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4.
Mediators Inflamm ; 2017: 6909415, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-29138532

RESUMEN

We investigated the regulatory roles of USP2 in mRNA accumulation of proinflammatory cytokines in macrophage-like cells after stimulation with a toll-like receptor (TLR) 4 ligand, lipopolysaccharide (LPS). Human macrophage-like HL-60 cells, mouse macrophage-like J774.1 cells, and mouse peritoneal macrophages demonstrated negative feedback to USP2 mRNA levels after LPS stimulation, suggesting that USP2 plays a significant role in LPS-stimulated macrophages. USP2 knockdown (KD) by short hairpin RNA in HL-60 cells promoted the accumulation of transcripts for 25 of 104 cytokines after LPS stimulation. In contrast, limited induction of cytokines was observed in cells forcibly expressing the longer splice variant of USP2 (USP2A), or in peritoneal macrophages isolated from Usp2a transgenic mice. An ubiquitin isopeptidase-deficient USP2A mutant failed to suppress LPS-induced cytokine expression, suggesting that protein ubiquitination contributes to USP2-mediated cytokine repression. Although USP2 deficiency did not accelerate TNF receptor-associated factor (TRAF) 6-nuclear factor-κB (NF-κB) signaling, it increased the DNA binding ratio of the octamer binding transcription factor (Oct)-1 to Oct-2 in TNF, CXCL8, CCL4, and IL6 promoters. USP2 decreased nuclear Oct-2 protein levels in addition to decreasing the polyubiquitination of Oct-1. In summary, USP2 modulates proinflammatory cytokine induction, possibly through modification of Oct proteins, in macrophages following TLR4 activation.


Asunto(s)
Proteínas de Drosophila/metabolismo , Proteasas Ubiquitina-Específicas/metabolismo , Animales , Línea Celular , Citocinas/metabolismo , Células HL-60 , Humanos , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , FN-kappa B/metabolismo , ARN Mensajero/metabolismo , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo
5.
FASEB J ; 27(12): 4940-53, 2013 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-24005904

RESUMEN

Macrophages play a critical role in chronic inflammation and metabolic diseases. We identified a longer splice variant of ubiquitin specific protease (USP) 2-69 as a novel molecule that modulates pathways implicated in metabolic disorders. Expression levels of aP2/FABP4 and PAI-1/SERPINE1 genes were increased by 4- and 1.8-fold, respectively, after short hairpin RNA-mediated knockdown (KD) of the USP2 gene, and such expression was alleviated by overexpression of USP2-69 in human myeloid cell lines. Supernatants derived from USP2-KD cells induced IL6 (∼6-fold) and SAA3 (∼15-fold) in 3T3-L1 adipocytes to suggest the anti-inflammatory properties of USP2. In addition, we observed a 30% decrease in the number of macrophages in mesenteric adipose tissue derived from USP2-69 transgenic mice fed a high-fat diet for 14 wk compared with that in their C57BL/6 littermates (P<0.01), which was consistent with a ∼40% decrease in transcription of aP2 and PAI-1. The aP2 locus exhibited elevated chromatin accessibility (>2.1-fold), methylation of histone H3 lysine 4 (>4.5-fold), and acetylation of histone H4 (>2.5-fold) in USP2-KD cells. Transfection of isopeptidase-mutated USP2-69 did not alter chromatin conformation on the aP2 locus in USP2-KD cells. Our results suggest that USP2-69 suppresses meta-inflammatory molecules involved in the development of type-2 diabetes.


