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1.
J Clin Biochem Nutr ; 74(2): 141-145, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38510680

RESUMO

We conducted a retrospective case-control study to assess the efficacy of personalized health guidance interventions on individuals with type 2 diabetes mellitus and obesity. A selection was made of individuals in regular visits to the Takagi Hospital for medical checkups between January 2017, and October 2021. Totally, 108 subjects (cases) with health guidance were divided into 2 groups: one group without pharmacotherapy for diabetes mellitus in medical institutions (n = 92) and another group with pharmacotherapy (n = 116). Cases were provided with personalized health guidance interventions by public health nurses for 30 min, in accordance with the Japanese clinical guidelines for the prevention of lifestyle-related diseases. Sex- and age-matched controls were chosen from individuals with diabetes mellitus without health guidance. The intervention without pharmacotherapy resulted in improvements in health indicators, including body weight, waist circumference, diastolic blood pressure, triglyceride levels, and γ-glutamyl trans-peptidase. These positive effects were not observed in the control group without health guidance. The therapeutic effects of health guidance were observed in cases where pharmacotherapy was administered. In conclusion, the implementation of individual health guidance interventions may prove to be effective for individuals with type 2 diabetes mellitus and obesity who regularly attend medical checkups.

2.
Rheumatology (Oxford) ; 62(11): 3757-3762, 2023 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-37144941

RESUMO

OBJECTIVES: AECAs are detected in multiple forms of vasculitis or vasculopathy, including JDM. High levels of tropomyosin alpha-4 chain (TPM4) gene expression in cutaneous lesions and TPM4 protein expression in some endothelial cells (ECs) have been proven. Furthermore, the presence of autoantibodies to tropomyosin proteins have been discovered in DM. We therefore investigated whether anti-TPM4 autoantibodies are an AECA in JDM and are correlated with clinical features of JDM. METHODS: The expression of TPM4 protein in cultured normal human dermal microvascular ECs was investigated by Western blotting. Plasma samples from 63 children with JDM, 50 children with polyarticular JIA (pJIA) and 40 healthy children (HC) were tested for the presence of anti-TPM4 autoantibodies using an ELISA. Clinical features were compared between JDM patients with and without anti-TPM4 autoantibodies. RESULTS: Autoantibodies to TPM4 were detected in the plasma of 30% of JDM, 2% of pJIA (P < 0.0001) and 0% of HC (P < 0.0001). In JDM, anti-TPM4 autoantibodies were associated with the presence of cutaneous ulcers (53%; P = 0.02), shawl sign rash (47%; P = 0.03), mucous membrane lesions (84%; P = 0.04) and subcutaneous edema (42%; P < 0.05). Anti-TPM4 autoantibodies significantly correlated with the use of intravenous steroids and IVIG therapy in JDM (both P = 0.01). The total number of medications received was higher in patients with anti-TPM4 autoantibodies (P = 0.02). CONCLUSION: Anti-TPM4 autoantibodies are detected frequently in children with JDM and are novel myositis-associated autoantibodies. Their presence correlates with vasculopathic and other cutaneous manifestations of JDM that may be indicative of more refractory disease.


Assuntos
Dermatomiosite , Miosite , Doenças Vasculares , Criança , Humanos , Células Endoteliais/patologia , Tropomiosina , Autoanticorpos , Proteínas do Citoesqueleto
3.
Rheumatology (Oxford) ; 61(7): 2969-2977, 2022 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-34791087

RESUMO

OBJECTIVES: JDM is an inflammatory myopathy characterized by prominent vasculopathy. AECAs are frequently detected in inflammatory and autoimmune diseases. We sought to determine whether AECAs correlate with clinical features of JDM, and thus serve as biomarkers to guide therapy or predict outcome. METHODS: Plasma samples from 63 patients with JDM, 49 patients with polyarticular JIA and 40 juvenile healthy controls were used to detect anti-heat shock cognate 71 kDa protein (HSC70) autoantibodies, a newly identified AECA, in ELISA assays. Clinical features were compared between JDM patients with and without anti-HSC70 autoantibodies. RESULTS: Anti-HSC70 autoantibodies were detected in 35% of patients with JDM, in 0% of patients with JIA (P < 0.0001) and in 0% of healthy donors (P < 0.0001). Both the presence of cutaneous ulcers (59% vs 17%, P < 0.002) and the use of wheelchairs and/or assistive devices (64% vs 27%, P < 0.007) were strongly associated with anti-HSC70 autoantibodies in JDM. High scores on the severity of myositis damage measures at the time of measurement of anti-HSC70 autoantibodies and an increased number of hospitalizations were also associated with anti-HSC70 autoantibodies. Intravenous immunoglobulin therapy was used more often in anti-HSC70 autoantibody-positive patients. CONCLUSION: Anti-HCS70 autoantibodies are detected frequently in children with JDM and are novel myositis-associated autoantibodies correlating with disease severity.


