RESUMEN
Flocculation has been recognized for hundreds of years as an important phenomenon in brewing and wastewater treatment. However, the underlying molecular mechanisms remain elusive. The lack of a distinct phenotype to differentiate between slow-growing mutants and floc-forming mutants prevents the isolation of floc-related gene by conventional mutant screening. To overcome this, we performed a two-step Escherichia coli mutant screen. The initial screen of E. coli for mutants conferring floc production during high salt treatment yielded a mutant containing point mutations in 61 genes. The following screen of the corresponding single-gene mutants identified two genes, mrcB, encoding a peptidoglycan-synthesizing enzyme and cpxA, encoding a histidine kinase of a two-component signal transduction system that contributed to salt tolerance and flocculation prevention. Both single mutants formed flocs during high salt shock, these flocs contained cytosolic proteins. ΔcpxA exhibited decreased growth with increasing floc production and addition of magnesium to ΔcpxA suppressed floc production effectively. In contrast, the growth of ΔmrcB was inconsistent under high salt conditions. In both strains, flocculation was accompanied by the release of membrane vesicles containing inner and outer membrane proteins. Of 25 histidine kinase mutants tested, ΔcpxA produced the highest amount of proteins in floc. Expression of cpxP was up-regulated by high salt in ΔcpxA, suggesting that high salinity and activation of CpxR might promote floc formation. The finding that ΔmrcB or ΔcpxA conferred floc production indicates that cell envelope stress triggered by unfavorable environmental conditions cause the initiation of flocculation in E. coli.
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Membrana Celular/metabolismo , Pared Celular/genética , Proteínas de Escherichia coli/metabolismo , Escherichia coli/genética , Proteínas de Unión a las Penicilinas/metabolismo , Peptidoglicano Glicosiltransferasa/metabolismo , Proteínas Quinasas/metabolismo , Tolerancia a la Sal/genética , D-Ala-D-Ala Carboxipeptidasa de Tipo Serina/metabolismo , Proteínas Bacterianas/metabolismo , Pared Celular/metabolismo , Citosol/metabolismo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Floculación , Proteínas de la Membrana/metabolismo , Proteínas de Unión a las Penicilinas/genética , Peptidoglicano Glicosiltransferasa/genética , Mutación Puntual , Proteínas Quinasas/genética , D-Ala-D-Ala Carboxipeptidasa de Tipo Serina/genéticaRESUMEN
BACKGROUND: It is very important to understand the acetabular morphology of the normal hip joint to assist in diagnosis and surgical planning of hip disorders. The purpose of the present study was to obtain gender-based reference values for the acetabular measurements of a normal hip using computed tomography data and investigate the effect of aging on the measurement values. METHODS: We measured acetabular parameters (center-edge angle, Sharp angle, vertical center anterior angle, acetabular anteversion) on computed tomography corrected for changing the obliquity, rotation, and tilt of the pelvis. We performed measurements in 245 patients (490 joints; 120 men [240 joints] and 125 women [250 joints]). The mean age was 64.7 ± 14.3 (31-88) years for men and 63.2 ± 15.2 (30-88) years for women. RESULTS: In men and women, the mean center-edge angle was 31.8° ± 6.4° and 30.6° ± 6.5°, the mean Sharp angle was 38.6° ± 3.2° and 40.6° ± 3.8°, the mean vertical center anterior angle was 44.3° ± 7.9° and 40.0° ± 8.5°, and the mean acetabular anteversion angle was 14.3° ± 5.2° and 18.8° ± 5.4°, respectively. All differences were statistically significant. The center-edge angle increased with age in women; however, such an effect was not observed in men. The other measurements showed a similar trend, such as larger vertical center anterior angle and smaller Sharp and acetabular anteversion angles, with aging in both men and women. CONCLUSIONS: We used computed tomography data to quantitatively assess the coverage and shape of the acetabulum in adult Japanese subjects and obtain the estimated reference ranges by gender. The results also proved that the measurements changed with aging in both sexes. These facts must be taken into account during the diagnosis of hip disease and planning of surgery.