Asunto(s)
Ensamble y Desensamble de Cromatina , Endopeptidasas/genética , Macrófagos/metabolismo , Transcripción Genética , Proteasas Ubiquitina-Específicas/genética , Adipocitos/metabolismo , Animales , Línea Celular , Cromatina/metabolismo , Endopeptidasas/metabolismo , Epigénesis Genética , Histonas/metabolismo , Humanos , Inflamación/genética , Inflamación/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Ratones , Ratones Endogámicos C57BL , Células Mieloides/metabolismo , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Proteína Amiloide A Sérica/genética , Proteína Amiloide A Sérica/metabolismo , Factor de Transcripción AP-2/genética , Factor de Transcripción AP-2/metabolismo , Ubiquitina Tiolesterasa , Proteasas Ubiquitina-Específicas/metabolismo
6.
Mol Biol Rep ; 40(12): 6747-55, 2013 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-24068436

RESUMEN

The common marmoset (Callithrix jacchus) is increasingly being used as a non-human primate animal model in biomedical research. To perform accurate quantitative analysis of gene expression by quantitative reverse transcription polymerase chain reaction, reliable reference genes should be selected. In this study, we evaluated the expressions of 11 widely used reference genes: ACTB, ATP5F1, B2M, GAPDH, HPRT1, PGK1, PPIA, RN18S1, RPLP0, TBP and UBC in 12 tissues and five brain areas of healthy common marmosets. NormFinder and geNorm indicated that the most suitable reference genes for cross-sectional studies of the 17 tissues were RN18S1 and RPLP0. Conversely, ACTB and PPIA were the most suitable for analyzing brain samples; however, the expression of PGK1 fluctuated among brain areas. These results indicate that suitable reference genes differ between the tissues examined. This study provides fundamental information for gene expression studies of the common marmoset and highlights the importance of validating reference genes before quantification of target mRNAs.


Asunto(s)
Callithrix/genética , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Especificidad de Órganos/genética , Animales , Femenino , Humanos , Masculino , ARN Mensajero/genética , ARN Mensajero/metabolismo , Estándares de Referencia , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Programas Informáticos
7.
Mol Clin Oncol ; 16(1): 18, 2022 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-34881038

RESUMEN

Brain metastasis (BM) in patients with non-small cell lung cancer (NSCLC) is usually associated with a poor prognosis. A 55-year-old Japanese man visited Tokyo Dental College Ichikawa General Hospital with complaints of motor aphasia and fatigue. Enhanced magnetic resonance imaging of the brain revealed multiple tumors. The patient's medical history included lung cancer surgery performed at another hospital 3 months prior to his visit to our hospital. Total resection of the left frontal tumor revealed BM from lung adenocarcinoma. Stereotactic radiosurgery (SRS) was performed for the remaining three BMs. At 9 months after SRS, another new BM was discovered, and SRS was again performed. More than 13 years have elapsed since the last SRS was performed, and the patient has remained relapse-free. To the best of our knowledge, this is the first case report describing a patient with NSCLC with multiple BMs who has remained relapse-free for >13 years with no neurological dysfunction, including cognitive deficit.

8.
J Dent Anesth Pain Med ; 21(5): 431-440, 2021 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-34703892

RESUMEN

BACKGROUND: Neurovascular compression (NVC) is a well-known cause of trigeminal neuralgia (TN). However, patients with idiopathic TN (ITN) do not have evidence of NVC on magnetic resonance imaging (MRI), and other patients may remain asymptomatic despite evidence of NVC on MRI. This suggests that there may be additional risk factors for TN development other than NVC. Although epidemiological factors, such as age and sex differences, are useful for understanding the pathophysiology of TN, detailed statistics for each TN subtype are currently unavailable. Therefore, this study aimed to classify patients with TN into the following groups based on data extracted from past medical records: classical TN (CTN), secondary TN, and ITN. METHODS: The characteristics of the groups and their differences were explored. RESULTS: CTN was more common in women than in men, as previously reported, whereas ITN was more common in men than in women. The ratio of pain sites located on the right side of the face was high in all groups. Patients with CTN were also prone to NVC on the asymptomatic side. CONCLUSION: By investigating TN subtype, it may be possible to elucidate the pathophysiology of TN. This would greatly improve treatment outcomes.