Assuntos
Doenças Autoimunes , Dermatomiosite , Miosite , Úlcera Cutânea , Autoanticorpos , Criança , Humanos , Imunoglobulinas Intravenosas
4.
Digestion ; 102(4): 630-639, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32932255

RESUMO

BACKGROUND/AIMS: For early gastric cancer (EGC) treated using endoscopic submucosal dissection (ESD) with poor curability defined by the Japanese Guidelines (non-curative EGC, N-EGC), additional gastrectomy has been recommended. However, N-EGC patients without additional gastrectomy often die of other diseases within a relatively short interval after ESD. It has been unclear whether additional gastrectomy is beneficial or not for such patients. The aim of this study was to clarify predictors for short-term survival of N-EGC patients without additional gastrectomy after ESD. METHODS: One hundred six N-EGC patients without additional gastrectomy were included in this study. Factors related to short-term survival, defined as death within 3 years after ESD, were evaluated using uni- and multivariate analyses by comparing patients with and without short-term survival (Groups S and C, respectively). RESULTS: During the mean follow-up period of 89 months, 39 patients died (14 patients died within 3 years, being Group S). The cause of death was gastric cancer for only 1 patient in the Group C. The 3- and 5-year overall survival rates were 86.8 and 81.8%, respectively, and the 3- and 5-years disease-specific survival rates were 100 and 98.9%, respectively. Univariate analyses showed that short-term survival was statistically associated with elevated morphology, high-risk status for lymph node metastases as defined by the eCura system, severe comorbidity (Charlson Comorbidity Index [CCI] ≥3), low level of activity in daily living (being unable to go out by oneself), habitation (a nursing home), and several poor nutritional prognostic indices (neutrophil to lymphocyte ratio ≥2.5, geriatric nutritional risk index <92, C-reactive protein ≥1.0). In the multivariate analysis, a high CCI (≥3) was the independent predictor for short-term survival after ESD (odds ratio, 8.1; 95% confidence interval, 1.53-43.0; p = 0.014). CONCLUSIONS: Severe comorbidity indicated by a high CCI score (≥3) was the independent predictor for short-term survival for EGC patients without additional gastrectomy after non-curative ESD. Since the cause of death for most patients was not gastric cancer, observational follow-ups without additional gastrectomy might be a reasonable option for patients with a poor general status indicated by a CCI ≥3.


Assuntos
Ressecção Endoscópica de Mucosa , Neoplasias Gástricas , Idoso , Ressecção Endoscópica de Mucosa/efeitos adversos , Gastrectomia/efeitos adversos , Mucosa Gástrica/cirurgia , Humanos , Prognóstico , Estudos Retrospectivos , Neoplasias Gástricas/cirurgia , Taxa de Sobrevida , Resultado do Tratamento
5.
Rheumatology (Oxford) ; 57(4): 671-676, 2018 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-29361142

RESUMO

Objective: Although generally classified within the group of inflammatory myopathies, JDM displays many pathological features of vasculitis. Previous work has shown that AECA are abundant in other forms of vasculitis. We therefore investigated whether such antibodies might also be detected in JDM. Methods: We screened plasma from children with JDM for the presence of AECA by western blotting and 2D gel electrophoresis (2DE) using proteins extracted from human aortic endothelial cells as the substrate. We performed mass spectrometry to identify candidate antigens from 2DE gels and used ELISA to confirm the presence of specific antibodies. Results: We identified 22 candidate target autoantigens for AECA probed with JDM plasma. Interestingly, 17 of these 22 target antigens were proteins associated with antigen processing and protein trafficking. ELISA confirmed the presence of antibodies to heat shock cognate 71 kDa protein in JDM plasma, particularly in children with active, untreated disease. Conclusion: Children with JDM express antibodies to autoantigens in endothelial cells. The clinical and pathological significance of such autoantibodies require further investigation.


Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Dermatomiosite/imunologia , Células Endoteliais/imunologia , Endotélio Vascular/patologia , Proteômica/métodos , Adolescente , Aorta/imunologia , Aorta/patologia , Autoanticorpos/sangue , Autoantígenos/sangue , Biomarcadores/sangue , Biópsia , Western Blotting , Células Cultivadas , Criança , Pré-Escolar , Cromatografia Líquida de Alta Pressão , Dermatomiosite/diagnóstico , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida , Células Endoteliais/patologia , Endotélio Vascular/imunologia , Ensaio de Imunoadsorção Enzimática , Feminino , Seguimentos , Humanos , Masculino , Estudos Retrospectivos
6.
Mol Pharm ; 13(2): 493-9, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26656401

RESUMO

The amorphization has been generally known to improve the absorption and permeation of poorly water-soluble drugs through the enhancement of the solubility. The present study focused on the direct contact of amorphous solid particles with the surface of the membrane using curcumin as a model for water-insoluble drugs. Amorphous nanoparticles of curcumin (ANC) were prepared with antisolvent crystallization method using a microreactor. The solubility of curcumin from ANC was two orders of magnitude higher than that of crystalline curcumin (CC). However, the permeation of curcumin from the saturated solution of ANC was negligible. The transepithelial permeation of curcumin from ANC suspension was significantly increased as compared to CC suspension, while the permeation was unlikely correlated with the solubility, and the increase in the permeation was dependent on the total concentration of curcumin in ANC suspension. The absorptive transport of curcumin (from apical to basal, A to B) from ANC suspension was much higher than the secretory transport (from basal to apical, B to A). In vitro transport of curcumin through air-interface monolayers is large from ANC but negligible from CC particles. These findings suggest that the direct contact of ANC with the absorptive membrane can play an important role in the transport of curcumin from ANC suspension. The results of the study suggest that amorphous particles may be directly involved in the transepithlial permeation of curcumin.


Assuntos
Antineoplásicos/química , Permeabilidade da Membrana Celular/efeitos dos fármacos , Curcumina/química , Células Epiteliais/efeitos dos fármacos , Nanopartículas/química , Animais , Antineoplásicos/farmacologia , Química Farmacêutica , Curcumina/farmacologia , Cães , Células Madin Darby de Rim Canino , Solubilidade , Suspensões , Água
7.
Dig Endosc ; 27(3): 303-9, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25357187

RESUMO

BACKGROUND AND AIM: To assess the usefulness of the thread-traction method (TT method) in esophageal endoscopic submucosal dissection (ESD). METHODS: A total of 40 lesions that were scheduled to be treated by esophageal ESD were included in the study. The TT method was used for 20 lesions (group TT) and conventional ESD was used for 20 lesions (group C) after randomization. The hook-knife method was used in all cases. In group TT, after circumferential mucosal incision, a clip with thread was attached to the oral edge of the lesion. RESULTS: ESD was carried out in all cases. Effective countertraction was created by the TT method, and it was possible to carry out an efficient dissection operation. Significant shortening of dissection time was achieved in group TT compared with group C (19.8 min vs 31.8 min, P = 0.044). Mean number of local injections during dissection was significantly less in group TT compared with that in group C (0.6 times vs 2.2 times, P < 0.001). As for the amount of local injection, group TT required significantly less compared with group C (2.6 mL vs 7.5 mL, P < 0.01). No complications were encountered. CONCLUSION: The TT method in esophageal ESD was safe and contributed to shortening of dissection time. The TT method is expected to become widespread as a safe and useful procedure.