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Acetábulo , Caracteres Sexuales , Acetábulo/diagnóstico por imagen , Adulto , Anciano , Envejecimiento , Femenino , Humanos , Japón , Masculino , Persona de Mediana Edad , Valores de Referencia , Tomografía Computarizada por Rayos XRESUMEN
The phototropic bacterium Synechocystis sp. strain PCC 6803 is able to adapt its morphology in order to survive in a wide range of harsh environments. Under conditions of high salinity, planktonic cells formed cell aggregates in culture. Further observations using crystal violet staining, confocal laser scanning microscopy, and field emission-scanning electron microscopy confirmed that these aggregates were Synechocystis biofilms. Polyamines have been implicated in playing a role in biofilm formation, and during salt stress the content of spermidine, the major polyamine in Synechocystis, was reduced. Two putative arginine decarboxylases, Adc1 and Adc2, in Synechocystis were heterologously expressed in Escherichia coli and purified. Adc2 had high arginine decarboxylase activity, whereas Adc1 was much less active. Disruption of the adc genes in Synechocystis resulted in decreased spermidine content and formation of biofilms even under nonstress conditions. Based on the characterization of the adc mutants, Adc2 was the major arginine decarboxylase whose activity led to inhibition of biofilm formation, and Adc1 contributed only minimally to the process of polyamine synthesis. Taken together, in Synechocystis the shift from planktonic lifestyle to biofilm formation was correlated with a decrease in intracellular polyamine content, which is the inverse relationship of what was previously reported in heterotroph bacteria.IMPORTANCE There are many reports concerning biofilm formation in heterotrophic bacteria. In contrast, studies on biofilm formation in cyanobacteria are scarce. Here, we report on the induction of biofilm formation by salt stress in the model phototrophic bacterium Synechocystis sp. strain PCC 6803. Two arginine decarboxylases (Adc1 and Adc2) possess function in the polyamine synthesis pathway. Inactivation of the adc1 and adc2 genes leads to biofilm formation even in the absence of salt. The shift from planktonic culture to biofilm formation is regulated by a decrease in spermidine content in Synechocystis This negative correlation between biofilm formation and polyamine content, which is the opposite of the relationship reported in other bacteria, is important not only in autotrophic but also in heterotrophic bacteria.
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Proteínas Bacterianas/genética , Biopelículas/crecimiento & desarrollo , Carboxiliasas/genética , Espermidina/análisis , Synechocystis/genética , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Regulación Enzimológica de la Expresión Génica , Silenciador del Gen , Synechocystis/enzimologíaRESUMEN
BACKGROUND: Intraoperative oxygen management is poorly understood. It was hypothesized that potentially preventable hyperoxemia and substantial oxygen exposure would be common during general anesthesia. METHODS: A multicenter, cross-sectional study was conducted to describe current ventilator management, particularly oxygen management, during general anesthesia in Japan. All adult patients (16 yr old or older) who received general anesthesia over 5 consecutive days in 2015 at 43 participating hospitals were identified. Ventilator settings and vital signs were collected 1 h after the induction of general anesthesia. We determined the prevalence of potentially preventable hyperoxemia (oxygen saturation measured by pulse oximetry of more than 98%, despite fractional inspired oxygen tension of more than 0.21) and the risk factors for potentially substantial oxygen exposure (fractional inspired oxygen tension of more than 0.5, despite oxygen saturation measured by pulse oximetry of more than 92%). RESULTS: A total of 1,786 patients were found eligible, and 1,498 completed the study. Fractional inspired oxygen tension was between 0.31 and 0.6 in 1,385 patients (92%), whereas it was less than or equal to 0.3 in very few patients (1%). Most patients (83%) were exposed to potentially preventable hyperoxemia, and 32% had potentially substantial oxygen exposure. In multivariable analysis, old age, emergency surgery, and one-lung ventilation were independently associated with increased potentially substantial oxygen exposure, whereas use of volume control ventilation and high positive end-expiratory pressure levels were associated with decreased potentially substantial oxygen exposure. One-lung ventilation was particularly a strong risk factor for potentially substantial oxygen exposure (adjusted odds ratio, 13.35; 95% CI, 7.24 to 24.60). CONCLUSIONS: Potentially preventable hyperoxemia and substantial oxygen exposure are common during general anesthesia, especially during one-lung ventilation. Future research should explore the safety and feasibility of a more conservative approach for intraoperative oxygen therapy.
