RESUMO
This study examined whether using an artificial neural network (ANN) helps beginners in diagnostic cardiac imaging to achieve similar results to experts when interpreting stress myocardial perfusion imaging (MPI). One hundred and thirty-eight patients underwent stress MPI with Tc-labeled agents. An expert and a beginner interpreted stress/rest MPI with or without the ANN and the results were compared. The myocardium was divided into 5 regions (the apex; septum; anterior; lateral, and inferior regions), and the defect score of myocardial blood flow was evaluated from 0 to 4, and SSS, SRS, and SDS were calculated. The ANN effect, defined as the difference in each of these scores between with and without the ANN, was calculated to investigate the influence of ANN on the interpreters' performance. We classified 2 groups (insignificant perfusion group and significant perfusion group) and compared them. In the same way, classified 2 groups (insignificant ischemia group and significant ischemia group) and compared them. Besides, we classified 2 groups (normal vessels group and multi-vessels group) and compared them. The ANN effect was smaller for the expert than for the beginner. Besides, the ANN effect for insignificant perfusion group, insignificant ischemia group and multi-vessels group were smaller for the expert than for the beginner. On the other hand, the ANN effect for significant perfusion group, significant ischemia group and normal vessels group were no significant. When interpreting MPI, beginners may achieve similar results to experts by using an ANN. Thus, interpreting MPI with ANN may be useful for beginners. Furthermore, when beginners interpret insignificant perfusion group, insignificant ischemia group and multi-vessel group, beginners may achieve similar results to experts by using an ANN.
Assuntos
Imagem de Perfusão do Miocárdio , Coração , Humanos , Redes Neurais de Computação , Perfusão , Valor Preditivo dos Testes , Tomografia Computadorizada de Emissão de Fóton ÚnicoRESUMO
Basal cell carcinoma (BCC) is a malignant skin tumor originating from cells of the epidermal basal layer and adnexal epithelium, especially in sun-exposed areas. Unlike squamous cell carcinoma (SCC), BCC has a propensity to grow only locally possibly due to differences in the surrounding microenvironment including the basement membrane (BM) and stroma. To investigate the components constituting the BM and surrounding connective tissue in BCC and SCC, we analyzed the expression of BM proteins, nidogen 1 (NID1) and type IV collagen (COL4). We compared the immunohistochemical expressions of NID1 and COL4 among tumor specimens from BCC, SCC and its precancerous condition, actinic keratosis (AK), (n = 5 each condition). The expressions of NID1 and COL4 were both decreased around the tumor nest of SCC. In contrast, the expressions of both NID1 and COL4 around the nest of BCC were much higher than in the peri-lesional normal skin not only at the BM, but also in the surrounding stromal tissue. Our findings imply that the surrounding stromal cells of BCC, but not SCC or AK, excessively produce NID1 and COL4, which may be involved in preventing BCC cells from destroying the BM and invading the dermis.
Assuntos
Carcinoma Basocelular/metabolismo , Carcinoma de Células Escamosas/metabolismo , Ceratose Actínica/metabolismo , Glicoproteínas de Membrana/biossíntese , Neoplasias Cutâneas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Membrana Basal/metabolismo , Colágeno Tipo IV/biossíntese , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-IdadeRESUMO
Although CD133 has been shown to be a marker for cancer stem cells in various tumours, its expression in pancreatic cancer has not yet been clinically reported. In this study, we investigated the relationship between CD133 expression and clinicopathological factors in pancreatic cancer. Pancreatic head carcinoma specimens from 80 patients who underwent surgical resection were immunohistochemically assessed for CD133, vascular endothelial growth factor (VEGF)-C, CXCR4, CD34, Ki-67, and cytokeratin (CK) expressions. Sixty percentage (48/80) of specimens were CD133-positive, with less than 15% cells per specimen expressing the marker. CD133-positive cells were found at the peripheral site of adenocarcinoma glandular structures and were negative for CK. There was a significant correlation between CD133 expression and clinicopathological factors, including histological type, lymphatic invasion, and lymph node metastasis (P=0.0215, 0.0023, and 0.0024, respectively). Vascular endothelial growth factor-C expression was also significantly correlated with CD133 expression (P=0.0002). Consequently, the 5-year survival rate of CD133-positive patients was significantly lower than that of CD133-negative patients (P=0.0002) and multivariate analysis revealed that CD133 expression was an independent prognostic factor (P=0.0103). These results suggest that CD133 expression in pancreatic cancer was significantly associated with lymphatic metastasis, VEGF-C expression, and prognosis.