Assuntos
Carcinoma de Células Escamosas/cirurgia , Dissecação/instrumentação , Neoplasias Esofágicas/cirurgia , Esofagoscopia/métodos , Mucosa/cirurgia , Idoso , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/patologia , Dissecação/métodos , Neoplasias Esofágicas/mortalidade , Neoplasias Esofágicas/patologia , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Procedimentos Cirúrgicos Minimamente Invasivos/efeitos adversos , Procedimentos Cirúrgicos Minimamente Invasivos/métodos , Mucosa/patologia , Duração da Cirurgia , Dor Pós-Operatória/fisiopatologia , Dor Pós-Operatória/terapia , Segurança do Paciente , Hemorragia Pós-Operatória/etiologia , Hemorragia Pós-Operatória/fisiopatologia , Hemorragia Pós-Operatória/terapia , Medição de Risco , Instrumentos Cirúrgicos , Taxa de Sobrevida , Resultado do Tratamento
8.
Rinsho Byori ; 62(3): 223-30, 2014 Mar.
Artigo em Japonês | MEDLINE | ID: mdl-24800496

RESUMO

The new classification criteria for systemic lupus erythematosus (SLE) by Systemic Lupus International Collaborating Clinics (SLICC) published in 2012 have defined positive level titer of antibody against double stranded (ds) DNA as more than double the reference range if tested by enzyme linked immunosorbent assay (ELISA). The aim of this study was to evaluate optimal cut-off value and diagnostic performance of the anti-dsDNA nucleosome-complexed (anti-dsDNA-NcX) ELISA, which uses salmon testis DNA complexed with purified nucleosomes as antigens, for diagnosis of SLE. Titers of antibodies against dsDNA-complexed nucleosome were measured in sera of 76 patients with SLE, 148 with other connective tissue diseases, and 323 healthy volunteers. Sensitivity, specificity, correlation and concordance rate were compared among anti-dsDNA-NcX, conventional ELISA methods (EIA) and radioimmunoassay (RIA). As a results, concordance rates and Spearman's coefficient of rank correlation (r(s)) of anti-dsDNA-NcX with EIA and RIA were 59.2%, r(s) = 0.40 and 53.9%, r(s) = 0.21, respectively. Receiver operating characteristic curve analysis showed that the titer of 44 IU/mL calculated as 99 percentile of 323 healthy volunteers is a better cut-off value for anti-dsDNA-NcX than the 100 IU/mL recommended by the manufacturer. By setting the cut-off value at 44 IU/mL, anti-dsDNA-NcX showed the highest sensitivity (75.0%) and specificity (90.5%) of the 3 assays. With SLICC criterion, positivity rates of anti-dsDNA-NcX, EIA and RIA for SLE patients were 50.0%, 30.3% and 50.0%, respectively. In conclusion, anti-dsDNA-NcX has a good diagnostic performance for SLE with our proposed cut-off value of 44 IU/mL.


Assuntos
Anticorpos Antinucleares/sangue , DNA/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Nucleossomos/metabolismo , Anticorpos Antinucleares/imunologia , Artrite Reumatoide/sangue , Artrite Reumatoide/diagnóstico , Artrite Reumatoide/imunologia , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/imunologia , Nucleossomos/imunologia
9.
Dig Endosc ; 24(5): 309-14, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22925281

RESUMO

AIM: Endoscopic diagnosis of the lateral extension of Barrett's cancer under the squamous epithelium (BCUS) is sometimes difficult because the cancer is unobservable in the esophageal lumen. The aim of the present study was to clarify the endoscopic features of the extension of BCUS and verify the usefulness of the acetic acid-spraying method (AAS) for diagnosis. METHODS: A total of 25 patients with Barrett's cancer who had undergone endoscopic resection were included in this study. Histological examination of patients' resected specimens was performed to identify the presence of BCUS. Then, the endoscopic images of the BCUS cases were reviewed to summarize the findings and to evaluate the feasibility of diagnosing the extent of BCUS with each imaging technique. RESULTS: Of the 25 patients, 10 (40%) had BCUS. With white-light imaging, subtle reddish change was observed in the area of BCUS in 80% of the patients, and a flat elevated lesion was recognized in 30%. With narrow band imaging, slight brownish change was observed in the area of BCUS in 86% of the patients. Slight white changes were visualized in all cases with AAS. The extension of BCUS was correctly diagnosed by white-light imaging, narrow band imaging and AAS in 50%, 43% and 100% of the cases, respectively. Histology verified the opening of cancerous glands, which extended under the squamous epithelium, into the esophagus in the area showing slight white changes by AAS. CONCLUSION: AAS can be useful for diagnosing the extension of BCUS.


