RESUMO
The aim of this study was to evaluate the suitability of size specific dose estimates (SSDE) to estimate patient dose in Fast kVp switching dual energy CT. An anthropomorphic phantom (RAN-110) was repeatedly scanned (chest, abdomen and the pelvis) using a 64 detector row MDCT (Discovery CT750 HD, GE Healthcare, Milwaukee, WI, USA) with various CT parameters, including Fast kVp switching. Dosimetry was performed using thermo-luminescent dosimeters, positioned both superficially and within the phantom. SSDE was calculated for all slices of the anthropomorphic phantom using both the localiser and axial images. In Fast kVp switching, SSDE underestimated the measured absorbed dose for the chest/abdomen region ~35% at the maximum, but were in closer agreement for the pelvic region about within 10%. In single energy techniques, SSDE could not be applied in the estimation of organ doses, but in Fast kVp switching dual energy techniques, SSDE could be applied for anatomical regions with larger thicknesses.
Assuntos
Abdome/efeitos da radiação , Pelve/efeitos da radiação , Imagens de Fantasmas , Monitoramento de Radiação , Imagem Radiográfica a Partir de Emissão de Duplo Fóton/métodos , Tomografia Computadorizada por Raios X/métodos , Humanos , Doses de Radiação , Radiografia TorácicaRESUMO
We encountered a 61-year-old woman with primary cardiac angiosarcoma in the left atrium. On echocardiography, the tumor extended into the atrial septum and mitral valve, and mitral valve stenosis and regurgitation were significant. We resected the tumor protruding into left atrium, and affecting mitral valve. The surgical procedure was not radical, but on postoperative echocardiography, function of the mitral valve was improved. Three months later, elevation of her right diaphragma was observed on chest X-ray and a giant adrenal tumor was detected by magnetic resonance imaging. Tumor biopsy indicated that this tumor was adrenal metastasis from cardiac angiosarcoma. In addition, echocardiography showed the recurrence of angiosarcoma in the left atrium and the presence of mitral stenosis and regurgitation. She died of heart failure 185 days postoperatively.
Assuntos
Neoplasias das Glândulas Suprarrenais/secundário , Neoplasias Cardíacas/patologia , Hemangiossarcoma/secundário , Neoplasias das Glândulas Suprarrenais/diagnóstico , Feminino , Átrios do Coração/diagnóstico por imagem , Neoplasias Cardíacas/diagnóstico por imagem , Hemangiossarcoma/diagnóstico por imagem , Humanos , Imageamento por Ressonância Magnética , Pessoa de Meia-Idade , UltrassonografiaRESUMO
The objective of this study was to determine whether anorectal malformations (ARMs) and anterior sacral myelomeningocele share the same embryogenic pathway in a mouse model. Etretinate (Ro 10-9359) was administrated to C57BL/6 mice on gestation day 9 (E9). Sections of embryos and fetuses from E9.5 to E18 were observed by HE staining. Immunohistochemical staining with anti-NeuN and anti-GFAP was also done to determine cell origins of a presacral mass. In etretinate-treated embryos, neuroepithelial cells proliferated in the presacral region on E9.5. On E12, a canal appeared between the ectopic proliferated neuroepithelium and hindgut. On E13, anorectum abnormally kept a canal with the ventral urogenital tract through a fistula. On E13.5, a huge mass formed in the presacral region. On E18, 76.9% (30/39) of fetuses had ARMs, 100% (39/39) had a presacral mass (71.8% were huge) and 100% (39/39) had a sacral defect. The types of ARMs were mainly rectourethral or rectocloacal fistula. The presacral mass was anterior sacral myelomeningocele. We thus established the first mouse model of the Currarino triad, congenital caudal anomalies, including ARM, sacral abnormality and presacral mass. These disorders share the same embryogenic pathway. The teratogenic target of etretinate is the tail bud. Abnormal differentiation of the tail bud mesenchyme leads to defects of the tailgut and caudal neural tube. The abnormal mass blocks normal descent of the dorsal cloaca through the most posterior part of the cloacal plate.
