RESUMO
It has been reported that anti-A and anti-B (ABO antibody) titers decrease with age, but little is known about the association between ABO antibody titers and physiologic/biochemical parameters such as body mass index (BMI), gamma-glutamyl transpeptidase (GGT), and total cholesterol (T-Cho). We investigated the present situation of ABO antibody titers among healthy blood donors in Japan and the physiologic/biochemical factors that may be associated with changes in ABO antibody titers. Plasma from 7450 Japanese blood donors was tested for ABO antibody titers using ABO reverse typing reagents by an automated microplate system; donor samples were classified into low, middle, and high titers according to the agglutination results obtained with diluted plasma samples. Multivariate regression analysis was performed to analyze the association between ABO antibody titers and age, gender, biochemical parameters (alanine transaminase [ALT], GGT, globulin, T-Cho, and glycosylated albumin [GA]), and BMI according to the ABO blood groups. A significant correlation between ABO antibody titers and age/gender, except for gender in anti-A of blood group B donors, was observed. BMI showed significant but negative correlations with anti-A and anti-B (ß = -0.085 and -0.062, respectively; p < 0.01) in blood group O donors. In addition, significant but negative correlations between GGT and T-Cho with anti-B of blood group A donors (ß = -0.055 and -0.047, respectively; p < 0.05) were observed. Although differences existed among the ABO blood groups, ABO antibody titers seem to be associated with physiologic and biochemical parameters of healthy individuals.
Assuntos
Sistema ABO de Grupos Sanguíneos , Doadores de Sangue , Humanos , Índice de Massa Corporal , Japão , Anticorpos , Incompatibilidade de Grupos SanguíneosRESUMO
OBJECTIVES: We investigated the effect of hydroxychloroquine (HCQ) on S100A8 and S100A9 serum levels in systemic lupus erythematosus (SLE) patients with low disease activity receiving immunosuppressants. METHODS: SELENA-SLEDAI, Cutaneous Lupus Erythematous Disease Area and Severity Index (CLASI) and serum levels of complement factors, anti-dsDNA antibodies, and white blood cell, lymphocyte, and platelet counts were used to evaluate disease activity, cutaneous disease activity, and immunological activity, respectively. Serum S100A8 and S100A9 were measured at HCQ administration and after 3 or 6 months using ELISA. RESULTS: S100A8 and S100A9 serum levels were elevated at baseline and the magnitude of decrease from baseline at 3 and 6 months after HCQ administration was greater in patients with renal involvement than in those without (baseline: S100A8, p = 0.034; S100A9, p = 0.0084; decrease: S100A8, p = 0.049; S100A9, p = 0.023). S100 modulation was observed in patients with (n = 17; S100A8, p = 0.0011; S100A9, p = 0.0002) and without renal involvement (n = 20; S100A8, p = 0.0056; S100A9, p = 0.0012), and was more apparent in patients with improved CLASI activity scores (improved: S100A8, p = 0.013; S100A9, p = 0.0032; unimproved: S100A8, p = 0.055; S100A9, p = 0.055). No associations were observed for immunological biomarkers. CONCLUSION: HCQ may improve organ involvement in SLE by modulating S100 protein levels, especially in patients with renal or skin involvement.
Assuntos
Antirreumáticos/uso terapêutico , Calgranulina A/sangue , Calgranulina B/sangue , Hidroxicloroquina/uso terapêutico , Lúpus Eritematoso Cutâneo/tratamento farmacológico , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Adulto , Biomarcadores/sangue , Feminino , Humanos , Lúpus Eritematoso Cutâneo/sangue , Lúpus Eritematoso Sistêmico/sangue , Nefrite Lúpica/sangue , Nefrite Lúpica/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
Intervertebral disc (IVD) cells are naturally exposed to high osmolarity and complex mechanical loading, which drive microenvironmental osmotic changes. Age- and degeneration-induced degradation of the IVD's extracellular matrix causes osmotic imbalance, which, together with an altered function of cellular receptors and signalling pathways, instigates local osmotic stress. Cellular responses to osmotic stress include osmoadaptation and activation of pro-inflammatory pathways. This review summarises the current knowledge on how IVD cells sense local osmotic changes and translate these signals into physiological or pathophysiological responses, with a focus on inflammation. Furthermore, it discusses the expression and function of putative membrane osmosensors (e.g. solute carrier transporters, transient receptor potential channels, aquaporins and acid-sensing ion channels) and osmosignalling mediators [e.g. tonicity response-element-binding protein/nuclear factor of activated T-cells 5 (TonEBP/NFAT5), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)] in healthy and degenerated IVDs. Finally, an overview of the potential therapeutic targets for modifying osmosensing and osmosignalling in degenerated IVDs is provided.