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Anestesia General/métodos , Monitoreo Intraoperatorio/métodos , Terapia por Inhalación de Oxígeno/métodos , Respiración Artificial/métodos , Ventiladores Mecánicos , Anciano , Anestesia General/efectos adversos , Anestesia General/normas , Estudios Transversales , Femenino , Humanos , Hiperoxia/inducido químicamente , Hiperoxia/prevención & control , Masculino , Persona de Mediana Edad , Monitoreo Intraoperatorio/normas , Ventilación Unipulmonar/efectos adversos , Ventilación Unipulmonar/métodos , Ventilación Unipulmonar/normas , Terapia por Inhalación de Oxígeno/efectos adversos , Terapia por Inhalación de Oxígeno/normas , Estudios Prospectivos , Respiración Artificial/efectos adversos , Respiración Artificial/normas , Ventiladores Mecánicos/normasRESUMEN
Intact ß2-glycoprotein I (iß2GPI) is a glycoprotein that regulates coagulation and fibrinolysis. Nicked ß2GPI (nß2GPI) possesses an angiogenic property at a relatively low concentration, and an antiangiogenic property at a high concentration. Here we investigated the functions of ßi 2GPI and nß2GPI in vascular endothelial growth factor (VEGF)-A-induced endothelial cell proliferation and tube formation. We used noninvasive PET imaging to analyze the ï½ï½ãï½ï½ï½ï½ distribution of intravenously injected ß2GPI variants in tumor lesions in mice. iß2GPI was incubated with plasmin to obtain nß2GPI, and its N-terminal sequence was analyzed. nß2GPI had at least one other cleavage site upstream of the ß2GPI's domain V, whereas the former plasmin-cleavage site locates between K317 and T318. Both of intact and nicked ß2GPI significantly inhibited the VEGF-A-induced cell proliferation and the tube formation of human umbilical vein endothelial cells (HUVECs). PET imaging visualized considerably distributed intensities of all tested ß2GPI variants in tumor lesions of pancreatic tumor cell-xenografts. These results indicate that ß2GPI may be physiologically and pathophysiologically important in the regulation of not only coagulation and fibrinolysis, but also angiogenesis.
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Inhibidores de la Angiogénesis/metabolismo , Neovascularización Patológica/metabolismo , Factores de Crecimiento Endotelial Vascular/metabolismo , beta 2 Glicoproteína I/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Ensayo de Inmunoadsorción Enzimática , Humanos , Ratones , Neoplasias Pancreáticas/metabolismo , Tomografía de Emisión de Positrones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We retrospectively assessed the effectiveness and the safety of thoracic paravertebral block(PVB) in patients ineligible for epidural block (EP). Eleven PVB patients and 33 EP patients were enrolled. Postoperative pain was evaluated using a numerical rating scale (NRS). The mean NRS ± standard deviation at rest 24 and 48 hours after surgery were 1.36 ± 1.63 and 0.55 ± 1.03 in the PVB group and 1.07 ± 1.47 and 1.38 ± 1.31 in the EP group, respectively. There were no statistically significant differences in the NRS scores. Approximately 10% of the EP patients had complications such as hypotension, nausea and vomiting, or urinary retention. On the other hand, there were no adverse events in the PVB group. PVB can provide pain relief comparable to EP with a better side-effect profile. There were no technical complications associated with PVB. Thoracic PVB is an effective and safe method of postoperative analgesia for patients undergoing thoracic surgery with ineligibilities for EP.
