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1.
Molecules ; 29(5)2024 Feb 29.
Article in English | MEDLINE | ID: mdl-38474598

ABSTRACT

Single-particle inductively coupled plasma mass spectrometry (spICP-MS) has been used for particle size measurement of diverse types of individual nanoparticles and micrometer-sized carbon-based particles such as microplastics. However, its applicability to the measurement of micrometer-sized non-carbon-based particles such as silica (SiO2) particles is unclear. In this study, the applicability of spICP-MS to particle size measurement of non-porous/mesoporous SiO2 microspheres with a nominal diameter of 5.0 µm or smaller was investigated. Particle sizes of these microspheres were measured using both spICP-MS based on a conventional calibration approach using an ion standard solution and scanning electron microscopy as a reference technique, and the results were compared. The particle size distributions obtained using both techniques were in agreement within analytical uncertainty. The applicability of this technique to the detection of metal-containing protein-binding mesoporous SiO2 microspheres was also investigated. Bound iron (Fe)-containing proteins (i.e., lactoferrin and transferrin) of mesoporous SiO2 microspheres were detected using Fe as a presence marker for the proteins. Thus, spICP-MS is applicable to the particle size measurement of large-sized and non-porous/mesoporous SiO2 microspheres. It has considerable potential for element-based detection and qualification of bound proteins of mesoporous SiO2 microspheres in a variety of applications.


Subject(s)
Plastics , Silicon Dioxide , Silicon Dioxide/chemistry , Particle Size , Microspheres , Mass Spectrometry/methods
2.
Medicina (Kaunas) ; 59(10)2023 Sep 25.
Article in English | MEDLINE | ID: mdl-37893433

ABSTRACT

Background and Objectives: Vasa previa (VP) is a significant perinatal complication that can have serious consequences for the fetus/neonate. Velamentous cord insertion (VCI) is a crucial finding in prenatal placental morphology surveillance as it is indicative of comorbid VP. Assisted reproductive technology (ART) has been identified as a risk factor for VCI, so identifying risk factors for VCI in ART could improve VP recognition. This study aims to evaluate the displacement of umbilical cord insertion (CI) from the placental center and to examine the relationship between the modes of conception. Materials and Methods: We conducted a retrospective study at the Obstetrics Department of Osaka Metropolitan University Hospital in Japan between May 2020 and June 2022. The study included a total of 1102 patients who delivered after 22 weeks of gestation. They were divided into three groups: spontaneous pregnancy, conventional in vitro fertilization (cIVF), and in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI). We recorded patient background information, perinatal complications, perinatal outcomes, and a numerical "displacement score", indicating the degree of separation between umbilical CI and the placental center. Results: The displacement score was significantly higher in the cIVF and IVF/ICSI groups compared with the spontaneous conception group. Additionally, the IVF/ICSI group showed a significantly higher displacement score than the cIVF group. Conclusions: Our study provides the first evidence that the methods of ART can affect the location of umbilical CI on the placental surface. Furthermore, we found that IVF/ICSI may contribute to greater displacement of CI from the placental center.


Subject(s)
Vasa Previa , Vascular Diseases , Infant, Newborn , Pregnancy , Humans , Male , Female , Sperm Injections, Intracytoplasmic/adverse effects , Vasa Previa/etiology , Retrospective Studies , Placenta , Semen , Umbilical Cord , Reproductive Techniques, Assisted
3.
J Autoimmun ; 135: 102995, 2023 02.
Article in English | MEDLINE | ID: mdl-36724643

ABSTRACT

Cutaneous arteritis (CA) is a single-organ vasculitis that exclusively affects the small to medium-sized arteries of the skin. Diagnosis depends on a histological investigation with skin biopsy, which could be burdensome for both patients and clinicians. Moreover, the pathogenesis of CA remains unstudied, and treatment has not yet been established. Herein, we applied our proteome-wide autoantibody screening method to explore autoantibodies in the serum of CA patients. As a result, anti-transcobalamin receptor (TCblR) antibodies (Abs) were specifically detected in 24% of CA patients. Patients with positive anti-TCblR Abs were spared from peripheral neuropathy compared to those with negative anti-TCblR Abs, showing characteristics as CA confined to the skin. In addition, we revealed that anti-TCblR Abs trigger the autocrine loop of interleukin-6 mediated by tripartite motif-containing protein 21 in human endothelial cells and induce periarterial inflammation in murine skin. Furthermore, we demonstrated that methylcobalamin, a ligand of TCblR, ameliorates inflammation caused by anti-TCblR Abs both in vitro and in vivo. Collectively, our investigation unveils the pathologic significance of anti-TCblR Abs in CA and their potential as a diagnostic marker and a pathophysiology-oriented therapeutic target.