Assuntos
Inflamação/patologia , Disco Intervertebral/patologia , Osmorregulação , Osmose , Transdução de Sinais , Animais , Humanos , Concentração OsmolarRESUMO
BACKGROUND: Pasteurella species, widely known as indigenous organisms in the oral and gastrointestinal floras of many wild and domestic animals, are important pathogens in both animals and humans. Human infections due to Pasteurella species are in most cases associated with infected injuries following animal bites. We encountered a rare case of dual infections caused by different two Pasteurella species occurred in a previously healthy 25-year-old female sustaining injury by a dog-bite. METHODOLOGY: Exudates from the open wound of her dog-bite site, together with the saliva of the dog were submitted for bacteriological examination. Predominantly appearing grayish-white smooth colonies with almost the same colonial properties but slightly different glistening grown on chocolate and sheep blood agar plates were characterized morphologically by Gram's stain, biochemically by automated instrument using Vitek 2 system using GN cards together with commercially available kit system, ID-Test HN-20 rapid panels, and genetically by sequencing the 16S rRNA genes of the organism using a Taq DyeDeoxy Terminator Cycle Sequencing and a model 3100 DNA sequencer instrument. RESULTS: The causative isolates from the dog-bite site were finally identified as P. canis and P. dagmatis from the findings of the morphological, cultural, and biochemical properties together with the comparative sequences of the 16S rRNA genes. Both the isolates were highly susceptible to many antibiotics and the patient was successfully treated with the administration of so-called the first generation cephalosporin, cefazolin followed by so-called the third generation cephalosporin, cefcapene pivoxil. The isolate from the dog was subsequently identified as P. canis, the same species as the isolate from the patient. CONCLUSIONS: To the best of our knowledge, this was the second report of a dual infection with Pasteurella species consisting of P. dagmatis and P. canis resulting from a dog-bite, followed by the first report of dual infections due to P. dagmatis and P. multocida in 1988. Our isolate finally identified as P. dagmatis was misidentified as P. pneumotripica by means of the Vitek 2 system. The species name "P. dagmatis" was not included in the database of the system. It is also important for routine clinical microbiology laboratories to know the limitation of the automated Vitek 2 system for the accurate identification of Pasteurella species especially P. dagmatis. It should be emphasized that there still exists much room for improvement in Vitek 2 system. Significant improvement of Vitek 2 system especially in the identification of Pasteurella species is urgently desired.
Assuntos
Técnicas de Tipagem Bacteriana/métodos , Mordeduras e Picadas/microbiologia , Infecções por Pasteurella/microbiologia , Pasteurella/isolamento & purificação , Infecção dos Ferimentos/microbiologia , Adulto , Animais , Antibacterianos/uso terapêutico , Mordeduras e Picadas/complicações , Cefazolina/uso terapêutico , Cefalosporinas/uso terapêutico , Cães , Feminino , Humanos , Pasteurella/classificação , Infecções por Pasteurella/tratamento farmacológico , Infecções por Pasteurella/etiologia , Infecção dos Ferimentos/etiologiaRESUMO
Estrogen deficiency causes bone loss, which can be prevented by estrogen replacement therapy. Using a recently developed technique for isolation of highly purified mammalian osteoclasts, we showed that 17 beta-estradiol (E2) was able to directly inhibit osteoclastic bone resorption. At concentrations effective for inhibiting bone resorption, E2 also directly induced osteoclast apoptosis in a dose- and time-dependent manner. ICI164,384 and tamoxifen, as pure and partial antagonists, respectively, completely or partially blocked the effect of E2 on both inhibition of osteoclastic bone resorption and induction of osteoclast apoptosis. These data suggest that the protective effects of estrogen against postmenopausal osteoporosis are mediated in part by the direct induction of apoptosis of the bone-resorbing osteoclasts by an estrogen receptor- mediated mechanism.