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Anestesia Raquidea , Columna Vertebral/efectos de los fármacos , Procedimientos Quirúrgicos Torácicos , Adulto , Anciano , Anciano de 80 o más Años , Anestesia Epidural , Anestesia Raquidea/métodos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
AIM: Radiofrequency ablation therapy (RFA) combined with transarterial chemoembolization (TACE) (combination therapy) is effective for early-stage hepatocellular carcinoma (HCC). The aim of this study was to compare the long-term effects of combination therapy with supportive care alone for intermediate HCC. METHODS: The study included 58 patients with intermediate HCC who received combination therapy (n = 34) or supportive care alone (n = 24). The inclusion criteria were a single nodule of more than 50 mm in diameter or two to three nodules, each measuring more than 30 mm in diameter, or more than three nodules, no vascular invasion and no extrahepatic metastasis. RESULTS: The overall survival rates at 1, 2, 3 and 5 years of the combination therapy group (91%, 65%, 53% and 27%, respectively) were significantly better (P < 0.0001) than those of the supportive care group (42%, 8%, 8% and 0%, respectively). Multivariate analysis identified treatment modality (combination therapy vs supportive care alone: P < 0.0001, risk ratio [RR] = 4.290 [95% confidence interval [CI] = 2.157-8.529]) and serum α-fetoprotein (P = 0.017, RR = 2.318 [95% CI = 1.166-4.610]) as independent and significant factors of overall survival. CONCLUSION: The combination of TACE and RFA is a safe and effective therapy in patients with intermediate HCC.
RESUMEN
AIM: The aim of this study was to evaluate the long-term outcome of elderly patients with hepatocellular carcinoma (HCC) aged 75 years or older. METHODS: The study included 422 patients with HCC, who were divided into two age groups: 75 years or older (n = 140) and younger than 75 (n = 282). Outcomes were compared between the two groups. RESULTS: The number of elderly patients treated with supportive care alone (33 patients; 24%) was significantly higher than younger patients (30 patients; 11%, P < 0.01). The 1-, 3-, 5- and 7-year overall survival rates of the elderly patients (81%, 55%, 39% and 23%, respectively) were worse than those of younger patients (85%, 64%, 49% and 36%, respectively, P = 0.042). However, the overall survival rate of the elderly group after excluding 63 patients treated with supportive care alone, was similar to that of the younger group (P = 0.615). Multivariate analysis identified age, total bilirubin levels, albumin levels, serum des-γ-carboxy prothrombin levels, tumor size, number of HCC nodules, vascular invasion, extrahepatic metastasis and treatment modality as independent and significant factors of overall survival. CONCLUSION: Advanced age is a negative prognostic factor in patients with HCC due to the tendency for frequent use of conservative treatment rather than locoregional or surgical treatment.
RESUMEN
To establish a strategy for the comprehensive identification of human N-myristoylated proteins, the susceptibility of human cDNA clones to protein N-myristoylation was evaluated by metabolic labeling and MS analyses of proteins expressed in an insect cell-free protein synthesis system. One-hundred-and-forty-one cDNA clones with N-terminal Met-Gly motifs were selected as potential candidates from approximately 2000 Kazusa ORFeome project human cDNA clones, and their susceptibility to protein N-myristoylation was evaluated using fusion proteins, in which the N-terminal ten amino acid residues were fused to an epitope-tagged model protein. As a result, the products of 29 out of 141 cDNA clones were found to be effectively N-myristoylated. The metabolic labeling experiments both in an insect cell-free protein synthesis system and in the transfected COS-1 cells using full-length cDNA revealed that 27 out of 29 proteins were in fact N-myristoylated. Database searches with these 27 cDNA clones revealed that 18 out of 27 proteins are novel N-myristoylated proteins that have not been reported previously to be N-myristoylated, indicating that this strategy is useful for the comprehensive identification of human N-myristoylated proteins from human cDNA resources.