9.
J Vet Med Sci ; 82(1): 14-22, 2020 Jan 10.
Artículo en Inglés | MEDLINE | ID: mdl-31776296

RESUMEN

Instrument cost is a major problem for the transduction of DNA fragments and proteins into cells. Water-in-oil droplet electroporation (droplet-EP) was recently invented as a low-cost and effective method for the transfection of plasmids into cultured human cells. We here applied droplet-EP to livestock animal cells. Although it is difficult to transfect plasmids into bovine fibroblasts using conventional lipofection methods, droplet-EP enabled us to introduce an enhanced green fluorescent protein (EGFP)-expressing plasmid into bovine earlobe fibroblasts. The optimal transfection condition was 3.0 kV, which allowed 19.1% of the cells to be transfected. For swine earlobe fibroblasts, the maximum transfection efficacy was 14.0% at 4.0 kV. After transfection with droplet-EP, 69.1% of bovine and 76.5% of swine cells were viable. Furthermore, droplet-EP successfully transduced Escherichia coli recombinant EGFP into frozen-thawed bovine sperm at 1.5 kV. Flow cytometry analysis revealed that 71.5% of spermatozoa exhibited green fluorescence after transfection. Overall, droplet-EP is suitable for the transfection of plasmids and proteins into cultured livestock animal cells.


Asunto(s)
Electroporación/veterinaria , Plásmidos , Espermatozoides , Transfección/veterinaria , Animales , Bovinos , Células Cultivadas , Electroporación/métodos , Fibroblastos , Proteínas Fluorescentes Verdes , Masculino , Ratones Endogámicos C57BL , Proteínas Recombinantes , Porcinos , Transfección/métodos
11.
Vet Immunol Immunopathol ; 198: 6-13, 2018 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-29571519

RESUMEN

The scavenger receptor CD163 is widely used as a cell signature for alternatively active "M2" macrophages in mammals. In this study, we generated two monoclonal antibodies, ABM-1A9 and ABM-2D6, against the extracellular region of bovine CD163. Conventional Western blotting using the antibodies yielded immunoreactive bands of bovine CD163 at 130 and 150 kDa in non-reduced and reduced spleen lysates, respectively. The minimum limit of detectable concentration of both antibodies was relatively lower (5.0 ng/mL) than that of the anti-human CD163 monoclonal antibody AM-3 K (>1.0 µg/mL), which has been used previously for the detection of bovine CD163. An immunohistochemical study using formalin-fixed paraffin-embedded sections revealed that ABM-1A9 and ABM-2D6 clearly stained some Iba1+ macrophages in the lymph nodes of cattle with mastitis. Moreover, the CD163-stained macrophages were frequently observed engulfing leukocytes. ELISA using ABM-2D6 distinguished levels of circulating soluble CD163 in healthy cattle (less than 16.9 pmol/mL) and cattle with mastitis (more than 33.7 pmol/mL). These new monoclonal antibodies can be used in the diagnosis and evaluation of inflammatory disease prognosis in cattle with immunohistological analyses and blood test applications.


Asunto(s)
Anticuerpos Monoclonales/biosíntesis , Antígenos CD/inmunología , Antígenos de Diferenciación Mielomonocítica/inmunología , Mastitis Bovina/inmunología , Receptores de Superficie Celular/inmunología , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/inmunología , Antígenos CD/análisis , Antígenos de Diferenciación Mielomonocítica/análisis , Western Blotting/veterinaria , Bovinos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Femenino , Humanos , Células Híbridas , Inmunohistoquímica/veterinaria , Ganglios Linfáticos/citología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Receptores de Superficie Celular/análisis
12.
Biochem Biophys Rep ; 9: 322-329, 2017 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-28956020

RESUMEN

We previously reported that ubiquitin-specific protease (USP) 2 in macrophages down-regulates genes associated with metabolic diseases, suggesting a putative anti-diabetic role for USP2 in macrophages. In this study, we evaluate this role at both cellular and individual levels. Isolated macrophages forcibly expressing Usp2a, a longer splicing variant of USP2, failed to modulate the insulin sensitivity of 3T3-L1 adipocytes. Similarly, macrophage-selective overexpression of Usp2a in mice (Usp2a transgenic mice) had a negligible effect on insulin sensitivity relative to wild type littermates following a three-month high-fat diet. However, Usp2a transgenic mice exhibited fewer M1 macrophages in their mesenteric adipose tissue. Following a six-month high-fat diet, Usp2a transgenic mice exhibited a retarded progression of insulin resistance in their skeletal muscle and liver, and an improvement in insulin sensitivity at an individual level. Although conditioned media from Usp2a-overexpressing macrophages did not directly affect the insulin sensitivity of C2C12 myotubes compared to media from control macrophages, they did increase the insulin sensitivity of C2C12 cells after subsequent conditioning with 3T3-L1 cells. These results indicate that macrophage USP2A hampers obesity-elicited insulin resistance via an adipocyte-dependent mechanism.