Subject(s)
Arteritis , Transcobalamins , Humans , Animals , Mice , Transcobalamins/metabolism , Proteome/metabolism , Autoantibodies/metabolism , Endothelial Cells/metabolism , Inflammation
4.
Medicina (Kaunas) ; 58(8)2022 Aug 11.
Article in English | MEDLINE | ID: mdl-36013550

ABSTRACT

Background and Objectives: There are no reports showing the hematopoietic effect of TJ-108 on pregnant women. The aim of this study was to investigate the effect of TJ-108 on the hemoglobin and hematocrit levels, and white blood cell and platelet counts of pregnant women complicated with placenta previa who were managed with autologous blood storage for cesarean section. Materials and Methods: We studied two groups of patients who were complicated with placenta previa and who underwent cesarean delivery. Group A consisted of women who were treated with oral iron medication (100 mg/day), and Group B consisted of women who were treated with TJ-108 at a dose of 9.0 g per day, in addition to oral iron medication, from the first day of blood storage until the day before cesarean delivery. To evaluate the effect of TJ-108, the patients' red blood cell (RBC); Hb; hematocrit (Ht); white blood cell (WBC); and platelet count (PLT) levels were measured 7 days after storage and at postoperative days (POD) 1 and 5. Results: The study included 65 individuals, 38 in group A and 27 in group B. At the initial storage, a 0.2 g/dL reduction in Hb levels was observed, as compared to the initial Hb levels, in the TJ-108 treated patients, whereas a 0.6 g/dL reduction in Hb levels was observed in the non-TJ-108 treated group. On the other hand, regarding the second and subsequent storages, no significant difference was found in the decrease in the Hb levels of both groups. Conclusions: This study is the first report showing the effect of TJ-108 on improving anemia in pregnant women, presumably by its boosting effect on myelohematopoiesis. Therefore, the combined administration of both iron and TJ-108 is effective as a strategy for pregnant women at a high risk of PPH due to complications such as placenta previa.


Subject(s)
Blood Preservation , Placenta Previa , Plant Preparations , Postpartum Hemorrhage , Cesarean Section/adverse effects , Female , Hemoglobins , Humans , Iron , Japan , Medicine, East Asian Traditional , Placenta Previa/etiology , Plant Preparations/therapeutic use , Plants, Medicinal , Postpartum Hemorrhage/surgery , Pregnancy , Pregnant Women , Retrospective Studies
5.
Front Immunol ; 13: 893086, 2022.
Article in English | MEDLINE | ID: mdl-35603173

ABSTRACT

Autoantibodies are found in various pathological conditions such as autoimmune diseases, infectious diseases, and malignant tumors. However their clinical implications have not yet been fully elucidated. Herein, we conducted proteome-wide autoantibody screening and quantification with wet protein arrays consisting of proteins synthesized from proteome-wide human cDNA library (HuPEX) maintaining their three-dimensional structure. A total of 565 autoantibodies were identified from the sera of three representative inflammatory disorders (systemic sclerosis, psoriasis, and cutaneous arteritis). Each autoantibody level either positively or negatively correlated with serum levels of C-reactive protein, the best-recognized indicator of inflammation. In particular, we discovered total levels of a subset of autoantibodies correlates with the severity of clinical symptoms. From the sera of malignant melanoma, 488 autoantibodies were detected. Notably, patients with metastases had increased overall autoantibody production compared to those with tumors limiting to the primary site. Collectively, proteome-wide screening of autoantibodies using the in vitro proteome can reveal the "autoantibody landscape" of human subjects and may provide novel clinical biomarkers.