Assuntos
Canal Anal/anormalidades , Meningomielocele/embriologia , Reto/anormalidades , Sacro/anormalidades , Canal Anal/embriologia , Animais , Etretinato , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Reto/embriologia , Sacro/embriologiaRESUMO
PURPOSE: To describe the surgical technique, and its usefulness, of temporary amniotic membrane patching (AMP) in the acute phase of ocular chemical injury. METHODS: Temporary AMP with modification in suture placement was performed on five eyes of five consecutive patients inflicted with acute chemical injury having a greater than grade II injury by the Roper-Hall classification. RESULTS: All patients reported herein presented with a large epithelial defect on the cornea and conjunctiva. Case 3 was classified as grade III while the other four cases were classified as grade II. The causative chemical agents were anhydrous acetic acid in Case 1, calcium oxide in Case 2, sodium hydroxide in Case 3, sodium silicate in Case 4, and sulphuric acid in Case 5. All cases experienced rapid relief of pain after AMP. Epithelialization of the cornea with improvement of visual acuity was observed in all cases when the amniotic membrane was removed within 2 weeks after surgery. During the mean follow-up of 19.6 months, the ocular surface remained stable and no cicatricial complications were noted. CONCLUSIONS: These results suggest that immediate AMP is quite useful for managing moderately severe acute ocular chemical injury by facilitating rapid epithelialization and pain relief, and securing ocular surface integrity.
Assuntos
Curativos Biológicos , Queimaduras Químicas/cirurgia , Queimaduras Oculares/cirurgia , Doença Aguda , Adulto , Queimaduras Químicas/patologia , Córnea/patologia , Queimaduras Oculares/patologia , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Cuidados Paliativos/métodosRESUMO
It is widely accepted that mature mammalian oocytes are induced to resume meiosis by a sperm-borne oocyte-activating factor(s) (sperm factor, SF) immediately after normal fertilization or intracytoplasmic sperm injection. The SF is most likely a soluble factor that is localized within the cytoplasm of mature spermatozoa, but the exact stage at which it appears during spermatogenesis and its localization after oocyte activation is not fully understood, except in the mouse. First, we injected mature spermatozoa and spermatogenic cells from cynomolgus monkeys into mouse oocytes to assess their oocyte-activating capacity. More than 90% of mouse oocytes were activated after injection of monkey spermatozoa. Round spermatids and primary spermatocytes (late pachytene to diplotene) also activated oocytes (93% and 79%, respectively). Injection of monkey spermatozoa and spermatids induces intracellular Ca(2+) oscillations in a pattern similar to that seen following normal fertilization. Most spermatocytes did not produce typical intracellular Ca(2+) oscillations. Second, we transferred pronuclei or cytoplasts from mouse oocytes that had been activated by monkey spermatozoa or spermatids into intact mature mouse oocytes by electrofusion in order to examine the localization of the SF after pronuclear formation. Some of the SF was localized within the pronuclei, but some stayed in the ooplasm. This study demonstrated that spermatogenic cells of cynomolgus monkeys acquire oocyte-activating capacity at much earlier stages than those of mice, and that the monkey SF has a pronucleus-directing nature, although to a lesser extent than the mouse SF.