Assuntos
Apoptose/efeitos dos fármacos , Reabsorção Óssea/prevenção & controle , Estrogênios/farmacologia , Osteoclastos/efeitos dos fármacos , Animais , Osteoclastos/fisiologia , Coelhos , Receptores de Estrogênio/fisiologiaRESUMO
Evans syndrome is a rare autoimmune disorder with unknown aetiology. Although corticosteroids and/or intravenous immunoglobulin (IVIG) are commonly used in its treatment, no standard strategy has been established. We report here a 44-year-old male with refractory Evans syndrome combined with systemic lupus erythematosus (SLE) who responded well to rituximab. He was admitted to our hospital with severe bleeding caused by worsening of Evans syndrome. Despite treatment with a high-dose corticosteroid and IVIG, his thrombocytopaenia and haemolytic anaemia did not improve. We started rituximab at a dose of 375 mg/m(2) once a week for a total of two doses. There was significant improvement in his thrombocytopaenia and anaemia 1 month after administration of rituximab. Although the total immunoglobulin G (IgG) level did not change, the titres of platelet-associated IgG (PA-IgG) and of an indirect antiglobulin test (IAT) decreased under the treatment with rituximab. It is suggested that rituximab would be a powerful candidate in the treatment of refractory Evans syndrome by depleting abnormal clone-producing autoantibody.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Antirreumáticos/uso terapêutico , Doenças Autoimunes/tratamento farmacológico , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Corticosteroides/uso terapêutico , Adulto , Anticorpos Monoclonais Murinos , Doenças Autoimunes/complicações , Relação Dose-Resposta a Droga , Resistência a Medicamentos , Humanos , Imunoglobulinas Intravenosas/uso terapêutico , Fatores Imunológicos/uso terapêutico , Lúpus Eritematoso Sistêmico/complicações , Masculino , Rituximab , SíndromeRESUMO
Mammalian chromatin remodeling factor, SWI/SNF complex contains a single molecule of either Brm or BRG1 as the ATPase catalytic subunit. Here, we show that the SWI/SNF complex forms a larger complex with neuron-restrictive silencer factor (NRSF) and its corepressors, mSin3A and CoREST, in human nonsmall cell lung carcinoma cell lines. We also demonstrate that the strong transcriptional suppression of such neuron-specific genes as synaptophysin and SCG10 by NRSF in these non-neural cells requires the functional SWI/SNF complex; these neuronal genes were elevated in cell lines deficient in both Brm and BRG1, whereas retrovirus vectors expressing siRNAs targeting integral components of SWI/SNF complex (Brm/BRG1 or Ini1) induced expression of these neuronal genes in SWI/SNF-competent cell lines. In cell lines deficient in both Brm and BRG1, exogenous Brm or BRG1 suppressed expression of these neuronal genes in an ATP-dependent manner and induced efficient and specific deacetylation of histone H4 around the NRSF binding site present in the synaptophysin gene by a large complex containing the recruited functional SWI/SNF complex. Patients with Brm/BRG1-deficient lung carcinoma have been reported to carry poor prognosis; derepression of NRSF-regulated genes including these neuron-specific genes could contribute to enhance tumorigenicity and also would provide selective markers for Brm/BRG1-deficient tumors.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Neurônios/metabolismo , Proteínas Repressoras/fisiologia , Fatores de Transcrição/fisiologia , Sequência de Bases , Western Blotting , Linhagem Celular Tumoral , Primers do DNA , Regulação da Expressão Gênica , Humanos , Imunoprecipitação , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Clarification of the interactions between carbon nanotubes (CNTs) and proteinogenic amino acids is a key approach to understanding CNT-protein interactions. Previous studies have addressed the mechanism of the physical adsorption of amino acids onto CNTs. However, little is known about their chemical reactions in aqueous solutions. Here, we established dispersant-free systems to clarify intrinsic CNT-thiol interactions. We demonstrated that the redox reaction of CNTs with cysteine, containing a thiol group, leads to disulfide bond formation between cysteine molecules, even under acidic conditions. The generality of the redox reaction is validated using other thiols such as dithiothreitol and glutathione. The present results suggest that structures of proteins and peptides containing free thiol groups are chemically modified and misfolded on CNT surfaces by this disulfide bond formation in biological systems.