Subject(s)
Autoimmune Diseases , Protein Array Analysis , Autoantibodies , Biomarkers , Humans , Protein Array Analysis/methods , Proteome
6.
Cancer Immunol Immunother ; 71(11): 2743-2755, 2022 Nov.
Article in English | MEDLINE | ID: mdl-35429246

ABSTRACT

The aim of this study was to determine the efficacy and the biomarkers of the CHP-NY-ESO-1 vaccine complexed with full-length NY-ESO-1 protein and a cholesteryl pullulan (CHP) in patients with esophageal squamous cell carcinoma (ESCC) after surgery. We conducted a randomized phase II trial. Fifty-four patients with NY-ESO-1-expressing ESCC who underwent radical surgery following cisplatin/5-fluorouracil-based neoadjuvant chemotherapy were assigned to receive either CHP-NY-ESO-1 vaccination or observation as control. Six doses of CHP-NY-ESO-1 were administered subcutaneously once every two weeks, followed by nine more doses once every four weeks. The endpoints were disease-free survival (DFS) and safety. Exploratory analysis of tumor tissues using gene-expression profiles was also performed to seek the biomarker. As there were no serious adverse events in 27 vaccinated patients, we verified the safety of the vaccine. DFS in 2 years were 56.0% and 58.3% in the vaccine arm and in the control, respectively. Twenty-four of 25 patients showed NY-ESO-1-specific IgG responses after vaccination. Analysis of intra-cohort correlations among vaccinated patients revealed that 5% or greater expression of NY-ESO-1 was a favorable factor. Comprehensive analysis of gene expression profiles revealed that the expression of the gene encoding polymeric immunoglobulin receptor (PIGR) in tumors had a significantly favorable impact on outcomes in the vaccinated cohort. The high PIGR-expressing tumors that had higher NY-ESO-1-specific IgA response tended to have favorable prognosis. These results suggest that PIGR would play a major role in tumor immunity in an antigen-specific manner during NY-ESO-1 vaccinations. The IgA response may be relevant.


Subject(s)
Cancer Vaccines , Esophageal Neoplasms , Esophageal Squamous Cell Carcinoma , Receptors, Polymeric Immunoglobulin , Antibodies, Neoplasm , Antigens, Neoplasm , Cisplatin , Esophageal Squamous Cell Carcinoma/drug therapy , Fluorouracil , Glucans , Humans , Immunoglobulin A , Immunoglobulin G , Membrane Proteins , Prognosis
7.
Heliyon ; 7(8): e07748, 2021 Aug.
Article in English | MEDLINE | ID: mdl-34395931

ABSTRACT

BACKGROUND: Antibody production is one of the primary mechanisms for recovery from coronavirus disease 2019 (COVID-19). It is speculated that massive clonal expansion of B cells, which can produce clinically meaningful neutralizing antibodies, occurs in patients who recover on the timing of acquiring adaptive immunity. METHODS: To evaluate fluctuations in clonal B cells and the size of the clones, we chronologically assessed the B-cell receptor (BCR) repertoire in three patients with COVID-19 who recovered around 10 days after symptom onset. RESULTS: We focused on the three dominant clonotypes (top 3) in each individual. The percentage frequencies of the top 3 clonotypes increased rapidly and accounted for 27.8 % on day 9 in patient 1, 10.4 % on day 12 in patient 2, and 10.8 % on day 11 in patient 3, respectively. The frequencies of these top 3 clonotypes rapidly decreased as the patients' clinical symptoms improved. Furthermore, BCR network analysis revealed that accumulation of clusters composed of similar complementarity-determining region 3 (CDR3) sequences were rapidly formed, grew, and reached their maximum size around 10 days after symptom onset. CONCLUSIONS: BCR repertoire analysis revealed that a massive surge of some unique BCRs occurs during the acquisition of adaptive immunity and recovery. The peaks were more prominent than expected. These results provide insight into the important role of BCRs in the recovery from COVID-19 and raise the possibility of developing neutralizing antibodies as COVID-19 immunotherapy.