Assuntos
Oócitos/fisiologia , Injeções de Esperma Intracitoplásmicas , Espermatozoides/fisiologia , Animais , Cálcio/metabolismo , Núcleo Celular/ultraestrutura , Feminino , Macaca fascicularis , Masculino , Camundongos , Oócitos/química , Oócitos/ultraestrutura , Espermátides/química , Espermátides/fisiologia , Espermatozoides/química , Espermatozoides/ultraestruturaRESUMO
Orexins (hypocretins) are a pair of neuropeptides implicated in energy homeostasis and arousal. Recent reports suggest that loss of orexin-containing neurons occurs in human patients with narcolepsy. We generated transgenic mice in which orexin-containing neurons are ablated by orexinergic-specific expression of a truncated Machado-Joseph disease gene product (ataxin-3) with an expanded polyglutamine stretch. These mice showed a phenotype strikingly similar to human narcolepsy, including behavioral arrests, premature entry into rapid eye movement (REM) sleep, poorly consolidated sleep patterns, and a late-onset obesity, despite eating less than nontransgenic littermates. These results provide evidence that orexin-containing neurons play important roles in regulating vigilance states and energy homeostasis. Orexin/ataxin-3 mice provide a valuable model for studying the pathophysiology and treatment of narcolepsy.
Assuntos
Proteínas de Transporte/metabolismo , Transtornos da Alimentação e da Ingestão de Alimentos/genética , Hipotálamo/fisiopatologia , Peptídeos e Proteínas de Sinalização Intracelular , Narcolepsia/genética , Proteínas do Tecido Nervoso/genética , Neurônios/fisiologia , Neuropeptídeos/metabolismo , Neurotransmissores/metabolismo , Obesidade/genética , Fases do Sono/genética , Animais , Ataxina-3 , Transtornos da Alimentação e da Ingestão de Alimentos/fisiopatologia , Feminino , Humanos , Hipotálamo/patologia , Doença de Machado-Joseph/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , Narcolepsia/fisiopatologia , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Neurônios/patologia , Proteínas Nucleares , Obesidade/fisiopatologia , Orexinas , Peptídeos/genética , Proteínas Repressoras , Deleção de Sequência , Fases do Sono/fisiologia , Sono REM/genética , Fatores de TranscriçãoRESUMO
The Biosafety Committee of the Japanese Association of Laboratory Animal Facilities of National Universities (JALAN) investigated recent episodes of microbiological contamination in genetically modified mice (GMM), and the countermeasures taken when the contaminated GMM were introduced into animal facilities, by questionnaires addressed to 53 animal facilities belonging to JALAN and serological tests. Although almost all of the contaminated GMM were accepted with conditions such as rederivation after or before reception and housing in designated rooms, contamination with a spectrum of microorganisms was demonstrated in GMM transferred domestically and from abroad. In serological tests, Mycoplasma pulmonis, mouse parvovirus, and mouse encephalomylitis virus were detected in GMM transferred from domestic facilities and from abroad. The present results of the questionnaires and serological tests suggest that GMM are highly and widely contaminated with microorganisms compared with mice from commercial breeders. Thus, we propose a microbiological requirement, including microbiological status--excellent, common, and minimum--as a guide for the transfer and procurement of mice and rats in Japan.
Assuntos
Transmissão de Doença Infecciosa/prevenção & controle , Controle de Infecções , Organismos Geneticamente Modificados/microbiologia , Animais , Guias como Assunto , Abrigo para Animais , Japão , Vírus Elberfeld do Camundongo/isolamento & purificação , Vírus Elberfeld do Camundongo/patogenicidade , Camundongos , Mycoplasma/isolamento & purificação , Mycoplasma/patogenicidade , Parvovirus/isolamento & purificação , Parvovirus/patogenicidade , Ratos , Medição de Risco , Testes SorológicosRESUMO
The non-structural protein NS1, encoded by the parvovirus minute virus of mice (MVM), is a potent regulator of viral gene expression in addition to prominent roles in viral replication and cytopathic effects associated with parvoviral infection. Although NS1 involves the modulation of viral and cellular transcription, the primary activation mechanism of MVM NS1 remains unclear. In the present study, we show here that the coactivator CREB binding protein, CBP, could potentiate NS1-mediated transcription as measured on the P38 promoter, which drives expression of the MVM capsid genes. NS1 bound to the two related cysteine-histidine-rich regions of CBP, referred to as C/H1 and C/H3, the former of which has an antagonistic function to CBP upon the NS1-transactivation. Furthermore, NS1 inhibited the synergistic transactivation by CBP and p53. These findings suggested that CBP as a transcriptional coactivator is required for NS1-mediated viral and cellular transcription in parvovirus-infected cells, resulting in cell proliferation and differentiation to achieve its lytic cycle.