Asunto(s)
Ácido Mirístico/análisis , Biosíntesis de Proteínas , Proteínas/análisis , Acilación , Secuencia de Aminoácidos , Animales , Línea Celular , Sistema Libre de Células/química , Chlorocebus aethiops , ADN Complementario/genética , Humanos , Datos de Secuencia Molecular , Peso Molecular , Ácido Mirístico/química , Ácido Mirístico/metabolismo , Proteínas/química , Proteínas/genética , Proteínas/metabolismo , SpodopteraRESUMEN
BACKGROUND AND AIMS: Dysregulated lipid metabolism has emerged as one of the major risk factors of atherosclerosis. Presently, there is a consensus that oxidized LDL (oxLDL) promotes development of atherosclerosis and downstream chronic inflammatory responses. Due to the dynamic metabolic disposition of lipoprotein, conventional approach to purify bioactive lipids for subsequent comprehensive analysis has proven to be inadequate for elucidation of the oxidized lipids species accountable for pathophysiology of atherosclerotic lesions. Herein, we aimed to utilize a novel mass microscopic imaging technology, coupled with mass spectrometry (MS) to characterize oxidized lipids in atherosclerotic lesions. METHODS: We attempted to use MALDI-TOF-MS and iMScope to identify selected oxidized lipid targets and visualize their respective localizations in study models of atherosclerosis. RESULTS: Based on the MS analysis, detection of 7-K under positive ionization through product ion peak at m/z 383 [M + H-H2O] indicated the distinctive presence of targeted lipid within Cu2+-oxLDL and Cu2+-oxLDL loaded macrophage-like J774A.1 cells, along with other cholesterol oxidation products. Moreover, the application of two-dimensional iMScope has successfully visualized the localization of lipids in aortic atherosclerotic plaques of the Watanabe heritable hyperlipidemic (WHHL) rabbit. Distinctive lipid distribution profiles were observed in atherosclerotic lesions of different sizes, especially the localizations of lysoPCs in atherosclerotic plaques. CONCLUSIONS: Taken together, we believe that both MALDI-TOF-MS and iMScope metabolomics technology may offer a novel proposition for future pathophysiological studies of lipid metabolism in atherosclerosis.
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Aterosclerosis , Placa Aterosclerótica , Animales , Lípidos , Lipoproteínas LDL , Espectrometría de Masas , Metabolómica , ConejosRESUMEN
A simple and efficient method for C-terminal sequencing of proteins has long been pursued because it would provide substantial information for identifying the covalent structure, including post-translational modifications. However, there are still significant impediments to both direct sequencing from C termini of proteins and specific isolation of C-terminal peptides from proteins. We describe here a highly successful, de novo C-terminal sequencing method of proteins by employing succinimidyloxycarbonylmethyl tris (2,4,6-trimethoxyphenyl) phosphonium bromide and mass spectrometry.
Asunto(s)
Caseínas/análisis , Citocromos c/análisis , Muramidasa/análisis , Ovalbúmina/análisis , Fragmentos de Péptidos/análisis , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Animales , Bovinos , Pollos , Caballos , Indicadores y Reactivos , Compuestos Organofosforados/metabolismo , Mapeo Peptídico , Procesamiento Proteico-Postraduccional , Proteínas Recombinantes/análisis , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
In proteomics, MS plays an essential role in identifying and quantifying proteins. To characterize mature target proteins from living cells, candidate proteins are often analyzed with PMF and MS/MS ion search methods in combination with computational search routines based on bioinformatics. In contrast to shotgun proteomics, which is widely used to identify proteins, proteomics based on the analysis of N- and C-terminal amino acid sequences (terminal proteomics) should render higher fidelity results because of the high information content of terminal sequence and potentially high throughput of the method not requiring very high sequence coverage to be achieved by extensive sequencing. In line with this expectation, we review recent advances in methods for N- and C-terminal amino acid sequencing of proteins. This review focuses mainly on the methods of N- and C-terminal analyses based on MALDI-TOF MS for its easy accessibility, with several complementary approaches using LC/MS/MS. We also describe problems associated with MS and possible remedies, including chemical and enzymatic procedures to enhance the fidelity of these methods.