8.
Genes Cells ; 26(3): 180-189, 2021 Mar.
Article in English | MEDLINE | ID: mdl-33527666

ABSTRACT

TRA98 is a rat monoclonal antibody (mAb) which recognizes a specific antigen in the nuclei of germ cells. mAb TRA98 has been used to understand the mechanism of germ cell development and differentiation in many studies. In mice, the antigen recognized by mAb TRA98 or GCNA1 has been reported to be a GCNA gene product, but despite the demonstration of the immunoreactivity of this mAb in human testis and sperm in 1997, the antigen in humans remains unknown, as of date. To identify the human antigen recognized by mAb TRA98, a human comprehensive wet protein array was developed containing 19,446 proteins derived from human cDNAs. Using this array, it was found that the antigen of mAb TRA98 is not a GCNA gene product, but nuclear factor-κB activating protein (NKAP). In mice, mAb TRA98 recognized both the GCNA gene product and NKAP. Furthermore, conditional knockout of Nkap in mice revealed a phenotype of Sertoli cell-only syndrome. Although NKAP is a ubiquitously expressed protein, NKAP recognized by mAb TRA98 in mouse testis was SUMOylated. These results suggest that NKAP undergoes modifications, such as SUMOylation in the testis, and plays an important role in spermatogenesis.


Subject(s)
Antibodies, Monoclonal/metabolism , Antigens/metabolism , Germ Cells/metabolism , Protein Array Analysis , Animals , Humans , Male , Mice , Repressor Proteins/genetics , Repressor Proteins/metabolism , Testis/metabolism
9.
Molecules ; 25(7)2020 Apr 04.
Article in English | MEDLINE | ID: mdl-32260359

ABSTRACT

Thrombopoietin (THPO) is a circulatory cytokine that plays an important role in platelet production. The presence of anti-THPO antibody relates to thrombocytopenia and is rarely seen in hematopoietic and autoimmune diseases. To date, there had been no reports that focused on the anti-THPO antibody in patients with type 2 diabetes mellitus (T2DM). To evaluate prevalence of the anti-THPO antibody in patients with T2DM and the relationship between anti-THPO antibody and platelet count, a cross-sectional study was performed on 82 patients with T2DM. The anti-THPO antibody was measured by ELISA using preserved sera and detected in 13 patients. The average platelet count was significantly lower in patients with the anti-THPO antibody than in those without the anti-THPO antibody. Multivariate linear regression analyses showed a significant relationship between the anti-THPO antibody and platelet count, after adjusting for other variables. To our best knowledge, this was the first report on the effect of the anti-THPO antibody on platelet count in patients with T2DM. Further investigation is needed to validate the prevalence and pathological significance of the anti-THPO antibody in patients with T2DM.


Subject(s)
Antibodies/blood , Diabetes Mellitus, Type 2/blood , Thrombopoietin/immunology , Aged , Aged, 80 and over , Cross-Sectional Studies , Diabetes Mellitus, Type 2/immunology , Female , Humans , Male , Middle Aged , Pilot Projects , Platelet Count , Regression Analysis
10.
Cancer Sci ; 111(2): 658-666, 2020 Feb.
Article in English | MEDLINE | ID: mdl-31823471

ABSTRACT

Metabolic reprogramming, including the Warburg effect, is a hallmark of cancer. Indeed, the diversity of cancer metabolism leads to cancer heterogeneity, but accurate assessment of metabolic properties in tumors has not yet been undertaken. Here, we performed absolute quantification of the expression levels of 113 proteins related to carbohydrate metabolism and antioxidant pathways, in stage III colorectal cancer surgical specimens from 70 patients. The Warburg effect appeared in absolute protein levels between tumor and normal mucosa specimens demonstrated. Notably, the levels of proteins associated with the tricarboxylic citric acid cycle were remarkably reduced in the malignant tumors which had relapsed after surgery and treatment with 5-fluorouracil-based adjuvant therapy. In addition, the efficacy of 5-fluorouracil also decreased in the cultured cancer cell lines with promotion of the Warburg effect. We further identified nine and eight important proteins, which are closely related to the Warburg effect, for relapse risk and 5-fluorouracil benefit, respectively, using a biomarker exploration procedure. These results provide us a clue for bridging between metabolic protein expression profiles and benefit from 5-fluorouracil adjuvant chemotherapy.