Assuntos
Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas não Estruturais Virais/metabolismo , Células 3T3 , Animais , Sítios de Ligação/genética , Ligação Competitiva , Proteína de Ligação a CREB , Linhagem Celular , DNA Recombinante , Regulação da Expressão Gênica , Humanos , Camundongos , Proteínas Nucleares/genética , Plasmídeos/genética , Testes de Precipitina , Ligação Proteica , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transativadores/genética , Ativação Transcricional , Transfecção , Proteína Supressora de Tumor p53/genética , Proteínas não Estruturais Virais/genéticaRESUMO
OBJECTIVES: The purpose of this study was to examine the effects of various cytokines and/or lipopolysaccharide (LPS) on nitric oxide (NO) production from USAC, a newly established clonal cell line derived from human osteogenic sarcoma that expressed chondrocytic phenotypes. MATERIALS AND METHODS: No production was measured by Griess method. Inducible nitric oxide synthase (iNOS) mRNA was detected by PCR analysis. Western blotting analysis and immunocytochemistry was used to detect iNOS protein. RESULTS: Although USAC cells treated without any stimulants produced only small amounts of NO, exposure to cytokines and/or LPS induced iNOS in USAC cells and produced high levels of NO. The stimulatory effects of cytokines and/or LPS on NO production required TNF-alpha. TNF-alpha alone neither induced iNOS in USAC cells nor caused production of NO, but addition of TNF-alpha to USAC cells pretreated with LPS and IFN-gamma enhanced the expression of iNOS mRNA, induced iNOS protein and produced NO. Dexamethasone inhibited the stimulatory effect of TNF-alpha. CONCLUSIONS: The responsiveness of USAC cells to cytokines and/or LPS and steroid hormone on NO production was quite different from that reported for rabbit and human articular cartilaginous cells. The differences in responsiveness between articular cartilaginous chondrocytes and USAC cells might have been because USAC cells were established from a malignant tumor.
Assuntos
Neoplasias Ósseas/patologia , Condrócitos/metabolismo , Citocinas/farmacologia , Óxido Nítrico/biossíntese , Osteossarcoma/patologia , Animais , Anti-Inflamatórios/farmacologia , Linhagem Celular Tumoral , Células Clonais , Dexametasona/farmacologia , Inibidores Enzimáticos/farmacologia , Humanos , Interferon gama/farmacologia , Interleucina-1/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Nus , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II , Coelhos , Fator de Necrose Tumoral alfa/farmacologia , ômega-N-Metilarginina/farmacologiaRESUMO
We have succeeded in transplanting a human osteogenic sarcoma of the mandible into athymic mice. The transplanted tumor showed marked chondrogenesis and mineralization. Recently, a cell line (USAC) with phenotypes of chondrocyte has been established from the transplanted tumor. USAC cells were stellate or spindle-shaped in sparse culture, but polygonal or spherical at sub-confluency to confluency. In long-term culture, the cells were condensed and calcified nodules were formed. Production of types I, II and X collagen were detected by immunohistochemical staining and Western blot analysis. Type I collagen was strongly expressed in the stellate or spindle-shaped cells. Although type II collagen was usually present in all cells during culture, it was strongly stained in polygonal cells at confluency. Type X collagen was seen in large polygonal cells around calcified nodules. Marked [35S]-sulfate uptake and metachromasia were seen at the confluent stage and in the nodule. The cells around the nodules were positive for alkaline phosphatase, and the center of the nodules was stained with alizarin red. The potentiality of cartilage formation was confirmed by in vivo experiments using a diffusion chamber in athymic mice. These observations indicate that USAC cells maintain characteristics of chondrocyte progenitor cells and thus may serve as a useful model to study the sequential events of chondrogenesis and the process of morbid endochondral calcification. This experiment also demonstrated that transplantation of tumor tissue into athymic mice is a convenient strategy for establishment of a cell line.