Assuntos
Antígenos CD/análise , Glicoproteínas/análise , Neoplasias Pancreáticas/química , Neoplasias Pancreáticas/patologia , Peptídeos/análise , Fator C de Crescimento do Endotélio Vascular/análise , Antígeno AC133 , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Imuno-Histoquímica , Queratinas/análise , Antígeno Ki-67/análise , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/irrigação sanguínea , Neoplasias Pancreáticas/mortalidade , Prognóstico , Receptores CXCR4/análiseRESUMO
Midkine (MK) is a heparin-binding growth factor and a product of a retinoic acid-responsive gene. Midkine is overexpressed in many carcinomas and thought to play an important role in carcinogenesis. However, no studies have been focussed on the role of MK in pancreatic carcinoma. This study sought to evaluate the clinical significance of MK expression in pancreatic head carcinoma, including the relationship between immunohistochemical expression and clinicopathologic factors such as prognosis. Immunohistochemical expression of MK and CD34 was evaluated in pancreatic head carcinoma specimens from 75 patients who underwent surgical resection. Midkine was expressed in 53.3% of patients. Midkine expression was significantly correlated with venous invasion, microvessel density, and liver metastasis (P=0.0063, 0.0025, and 0.0153, respectively). The 5-year survival rate was significantly lower for patients positive for MK vs patients negative for MK (P=0.0073). Multivariate analysis revealed that MK expression was an independent prognostic factor (P=0.0033). This is the first report of an association between MK expression and pancreatic head carcinoma. Midkine may play an important role in the progression of pancreatic head carcinoma, and evaluation of MK expression is useful for predicting malignant properties of pancreatic head carcinoma.
Assuntos
Citocinas/metabolismo , Neoplasias Pancreáticas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Western Blotting , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Hepáticas/secundário , Masculino , Pessoa de Meia-Idade , Midkina , Neoplasias Pancreáticas/patologia , Prognóstico , Análise de Sobrevida , Taxa de SobrevidaRESUMO
In this phase-I trial, we evaluated the safety of S-1, a novel oral fluoropyrimidine anticancer agent, combined with external-beam radiotherapy (EBRT) to determine the maximum-tolerated dose and dose-limiting toxicity (DLT) in unresectable pancreatic cancer patients. Patients had histologically proven unresectable locally advanced or metastatic pancreatic cancer. S-1 was administered orally twice daily. External-beam radiotherapy was delivered in fractions of 1.25 Gy x 2 per day, totalling 50 Gy per 40 fractions for 4 weeks. S-1 was given at five dose levels: 60 mg m(-2) day(-1) on days 1-7 and 15-21 (level 1), 1-14 (level 2), and 1-21 (level 3a) and 80 mg m(-2) day(-1) on days 1-21 (level 3b) and 1-28 (level 4). We studied 17 patients: dose levels 1 (four patients), 2 (four patients), 3a (three patients), 3b (three patients), and 4 (three patients). One patient in level 1 (grade 3 vomiting) and two patients in level 4 (grade 4 neutropenia and grade 3 anorexia) showed DLT. No DLT was seen for levels 2, 3a, and 3b. Clinical effects by computed tomography included 5 partial responses (35%), 11 cases of stable disease, and one case of progressive disease. CA19-9 levels of less than half the starting values were observed in 8 of 16 (50%) patients. S-1 at a dose of 80 mg m(-2) day(-1) given on days 1-21 is safe and recommended for phase-II study in patients with locally advanced and unresectable pancreatic cancer when given with EBRT.
Assuntos
Antimetabólitos Antineoplásicos/toxicidade , Ácido Oxônico/toxicidade , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/radioterapia , Radioterapia/métodos , Tegafur/toxicidade , Adulto , Idoso , Terapia Combinada , Relação Dose-Resposta a Droga , Combinação de Medicamentos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/mortalidade , Análise de SobrevidaRESUMO
In addition to its role in the adult mammalian nervous system as an inhibitory neurotransmitter, gamma-aminobutyric acid (GABA) is involved in the proliferation, differentiation, and migration of several kinds of cells including cancer cells. GABA is synthesized predominantly from glutamate by glutamate decarboxylase and exerts its effects via ionotropic GABA(A) receptors and/or metabotropic GABA(B) receptors. In this review, the current state of knowledge regarding the role of the GABAergic system in peripheral nonneuronal cell proliferation is described, and recent advances in elucidation of the mechanisms leading to cell proliferation are discussed.