Subject(s)
Antioxidants/metabolism , Carbohydrate Metabolism , Colorectal Neoplasms/drug therapy , Fluorouracil/administration & dosage , Adult , Aged , Chemotherapy, Adjuvant , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Female , Fluorouracil/pharmacology , Fluorouracil/therapeutic use , Gene Expression Regulation, Neoplastic/drug effects , HCT116 Cells , Humans , Male , Middle Aged , Neoplasm Staging , Signal Transduction/drug effects , Treatment Outcome
11.
Genes Cells ; 25(1): 41-53, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31733161

ABSTRACT

Proteomic analysis requires protein tags that enable high-throughput handling; however, versatile tags that can be used in in vitro expression systems are currently lacking. In this study, we developed an insoluble protein tag, INSOL-tag, derived from human transcription factor MafG. The INSOL-tagged target protein is expressed in a eukaryotic in vitro expression system and recovered as a pellet following centrifugation at 19,000 × g for 20 min. Comparisons of the target protein recovery rates of GST-tag and INSOL-tag using 111 cytoplasmic proteins revealed a fourfold increase in the yield of INSOL-tagged proteins. Using 267 cancer antigens purified with INSOL-tag, we subsequently developed an INSOL-CTA array method, for profiling autoantibodies in sera of cancer patients. The detection limit of the array was approximately 11.1 pg IgG, and the correlation with ELISA was high (R2  = .993, .955). Moreover, when autoantibody profiling of digestive cancer patient sera was performed, antigen spreading was observed. These data suggest that INSOL-tag is a versatile tag that can insolubilize a wide range of target proteins. It is therefore expected to become a powerful tool in comprehensive protein preparation for protein arrays, antibody production, and mass spectrometry.


Subject(s)
MafG Transcription Factor/isolation & purification , MafG Transcription Factor/metabolism , Proteomics/methods , Repressor Proteins/isolation & purification , Repressor Proteins/metabolism , High-Throughput Screening Assays/methods , Humans , MafG Transcription Factor/genetics , Mass Spectrometry/methods , Protein Array Analysis/methods , Protein Engineering/methods , Proteome/genetics , Repressor Proteins/genetics , Transcription Factors/metabolism
12.
Am J Pathol ; 188(6): 1328-1333, 2018 06.
Article in English | MEDLINE | ID: mdl-29751934

ABSTRACT

Our aim was to develop a serodiagnostic marker for lung cancer. Monoclonal antibodies were generated, and one antibody designated as KU-Lu-1, recognizing cytoskeleton-associated protein 4 (CKAP4), was studied further. To evaluate the utility of KU-Lu-1 antibody as a serodiagnostic marker for lung cancer, reverse-phase protein array analysis was performed with sera of 271 lung cancer patients and 100 healthy controls. CKAP4 was detected in lung cancer cells and tissues, and its secretion into the culture supernatant was also confirmed. The serum CKAP4 levels of lung cancer patients were significantly higher than those of healthy controls (P < 0.0001), and the area under the curve of receiver-operating characteristic curve analysis was 0.890, with 81.1% sensitivity and 86.0% specificity. Furthermore, the serum CKAP4 levels were also higher in patients with stage I adenocarcinoma or squamous cell carcinoma than in healthy controls (P < 0.0001). Serum CKAP4 levels may differentiate lung cancer patients from healthy controls, and they may be detected early even in stage I non-small cell lung cancer. Serum CKAP4 levels were also significantly higher in lung cancer patients than in healthy controls in the validation set (P < 0.0001). The present results provide evidence that CKAP4 may be a novel early serodiagnostic marker for lung cancer.