Assuntos
Linhagem Celular , Condrócitos/citologia , Osteossarcoma , Análise de Variância , Animais , Divisão Celular , Condrócitos/metabolismo , Condrogênese , Células Clonais , Colágeno/biossíntese , Humanos , Cariotipagem , Neoplasias Mandibulares , Camundongos , Camundongos Nus , Fenótipo , Proteoglicanas/biossínteseRESUMO
To define a gene expression mechanism, it is often advantageous to use a reporter gene and transgenic mouse. The lacZ reporter gene is particularly useful for studies of the cis-regulatory element for tissue-specific expression in transgenic mice because of the ease of the enzyme assay and visualization on sections. In this report, we describe our method for examining the cis-regulatory element in transgenic mice, including choice of the lacZ gene, generation of transgenic mice, and analysis of beta-galactosidase activity.
Assuntos
Escherichia coli/genética , Expressão Gênica , Genes Reporter/genética , Óperon Lac/genética , Camundongos Transgênicos/genética , Glândulas Suprarrenais/metabolismo , Animais , Encéfalo/citologia , Encéfalo/metabolismo , Canais de Cálcio Tipo P/genética , Canais de Cálcio Tipo P/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Camundongos Transgênicos/metabolismo , Neurônios/metabolismo , Coloração e Rotulagem , Distribuição Tecidual , beta-Galactosidase/metabolismoRESUMO
The P/Q-type Ca(2+) channel alpha(1A) subunit is expressed in spinal cord including ventral motor neurons and interneurons and dorsal horn. To identify the transcriptional mechanisms of the mouse alpha(IA) subunit gene in spinal cord, transgenic mice carrying a 0.5, 1.5, 3.0 or 6.3-kb 5'-upstream region fused to the Escherichia coli lacZ reporter gene were examined. Transgenic mice carrying the 3.0-kb region expressed the reporter gene in dorsal horn and interneurons of ventral horn, although those with the 0.5-kb, 1.5-kb or 6.3-kb region did not. No transgenic mice expressed the reporter gene in motor neurons of ventral horn. These results suggest that in spinal cord, the expression mechanisms of the alpha(1A) subunit gene are complex, involving both positive and negative cis-regulatory elements, and the 6.3-kb 5'-upstream region alone is not sufficient for the expression.
Assuntos
Regiões 5' não Traduzidas/fisiologia , Canais de Cálcio Tipo P/genética , Canais de Cálcio Tipo P/metabolismo , Canais de Cálcio Tipo Q/genética , Canais de Cálcio Tipo Q/metabolismo , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica/fisiologia , Genes Reporter/fisiologia , Óperon Lac/fisiologia , Medula Espinal/metabolismo , Animais , Perfilação da Expressão Gênica , Camundongos , Camundongos Transgênicos , Neurônios/citologia , Neurônios/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Medula Espinal/citologiaRESUMO
The mechanisms underlying oligodendrocyte (OLG) loss and the precise roles played by OLG death in human demyelinating diseases such as multiple sclerosis (MS), and in the rodent model of MS, experimental autoimmune encephalomyelitis (EAE), remain to be elucidated. To clarify the involvement of OLG death in EAE, we have generated transgenic mice that express the baculovirus anti-apoptotic protein p35 in OLGs through the Cre-loxP system. OLGs from cre/p35 transgenic mice were resistant to tumor necrosis factor-alpha-, anti-Fas antibody- and interferon-gamma-induced cell death. cre/p35 transgenic mice were resistant to EAE induction by immunization with the myelin oligodendrocyte glycoprotein. The numbers of infiltrating T cells and macrophages/microglia in the EAE lesions were significantly reduced, as were the numbers of apoptotic OLGs expressing the activated form of caspase-3. Thus, inhibition of apoptosis in OLGs by p35 expression alleviated the severity of the neurological manifestations observed in autoimmune demyelinating diseases.