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Espectrometría de Masas/métodos , Proteínas/química , Análisis de Secuencia de Proteína/métodos , Péptido Hidrolasas/metabolismo , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
BACKGROUND: Macroscopic vascular invasion is known to be a poor prognostic factor in hepatocellular carcinoma (HCC). The aim of this study was to determine the outcomes and predictive factors after hepatic resection for HCC with microvascular invasion (MVI). METHODS: One hundred ten patients who underwent curative resection for HCC without macroscopic vascular invasion were included in this retrospective study. The risk factors of these patients for recurrence-free and disease-specific survival were investigated, and the clinicopathological factors predicting the presence of MVI were also determined. RESULTS: Of the 110 resected specimens, 49 (45%) had evidence of MVI. By univariate analysis, MVI was found to be statistically significantly associated with greater tumor size, gross classification, histological grade, and intrahepatic micrometastasis. Gross classification proved to be the only independent predictive factor for MVI by multiple logistic regression analysis. By multivariate analysis, cirrhosis and MVI were identified as independent risk factors for recurrence-free survival. The 5-year recurrence-free survival rates for patients with and without MVI were 20.8% and 52.6%, respectively. By multivariate analysis, the number of tumors, presence of MVI, and intrahepatic micrometastasis were identified as independent predictors of disease-specific survival. The 5-year disease-specific survival rates for patients with and without MVI were 59.3% and 92.0%, respectively. CONCLUSIONS: The presence of MVI was the most important risk factor affecting recurrence and survival in HCC patients after curative resection. Furthermore, this study showed that gross classification of HCC can be very helpful in predicting the presence of MVI.
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Carcinoma Hepatocelular/patología , Hepatectomía , Neoplasias Hepáticas/patología , Hígado/irrigación sanguínea , Recurrencia Local de Neoplasia/patología , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/cirugía , Progresión de la Enfermedad , Femenino , Estudios de Seguimiento , Humanos , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/cirugía , Masculino , Microcirculación , Persona de Mediana Edad , Invasividad Neoplásica , Cuidados Preoperatorios , Pronóstico , Estudios Retrospectivos , Factores de Riesgo , Tasa de SupervivenciaRESUMEN
ß2-Glycoprotein I (ß2GPI) is a highly-glycosylated plasma protein composed of five homologous domains which regulates coagulation, fibrinolysis, and/or angiogenesis by interacting to negatively charged hydrophobic molecules and/or with plasminogen and its metabolites. The present study focused on structural and functional characterization of ß2GPI's domain I (DI) and V (DV). Through N-terminal amino acid sequencing, a novel plasmin-cleaved site at K287C288 was identified in DV. We further modified the intact DV by altering two amino acids at specific proteolytic cleavage sites to generate three stable DV mutants: DV(PP), (PE), and (AA). Results of both SDS-PAGE and MALDI-TOF-MS showed that all three DV mutants were more stable than the intact DV, and DV(PE) was predominantly resistant to proteolysis. Competitive ELISA assessed affinities of intact ß2GPI and those mutants to cardiolipin. In culture system, all DV and DI mutants potently inhibited HUVEC's proliferation by 18-30% as compared to control. Only DI and nicked ß2GPI showed significant inhibition in HUVEC's tube formation. Moreover, DV(PE)-coated affinity columns demonstrated its binding property towards anionic lipids and could substantially isolate anionic DOPS from zwitterionic DOPC as a purification model. In summary, the proteolytic resistant and unhindered phospholipid (PL) binding properties of DV(PE) have made it an appealing element for subsequent prospective studies. Future in-depth characterization and optimized applications of cleavage-resistant DV(PE) would complement its full capacity as a novel clinical modality in the field of vascular imaging and/or lipidomics studies.
