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1.
Biochem Biophys Res Commun ; 482(4): 1013-1018, 2017 Jan 22.
Artículo en Inglés | MEDLINE | ID: mdl-27908722

RESUMEN

Maternal diabetes is one of major causes of congenital malformations in offspring, but the underlying mechanism is still unclear. MiRNAs play an important role in transcriptional and post-transcriptional regulation of gene expression. However, no miRNA expression profiling of hyperglycemic offspring are thus far available. Female mice were made diabetic with streptozotocin, treated with slow-release insulin tablets, and mated. MiRNA expression profiling with Next Generation Sequencing on the SOLiD5 platform was performed on 8 control and 5 hyperglycemic embryonic day (ED)8.5 and 9 control and 6 hyperglycemic ED9.5 embryos. Differential expression was analyzed with the Wald test. On ED8.5, the abundance of expressed miRNAs was similar in control and hyperglycemic ED8.5 embryos. The spectrum of expressed miRNAs had not changed in ED9.5 embryos, but the abundance of most miRNAs increased ∼5-fold in control embryos. However, hyperglycemic D9.5 embryos were unable to mount this increase in prevalence. Only 3 miRNAs were differentially expressed in control and hyperglycemic ED9.5 embryos, but their putative target genes were underrepresented in the Jackson database of genes causing cardiovascular or neural malformations.


Asunto(s)
Regulación del Desarrollo de la Expresión Génica , MicroARNs/genética , Embarazo en Diabéticas , Somitos/embriología , Animales , Embrión de Mamíferos/metabolismo , Femenino , Hiperglucemia/etiología , Hiperglucemia/genética , Masculino , Ratones , Embarazo , Embarazo en Diabéticas/etiología , Somitos/metabolismo , Transcriptoma
2.
Neurogastroenterol Motil ; 30(6): e13299, 2018 06.
Artículo en Inglés | MEDLINE | ID: mdl-29383802

RESUMEN

BACKGROUND: Irritable bowel syndrome (IBS) is a common gastrointestinal disorder associated with altered gastrointestinal microflora and increased nociception to colonic distension. This visceral hypersensitivity can be reversed in our rat maternal separation model by fungicides. Menthacarin® is a proprietary combination of essential oils from Mentha x piperita L. and Carum carvi. Because these oils exhibit antifungal and antibacterial properties, we investigated whether Menthacarin® can reverse existing visceral hypersensitivity in maternally separated rats. METHODS: In non-handled and maternally separated rats, we used the visceromotor responses to colorectal distension as measure for visceral sensitivity. We evaluated this response before and 24 hours after water-avoidance stress and after 7 days treatment with Menthacarin® or control. The pre- and post-treatment mycobiome and microbiome were characterized by sequencing of fungal internal transcribed spacer 1 (ITS-1) and bacterial 16s rDNA regions. In vitro antifungal and antimicrobial properties of Menthacarin® were studied with radial diffusion assay. KEY RESULTS: Menthacarin® inhibited in vitro growth of yeast and bacteria. Water-avoidance caused visceral hypersensitivity in maternally separated rats, and this was reversed by treatment. Multivariate analyses of ITS-1 and 16S high throughput data showed that maternal separation, induced changes in the myco- and microbiome. Menthacarin® treatment of non-handled and maternally separated rats shifted the mycobiomes to more similar compositions. CONCLUSIONS & INFERENCES: The development of visceral hypersensitivity in maternally separated rats and the Menthacarin® -mediated reversal of hypersensitivity is associated with changes in the mycobiome. Therefore, Menthacarin® may be a safe and effective treatment option that should be tested for IBS.