Assuntos
Inibidores de Caspase , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/prevenção & controle , Marcação de Genes , Oligodendroglia/metabolismo , Proteínas Virais/genética , Animais , Apoptose , Doenças Autoimunes Desmielinizantes do Sistema Nervoso Central/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Inibidores Enzimáticos/metabolismo , Expressão Gênica , Imuno-Histoquímica , Proteínas Inibidoras de Apoptose , Interferon gama/farmacologia , Camundongos , Camundongos Transgênicos , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , RNA Mensageiro/metabolismo , Medula Espinal/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Receptor fas/farmacologiaRESUMO
Renin plays a key role in controlling blood pressure through its specific cleavage of angiotensinogen to generate angiotensin I (AI). Although possible existence of the other angiotensin forming enzymes has been discussed to date, its in vivo function remains to be elucidated. To address the contribution of renin, we generated renin knockout mice. Homozygous mutant mice show neither detectable levels of plasma renin activity nor plasma AI, lowered blood pressure 20-30 mm Hg less than normal, increased urine and drinking volume, and altered renal morphology as those observed in angiotensinogen-deficient mice. We recently found the decreased density in granular layer cells of hippocampus and the impaired blood-brain barrier function in angiotensinogen-deficient mice. Surprisingly, however, such brain phenotypes were not observed in renin-deficient mice. Our results demonstrate an indispensable role for renin in the circulating angiotensin generation and in the maintenance of blood pressure, but suggest a dispensable role for renin in the blood-brain barrier function.
Assuntos
Angiotensina I/sangue , Barreira Hematoencefálica/fisiologia , Encéfalo/fisiologia , Fenômenos Fisiológicos Cardiovasculares , Renina/deficiência , Angiotensinogênio/sangue , Animais , Pressão Sanguínea , Homeostase , Homozigoto , Rim/patologia , Camundongos , Camundongos Knockout , Sistema Renina-AngiotensinaRESUMO
Matrix GLA protein (MGP), a gamma-carboxyglutamic acid (GLA)-rich, vitamin K-dependent and apatite-binding protein, is a regulator of hypertrophic cartilage mineralization during development. However, MGP is produced by both hypertrophic and immature chondrocytes, suggesting that MGP's role in mineralization is cell stage-dependent, and that MGP may have other roles in immature cells. It is also unclear whether MGP regulates the quantity of mineral or mineral nature and quality as well. To address these issues, we determined the effects of manipulations of MGP synthesis and expression in (a) immature and hypertrophic chondrocyte cultures and (b) the chick limb bud in vivo. The two chondrocyte cultures displayed comparable levels of MGP gene expression. Yet, treatment with warfarin, a gamma-carboxylase inhibitor and vitamin K antagonist, triggered mineralization in hypertrophic but not immature cultures. Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment. Scanning electron microscopy, x-ray microanalysis, and Fourier-transform infrared spectroscopy revealed that mineral forming in control and warfarin-treated hypertrophic cell cultures was similar and represented stoichiometric apatite. Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization. Surprisingly, MGP overexpression in the developing limb not only inhibited cartilage mineralization, but also delayed chondrocyte maturation and blocked endochondral ossification and formation of a diaphyseal intramembranous bone collar. The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Condrócitos/fisiologia , Proteínas da Matriz Extracelular , Osteogênese/fisiologia , Ácido 1-Carboxiglutâmico/biossíntese , Ácido 1-Carboxiglutâmico/genética , Ácido 1-Carboxiglutâmico/metabolismo , Animais , Osso e Ossos/metabolismo , Calcificação Fisiológica/fisiologia , Proteínas de Ligação ao Cálcio/biossíntese , Proteínas de Ligação ao Cálcio/genética , Células Cultivadas , Embrião de Galinha , Regulação da Expressão Gênica no Desenvolvimento , Botões de Extremidades/embriologia , Microscopia Eletrônica de Varredura , Minerais/metabolismo , Osteogênese/efeitos dos fármacos , Vitamina K/biossíntese , Vitamina K/genética , Vitamina K/metabolismo , Varfarina/farmacologia , Proteína de Matriz GlaRESUMO