Asunto(s)
Análisis de Secuencia de Proteína/métodos , beta 2 Glicoproteína I/uso terapéutico , Humanos , Estudios Prospectivos , beta 2 Glicoproteína I/farmacologíaRESUMEN
We have developed a method for on-membrane direct identification of phosphoproteins, which are detected by a phosphate-binding tag (Phos-tag) that has an affinity to phosphate groups with a chelated Zn2+ ion. This rapid profiling approach for phosphoproteins combines chemical inkjet technology for microdispensing of reagents onto a tiny region of target proteins with mass spectrometry for on-membrane digested peptides. Using this method, we analyzed human epidermoid carcinoma cell lysates of A-431 cells stimulated with epidermal growth factor, and identified six proteins with intense signals upon affinity staining with the phosphate-binding tag. It was already known that these proteins are phosphorylated, and our new approach proved to be effective at rapid profiling of phosphoproteins. Furthermore, we tried to determine their phosphorylation sites by MS/MS analysis after in-gel digestion of the corresponding spots on the 2DE gel to the rapid on-membrane identifications. As one example of use of information gained from the rapid-profiling approach, we successfully characterized a phosphorylation site at Ser-113 on prostaglandin E synthase 3.
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Membrana Celular/química , Tinta , Proteínas de la Membrana/química , Mapeo Peptídico , Fosfatos/química , Fosfoproteínas/química , Zinc , Secuencia de Aminoácidos , Línea Celular Tumoral , Membrana Celular/metabolismo , Humanos , Proteínas de la Membrana/metabolismo , Datos de Secuencia Molecular , Mapeo Peptídico/métodos , Fosfatos/metabolismo , Fosfopéptidos/química , Fosfopéptidos/metabolismo , Fosfoproteínas/metabolismo , Fosforilación , Unión Proteica , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Espectrometría de Masas en Tándem , Zinc/metabolismoRESUMEN
Glucose intolerance frequently is found in hepatocellular carcinoma (HCC) patients with hepatitis C virus (HCV) infection; however, the significance of glucose intolerance remains unclear. In addition, SH2 domain-containing inositol phosphatase (SHIP) 2 is a negative regulator of intracellular insulin signaling; however, changes in SHIP2 expression have not been investigated in HCC. To assess the significance of glucose intolerance, we analyzed 118 HCC patients with HCV infection. Twenty HCC specimens were used for immunoblotting and immunostaining for SHIP2. Patients were classified into two groups: a glucose intolerance group (n=39) and a normal glucose tolerance group (n=79). There was no significant difference in the disease-free survival (P=0.838) or long-term survival (P=0.091) between the groups. However, for males, the cumulative survival rate was significantly lower in the glucose intolerance group (n=22) than that in the normal glucose tolerance group (n=52) (P=0.036). In multivariate analysis, Child-Pugh class (P=0.0003) and glucose intolerance (P=0.036) were identified as statistically significant and independent prognostic factors in males. SHIP2 expression level decreased in HCC compared to that in nontumor tissues. In conclusion, this study is the first to demonstrate the significance of glucose intolerance in prognosis of male HCC patients and down-regulation of SHIP2 expression in HCC.
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Carcinoma Hepatocelular/genética , Regulación Enzimológica de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Intolerancia a la Glucosa/complicaciones , Hepatitis C/complicaciones , Neoplasias Hepáticas/genética , Monoéster Fosfórico Hidrolasas/metabolismo , Anciano , Anciano de 80 o más Años , Carcinoma Hepatocelular/sangre , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/virología , Supervivencia sin Enfermedad , Femenino , Prueba de Tolerancia a la Glucosa , Hepatitis C/sangre , Humanos , Inmunohistoquímica , Neoplasias Hepáticas/sangre , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/virología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatasas , Monoéster Fosfórico Hidrolasas/genética , Estudios RetrospectivosRESUMEN
We have identified tyrosine-phosphorylated proteins on membrane from A-431 human epidermoid carcinoma cells by using detection with anti-phosphotyrosine antibody followed by PMF analysis. In there, on-membrane digestion for these protein spots was carried out on microscale region using chemical inkjet technology and the resulting tryptic digests were directly analyzed by MALDI-TOF MS. Proteins identified by a database search included phosphoproteins that are known to be markedly phosphorylated on tyrosine sites after the cells are treated with epidermal growth factor (EGF). This procedure is a rapid and easily handled approach that enables both detection and identification of phosphoproteins on a single blot membrane.