Subject(s)
Adenocarcinoma/diagnosis , Biomarkers, Tumor/blood , Carcinoma, Non-Small-Cell Lung/diagnosis , Carcinoma, Squamous Cell/diagnosis , Lung Neoplasms/diagnosis , Membrane Proteins/blood , Adenocarcinoma/blood , Carcinoma, Non-Small-Cell Lung/blood , Carcinoma, Squamous Cell/blood , Case-Control Studies , Humans , Lung Neoplasms/blood , Prognosis , Protein Array Analysis , Tumor Cells, Cultured
13.
Pathol Int ; 68(4): 232-240, 2018 Apr.
Article in English | MEDLINE | ID: mdl-29431238

ABSTRACT

We established the KU-Lu-8 monoclonal antibody (MoAb) using a lung cancer cell line as an antigen and a random immunization method. The KU-Lu-8 MoAb recognizes basigin (BSG), which is a transmembrane-type glycoprotein that is strongly expressed on the cell membranes of lung cancer cells. This study aimed to clarify the relationships between BSG expression and clinicopathological parameters and determine the prognostic significance of BSG expression in pulmonary adenocarcinoma (AC) patients. To evaluate the significance of BSG expression in lung cancer, we immunohistochemically analyzed 113 surgically resected pulmonary adenocarcinomas, and the associations between BSG expression and various clinicopathological parameters were evaluated. Kaplan-Meier survival analysis and Cox proportional hazards models were used to investigate the effects of BSG expression on survival. Clinicopathologically, BSG expression was significantly associated with tumor differentiation, vascular invasion, lymphatic invasion, and a poor prognosis. In particular, BSG expression was significantly correlated with poorer survival in patients with stage I AC. The high BSG expression group (compared with the low BSG expression group) exhibited adjusted hazard ratios for mortality of 4.694. BSG expression is indicative of a poor prognosis in AC patients, particularly in those with stage I disease.


Subject(s)
Adenocarcinoma/pathology , Basigin/biosynthesis , Biomarkers, Tumor/analysis , Lung Neoplasms/pathology , Adenocarcinoma/metabolism , Adenocarcinoma/mortality , Adenocarcinoma of Lung , Adult , Aged , Aged, 80 and over , Basigin/analysis , Disease-Free Survival , Female , Humans , Kaplan-Meier Estimate , Lung Neoplasms/metabolism , Lung Neoplasms/mortality , Male , Middle Aged , Prognosis , Proportional Hazards Models
14.
Anticancer Res ; 37(12): 6705-6714, 2017 12.
Article in English | MEDLINE | ID: mdl-29187447

ABSTRACT

BACKGROUND/AIM: Bladder cancer (BC) has a high recurrence rate and may progress to being a muscle-invasive lesion, that is potentially associated with a poor prognosis. We identified tumor-associated proteins that were recognized by autoantibodies in sera from patients with high-grade non-muscle-invasive bladder cancer (HG-NMIBC) by proteomic analysis. MATERIALS AND METHODS: The serum levels of these autoantibodies against identified proteins were validated by dot blot analysis with sera from 95 patients with BC and 35 healthy controls. The expression of identified proteins was immunohistochemically analyzed in 115 BC tissues. RESULTS: Autoantibody against protein phosphatase 1, catalytic subunit, alpha isoform (PPP1CA) protein was detected in pretreated sera from patients with HG-NMIBC who showed progression. The serum IgG level of anti-PPP1CA autoantibody was significantly correlated with pathological stage, grade, lymphovascular invasion, and prognosis. The immunoreactions for PPP1CA protein in BC was significantly correlated with pathological stage, grade, and lymphovascular invasion. CONCLUSION: PPP1CA is a candidate sero-diagnostic and prognostic marker for patients with BC.


Subject(s)
Autoantibodies/immunology , Proteome/immunology , Proteomics/methods , Urinary Bladder Neoplasms/immunology , Adult , Aged , Aged, 80 and over , Autoantibodies/blood , Disease Progression , Female , Humans , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunohistochemistry , Male , Middle Aged , Neoplasm Grading , Prognosis , Protein Phosphatase 1/immunology , Protein Phosphatase 1/metabolism , Urinary Bladder Neoplasms/blood , Urinary Bladder Neoplasms/metabolism
15.
J Proteome Res ; 16(10): 3576-3584, 2017 10 06.
Article in English | MEDLINE | ID: mdl-28810742