Angiotensinogen-deleted mice (Agt-KO) show phenotypes of hypotension and renal atrophy. To investigate whether an alternative pathway other than angiotensin II (AII), i.e., processed angiotensin fragments, may play a biological role in nephrogenesis, we analyzed a congenic line of Agt-KO fetuses and neonates derived from two sources: one (Agt-KO/He) from mating with heterozygous angiotensinogen-deleted mice and the other (Agt-KO/Ho) from mating homozygous angiotensinogen-deleted mice. Although Agt-KO/He did not show a typical phenotype at birth, these mice showed papillary atrophy 2 weeks later and thereafter, a marked increase in renal size, i.e., pelvic dilatation. In contrast, Agt-KO/Ho showed renal abnormalities at birth and subsequently died. TUNEL staining and electron microscopy revealed that accelerated papillary apoptosis was present at birth in Agt-KO/Ho and caused abnormal papillary development; however, apoptosis was not detected in Agt-KO/He, suggesting that different mechanisms for the abnormal renal development exist in Agt-KO/He and Agt-KO/Ho. Two-week administration of an angiotensin fragment (3-8), angiotensin IV (AIV), to Agt-KO/He markedly attenuated the renal atrophy, decreasing the incidence from 81% to 14%. However, administration of AIV to fetal Agt-KO/Ho through the mother did not decrease the incidence. This is marked contrast to AII, which prevented renal atrophy in both fetal and neonatal periods. It is therefore suggested that AIV is involved in nephrogenesis in a developmental stage-specific manner.
Assuntos
Angiotensinas/fisiologia , Rim/efeitos dos fármacos , Rim/crescimento & desenvolvimento , Angiotensina II/análogos & derivados , Angiotensina II/farmacologia , Angiotensina II/fisiologia , Angiotensinogênio/genética , Angiotensinas/genética , Animais , Apoptose , Marcação In Situ das Extremidades Cortadas , Rim/patologia , Rim/ultraestrutura , Camundongos , Camundongos Knockout , Microscopia Eletrônica , Fatores de TempoRESUMO
We previously identified various upstream and downstream regulatory elements and factors important for hepatic expression of the human angiotensinogen (ANG) gene, the precursor of vasoactive octapeptide angiotensin II. In the present study, to further investigate the molecular mechanism of human ANG transcriptional regulation, we generated transgenic mice carrying the fusion gene composed of the 1. 3-kilobase promoter of the human ANG gene, its downstream enhancer, and the chloramphenicol acetyltransferase reporter gene. Because expression of the chloramphenicol acetyltransferase gene was observed strongly in the liver and weakly in the kidney, we suspected that hepatocyte nuclear factor (HNF) 4 with a tissue expression pattern similar to that of the reporter gene would regulate ANG transcription. In vitro assays indicated that HNF4 bound to the promoter elements and strongly activated the ANG transcription, but that chicken ovalbumin upstream promoter transcription factor (COUP-TF), a transcriptional repressor, dramatically repressed human ANG transcription through the promoter elements and the downstream enhancer core elements. Furthermore, COUP-TF dramatically decreased the human ANG transcription in the mouse liver by the Helios Gene Gun system in vivo. These results suggest that an interplay between HNF4 and COUP-TF could be important in hepatic human ANG transcription.