Assuntos
Ácido Acético , Esôfago de Barrett/patologia , Neoplasias Esofágicas/patologia , Esôfago/patologia , Mucosa Intestinal/patologia , Lesões Pré-Cancerosas , Idoso , Diagnóstico Diferencial , Epitélio/patologia , Esofagoscopia/métodos , Feminino , Seguimentos , Humanos , Indicadores e Reagentes/farmacologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos
10.
Yeast ; 28(4): 265-78, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21456053

RESUMO

Yeast-based reporter assays are useful for detecting various genotoxic chemicals. We established a genotoxicity assay using recombinant strains of Saccharomyces cerevisiae, each containing a reporter plasmid with the secretory luciferase gene from Cypridina noctiluca, driven by a DNA damage-responsive promoter of the yeast RNR3 gene. This system detected the genotoxicity of methyl methanesulphonate (MMS) as sensitively as conventional yeast-based reporter assays, using the ß-galactosidase gene in a concentration-dependent manner; it also detects four other genotoxic chemicals, allowing us to monitor DNA damage easily by skipping the cell extraction process for the assay. We examined Cypridina luciferase levels induced by MMS and three antitumour agents using a set of BY4741-derived deletion mutants, each defective in a DNA repair pathway or DNA damage checkpoint. Luciferase activities were particularly enhanced in mutant strains with mms2 Δ and mag1 Δ by exposure to MMS, rad59 Δ and mlh1 Δ to camptothecin and mms2 Δ and mlh1 Δ to mitomycin C, respectively, compared with their parent strains. Enhanced reporter activities were also found in some DNA repair mutants with cisplatin. These observations suggest that this Cypridina secretory luciferase reporter assay using yeast DNA repair mutants offers convenient and sensitive detection of the potential genotoxicity of numerous compounds, including antitumour drugs and studying the mechanisms of DNA damage response in yeast.


Assuntos
Reparo do DNA/efeitos dos fármacos , Luciferases/metabolismo , Testes de Mutagenicidade/métodos , Mutagênicos/toxicidade , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Genes Reporter , Luciferases/genética , Testes de Mutagenicidade/instrumentação , Saccharomyces cerevisiae/metabolismo
11.
Biosci Biotechnol Biochem ; 75(1): 26-33, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21228494

RESUMO

Rhodococcus jostii RHA1 is a polychlorinated biphenyl degrader. Multi-component biphenyl 2,3-dioxygenase (BphA) genes of RHA1 encode large and small subunits of oxygenase component and ferredoxin and reductase components. They did not express enzyme activity in Escherichia coli. To obtain BphA activity in E. coli, hybrid BphA gene derivatives were constructed by replacing ferredoxin and/or reductase component genes of RHA1 with those of Pseudomonas pseudoalcaligenes KF707. The results obtained indicate a lack of catalytic activity of the RHA1 ferredoxin component gene, bphAc in E. coli. To determine the cause of inability of RHA1 bphAc to express in E. coli, the bphAc gene was introduced into Rosetta (DE3) pLacI, which has extra tRNA genes for rare codons in E. coli. The resulting strain abundantly produced the bphAc product, and showed activity. These results suggest that codon usage bias is involved in inability of RHA1 bphAc to express its catalytic activity in E. coli.


Assuntos
Escherichia coli/genética , Proteínas Ferro-Enxofre/genética , Oxigenases/genética , Bifenilos Policlorados/metabolismo , Engenharia de Proteínas/métodos , Rhodococcus/enzimologia , Rhodococcus/genética , Sequência de Bases , Biocatálise , Códon/genética , Expressão Gênica , Proteínas Ferro-Enxofre/biossíntese , Proteínas Ferro-Enxofre/metabolismo , Oxigenases/biossíntese , Oxigenases/metabolismo , RNA de Transferência/genética
12.
ACS Chem Biol ; 16(6): 1059-1069, 2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34080843

RESUMO

C-Glycosylation in the biosynthesis of bioactive natural products is quite unique, which has not been studied well. Medermycin, as an antitumor agent in the family of pyranonaphthoquinone antibiotics, is featured with unique C-glycosylation. Here, a new C-glycosyltransferase (C-GT) Med-8 was identified to be essential for the biosynthesis of medermycin, as the first example of C-GT to recognize a rare deoxyaminosugar (angolosamine). med-8 and six genes (med-14, -15, -16, -17, -18, and -20 located in the medermycin biosynthetic gene cluster) predicted for the biosynthesis of angolosamine were proved to be functional and sufficient for C-glycosylation. A C-glycosylation cassette composed of these seven genes could convert a proposed substrate into a C-glycosylated product. In conclusion, these genes involved in the C-glycosylation of medermycin were functionally identified and biosynthetically engineered, and they provided the possibility of producing new C-glycosylated compounds.