Assuntos
Neoplasias/metabolismo , Neoplasias/patologia , Ácido gama-Aminobutírico/metabolismo , Animais , Diferenciação Celular , Movimento Celular , Proliferação de Células , Humanos , Modelos Biológicos , Neurônios/metabolismo , Neurotransmissores/metabolismo , Transdução de SinaisRESUMO
Glutamate decarboxylase (GAD), which has two isoforms, GAD65, and GAD67, is responsible for synthesis of the major inhibitory neurotransmitter, gamma-aminobutyric acid. GAD is expressed predominantly in the central nervous system; recent reports suggest that GAD is also expressed in non-neuronal organs including the pancreas. In the pancreatic islets, GAD serves as one of the autoantigens in type I diabetes mellitus. Recent flow cytometric analyses have shown that a variety of self-antigens, including GAD, are ectopically transcribed and expressed in particular cell populations of the thymus, although consensus concerning the cellular phenotype has not been obtained. The aim of this study was to clarify the localization and cellular phenotype of GAD67-expressing cells in the thymus at a cellular level with a novel approach using GAD67-green fluorescent protein (GFP) knock-in mice, in which GFP is expressed specifically in GAD67-positive cells. GFP-positive cells were detected in the thymic medulla and were identified as epithelial cells by immunohistochemistry. Almost all GFP-positive cells were positive for major histocompatibility complex (MHC) class II antigen staining and were positive for both cytokeratin and Ulex Europaeus Agglutinin I, markers of medullary thymic epithelial cells, but were negative for CD11c, Gr-1, and CD45, markers of dendritic cells, macrophages, and B-lymphocytes, respectively.
Assuntos
Células Apresentadoras de Antígenos/enzimologia , Células Epiteliais/metabolismo , Glutamato Descarboxilase/imunologia , Isoenzimas/metabolismo , Timo/enzimologia , Animais , Técnica Indireta de Fluorescência para Anticorpo , Corantes Fluorescentes , Expressão Gênica , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Imuno-Histoquímica , Queratinas/metabolismo , Camundongos , Camundongos Transgênicos , Microscopia de Fluorescência , Lectinas de Plantas/metabolismo , Proinsulina/metabolismo , Compostos de Quinolínio , Timo/citologiaRESUMO
We investigated the GABAergic system within the Sprague-Dawley rat (2-3-weeks old) trigeminal ganglion (TG). Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed expression of glutamate decarboxylase (GAD) 65 and GAD67 mRNAs and mRNAs encoding GABA(A) receptor subunits alpha1-6, beta1-3, gamma1-3, and delta. In situ hybridization revealed that GAD65 and GAD67 mRNAs were expressed in neuronal cell bodies but not satellite cells. Immunohistochemical analysis showed that only GAD65 was expressed in all neuronal cell bodies, and approximately 70% of all neurons exhibited GABA immunoreactivity. Satellite cells were strongly immunopositive for GABA. GABA(A) receptor alpha1, alpha5, beta2/3 and gamma1/2/3 subunit immunoreactivities were observed in the majority of neurons, but no immunoreactivity for alpha2 was observed. Two types of cells were identified in TG based on cell size and morphology, type A and B. The percentage of cells expressing alpha3, alpha4, alpha6, and delta subunits appeared to be dependent on cell size, as delta and alpha6 expression were only observed in small (B-type) neurons. In whole-cell patch clamp experiments, GABA application induced inward Cl- currents in all neurons examined. The EC50 for GABA varied from 5.3 to 240 microm, and the Hill Coefficient (nH) varied between 0.98 and 2.6 at -60 mV. We found that GABA was released from TG cells by increasing extracellular K+ concentration to 100 mm. We speculate that GABA acts as a nonsynaptically released diffusible neurotransmitter, which may modulate somatic inhibition of neurons within the TG.