Assuntos
Angiotensinas/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Fosfoproteínas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Angiotensinas/biossíntese , Animais , Sequência de Bases , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Sítios de Ligação , Fator I de Transcrição COUP , Núcleo Celular/metabolismo , Cloranfenicol O-Acetiltransferase/metabolismo , Relação Dose-Resposta a Droga , Deleção de Genes , Fator 4 Nuclear de Hepatócito , Humanos , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Camundongos Transgênicos , Dados de Sequência Molecular , Ligação Proteica , Elementos de Resposta , Transcrição Gênica , Células Tumorais CultivadasRESUMO
The nonstructural protein 2 (NS2) from parvovirus minute virus of mice (MVMp) is a 25-kDa polypeptide which localizes preferentially to the cytoplasm and associates with cellular proteins in cytoplasm. These lines of evidence suggest that NS2 is positively exported from the nucleus to cytoplasm and functions in cytoplasm. We report here that nuclear export of NS2 is inhibited by leptomycin B (LMB), a drug that specifically blocks nuclear export signal (NES)-chromosomal region maintenance 1 (CRM1) interactions. CRM1 binds specifically to the 81- to 106-amino-acid (aa) region of NS2, and the region of NS2 actually functions as a NES. Interestingly, this region appears to be distinct from a typical NES sequence, which consists of leucine-rich sequences. These results indicate that NS2 protein is continuously exported from the nucleus by a CRM1-dependent mechanism and suggest that CRM1 also exports to distinct type of NESs.
Assuntos
Proteínas de Transporte/metabolismo , Núcleo Celular/metabolismo , Carioferinas , Vírus Miúdo do Camundongo/metabolismo , Receptores Citoplasmáticos e Nucleares , Proteínas não Estruturais Virais/metabolismo , Sequência de Aminoácidos , Animais , Transporte Biológico/efeitos dos fármacos , Núcleo Celular/virologia , Citoplasma/efeitos dos fármacos , Citoplasma/metabolismo , Ácidos Graxos Insaturados/farmacologia , Células HeLa , Humanos , Camundongos , Vírus Miúdo do Camundongo/efeitos dos fármacos , Dados de Sequência Molecular , Homologia de Sequência de Aminoácidos , Transdução de Sinais/efeitos dos fármacos , Proteína Exportina 1RESUMO
Human T-cell leukemia virus type 1 (HTLV-1) infection is shown to be closely associated with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the occurrence of HAM/TSP was reported to be associated with MHC class II, the mechanism is still unclear. The WKA(RT1k) strain of rats was reported to develop HAM/TSP-like paraparesis after HTLV-1 infection, and was suggested to be an animal model of HAM/TSP. We asked whether MHC k-haplotype is specifically involved in the pathogenesis of paraparesis of WKA(RT1k) rats. We injected the HTLV-1 producing human T cells (MT-2 cells) intravenously into WKA(RT1k) rats and MHC congenic WKA.1L(RT1l) rats which have MHC l-haplotype of LEW rats on the WKA background. Positive antibody response to HTLV-1 antigens and presence of provirus in peripheral blood mononuclear cells confirmed that MT-2 cell-injected rats were infected with HTLV-1. Two of 13 MT-2 cell-injected WKA(RT1k) rats and five of 13 MT-2 cell-injected WKA.1L(RT1l) rats developed HAM/TSP-like hindlimb paraparesis between 16 and 26 months old. Interestingly, three of 14 MT-2 cell-uninjected WKA(RT1k) rats and four of 13 MT-2 cell-uninjected WKA.1L(RT1l) rats showed similar paraparesis between 15 and 26 months old. MHC k-haplotype is not specific to the development of paraparesis in WKA(RT1k) rats. The role of aging, genetic background, HTLV-1 infection and other factors on the development of HAM/TSP-like paraparesis in rats are discussed.