Assuntos
Proteínas de Bactérias/metabolismo , Vias Biossintéticas , Glicosiltransferases/metabolismo , Streptomyces/metabolismo , Proteínas de Bactérias/genética , Genes Bacterianos , Glicosiltransferases/genética , Modelos Moleculares , Família Multigênica , Naftoquinonas/metabolismo , Filogenia , Streptomyces/genética
13.
Clin Endosc ; 54(3): 340-347, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33302328

RESUMO

BACKGROUND/AIMS: The aim of this study was to evaluate the safety of sedation with propofol as an alternative to benzodiazepine drugs in outpatient endoscopy. METHODS: In this prospective study, examinees who underwent outpatient endoscopy under propofol sedation and submitted a nextday questionnaire with providing informed consent were evaluated. Periprocedural acute responses, late adverse events within 24 hours, and examinee satisfaction were evaluated. RESULTS: Among the 4,122 patients who received propofol in the 17,978 outpatient-based endoscopic examinations performed between November 2016 and March 2018, 2,305 eligible examinees (esophagogastroduodenoscopy for 1,340, endoscopic ultrasonography for 945, and total colonoscopy for 20) were enrolled, and their responses to a questionnaire were analyzed. The mean propofol dose was 69.6±24.4 mg (range, 20-200 mg). Diazepam, midazolam, and/or pentazocine in combination with propofol was administered to 146 examinees. Mild oxygen desaturation was observed in 59 examinees (2.6%); and mild bradycardia, in 2 (0.09%). Other severe reactions or late events did not occur. After eliminating 181 invalid responses, 97.7% (2,065/2,124) of the patients desired propofol sedation in future examinations. CONCLUSION: Propofol sedation was found to be safe-without severe adverse events or accidents-for outpatient endoscopy on the basis of the patients' next-day self-evaluation. Given the high satisfaction level, propofol sedation might be an ideal tool for painless endoscopic screening.

14.
Anticancer Drugs ; 21(10): 907-16, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20847643

RESUMO

Glypican 3 (GPC3), a glycosylphosphatidylinositol-anchored heparan sulfate proteoglycan, is expressed in a majority of hepatocellular carcinoma tissues. The murine monoclonal antibody GC33 that specifically binds to the COOH-terminal part of GPC3 causes strong antibody-dependent cellular cytotoxicity against hepatocellular carcinoma cells and exhibits strong antitumor activity in the xenograft models. To apply GC33 for clinical use, we generated a humanized GC33 from complementarity-determining region grafting with the aid of both the hybrid variable region and two-step design methods. The humanized antibody bound to GPC3 specifically and induced antibody-dependent cellular cytotoxicity as effectively as a chimeric GC33 antibody. To improve stability of the humanized GC33, we further optimized humanized GC33 by replacing the amino acid residues that may affect the structure of the variable region of a heavy chain. Substitution of Glu6 with Gln in the heavy chain significantly improved the stability under high temperatures. GC33 also has the risk of deamidation of the -Asn-Gly- sequence in the complementarity-determining region 1 of the light chain. As substitution of Asn diminished the antigen binding, we changed the neighboring Gly to Arg to avoid deamidation. The resulting humanized anti-GPC3 antibody was as efficacious as chimeric GC33 against the HepG2 xenograft and is now being evaluated in clinical trials.