Assuntos
Inibição Neural/fisiologia , Neurônios/metabolismo , Gânglio Trigeminal/citologia , Ácido gama-Aminobutírico/metabolismo , Animais , Animais Recém-Nascidos , Bicuculina/farmacologia , Contagem de Células/métodos , Células Cultivadas , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Capacitância Elétrica , Estimulação Elétrica/métodos , Epistasia Genética , Agonistas GABAérgicos/farmacologia , Antagonistas GABAérgicos/farmacologia , Glutamato Descarboxilase/metabolismo , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Isoenzimas/metabolismo , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos da radiação , Modelos Neurológicos , Muscimol/farmacologia , Inibição Neural/efeitos dos fármacos , Inibição Neural/efeitos da radiação , Neurônios/classificação , Neurônios/efeitos dos fármacos , Neurônios/efeitos da radiação , Técnicas de Patch-Clamp , Potássio/farmacologia , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de GABA-A/genética , Receptores de GABA-A/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Ácido gama-Aminobutírico/farmacologiaRESUMO
Expression of the EP4 receptor, a prostaglandin (PG)E2 receptor subtype, as well as disease suppression by the administration of a selective EP4 agonist (ONO-AE1-329) was investigated in the colorectal mucosa of rats with dextran sodium sulphate (DSS)-induced colitis. Rats were given drinking water containing 3% DSS for 2 weeks. Expression of EP4 receptor mRNA was barely detectable under normal conditions according to reverse transcription-polymerase chain reaction (RT-PCR). By 1 week after the initial administration of DSS, the receptor mRNA was strongly expressed. After ONO-AE1-329 was administered intracolonically to rats with DSS colitis for 7 consecutive days, erosion and ulceration decreased. Peripheral white blood cell (WBC) counts became less elevated. Interleukin (IL)-1beta and growth-regulated gene product/cytokine-induced neutrophil chemoattractant (GRO/CINC-1) concentrations in colorectal mucosa were lower than in colitis control group (IL-1beta: 12.8 +/- 4.6 and 30.8 +/- 6.2 microg/mg protein, P < 0.05; GRO/CINC-1: 15.5 +/- 3.0 and 39.2 +/- 5.4 microg/mg protein, P < 0.05), and the expression of the corresponding cytokine mRNA was strongly suppressed. IL-10 concentration was higher than in control group (14.5 +/- 1.7 and 7.9 +/- 1.2 microg/mg, P < 0.05), and the mRNA was more strongly expressed. These results suggest that the EP4 receptor is important in colonic inflammation, and that PGE2 suppresses DSS colitis at least partly via the EP4 receptor and the above cytokine changes. Intracolonic administration of selective EP4 agonist might have therapeutic applicability in inflammatory bowel disease such as ulcerative colitis.
Assuntos
Antiulcerosos/farmacologia , Quimiocinas CXC , Colite/imunologia , Dinoprostona/metabolismo , Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular , Éteres Metílicos/farmacologia , Receptores de Prostaglandina E/genética , Doença Aguda , Animais , Antiulcerosos/administração & dosagem , Células CHO , Quimiocina CXCL1 , Fatores Quimiotáticos/biossíntese , Fatores Quimiotáticos/genética , Fatores Quimiotáticos/imunologia , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Colo/enzimologia , Colo/imunologia , Colo/patologia , Cricetinae , Sulfato de Dextrana/efeitos adversos , Relação Dose-Resposta a Droga , Substâncias de Crescimento/biossíntese , Substâncias de Crescimento/genética , Substâncias de Crescimento/imunologia , Interleucina-1/biossíntese , Interleucina-1/genética , Interleucina-1/imunologia , Interleucina-10/biossíntese , Interleucina-10/genética , Interleucina-10/imunologia , Contagem de Leucócitos , Masculino , Éteres Metílicos/administração & dosagem , Peroxidase/metabolismo , RNA Mensageiro , Ratos , Ratos Sprague-Dawley , Receptores de Prostaglandina E/agonistas , Receptores de Prostaglandina E Subtipo EP2 , Receptores de Prostaglandina E Subtipo EP4 , Fatores de TempoRESUMO
The recent development of endothelin-1 (ET-1) antagonists and their potential use in the treatment of human disease raises questions as to the role of ET-1 in the pathophysiology of such cardiovascular ailments as hypertension, heart failure, renal failure and atherosclerosis. It is still unclear, for example, whether activation of an endogenous ET-1 system is itself the primary cause of any of these ailments. In that context, the phenotypic manifestations of chronic ET-1 overproduction may provide clues about the tissues and systems affected by ET-1. We therefore established two lines of transgenic mice overexpressing the ET-1 gene under the direction of its own promoter. These mice exhibited low body weight, diminished fur density and two- to fourfold increases in the ET-1 levels measured in plasma, heart, kidney and aorta. There were no apparent histological abnormalities in the visceral organs of young (8 weeks old) transgenic mice, nor was their blood pressure elevated. In aged (12 months old) transgenic mice, however, renal manifestations, including prominent interstitial fibrosis, renal cysts, glomerulosclerosis and narrowing of arterioles, were detected. These pathological changes were accompanied by decreased creatinine clearance, elevated urinary protein excretion and salt-dependent hypertension. It thus appears that mild, chronic overproduction of ET-1 does not primarily cause hypertension but triggers damaging changes in the kidney which lead to the susceptibility to salt-induced hypertension.