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Extractos Celulares/química , Electroforesis en Gel Bidimensional/métodos , Mapeo Peptídico , Fosfoproteínas/análisis , Fosfotirosina/análisis , Proteínas/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Anticuerpos Fosfo-Específicos/análisis , Western Blotting/métodos , Carcinoma de Células Escamosas , Línea Celular Tumoral/química , Línea Celular Tumoral/efectos de los fármacos , Membrana Celular/química , Factor de Crecimiento Epidérmico/farmacología , Humanos , Peso Molecular , Fosfoproteínas/inmunología , Fosfotirosina/inmunología , Espectrometría de Masas en Tándem/métodosRESUMEN
A 64-year-old man was admitted for further examinations of a liver tumor. The patient was diagnosed as chronic hepatitis C complicated with advanced hepatocelluar carcinoma (HCC) with left portal vein tumor thrombosis. As he refused surgical treatment, hepatic arterial infusion chemotherapy (HAIC) using cisplatin and 5-fluorouracil was performed initially. Administration of ursodesoxycholic acid (UDCA) was also started. Following HAIC, microwave coagulation therapy for residual tumor was added. Consequently, viable lesions of HCC disappeared completely. At present, after more than 8 years, neither signs of tumor recurrence, nor elevation of hepatic enzymes has been observed. Although the precise reason for long survival of this patient is not known, we speculate that suppression of levels of hepatic enzymes, as well as HAIC for subclinical intrahepatic metastasis, contributed to the good outcome. Therapeutic strategy for hepatic inflammation seems to be important for long-term prevention of hepatocarcinogenesis.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma Hepatocelular/tratamiento farmacológico , Hepatitis C Crónica/complicaciones , Bombas de Infusión Implantables , Neoplasias Hepáticas/tratamiento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Carcinoma Hepatocelular/etiología , Cisplatino/administración & dosificación , Esquema de Medicación , Fluorouracilo/administración & dosificación , Arteria Hepática , Humanos , Infusiones Intraarteriales , Neoplasias Hepáticas/etiología , Masculino , Persona de Mediana Edad , Inducción de Remisión , SobrevivientesRESUMEN
BACKGROUND: Thoracic epidural analgesia (EDA) is the gold standard for pain control after thoracotomy. However, because of its severe side effects, it is contraindicated in patients taking anticoagulant or antiplatelet drugs. In addition, some patients' anatomy can make epidural catheter insertion challenging. We therefore investigated the safety and efficacy of paravertebral block (PVB) using a thoracoscopic insertion technique, which avoids damage to the parietal pleura, for postoperative pain after thoracotomy. METHODS: Patients who underwent thoracotomy with thoracic PVB in our hospital between March 2013 and March 2014 were examined retrospectively. Prior to creating the thoracotomy incision, a catheter for PVB was inserted percutaneously into the paravertebral space under thoracoscopic guidance. A matched-pair control group was selected at a 1:2 ratio from patients who underwent thoracotomy with thoracic EDA in our hospital from April 2011 to February 2013. Pain control and side effects were compared between groups and the results statistically analyzed. RESULTS: Thoracic PVB was performed in 56 patients during this period, and 112 patients were selected as matched controls. Numeric Rating Scale scores on postoperative day 2 did not differ significantly between the PVB group (3.25 ± 1.80) and the EDA group (3.56 ± 2.05) (p = 0.334). In terms of side effects, urinary retention occurred less frequently in thoracic PVB patients (p = 0.03). CONCLUSION: Under the conditions of the present study, continuous thoracic PVB was at least as effective as epidural analgesia for postoperative pain control after thoracotomy with lung resection.