Assuntos
Anticorpos Monoclonais Murinos/imunologia , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/terapia , Glipicanas/imunologia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/terapia , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Anticorpos Monoclonais Murinos/química , Anticorpos Monoclonais Murinos/uso terapêutico , Citotoxicidade Celular Dependente de Anticorpos , Carcinoma Hepatocelular/patologia , Regiões Determinantes de Complementaridade/imunologia , Desenho de Fármacos , Humanos , Região Variável de Imunoglobulina/imunologia , Neoplasias Hepáticas/patologia , Camundongos , Estabilidade Proteica , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Front Cell Dev Biol ; 8: 855, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33042986

RESUMO

Plasmalogens are a subclass of ether glycerophospholipids characterized by a vinyl-ether bond at the sn-1 position of the glycerol backbone. Plasmalogen biosynthesis is initiated in peroxisomes. At the third step of plasmalogen synthesis, alkyl-dihydroxyacetonephosphate (DHAP) is enzymatically reduced to 1-alkyl-sn-glycero-3-phospate by acyl/alkyl DHAP reductase (ADHAPR), whose activity is found in both peroxisomes and microsomes. We herein show that knockdown of ADHAPR in HeLa cells reduced the synthesis of ethanolamine plasmalogen (PlsEtn), similar to the Chinese hamster ovary cell mutant FAA.K1B deficient in ADHAPR activity. Endogenous ADHAPR and ectopically expressed FLAG-tagged ADHAPR were localized to peroxisomes and endoplasmic reticulum (ER) as a type I integral membrane protein in HeLa cells. ADHAPR targets to peroxisomes via a Pex19p-dependent class I pathway. In addition, it is also inserted into the ER via the SRP-dependent mechanism. The ADHAPR mutant lacking the N-terminal domain preferentially targets to the ER, restoring the reduced level of PlsEtn synthesis in FAA.K1B cell. In contrast, the expression of full-length ADHAPR in the mutant cells elevates the synthesis of phosphatidylethanolamine, but not PlsEtn. Taken together, these results suggest that the third step of plasmalogen synthesis is mediated by ER-localized ADHAPR.

16.
J Food Prot ; 83(7): 1234-1240, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32577758

RESUMO

ABSTRACT: Hygiene management of domestic refrigerators is an important aspect of food poisoning prevention. The aim of the present study was to confirm the relationship between microbial contamination and hygiene management by measuring microbial levels and investigating temperature and cleaning frequency and method of domestic refrigerators in Japan. We analyzed three internal sections (the egg compartment, bottom shelf, and vegetable drawer) of 100 domestic refrigerators in Japan. Salmonella, Listeria monocytogenes, and Yersinia enterocolitica were not found in any of the refrigerators, but coliforms and Escherichia coli were detected in more than one household, and Staphylococcus aureus was the most frequently isolated pathogen. The prevalences of these microorganisms had similar tendencies in all three sections sampled and were highest in the vegetable drawer. The temperature distribution in the refrigerators was also investigated, and a temperature >6.1°C (improper temperature) was found in 46.2% of the areas surveyed. Only 17% of the respondents cleaned their refrigerators monthly or more often, and this frequency was lower than that reported in other countries. Fifty percent of the respondents used only water to clean the refrigerator, 10% used only an alcohol or disinfecting wipe, and 8% used only a dry cloth. Although no significant correlations were found between microbial contamination and temperatures in refrigerators, correlations were found between microbial contamination and refrigerator cleaning frequency and/or method. To our knowledge, this is the first detailed survey concerning relationships between microbial contamination and hygiene management in domestic refrigerators in Japan. The data obtained can be used to promote food poisoning management in Japanese households.


Assuntos
Listeria monocytogenes , Refrigeração , Contagem de Colônia Microbiana , Contaminação de Alimentos/análise , Microbiologia de Alimentos , Japão , Salmonella , Temperatura
17.
Biosci Biotechnol Biochem ; 73(1): 109-16, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19129644

RESUMO

The beta-1,4-mannanase 5C gene (man5C) of Vibrio sp. strain MA-138 was cloned and expressed in Escherichia coli. The man5C gene consisted of 2,010 bp nucleotides encoding a protein of 669 amino acids with a predicted molecular weight of 76,309. beta-1,4-Mannanase (Man5C) is a modular enzyme composed of a catalytic module belonging to glycoside hydrolase family 5, a linker region, and a putative carbohydrate-binding module (CBM) belonging to family 27. Recombinant Man5C exhibited maximal activity at 50 degrees C at pH 7.0, and it had a K(m) of 0.6 mg ml(-1) and a V(max) of 556.2 micromol min(-1) mumol(-1) for glucomannan. Binding studies revealed that the C-terminal putative CBM27 had the ability to bind soluble beta-mannans and contributed to increasing the rate of depolymerization by binding to the polymeric substrate. Man5C of Vibrio sp. MA-138 is the first non-extremophile enzyme to be identified as a beta-mannanase possessing CBM27.