13.
Exp Anim ; 65(4): 465-471, 2016 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-27356856

RESUMEN

The common marmoset is a non-human primate that has increasingly employed in the biomedical research including the fields of neuroscience and behavioral studies. Cytochrome P450 (CYP) 2D has been speculated to be involved in psycho-neurologic actions in the human brain. In the present study, to clarify the role of CYP2D in the marmoset brain, we investigated the expression patterns of CYP2D mRNA in the brain using in situ hybridization (ISH). In addition, to identify the gene location of CYP2D19, a well-studied CYP2D isoform in the common marmoset, a fluorescence in situ hybridization (FISH) study was performed. Consistent with findings for the human brain, CYP2D mRNA was localized in the neuronal cells of different brain regions; e.g., the cerebral cortex, hippocampus, substantia nigra, and cerebellum. FISH analysis showed that the CYP2D19 gene was located on chromosome 1q, which is homologous to human chromosome 22 on which the CYP2D6 gene exists. These results suggest that CYP2D in the marmoset brain may play the same role as human CYP2D6 in terms of brain actions, and that the CYP2D19 gene is conserved in a syntenic manner. Taken together, these findings suggest that the common marmoset is a useful model for studying psychiatric disorders related to CYP2D dysfunction in the brain.


Asunto(s)
Hidrocarburo de Aril Hidroxilasas/genética , Callithrix/crecimiento & desarrollo , Sistema Enzimático del Citocromo P-450/genética , Expresión Génica , Animales , Hidrocarburo de Aril Hidroxilasas/metabolismo , Encéfalo/metabolismo , Callithrix/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Hibridación Fluorescente in Situ , Masculino , ARN Mensajero/metabolismo
14.
Biomed Res ; 36(5): 343-6, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26522151

RESUMEN

We investigated the anti-obesity effects of Brazilian green propolis ethanol extract using a mouse model of obesity. Repeated intraperitoneal injection of propolis (100 mg/kg twice a week) caused feeding suppression in C57BL/6 mice, whereas this treatment had negligible effects on C57BL/6 ob/ob mice. Since C57BL/6 ob/ob mice have a missense mutation in the Lep gene, leptin is likely to contribute to the propolis-induced feeding suppression. We found that propolis treatment indeed clearly increased leptin mRNA production in the visceral adipose tissues. Moreover, propolis extract directly elevated leptin expression in differentiated 3T3-L1 adipocytes. Artepillin C, an important organic compound found in Brazilian green propolis, failed to induce leptin mRNA in 3T3-L1 cells. Compounds other than artepillin C in Brazilian propolis must thus cause leptin induction in adipocytes, possibly resulting in the suppression of feeding and obesity.


Asunto(s)
Adipocitos/metabolismo , Leptina/metabolismo , Própolis , Animales , Brasil , Ratones
15.
J Vet Med Sci ; 76(8): 1147-51, 2014 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-24784440

RESUMEN

To examine the effect of Babesia infection on the level of the drug-metabolizing enzyme hepatic cytochrome P450 (CYP) 2D, we intraperitoneally inoculated Babesia microti into male ICR mice. CYP2D protein and CYP2D9 mRNA were significantly decreased at 12 days after infection with B. microti. The activity of bunitrolol 4-hydroxylase, which is catalyzed by CYP2D, was also significantly decreased. The mRNA levels of transcriptional regulators of CYP2D9, hepatocyte nuclear factor 4α and signal transducer and activator of transcription 5b, were markedly suppressed. These results suggest that Babesia infection represses CYP2D expression in the mouse liver. The decline in CYP2D-dependent drug metabolism might be involved in the incidence of adverse drug reactions in patients with babesiosis.