Assuntos
Envelhecimento/genética , Endotelina-1/biossíntese , Hipertensão/genética , Hipertensão/fisiopatologia , Nefropatias/genética , Nefropatias/fisiopatologia , Cloreto de Sódio na Dieta/metabolismo , Animais , Pressão Sanguínea/genética , Pressão Sanguínea/fisiologia , Creatinina/sangue , Creatinina/metabolismo , Endotelina-1/sangue , Endotelina-1/genética , Coração/fisiopatologia , Frequência Cardíaca/genética , Frequência Cardíaca/fisiologia , Hipertensão/sangue , Rim/irrigação sanguínea , Rim/fisiopatologia , Rim/ultraestrutura , Nefropatias/sangue , Masculino , Taxa de Depuração Metabólica/genética , Taxa de Depuração Metabólica/efeitos da radiação , Camundongos , Camundongos Transgênicos , Microinjeções/métodos , Microscopia Eletrônica de Varredura , Óvulo/química , Óvulo/crescimento & desenvolvimento , Óvulo/metabolismo , Fenótipo , Transgenes/genéticaRESUMO
The epiphyseal growth plate, where chondrocytes proliferate and differentiate, is the major site for longitudinal bone growth, matrix synthesis and mineralization. Glucose is an important energy source for the metabolism and growth of chondrocytes. The family of facilitative glucose transporters (GLUTs) mediates glucose transport across the plasma membrane in mammalian cells. We used immunocytochemical methods with anti-GLUT antibodies to investigate the localization of GLUTs in chondrocytes of the epiphyseal growth plate in 3 age groups of rats (3, 7, and 28 days after birth). Intense immunoreactivity of GLUT isoforms 1-5 was detected in chondrocytes of 3-day and 7-day old rats, and all GLUTs were localized in the maturation zone of the hypertrophic zone. On postnatal day 28, chondrocytes in the maturation zone showed intense GLUT1, 4 and 5 immunoreactivity, and weak GLUT2 and 3 immunoreactivity. In addition to chondrocytes in the maturation zone, those in the degenerative zone and in the zone of provisional calcification showed strong GLUT4 and 5 immunoreactivity. Autoradiography of bone sections from 4-week old mice injected with 14C-2-deoxyglucose showed high silver grain density within matrix tissue in the reserve and proliferative zones but not around chondrocytes. However, in the hypertrophic zone, silver grain density was high in matrix and chondrocytes. These data indicate that chondrocytes in the hypertrophic zones use glucose as energy source. High levels of GLUT4 expression imply that glucose use in chondrocytes is regulated by insulin. Expression of GLUT5 in chondrocytes suggests that fructose is also used as an energy source.
Assuntos
Condrócitos/metabolismo , Desoxiglucose/farmacocinética , Glucose/metabolismo , Lâmina de Crescimento/metabolismo , Proteínas de Transporte de Monossacarídeos/genética , Transcrição Gênica , Envelhecimento , Animais , Autorradiografia/métodos , Radioisótopos de Carbono , Transportador de Glucose Tipo 1 , Transportador de Glucose Tipo 2 , Imuno-Histoquímica/métodos , Masculino , Camundongos , Ratos , Ratos Wistar , Distribuição TecidualRESUMO
Heme oxygenase (HO)-1 degrades the pro-oxidant heme and generates carbon monoxide and antioxidant bilirubin. We have previously shown that in response to hypoxia, HO-1-null mice develop infarcts in the right ventricle of their hearts and that their cardiomyocytes are damaged by oxidative stress. To test whether HO-1 protects against oxidative injury in the heart, we generated cardiac-specific transgenic mice overexpressing different levels of HO-1. By use of a Langendorff preparation, hearts from transgenic mice showed improved recovery of contractile performance during reperfusion after ischemia in an HO-1 dose-dependent manner. In vivo, myocardial ischemia and reperfusion experiments showed that infarct size was only 14.7% of the area at risk in transgenic mice compared with 56.5% in wild-type mice. Hearts from these transgenic animals had reduced inflammatory cell infiltration and oxidative damage. Our data demonstrate that overexpression of HO-1 in the cardiomyocyte protects against ischemia and reperfusion injury, thus improving the recovery of cardiac function.