ABSTRACT

Wnt/ß-catenin signaling plays important roles in both ontogenesis and development. In the absence of a Wnt stimulus, ß-catenin is degraded by a multiprotein "destruction complex" that includes Axin, APC, GSK3B, and FBXW11. Although the key molecules required for transducing Wnt signals have been identified, a quantitative understanding of this pathway has been lacking. Here, we calculated the absolute number of ß-catenin destruction complexes by absolute protein quantification using LC-MS/MS. Similar amounts of destruction complex-constituting proteins and ß-catenin interacted, and the number of destruction complexes was calculated to be about 1468 molecules/cell. We demonstrated that the calculated number of destruction complexes was valid for control of the ß-catenin destruction rate under steady-state conditions. Interestingly, APC had the minimum expression level among the destruction complex components at about 2233 molecules/cell, and this number approximately corresponded to the calculated number of destruction complexes. Decreased APC expression by siRNA transfection decreased the number of destruction complexes, resulting in ß-catenin accumulation and stimulation of the transcriptional activity of T-cell factor. Taken together, our results suggest that the amount of APC expression is the rate-limiting factor for the constitution of ß-catenin destruction complexes.


Subject(s)
Adenomatous Polyposis Coli Protein/genetics , Axin Signaling Complex/genetics , Wnt Signaling Pathway/genetics , beta Catenin/genetics , Axin Protein/genetics , Axin Signaling Complex/chemistry , Axin Signaling Complex/metabolism , Gene Expression Regulation/genetics , Glycogen Synthase Kinase 3 beta/genetics , HCT116 Cells , Humans , Phosphorylation , RNA, Small Interfering/genetics , Ubiquitin-Protein Ligases/genetics , beta Catenin/isolation & purification , beta-Transducin Repeat-Containing Proteins/genetics
16.
Nat Methods ; 14(3): 251-258, 2017 03.
Article in English | MEDLINE | ID: mdl-28267743

ABSTRACT

Targeted proteomics approaches are of value for deep and accurate quantification of protein abundance. Extending such methods to quantify large numbers of proteins requires the construction of predefined targeted assays. We developed a targeted proteomics platform-in vitro proteome-assisted multiple reaction monitoring (MRM) for protein absolute quantification (iMPAQT)-by using >18,000 human recombinant proteins, thus enabling protein absolute quantification on a genome-wide scale. Our platform comprises experimentally confirmed MRM assays of mass tag (mTRAQ)-labeled peptides to allow for rapid and straightforward measurement of the absolute abundance of predefined sets of proteins by mass spectrometry. We applied iMPAQT to delineate the quantitative metabolic landscape of normal and transformed human fibroblasts. Oncogenic transformation gave rise to relatively small but global changes in metabolic pathways resulting in aerobic glycolysis (Warburg effect) and increased rates of macromolecule synthesis. iMPAQT should facilitate quantitative biology studies based on protein abundance measurements.


Subject(s)
Genome, Human/genetics , Mass Spectrometry/methods , Proteome/analysis , Proteomics/methods , Cell Line, Transformed , Fibroblasts/metabolism , Glycolysis/physiology , Humans , Peptide Library , Recombinant Proteins/analysis
17.
Nat Prod Commun ; 12(1): 27-29, 2017 Jan.
Article in English | MEDLINE | ID: mdl-30549818

ABSTRACT

The roots and stolons of some Glycyrrhiza species are used worldwide for traditional folk medicines and commercial pharmaceuticals. Phenolic constituents such as flavonoids and coumarins are medicinal and vary according to species. Therefore, species identification is important for quality analysis. In order to identify Glycyrrhiza species by chemical fingerprinting, methanol extracts of the root bark of Glycyrrhiza uralensis Fischer and G. glabra Linn6 were analyzed using EI-MS. Differences in kinds and quantity of components are reflected in complex EI-MS data and determining characteristic peaks for each species is straightforward.. The chaiacteristic peaks were determined statistically by volcano plot, a multivariate analysis method. EI-MS data of G. uralensis and G. glabra showed differential patterns, and the notable peaks in each pattern were identified. Peaks at m/z 153 and 221 are signature peaks of G. uralensis, and at 11/z 173, 309, and 324 are those of G. glabra. In conclusion, we found species-specific patterns by EI-MS that distinguish G. uralensis and G. glabra. This method based on chemical constituent patterns can be applied to identify other Glycyrrhiza species and similar natural products.