RESUMO
Mutations of calreticulin (CALR) are detected in 25-30% of patients with essential thrombocythemia (ET) or primary myelofibrosis and cause frameshifts that result in proteins with a novel C-terminal. We demonstrate that CALR mutations activated signal transducer and activator of transcription 5 (STAT5) in 293T cells in the presence of thrombopoietin receptor (MPL). Human megakaryocytic CMK11-5 cells and erythroleukemic F-36P-MPL cells with knocked-in CALR mutations showed increased growth and acquisition of cytokine-independent growth, respectively, accompanied by STAT5 phosphorylation. Transgenic mice expressing a human CALR mutation with a 52 bp deletion (CALRdel52-transgenic mice (TG)) developed ET, with an increase in platelet count, but not hemoglobin level or white blood cell count, in association with an increase in bone marrow (BM) mature megakaryocytes. CALRdel52 BM cells did not drive away wild-type (WT) BM cells in in vivo competitive serial transplantation assays, suggesting that the self-renewal capacity of CALRdel52 hematopoietic stem cells (HSCs) was comparable to that of WT HSCs. Therapy with the Janus kinase (JAK) inhibitor ruxolitinib ameliorated the thrombocytosis in TG mice and attenuated the increase in number of BM megakaryocytes and HSCs. Taken together, our study provides a model showing that the C-terminal of mutant CALR activated JAK-STAT signaling specifically downstream of MPL and may have a central role in CALR-induced myeloproliferative neoplasms.
Assuntos
Calreticulina/genética , Animais , Autorrenovação Celular , Células HEK293 , Células-Tronco Hematopoéticas , Humanos , Janus Quinases/antagonistas & inibidores , Camundongos , Camundongos Transgênicos , Transtornos Mieloproliferativos/induzido quimicamente , Transtornos Mieloproliferativos/etiologia , Nitrilas , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas , Receptores de Trombopoetina , Fator de Transcrição STAT5/metabolismo , Trombocitemia Essencial/tratamento farmacológico , Trombocitemia Essencial/genéticaRESUMO
Glucocorticoid receptor (GR) has been shown to suppress activator protein 1 (AP-1)-mediated transcription by several molecular mechanisms. We previously showed that activation of endogenous AP-1 is essential for cellular transformation induced by oncogenes, such as v-src and c-Ha-ras. In the present study, we have analyzed whether high levels of GR expression suppress cellular transformation caused by these oncogenes. To eliminate the ligand effects that induce the transcriptional stimulation via glucocorticoid response elements, we constructed two GR mutants: CD-GR-1, lacking the COOH-terminal portion, including both the ligand and Hsp90-binding domains, and tau1TDCD-GR-1, a derivative of CD-GR-1 lacking the tau1 transactivation domain. When these GR mutants were expressed in chicken embryonic fibroblasts by retroviral vectors, they translocated into the nucleus without addition of glucocorticoid to the culture medium, and they suppressed cellular transformation caused by v-src and c-Ha-ras, as well as by c-fos and c-jun. Cellular transformation by v-myc was not suppressed by these mutants. Such suppressive effects of these GR mutants have a very similar oncogene dependency to that of dominant-negative mutants of AP-1. This suggests that GR can be altered to suppress cellular transformation by inhibiting endogenous AP-1 activity without activating glucocorticoid response elements.