Asunto(s)
Hiperalgesia/tratamiento farmacológico , Micobioma/efectos de los fármacos , Aceites Volátiles/administración & dosificación , Aceites de Plantas/administración & dosificación , Dolor Visceral/tratamiento farmacológico , Animales , Animales Recién Nacidos , Antibacterianos/administración & dosificación , Antibacterianos/aislamiento & purificación , Antifúngicos/administración & dosificación , Antifúngicos/aislamiento & purificación , Combinación de Medicamentos , Hiperalgesia/microbiología , Hiperalgesia/psicología , Masculino , Privación Materna , Mentha piperita , Micobioma/fisiología , Aceites Volátiles/aislamiento & purificación , Aceites de Plantas/aislamiento & purificación , Ratas , Ratas Long-Evans , Dolor Visceral/microbiología , Dolor Visceral/psicología
3.
J Crohns Colitis ; 11(7): 831-839, 2017 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-28158397

RESUMEN

BACKGROUND AND AIM: T cells are key players in the chronic intestinal inflammation that characterises Crohn's disease. Here we aim to map the intestinal T-cell receptor [TCR] repertoire in patients with Crohn's disease, using next-generation sequencing technology to examine the clonality of the T-cell compartment in relation to mucosal inflammation and response to therapy. METHODS: Biopsies were taken from endoscopically inflamed and uninflamed ileum and colon of 19 patients with Crohn's disease. From this cohort, additional biopsies were taken after 8 weeks of remission induction therapy from eight responders and eight non-responders. Control biopsies from 11 patients without inflammatory bowel disease [IBD] were included. The TCRß repertoire was analysed by next-generation sequencing of biopsy RNA. RESULTS: Both in Crohn's disease patients and in non-IBD controls, a broad intestinal T-cell repertoire was found, with a considerable part consisting of expanded clones. Clones in Crohn's disease were more expanded [p = 0.008], with the largest clones representing up to as much as 58% of the total repertoire. There was a substantial overlap of the repertoire between inflamed and uninflamed tissue and between ileum and colon. Following therapy, responders showed larger changes in the T-cell repertoire than non-responders, although a considerable part of the repertoire remained unchanged in both groups. CONCLUSIONS: The intestinal T-cell repertoire distribution in Crohn's disease is different from that in the normal gut, containing profoundly expanded T-cell clones that take up a large part of the repertoire. The T-cell repertoire is fairly stable regardless of endoscopic mucosal inflammation or response to therapy.


Asunto(s)
Enfermedad de Crohn/inmunología , Enfermedad de Crohn/patología , Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T/metabolismo , Linfocitos T/patología , Adalimumab/uso terapéutico , Adulto , Antiinflamatorios/uso terapéutico , Biopsia , Budesonida/uso terapéutico , Proteína C-Reactiva/metabolismo , Estudios de Casos y Controles , Células Clonales/efectos de los fármacos , Colon/patología , Enfermedad de Crohn/tratamiento farmacológico , Femenino , Fármacos Gastrointestinales/uso terapéutico , Humanos , Íleon/patología , Inflamación/inmunología , Inflamación/patología , Infliximab/uso terapéutico , Masculino , Persona de Mediana Edad , Índice de Severidad de la Enfermedad , Linfocitos T/efectos de los fármacos , Adulto Joven
4.
Biochimie ; 88(9): 1255-64, 2006 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-16839656

RESUMEN

Glutamine synthetase (GS) is expressed at various levels in a wide range of tissues, suggesting that a complex network of modules regulates its expression. We explored the interactions between the upstream enhancer, regulatory regions in the first intron, and the 3'-untranslated region and immediate downstream genomic sequences of the GS gene (the GS "tail"), and compared the results with those obtained previously in conjunction with the bovine growth hormone (bGH) tail. The statistical analysis of these interactions revealed that the GS tail was required for full enhancer activity of the combination of the upstream enhancer and either the middle or the 3'-intron element. The GS tail also prevented a productive interaction between the upstream enhancer and the 5'-intron element, whereas the bGH tail did not, suggesting that the 5'-intron element is a regulatory element that needs to be silenced for full GS expression. Using the CMV promoter/enhancer and transfection experiments, we established that the 2.8 kb GS mRNA polyadenylation signal is approximately 10-fold more efficient than the 1.4 kb mRNA signal. Because the steady-state levels of both mRNAs are similar, the intervening conserved elements destabilize the long mRNA. Indeed, one but not all constructs containing these elements had a shorter half life in FTO-2B cells. A construct containing only 300 bases before and 100 bases after the 2.8 kb mRNA polyadenylation site sufficed for maximal expression. A stretch of 21 adenines inside this fragment conferred, in conjunction with the upstream enhancer and the 3'-part of the first intron, sensitivity of GS expression to ambient glutamine.