Assuntos
Heme Oxigenase (Desciclizante)/genética , Isquemia Miocárdica/prevenção & controle , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/enzimologia , Animais , Regulação Enzimológica da Expressão Gênica/fisiologia , Genótipo , Coração/fisiopatologia , Heme Oxigenase (Desciclizante)/metabolismo , Heme Oxigenase-1 , Humanos , Proteínas de Membrana , Camundongos , Camundongos Transgênicos , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/prevenção & controle , Isquemia Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologiaRESUMO
Aortic carboxypeptidase-like protein (ACLP) is a member of a diverse group of proteins that contain a domain with similarity to that of the Dictyostelium discoideum protein discoidin I. The discoidin domain has been identified in mammalian milk fat globule membrane proteins, blood coagulation factors, and receptor tyrosine kinases, where it may facilitate cell aggregation, adhesion, or cell-cell recognition. Here we show that ACLP is a secreted protein that associates with the extracellular matrix (ECM). During mouse embryogenesis, ACLP is abundantly expressed in the ECM of collagen-rich tissues, including the vasculature, dermis, and the developing skeleton. We deleted the ACLP gene in mice by homologous recombination. The majority of ACLP(-/-) mice die perinatally due to gastroschisis, a severe disruption of the anterior abdominal wall and herniation of the abdominal organs. ACLP(-/-) mice that survived to adulthood developed nonhealing skin wounds. Following injury by a dermal punch biopsy, ACLP(-/-) mice exhibited deficient wound healing compared with controls. In addition, dermal fibroblasts isolated from ACLP(-/-) 18.5-day-postconception embryos exhibited a reduced proliferative capacity compared with wild-type cells. These results indicate that ACLP is an ECM protein that is essential for embryonic development and dermal wound healing processes.
Assuntos
Músculos Abdominais/embriologia , Proteínas/genética , Proteínas/fisiologia , Cicatrização , Animais , Desenvolvimento Ósseo , Carboxipeptidases , Adesão Celular , Agregação Celular , Divisão Celular , Células Cultivadas , Clonagem Molecular , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Imuno-Histoquímica , Camundongos , Microscopia de Fluorescência , Modelos Genéticos , Músculo Liso/citologia , Mutagênese Sítio-Dirigida , Fenótipo , Estrutura Terciária de Proteína , Recombinação Genética , Proteínas Repressoras , Pele/metabolismo , Pele/patologia , Frações Subcelulares , Fatores de TempoRESUMO
Heme oxygenase (HO) is a cytoprotective enzyme that degrades heme (a potent oxidant) to generate carbon monoxide (a vasodilatory gas that has anti-inflammatory properties), bilirubin (an antioxidant derived from biliverdin), and iron (sequestered by ferritin). Because of properties of HO and its products, we hypothesized that HO would be important for the regulation of blood pressure and ischemic injury. We studied chronic renovascular hypertension in mice deficient in the inducible isoform of HO (HO-1) using a one kidney-one clip (1K1C) model of disease. Systolic blood pressure was not different between wild-type (HO-1(+/+)), heterozygous (HO-1(+/-)), and homozygous null (HO-1(-/-)) mice at baseline. After 1K1C surgery, HO-1(+/+) mice developed hypertension (140+/-2 mm Hg) and cardiac hypertrophy (cardiac weight index of 5.0+/-0.2 mg/g) compared with sham-operated HO-1(+/+) mice (108+/-5 mm Hg and 4.1+/-0.1 mg/g, respectively). However, 1K1C produced more severe hypertension (164+/-2 mm Hg) and cardiac hypertrophy (6.9+/-0.6 mg/g) in HO-1(-/-) mice. HO-1(-/-) mice also experienced a high rate of death (56%) within 72 hours after 1K1C surgery compared with HO-1(+/+) (25%) and HO-1(+/-) (28%) mice. Assessment of renal function showed a significantly higher plasma creatinine in HO-1(-/-) mice compared with HO-1(+/-) mice. Histological analysis of kidneys from 1K1C HO-1(-/-) mice revealed extensive ischemic injury at the corticomedullary junction, whereas kidneys from sham HO-1(-/-) and 1K1C HO-1(+/-) mice appeared normal. Taken together, these data suggest that chronic deficiency of HO-1 does not alter basal blood pressure; however, in the 1K1C model an absence of HO-1 leads to more severe renovascular hypertension and cardiac hypertrophy. Moreover, renal artery clipping leads to an acute increase in ischemic damage and death in the absence of HO-1.