Assuntos
Transformação Celular Neoplásica/genética , Genes ras , Genes src , Mutação , Receptores de Glucocorticoides/fisiologia , Fator de Transcrição AP-1/genética , Animais , Linhagem Celular Transformada , Embrião de Galinha , Regulação Neoplásica da Expressão Gênica , Vetores Genéticos , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-jun/genética , Retroviridae , Transcrição GênicaRESUMO
The expression of epidermal growth factor (EGF) receptor was examined immunohistochemically in a total of 122 gastric and 61 colonic carcinomas, out of which 16 gastric and 8 colonic carcinomas were also examined by 125I-labeled EGF binding analysis and Western blotting. The values of EGF binding were 12.68 +/- 1.98 (SE; n = 16) fmol/mg protein in gastric carcinomas and 5.72 +/- 2.15 (n = 8) fmol/mg protein in nonneoplastic gastric mucosa, the difference being significant (P less than 0.01). In the colonic tissue, the binding capacities in carcinomas and nonneoplastic mucosa were 13.29 +/- 4.17 (n = 8) and 10.68 +/- 0.41 (n = 3) fmol/mg protein, respectively. Scatchard analysis of 125I-labeled EGF binding indicated a single class of receptors in gastric and colonic carcinomas with an apparent Kd value of from 111 to 277 (n = 4) and from 87.4 to 341 fM (n = 5), respectively, except for one gastric carcinoma having two classes of receptors (Kd = 15.9 and 896 fM). In Western blotting using monoclonal anti-EGF receptor antibody, various levels of EGF receptor expression were detected in 12 (85.7%) of the 14 gastric carcinomas and in 7 (87.5%) of the 8 colonic carcinomas. Immunohistochemically, EGF receptor immunoreactivity was detected in one (3.8%) of the 26 early gastric carcinomas, while it was observed in 33 (34.4%) of the 96 advanced gastric carcinomas, the incidence between the two being significantly different (P less than 0.01). In the colonic carcinomas, 47 (77.1%) of the 61 cases showed positive immunoreactivity to EGF receptor, which did not differ by histological type.
Assuntos
Carcinoma/análise , Neoplasias do Colo/análise , Receptores ErbB/análise , Neoplasias Gástricas/análise , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/imunologia , Humanos , Imuno-Histoquímica , Radioisótopos do Iodo , Coloração e RotulagemRESUMO
The expression of p185ERBB2 in a total of 34 human gastric carcinoma tissues as well as in corresponding normal mucosa was examined by Western blotting. More than 70% of both tumor tissues and normal mucosa showed p185ERBB2 expression at various levels. Eighteen (55%) cases revealed higher levels of p185ERBB2 in the tumor than in normal mucosa, while 13 (38%) cases showed lower levels in the tumor tissues. Higher expression of p185ERBB2 was frequently observed in well differentiated adenocarcinomas, with the incidence between well differentiated type and poorly differentiated type being significantly different (P less than 0.05). Comparative immunohistochemical analysis revealed the consistent results with p185ERBB2 expression obtained by Western blotting in well differentiated adenocarcinomas. Of the 34 cases, three well differentiated adenocarcinomas had extremely high levels of p185ERBB2. ERBB2 gene was amplified in two of the three tumors, but the amplification differed by the tumor site from where the sample was obtained. Another tumor which showed an extremely high level of p185ERBB2 but no gene amplification demonstrated a high level of binding protein to the TATA box that is located in the promoter of the ERBB2 gene. A high level of TATA-binding protein was also detected in gastric carcinoma cell lines which contain a single copy of ERBB2 gene and a high expression of p185ERBB2.
Assuntos
Amplificação de Genes , Proteínas Proto-Oncogênicas/genética , Proto-Oncogenes , Neoplasias Gástricas/genética , Northern Blotting , Linhagem Celular , Expressão Gênica , Humanos , Immunoblotting , Estadiamento de Neoplasias , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas/isolamento & purificação , Proteínas Proto-Oncogênicas/metabolismo , RNA Mensageiro/genética , Receptor ErbB-2 , Neoplasias Gástricas/enzimologia , Neoplasias Gástricas/patologia , Transcrição GênicaRESUMO
fra-2 (fos-related antigen-2) expression is detected at a basal level even in growth-arrested chicken embryo fibroblasts (CEF), but upon serum-stimulation high levels of its transcripts are transiently observed. This induction is delayed and prolonged compared to that of c-fos. Transient expression experiments in CEF using a series of constructs of chicken fra-2 promoter region linked to the CAT reporter gene indicated previously that serum response element (SRE) is not required for full serum inducibility. In this report, we show that constructs in which the CRE-like sequence and both AP-1 binding sites are disrupted lack serum inducibility, suggesting that either of these enhancers is important in serum induction of fra-2. In growth-arrested CEF, small amounts of Fra-2/c-Jun complex bind to the AP-1 consensus sequences in fra-2 promoter, while a significant part of the enhanced AP-1 binding activity after 60-120 min of serum stimulation is attributable to c-Fos/c-Jun heterodimer. At later times Fra-2/c-Jun again becomes the main complex. Transient expression assays in F9 cells indicated that c-Fos/c-Jun heterodimers have strong stimulatory effects on fra-2 promoter activity, while Fra-2/c-Jun complex has lower transcriptional activity than that of c-Jun homodimer. These results suggest that c-Fos (induced at earlier times) and c-Jun proteins are at least partly responsible for serum-induced expression of fra-2.