Assuntos
Vibrio/enzimologia , beta-Manosidase/metabolismo , Sequência de Aminoácidos , Clonagem Molecular , Cinética , Mananas/metabolismo , Peso Molecular , Ligação Proteica , beta-Manosidase/genética
19.
Microbes Environ ; 34(2): 191-198, 2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31080215

RESUMO

We investigated whether ectomycorrhizal (ECM) fungal species exhibit antibacterial activity towards culturable bacterial communities in mycorrhizospheres. Four hundred and thirty bacterial strains were isolated from the ECM root tips of Pinus densiflora and bulk soil, and 21 were co-cultured with six ECM fungal species. Three hundred and twenty-nine bacterial 16S rDNA sequences were identified in ECM roots (n=185) and bulk soil (n=144). Mycorrhizosphere isolates were dominated by Gram-negative Proteobacteria from 16 genera, including Burkholderia, Collimonas, Paraburkholderia, and Rhizobium. Paraburkholderia accounted for approximately 60%. In contrast, bulk soil isolates contained a high number of Gram-positive Firmicutes, particularly from Bacillus. Paraburkholderia accounted for ≤20% of the bacterial isolates from bulk soil, which was significantly lower than its percentage in ECM root tips. Co-cultures of six ECM fungal species with the 21 bacterial strains revealed that eight strains of three Gram-positive genera-Arthrobacter, Bacillus, and Lysinibacillus-were sensitive to the antibacterial activity of the fungi. In contrast, the Gram-negative strains, including five Paraburkholderia strains, two Burkholderia strains, and a Rhizobium sp., were not sensitive. The strength of fungal antibacterial activity varied in a species-dependent manner, but consistently affected Gram-positive bacteria. These results suggest that Gram-positive bacteria are excluded from the mycorrhizosphere by the antibacterial activity of ECM fungi, which develops specific soil bacterial communities in the mycorrhizosphere.


Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Microbiota , Micorrizas/metabolismo , Rizosfera , Antibacterianos/metabolismo , Bactérias/classificação , Bactérias/crescimento & desenvolvimento , Bactérias/isolamento & purificação , Técnicas de Cocultura , Fungos/classificação , Fungos/crescimento & desenvolvimento , Fungos/metabolismo , Microbiota/efeitos dos fármacos , Microbiota/genética , Micélio/crescimento & desenvolvimento , Micorrizas/classificação , Micorrizas/crescimento & desenvolvimento , Pinus/microbiologia , Raízes de Plantas/microbiologia , Microbiologia do Solo
20.
Ann Gastroenterol Surg ; 3(2): 209-216, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30923791

RESUMO

AIM: Endoscopic decompression using the self-expandable metallic colonic stent (SEMS) or transanal decompression tube (TDT) can convert emergency surgery into elective one-stage surgery for obstructive colorectal cancer (OCRC). The aim of the present study was to clarify the effect of SEMS and TDT on long-term oncological outcomes. METHODS: We retrospectively analyzed 76 consecutive pathological stage II and III OCRC patients who were inserted with SEMS or TDT as a bridge to curative surgery between 2009 and 2018. RESULTS: There were 53 SEMS cases and 23 TDT cases. The tumor was located in the left colon in 58 cases and in the right colon in 18 cases. The interval between the decompression and the surgery was 16.5 days in the SEMS group and 13.0 days in the TDT group (P = 0.09). Technical and clinical success rates were 100% and 100% for SEMS, and 95% and 91% for TDT, respectively. Stoma was created in four patients in the SEMS group, and in five in the TDT group (P = 0.08). Three-year overall survival rates of the SEMS and TDT groups were 82% and 86% (P = 0.94), and disease-free survival rates were 68% and 62% (P = 0.79), respectively. The recurrence pattern was not significantly different. CONCLUSION: This study found no statistically significant differences between the effects of SEMS and TDT for OCRC as a bridge to surgery on long-term outcomes.

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