morning, brown hens occupied more nest space and laid more nest eggs than white hens (e.g., HB vs. DW: 82.97 and 34.66% of space; 91.35 and 68.73% of nest eggs; P ≤ 0.05). At midday, white hens occupied more nest space and laid more nest eggs than brown hens (e.g., HB vs. DW: 28.47 and 15.81% of space; 27.39 and 8.29% of nest eggs; P ≤ 0.05). Brown hens preferred right nest compartments and laid more eggs there, whereas white hens preferred left compartments and W36 laid more eggs there (P ≤ 0.05). These findings indicate that different strains of hens have different patterns of nest use and laying behavior. In brown hens, heavy morning nest use was related to laying eggs outside nests and more damaged eggs, suggesting insufficient space for oviposition in nests. Specific facility design should be matched to hens' preferences to accommodate behavioral needs of different strains.

Effect of Helicobacter pylori eradication therapy in rosacea patients.

OBJECTIVE: The causal relation between rosacea and Helicobacter pylori infection is discussed. We evaluated the clinical evolution of rosacea after infection eradication. PATIENTS AND METHODS: We have prospectively studied 44 patients diagnosed with rosacea. Helicobacter pylori infection was determined, and infected patients were treated with eradication therapy. The evolution of dermatological symptoms in a subgroup of 29 infected patients in whom eradication had been achieved was followed during 16.8 (+/- 17.8) months. Median age was 50.6 (+/- 14.1) years for 22 women (75.9%) and 7 men (24.1%). Clinical response according to gender and clinical subtype of rosacea was evaluated. RESULTS: Complete improvement was observed in 10 patients (34.5%; 95% CI: 18.6-54.3%), relevant improvement in 9 (31.1%; 95% CI: 16-51%), poor improvement in 5 (17.2%; 95% CI: 6.5-36.4%), and absence of improvement in 5 cases (17.2%; 95% CI: 6.5-36.4%). No significant differences in dermatological evolution according to sex were observed. Regarding subtype of rosacea there was a relevant improvement in 83.3% (95% CI: 64.1-93.8%) of cases with papulopustular type as opposed to 36.5% (95% CI: 20-56.1%) of cases with erythematous predominance, p = 0.02. CONCLUSIONS: Based on these results, the relation between Helicobacter pylori and rosacea is supported, and infection should be investigated in these patients because an appreciable percentage of patients diagnosed with rosacea and Helicobacter pylori infection can benefit from eradication therapy, mainly in the papulopustular subtype.

Pharmacokinetics of prednisolone in man during acute and chronic exposure to high altitude.

INTRODUCTION: The exposure to high altitude (H) produces several physiologic alterations which may induce changes in the pharmacokinetics of drugs. This hypothesis has been confirmed in previous studies which suggest that drugs which are highly bound to plasma proteins are most likely to exhibit altered pharmacokinetics. OBJECTIVES: To further elucidate the influence of H on pharmacokinetics, prednisolone was selected, since it is highly bound to plasma proteins, renally excreted and poorly bound to red blood cells. SUBJECTS, MATERIALS AND METHODS: Prednisolone (80 mg) was given orally to three groups of young healthy volunteers. One group was residing at sea level (L): the same volunteers were studied again after 15 hours of exposure to high altitude (3600 m, HA group), and volunteers living at H for at least six months (group HC). RESULTS: There were no statistically significant differences in the pharmacokinetic parameters calculated from plasma data in the three situations studied. When calculated from whole blood data, however, AUC and Cmax were increased and both volume of distribution and clearance diminished after exposure to H, either acute or chronically. Binding to proteins increased significantly after H exposure from 57% in group L to 75% and 94% in group H and HC, respectively. Binding to erythrocytes also increased with H exposure from 43.7% in group L to 50.6% and 61.6% in group HA and HC, respectively. The prednisolone/prednisone ratio in urine was 11.1 in group L, 7.3 in group HA and 45.6 in group HC. CONCLUSION: Since prednisone has very little intrinsic glucocorticoid activity and has to be converted to prednisolone for therapeutic effect, the alteration of the prednisolone/prednisone ratio, as a result of high altitude exposure could be clinically relevant. Additional experiments are desirable to further evaluate this observation.

The chemical nature of the main central excitatory transmitter: a critical appraisal based upon release studies and synaptic vesicle localization.