Assuntos
Proteínas de Ligação a DNA/genética , Proteínas Proto-Oncogênicas c-fos/genética , Fator de Transcrição AP-1/fisiologia , Fatores de Transcrição/genética , Animais , Sequência de Bases , Sítios de Ligação , Ciclo Celular , Células Cultivadas , Embrião de Galinha , Análise Mutacional de DNA , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/fisiologia , Antígeno 2 Relacionado a Fos , Expressão Gênica , Genes fos , Genes jun , Técnicas In Vitro , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-fos/biossíntese , Fatores de Transcrição/biossínteseRESUMO
A pituitary glycoprotein hormone FSH stimulates ovarian granulosa cells to induce ovarian follicular development. In this study we identified rat ovarian genes that were rapidly induced by FSH in the cultured rat granulosa cells by means of subtraction cloning. Complementary DNA clones encoding cAMP responsive element binding modulator (CREM) were identified as one of the FSH inducible genes. Northern blotting and reverse transcription and polymerase chain reaction (RT-PCR) analyses revealed that only the repressor type of CREM gene products, ICER (inducible cAMP early repressor) isoforms, were induced by FSH treatment in cultured rat granulosa cells. The induction of ICER by FSH was mimicked by reagents known to increase intracellular cAMP levels, indicating that the induction is through cAMP and protein kinase A signal transduction system. Induction of ICER was also confirmed as the protein levels. Electrophoretic mobility shift assay of granulosa cell extracts with a radiolabeled double stranded oligonucleotide corresponding to somatostatin cAMP responsive element also revealed that only the ICER proteins were induced by FSH treatment, whereas levels of CREM proteins were nearly constant regardless of the FSH treatment. Our present study demonstrates that FSH-induced and cAMP-mediated induction and attenuation of transcriptional responses by CREM gene products may be a key mechanistic component for the granulosa cell differentiation and proliferation.
Assuntos
Proteínas de Ligação a DNA/genética , Células da Granulosa/metabolismo , Ovário/metabolismo , Proteínas Repressoras , Animais , Northern Blotting , Células Cultivadas , Clonagem Molecular , AMP Cíclico/metabolismo , Modulador de Elemento de Resposta do AMP Cíclico , Feminino , Hormônio Foliculoestimulante/farmacologia , Regulação da Expressão Gênica , Isoformas de Proteínas/genética , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Our studies using immature rat granulosa cells cultured in serum-free medium on collagen-coated dishes indicated that FSH receptor mRNA levels do not change for at least 4 days of culture in the absence of hormone treatment. Addition of FSH (30 ng ml[-1]) led to a reduction of FSH receptor mRNA for a short time (6 h), followed by an increase in FSH receptor mRNA levels that reached maximum of around 200% of the initial level within 2-3 days after the addition of FSH. Following the addition of 10 nM PMA, FSH receptor mRNA levels were decreased to 50% of the pretreatment levels. During prolonged exposure to PMA, gradual recovery of the FSH receptor mRNA level was observed, and it was significantly higher than the control level at 48 h. The inactive phorbol ester 4 alpha-phorbol-12,13-didecanoate did not depress FSH receptor mRNA levels. Downregulation of the FSH receptor mRNA was detectable at a PMA concentration of 1 nM. The two predominant FSH receptor mRNA transcripts, ca. 5.5 and 2.4 kb, respectively, appeared to be equally affected by SH and PMA treatments. To examine the role of PKC mediation of the effect of FSH on FSH receptor mRNA levels, granulosa cells were treated with the PKC inhibitor, H-7, and FSH. Although, FSH receptor mRNA levels decreased to 50% of control in the cells treated with FSH alone, the addition of H-7 (0.1 nM) caused no decline in FSH receptor mRNA levels relative to the control in the cells treated with FSH. On the other hand, inhibition of FSH receptor mRNA by FSH was partially suppressed by the PKC-selective inhibitor bisindolylmaleimide. The mRNA turnover experiments showed that the half-life of FSH receptor transcripts was unaffected by PMA exposure.