Assuntos
Injúria Renal Aguda/patologia , Heme Oxigenase (Desciclizante)/deficiência , Hipertensão Renovascular/genética , Injúria Renal Aguda/sangue , Injúria Renal Aguda/complicações , Animais , Pressão Sanguínea/genética , Cardiomegalia/etiologia , Cardiomegalia/patologia , Doença Crônica , Creatinina/sangue , Modelos Animais de Doenças , Antagonistas dos Receptores de Endotelina , Endotelina-1/genética , Endotelina-1/metabolismo , Heme Oxigenase (Desciclizante)/genética , Heme Oxigenase (Desciclizante)/metabolismo , Heme Oxigenase-1 , Heterozigoto , Homozigoto , Hipertensão Renovascular/sangue , Hipertensão Renovascular/complicações , Imuno-Histoquímica , Rim/patologia , Proteínas de Membrana , Camundongos , Camundongos Knockout , Nefrectomia , Tamanho do Órgão , RNA Mensageiro/metabolismo , Receptor de Endotelina A , Obstrução da Artéria Renal/complicações , Índice de Gravidade de Doença , Taxa de SobrevidaRESUMO
Inactivation of glycogen synthase kinase 3beta (GSK3beta) and the resulting stabilization of free beta-catenin are critical steps in the activation of Wnt target genes. While Akt regulates GSK3alpha/beta in the phosphatidylinositide 3-OH kinase signaling pathway, its role in Wnt signaling is unknown. Here we report that expression of Wnt or Dishevelled (Dvl) increased Akt activity. Activated Akt bound to the Axin-GSK3beta complex in the presence of Dvl, phosphorylated GSK3beta and increased free beta-catenin levels. Furthermore, in Wnt-overexpressing PC12 cells, dominant-negative Akt decreased free beta-catenin and derepressed nerve growth factor-induced differentiation. Therefore, Akt acts in association with Dvl as an important regulator of the Wnt signaling pathway.
Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras , Transdução de Sinais/fisiologia , Transativadores , Proteínas de Peixe-Zebra , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteína Axina , Linhagem Celular , Proteínas do Citoesqueleto/metabolismo , Proteínas Desgrenhadas , Ativação Enzimática , Genes Reporter , Quinase 3 da Glicogênio Sintase , Quinases da Glicogênio Sintase , Humanos , Camundongos , Células PC12 , Fosforilação , Proteínas Tirosina Quinases/metabolismo , Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Ratos , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transfecção , Proteínas Wnt , beta CateninaRESUMO
In order to investigate the mucosal injury mechanism in UC, we made dextran sulfate sodium (DSS)-induced colitis in rat and examined pathological findings, MPO activity, PGE(2) level, and local mRNA expression and secretion of IL-1 beta, TNF-alpha, GRO/CINC-1 and IL-10 in DSS colitis mucosa. Moreover, we estimated the correlation between the severity of mucosal damage and changes of these local inflammatory mediators' values. Neutrophil infiltration was marked and MPO activity was locally increased in proportion to the severity of mucosal damage. The mRNA expression and secretion of IL-1 beta, GRO/CINC-1 and IL-10 were increased. Especially, the secretions of IL-1 beta and GRO/CINC-1 were increased in proportion to the severity of mucosal damage. However, those of TNF-alpha were not increased in the colitis mucosa. An abnormal macrophage function and the presence of macrophage subtypes producing different cytokines would be predicted from our TNF-alpha data. The lesion was less severe in the colonic mucosa with higher levels of endogenous PGE(2), while it was more severe in the colonic mucosa with lower levels of endogenous PGE(2), implicating this compound as an inhibitory factor against the development of inflammation in the affected mucosa. Our results suggest that PGE(2) might have therapeutic applicability to UC.