The chemical nature of the central transmitter responsible for fast excitatory events and other related phenomena is analysed against the historical background that has progressively clarified the structure and function of central synapses. One of the problems posed by research in this field has been whether one or more of the numerous excitatory substances endogenous to the brain is responsible for fast excitatory synaptic transmission, or if such a substance is, or was, a previously unknown one. The second question is related to the presence in the CNS of three main receptor types related to fast excitatory transmission, the so-called alpha-amino-3-hydroxy-5-methylisoxazole propionic acid, kainate and N-methyl-D-aspartate receptors. This implies the possibility that each receptor type might have its own endogenous agonist, as has sometimes been suggested. To answer such questions, an analysis was done of how different endogenous substances, including L-glutamate, L-aspartate, L-cysteate, L-homocysteate, L-cysteine sulfinate, L-homocysteine sulfinate, N-acetyl-L-aspartyl glutamate, quinolinate, L-sulfoserine, S-sulfo-L-cysteine, as well as possible unknown compounds, were able to fulfil the more important criteria for transmitter identification, namely identity of action, induced release, and presence in synaptic vesicles. The conclusion of this analysis is that glutamate is clearly the main central excitatory transmitter, because it acts on all three of the excitatory receptors, it is released by exocytosis and, above all, it is present in synaptic vesicles in a very high concentration, comparable to the estimated number of acetylcholine molecules in a quantum, i.e. 6000 molecules. Regarding a possible transmitter role for aspartate, for which a large body of evidence has been presented, it seems, when this evidence is carefully scrutinized, that it is either inconclusive, or else negative. This suggests that aspartate is not a classical central excitatory transmitter. From this analysis, it is suggested that the terms alpha-amino-3-hydroxy-5-methylisoxazole propionic acid, kainate and N-methyl-D-aspartate receptors, should be changed to that of glutamate receptors, and, more specifically, to GLUA, GLUK and GLUN receptors, respectively. When subtypes are described, a Roman numeral may be added, as in GLUNI, GLUNII, and so on.

A study in rat brain cortex synaptic vesicles of endogenous ligands for N-methyl-D-aspartate receptors.

The presence of endogenous ligands for the N-methyl-D-aspartate receptor was looked for in highly purified rat brain cortex synaptic vesicles, the contents of which were extracted and fractionated by gel filtration on Sephadex G-10, or by three different high-voltage electrophoresis procedures. The presence of endogenous ligands was detected by their ability to compete with 50 nM L-[3H]glutamate for binding to whole rat brain N-methyl-D-aspartate receptors. The receptor preparations used were those present in purified postsynaptic densities, in which the quisqualate receptors were blocked by 10 microM quisqualate. Synaptic vesicles had a high content of N-methyl-D-aspartate receptor ligands, which on fractionation always coincided with glutamate or aspartate. A variable and very small amount of a highly acidic endogenous ligand was also found. The latter substance did not coincide in the electrophoresis with homocysteic, cysteic, quinolinic, cysteine sulphinic or homocysteine sulphinic acids, or with N-acetyl-aspartyl-glutamic acid, S-sulphocysteine or sulphoserine. We also found that a single centrifugation, in 0.25 M sucrose, 25 mM Tris-citrate, pH 7.1, of purified synaptic vesicles, at 135,000 gmax for 45 min, led to a 51% loss of endogenous glutamate, but did not change their aspartate content. Thus, in uncentrifuged vesicles the glutamate/aspartate ratio was 9.4, while in centrifuged ones the ratio was 3.9 ATP markedly enhanced L-[3H]glutamate uptake into synaptic vesicles, but did not change the binding of L-[3H]aspartate. Differences in labelled aspartate and glutamate efflux from the vesicles were also found.(ABSTRACT TRUNCATED AT 250 WORDS)

Glycoprotein synthesis at the synapse: fractionation of polypeptides synthesized within isolated dendritic fragments by concanavalin A affinity chromatography.

The synthesis of glycosylated proteins at postsynaptic sites was evaluated by combining metabolic labeling of isolated pinched-off dendritic fragments (synaptodendrosomes) with glycoprotein isolation by Con A affinity chromatography. Three major labeled proteins were detected (apparent molecular weights of 128, 42 and 19 kDa) along with seven minor polypeptides. Treatment of the glycoprotein fraction with N-glycosidase F led to shift in the apparent molecular weight of the bands. Also, label incorporation into glycoprotein species was blocked by tunicamycin. Thus, the three prominent polypeptides and most of the minor components of this fraction corresponded to bona fide N-glycoproteins. Incubation of synaptodendrosomes with cycloheximide also inhibited label incorporation into the isolated glycoproteins, indicating that the labeling resulted from local de novo synthesis. Subcellular fractionation revealed that the labeled glycoproteins were present in soluble and particulate fractions, mainly microsomes and synaptic membranes, and one of the species (42 kDa) appeared in the incubation medium, indicating secretion. In addition, these glycoproteins were dissimilarly distributed in several brain regions, and were expressed differentially during development, reaching their highest level of synthesis during the period of synaptogenesis. These results provide evidence for local dendritic synthesis of particular glycoprotein components of the synapse.