Assuntos
Hormônio Foliculoestimulante/farmacologia , Regulação da Expressão Gênica/genética , Células da Granulosa/metabolismo , Receptores do FSH/metabolismo , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina/farmacologia , Animais , Northern Blotting , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Sondas de DNA , Regulação para Baixo/fisiologia , Feminino , Células da Granulosa/efeitos dos fármacos , Indóis/farmacologia , Maleimidas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Receptores do FSH/genética , Acetato de Tetradecanoilforbol/farmacologia , Transcrição Gênica/genéticaRESUMO
OBJECTIVES: In our institute, internal mammary arteries (IMAs) have been preferred for coronary artery bypass grafting (CABG) in diabetic patients. The purpose of this study was to evaluate the influence of diabetes and IMA grafting on survival after CABG. BACKGROUND: The influence of diabetes on the results of CABG is not well documented, and there is controversy about whether the use of IMAs conveys greater survival benefits to diabetic patients. METHODS: A total of 420 consecutive patients who underwent CABG from April 1990 to July 1998 were reviewed; 211 of these patients had diabetes mellitus at the time of surgery. Internal mammary artery grafts have been used with increasing frequency, and bilateral IMAs have been used when possible since 1993. Internal mammary artery grafts were used in 164 nondiabetic patients (78%) and in 155 diabetic patients (73%). Seventy-eight nondiabetic patients and 74 diabetic patients received bilateral IMA grafts. RESULTS: The postoperative mortality was 2.4% in the nondiabetic and 2.8% in the diabetic group. With regard to postoperative complications, diabetic patients had a significantly higher rate of chest wound infection (p < 0.05), irrespective of whether IMAs were used or not. The use of bilateral IMAs did not increase the risk of chest wound infection in nondiabetic or diabetic patients. Overall survival curve, cardiac death-free curve and cardiac event-free curve were not affected adversely by diabetes, and in diabetic patients, CABG with saphenous veins alone conveyed significantly (p < 0.01) less long-term benefit than did CABG with at least one IMA graft. CONCLUSIONS: It was suggested that IMA grafts should be preferred in diabetic patients.
Assuntos
Ponte de Artéria Coronária/métodos , Complicações do Diabetes , Artéria Torácica Interna/transplante , Adulto , Idoso , Idoso de 80 Anos ou mais , Ponte de Artéria Coronária/efeitos adversos , Ponte de Artéria Coronária/mortalidade , Doença das Coronárias/complicações , Doença das Coronárias/cirurgia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Infecção da Ferida Cirúrgica , Taxa de SobrevidaRESUMO
The structure of a crystalline form of Bombyx mori silk fibroin, commonly found before the spinning process (known as silk I), was proposed by combining data obtained from two-dimensional spin-diffusion nuclear magnetic resonance under off magic angle spinning, rotational-echo double-resonance (REDOR), previously reported X-ray diffraction analyses and 13C NMR chemical shifts. Instead of B. mori silk fibroin with silk I structure, we used the sequential model peptide (Ala-Gly)15. The structure of the sequential model peptide is characterized as silk I after dissolving the peptide in 9 M LiBr and then dialyzing against water. Moreover, 13C or 15N-labeled sites may be introduced easily at any position in (Ala-Gly)(15) by the solid phase synthesis method for these NMR experiments. The torsional angles of (Ala-Gly)15 with silk I structure were determined as (-60(+/-5) degrees, 130(+/-5) degrees ) and (70(+/-5) degrees, 30(+/-5) degrees ) for Ala and Gly residues, respectively. The formation of the intra-molecular hydrogen bonding along the chain was confirmed from REDOR NMR by determination of the inter-atomic distance between the nitrogen and carbon atoms comprising the intra-molecular hydrogen bonding. The structure is named a repeated beta-turn type II-like structure.