Asunto(s)
Regiones no Traducidas 3'/metabolismo , Glutamato-Amoníaco Ligasa/genética , Glutamina/metabolismo , Estabilidad del ARN , ARN Mensajero/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , Animales , Secuencia de Bases , Bovinos , Regulación de la Expresión Génica , Glutamato-Amoníaco Ligasa/efectos de los fármacos , Glutamato-Amoníaco Ligasa/metabolismo , Hormona del Crecimiento/metabolismo , Datos de Secuencia Molecular , ARN Mensajero/biosíntesis , Ratas
5.
Genes Brain Behav ; 8(3): 290-5, 2009 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-19170755

RESUMEN

Glutamine synthetase (GS) is a pivotal glial enzyme in the glutamate-glutamine cycle. GS is important in maintaining low extracellular glutamate concentrations and is downregulated in the hippocampus of temporal lobe epilepsy patients with mesial-temporal sclerosis, an epilepsy syndrome that is frequently associated with early life febrile seizures (FS). Human congenital loss of GS activity has been shown to result in brain malformations, seizures and death within days after birth. Recently, we showed that GS knockout mice die during embryonic development and that haploinsufficient GS mice have no obvious abnormalities or behavioral seizures. In the present study, we investigated whether reduced expression/activity of GS in haploinsufficient GS mice increased the susceptibility to experimentally induced FS. FS were elicited by warm-air-induced hyperthermia in 14-day-old mice and resulted in seizures in most animals. FS susceptibility was measured as latencies to four behavioral FS characteristics. Our phenotypic data show that haploinsufficient mice are more susceptible to experimentally induced FS (P < 0.005) than littermate controls. Haploinsufficient animals did not differ from controls in hippocampal amino acid content, structure (Nissl and calbindin), glial properties (glial fibrillary acidic protein and vimentin) or expression of other components of the glutamate-glutamine cycle (excitatory amino acid transporter-2 and vesicular glutamate transporter-1). Thus, we identified GS as a FS susceptibility gene. GS activity-disrupting mutations have been described in the human population, but heterozygote mutations were not clearly associated with seizures or epilepsy. Our results indicate that individuals with reduced GS activity may have reduced FS seizure thresholds. Genetic association studies will be required to test this hypothesis.


Asunto(s)
Predisposición Genética a la Enfermedad/genética , Glutamato-Amoníaco Ligasa/genética , Ácido Glutámico/metabolismo , Haplotipos/genética , Convulsiones Febriles/genética , Animales , Biomarcadores/análisis , Biomarcadores/metabolismo , Encéfalo/enzimología , Encéfalo/fisiopatología , Química Encefálica/genética , Modelos Animales de Enfermedad , Regulación hacia Abajo/genética , Transportador 2 de Aminoácidos Excitadores/análisis , Transportador 2 de Aminoácidos Excitadores/metabolismo , Proteína Ácida Fibrilar de la Glía/análisis , Proteína Ácida Fibrilar de la Glía/metabolismo , Ratones , Ratones Noqueados , Tiempo de Reacción/genética , Convulsiones Febriles/enzimología , Convulsiones Febriles/fisiopatología , Proteína 1 de Transporte Vesicular de Glutamato/análisis , Proteína 1 de Transporte Vesicular de Glutamato/metabolismo , Vimentina/análisis , Vimentina/metabolismo
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