Asunto(s)
Hidrocarburo de Aril Hidroxilasas/metabolismo , Babesia microti , Babesiosis/metabolismo , Regulación Enzimológica de la Expresión Génica/fisiología , Hígado/metabolismo , Animales , Cartilla de ADN/genética , Factor Nuclear 4 del Hepatocito , Masculino , Ratones , Ratones Endogámicos ICR , Modelos Biológicos , Reacción en Cadena en Tiempo Real de la Polimerasa , Factor de Transcripción STAT5/metabolismo
16.
Adipocyte ; 2(4): 227-36, 2013 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-24052898

RESUMEN

The anti-diabetic effects of Brazilian propolis were examined using ob/ob mice. Although repeated injection of an ethanol extract of Brazilian propolis (100 mg/kg, ip, twice a week for 12 weeks) did not affect body weight gain and food intake of ob/ob mice, blood glucose and plasma cholesterol levels were significantly attenuated. Moreover, the propolis extract partially restored glucose tolerance and insulin resistance, indicating anti-diabetic properties of the extract. The propolis-treated mice exhibited lower weight gain in mesenteric adipose tissue, while weight gains in inguinal and epididymal adipose tissues were not modulated. Flow cytometric and microscopic analyses suggested that the extract promoted accumulation of eosinophils into mesenteric and epididymal adipose tissues. Alternatively, the ratio of M1-like macrophages to M2-like macrophages in mesenteric adipose tissue was reduced by the propolis injection, coincident with the decrement of the number of interleukin-12A(+) cells. Levels of M1 macrophage markers, such as Itgax and Il12b transcripts, were decreased in the vascular stromal fraction of mesenteric adipose tissue, whereas those of pan-macrophage markers Emr1 and Cd68 were not influenced. Microarray and subsequent gene ontology term analyses suggested that propolis attenuated immune activation in mesenteric adipose tissues. Taken together, this indicates that Brazilian propolis improves diabetes in ob/ob mice, presumably through modification of immune cells in mesenteric adipose tissues.

17.
J Vet Med Sci ; 74(2): 241-5, 2012 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-21959890

RESUMEN

To investigate effects of Babesia infection on drug metabolism, we intraperitoneally inoculated B. microti into ICR mice and measured the expression and activity of hepatic cytochrome P450 (CYP) 3A, a major drug-metabolizing enzyme. Twelve days after infection, CYP3A11 mRNA, CYP3A protein and activity and mRNAs of nuclear receptors, which participate in CYP3A expression, were significantly reduced. These results suggest that B. microti infection suppresses CYP3A-dependent drug metabolism. Additionally, tumor necrosis factor (TNF)-α and nitric oxide synthase (NOS) 2 mRNAs were induced in the infected mouse liver. Since TNF-α is one of the potent mediators that induce NOS2 and repress CYP3A transcription, the possible involvement of TNF-α in this downregulation of CYP3A was discussed.


Asunto(s)
Babesia microti/aislamiento & purificación , Babesiosis/enzimología , Citocromo P-450 CYP3A/biosíntesis , Hígado/enzimología , Animales , Babesiosis/parasitología , Western Blotting , Citocromo P-450 CYP3A/genética , Citocromo P-450 CYP3A/metabolismo , Regulación hacia Abajo , Masculino , Ratones , Ratones Endogámicos ICR , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , ARN Mensajero/química , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo
18.
Endocr Pathol ; 2(4): 230-234, 1991 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-32357637

RESUMEN

The case of a 51 -year-old man with recurrent nonfunctioning pituitary adenoma is presented. Despite clinically and endocrinologically normal pituitary function in regard to growth hormone and prolactin, many growth hormone- and prolactin-positive cells were immunohis-tochemically detected in adenoma tissue. Furthermore, a quite rare tumor of silent mixed growth hormone cell-prolactin cell pituitary adenoma was confirmed by the double-labeling immunoelectron-microscopical study.

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