Assuntos
Bombyx/química , Fibroínas/química , Proteínas de Insetos/química , Espectroscopia de Ressonância Magnética/métodos , Peptídeos/química , Alanina/química , Alanina/metabolismo , Sequência de Aminoácidos , Animais , Diálise , Difusão , Glicina/química , Glicina/metabolismo , Ligação de Hidrogênio , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Secundária de Proteína , Rotação , Seda , Água/metabolismo , Difração de Raios XRESUMO
Activin receptors (ActRs) and gonadotropin receptor mRNA expression were investigated in 18 human ovarian epithelial neoplasms. Northern blot analysis showed the presence of 3.0-kb type Ia ActR, 6.0- and 3.0-kb type IIa ActR, and 5.0-kb type IIb ActR mRNA transcripts in total RNA prepared from the cancer tissues. One carcinoma showed two major transcripts of a follicle-stimulating hormone receptor (FSH-R) gene, 4.1 and 2.4 kb, whereas the other two carcinomas showed two major transcripts of the luteinizing hormone/human chorionic gonadotropin receptor (LH-R) gene, 5.4 and 2.4 kb. These results were further analyzed by studying the corresponding PCR-amplified FSH and LH-R cDNA obtained by reverse transcription of total RNA. Expression of FSH-R mRNA was confirmed in about half of the cancer tissues. The size of the FSH-R reverse transcription-PCR product was the same as in normal ovarian follicles. Similarly, expression of LH-R mRNA was also detected in about half of the cancers. Normal ovaries and cancer tissues were homogenized, and activin concentrations were measured in extracts. Activin levels in normal ovarian tissue were around 0.59 +/- 0.01 ng/mg protein (mean +/- SE; n = 5), and activin production was detected in every cancer tissue, except one--serous adenocarcinoma. The findings in this study demonstrated that activin and ActRs are present in and synthesized by human ovarian epithelial neoplasms. Thus, activin seems to be available as an autocrine/paracrine factor in epithelial neoplasms and may contribute to the expression of FSH-R, although the roles of activin and gonadotropin in tumorigenesis has yet to be defined.
Assuntos
Carcinoma/genética , Neoplasias Ovarianas/genética , Receptores da Gonadotropina/genética , Receptores de Fatores de Crescimento/genética , Transcrição Gênica , Receptores de Ativinas , Ativinas , Adulto , Idoso , Northern Blotting , Carcinoma/química , Carcinoma/classificação , Carcinoma/patologia , Feminino , Humanos , Inibinas/análise , Pessoa de Meia-Idade , Neoplasias Ovarianas/química , Neoplasias Ovarianas/patologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
To address the relation between osteoblast growth and cell-to-cell communication, we examined the effects of basic fibroblast growth factor (bFGF) and 12-O-tetradecanoylphorbol-13-acetate (TPA), both potent stimulators of osteoblastic proliferation, on gap junctional intercellular communication between osteoblastic MC3T3-E1 cells. The level of intercellular communication was estimated by a photobleaching method. TPA inhibited the degree of intercellular communication in two different time-dependent manners. The early (< 1 h) inhibition by TPA was consistent with an increase in the phosphorylation of connexin 43 (Cx43). The later inhibition was caused by reduction in the total amount of Cx43 on the plasma membrane, due to the decrease in the level of Cx43 transcripts. These qualitative and quantitative modulations by TPA were inhibited by a selective inhibitor of protein kinase C, GF109203X. bFGF also attenuated the gap junctional intercellular communication. However, short exposure (< 5 h) to bFGF did not affect the communication. The fact that the growth factor immediately stimulated the phosphorylation of Cx43 indicates that the phosphorylation site(s) affected by bFGF was not involved in the inhibition of communication. The decrease in the intercellular communication level was detected by the longer exposure (> 8 h) to bFGF and paralleled the decline in the Cx-mRNA level. This inhibitory effect of bFGF was abolished by the addition of a tyrosine kinase inhibitor, herbimycin A. Thus, gap junctional intercellular communication between osteoblasts was down-regulated by osteoblastic mitogens through different mechanisms of the modulation of Cx43.