Assuntos
Colite/imunologia , Citocinas/metabolismo , Dinoprostona/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Animais , Anticoagulantes/administração & dosagem , Colite/fisiopatologia , Citocinas/imunologia , Sulfato de Dextrana/administração & dosagem , Dinoprostona/farmacologia , Modelos Animais de Doenças , Macrófagos/imunologia , Masculino , Infiltração de Neutrófilos , Peroxidase/metabolismo , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Índice de Gravidade de DoençaRESUMO
Adrenomedullin (AM) has been implicated as having hypotensive as well as protective effects on organs and vessels against different kinds of injuries. To elucidate the in vivo pathophysiological roles of adrenomedullin, we established transgenic mice (AMTg) overexpressing adrenomedullin driven by preproendothelin-1 promoter and adrenomedullin knockout mice (AMKO). Blood pressure in AMTg was significantly lower than that in wild-type mice, and AMTg was significantly resistant to lipopolysaccharide-induced septic shock and vascular injuries. On the other hand, heterozygotes of AMKO, AM(+/-), were fully viable and hypertensive as compared with wild littermates. Mice homozygous for adrenomedullin null mutation (AM-/-) were embryonic lethal, and no embryos could survive beyond the midterm of gestation. Collectively, our findings indicate the indispensable role of adrenomedullin in circulatory homeostasis and the organ protection as well as the fetal morphogenesis and the maintenance of pregnancy.
Assuntos
Peptídeos/genética , Adrenomedulina , Animais , Arteriosclerose/etiologia , Arteriosclerose/genética , Engenharia Genética/métodos , Camundongos , Camundongos Transgênicos , Peptídeos/efeitos adversos , Peptídeos/fisiologiaRESUMO
We recently isolated a novel bHLH/PAS protein, CLIF (cycle like factor), by yeast two-hybrid screening of human umbilical endothelial cell cDNA library. CLIF is preferentially expressed in endothelial and neuronal cells. Because CLIF is expressed in vascular endothelial cells and forms a heterodimer with CLOCK, the key transcription factor controlling the circadian rhythm, we hypothesized that CLIF regulates the circadian oscillation of PAI-1 gene expression in endothelial cells. Northern blot analysis of mouse organs showed circadian oscillations of PAI-I mRNA levels. In addition, the clock-related genes also showed circadian oscillation in peripheral tissues. In endothelial cells, the heterodimer of CLIF and CLOCK upregulated the PAI-1 gene expression through E-box sites. Furthermore, Period and Cryptochrome, which are negative regulators in the feedback loop of the biological clock, inhibited PAI-1 promoter activation by the CLOCK:CLIF heterodimer. These results suggest that the peripheral tissues have their own biological clock and CLIF regulates the circadian oscillation of PAI-1 gene expression in endothelial cells. This study suggests a novel molecular mechanism of the morning onset of myocardial infarction. Here we review our recent work and literature.
Assuntos
Ritmo Circadiano/fisiologia , Infarto do Miocárdio/fisiopatologia , Fatores de Transcrição ARNTL , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Regulação da Expressão Gênica , Humanos , Modelos Biológicos , Infarto do Miocárdio/genética , Inibidor 1 de Ativador de Plasminogênio/genética , Biossíntese de Proteínas , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transcrição GênicaRESUMO
We previously reported that intracolonic administration of enprostil, a prostaglandin-E(2) (PGE(2)) analogue, had therapeutic effects on acute colitis induced in rodents by dextran sulfate sodium (DSS). In addition, production of growth-regulated gene product/cytokine-induced neutrophil chemoattractant-1 [GRO/CINC-1; an interleukin(IL)-8 like cytokine] was suppressed in the inflamed tissues. In the present study we used a human colon cancer cell line (HT-29) to investigate enprostil effects on the IL-8 production of intestinal epithelial cells stimulated by various stimulants. In a MTT assay, concentrations of enprostil >10(-5)M had cytotoxitic effects on HT-29 cells. Furthermore, 10(-6) M enprostil suppressed IL-8 production in HT-29 cells, SW620 and CaCo2 stimulated with interleukin-1 beta (IL-1 beta) or lipopolysaccharide (LPS), but did not suppress this response when cells were stimulated with tumour necrosis factor (TNF)-alpha. These results suggest that enprostil affects a point in the pathway between the IL-1 receptor or LPS receptor and nuclear factor-kappa B(NF-kappa B), without affecting the pathway between the TNF receptor and NF-kappa B, with the latter factor being required for the IL-8 gene transcription. The therapeutic effect of exogenous enprostil on DSS colitis may involve the inhibition of IL-8 production in colonic epithelial cells stimulated by IL-1 beta or LPS.