Subject(s)
Glycyrrhiza/chemistry , Chromatography, High Pressure Liquid , Chromatography, Ion Exchange , Metabolomics , Natural Resources , Peptide Mapping , Plant Extracts/chemistry , Plant Roots/chemistry , Species Specificity , Spectrometry, Mass, Electrospray Ionization
18.
Phys Rev E ; 94(5-1): 052209, 2016 Nov.
Article in English | MEDLINE | ID: mdl-27967080

ABSTRACT

In the present study, we analyzed field measurement data obtained for a Japanese expressway and used it as a data set for the validation of microscopic simulation models. Consequently, in accordance with previous studies, we confirmed the common features depicted by the fundamental diagram (flux vs density relation) and lane-usage ratio vs density diagram. We found two things regarding lane-changing behavior: (1) a lane change occurs asymmetrically, where a lane change from a slow to a fast lane differs from that from a fast to a slow lane; and (2) the so-called incentive criterion in the case of small gaps between the preceding vehicles in both slow and fast lanes refers to the velocities and /or the relative velocities with respect to the preceding vehicles, whereas that for relatively large gaps refers to the distances to the preceding vehicles is cast into the above incentive criterion in addition to the two factors mentioned above.

19.
Phys Rev E ; 94(2-1): 022114, 2016 Aug.
Article in English | MEDLINE | ID: mdl-27627253

ABSTRACT

The existence of a zealot who stays a cooperator irrespective of the result of an interaction has been reported to add "social viscosity" to a population and thereby helps increase the cooperation level in prisoner's dilemma games, which premises the so-called well-mixed situation of a population. We found that this is not always true when a spatial structure, i.e., connecting agent, is introduced. Deploying zealots is counterproductive, especially when the underlying topology is homogenous, similar to that of a lattice. Our simulation reveals how the existence of never-converting cooperators destroys rather than boosts cooperation. We explain detailed mechanisms behind this interesting finding by referring to our previously presented concepts with respect to evolutionary dynamic processes for spatial games under the names enduring and expanding periods.

20.
Asian Pac J Cancer Prev ; 16(17): 7959-65, 2015.
Article in English | MEDLINE | ID: mdl-26625826

ABSTRACT

BACKGROUND: Identification of predictive markers for the efficacy of platinum-based chemotherapy is necessary to improve the quality of the life of cancer patients. MATERIALS AND METHODS: We detected proteins recognized by autoantibodies in pretreated sera from patients with lung adenocarcinoma (AC) evaluated as showing progressive disease (PD) or a partial response (PR) after cisplatin-based chemotherapy by proteomic analysis. Then, the levels of the candidate autoantibodies in the pretreated serum were validated by dot-blot analysis for 22 AC patients who received platinum-based chemotherapy, and the expression of identified proteins was immunohistochemically analyzed in 40 AC biopsy specimens. RESULTS: An autoantibody against galectin-3 (Gal-3) was detected in pretreated sera from an AC patient with PD. Serum IgG levels of anti-Gal-3 autoantibody were significantly higher in patients evaluated with PD than in those with PR and stable disease (SD) (p = 0.0084). Furthermore, pretreated biopsy specimens taken from patients evaluated as showing PD following platinum- based chemotherapy showed a tendency to have a higher positive rate of Gal-3 than those with PR and SD (p = 0.0601). CONCLUSIONS: These results suggest that serum IgG levels of anti-Gal-3 autoantibody may be useful to predict the efficacy of platinum-based chemotherapy for patients with lung AC.


Subject(s)
Adenocarcinoma/drug therapy , Antineoplastic Agents/therapeutic use , Autoantibodies/blood , Autoantibodies/immunology , Cisplatin/therapeutic use , Galectin 3/immunology , Lung Neoplasms/drug therapy , Adenocarcinoma of Lung , Biomarkers, Tumor/blood , Biomarkers, Tumor/immunology , Blood Proteins , Cell Line, Tumor , Galectins , Humans , Immunoglobulin G/blood , Immunoglobulin G/immunology , Quality of Life , Treatment Outcome
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