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1.
Folia Morphol (Warsz) ; 77(2): 286-292, 2018.
Article in English | MEDLINE | ID: mdl-29064554

ABSTRACT

BACKGROUND: Anatomical knowledge of the duodenojejunal flexure is necessary for abdominal surgeries, and also important for physiologic studies about the duodenum. But little is known about the anatomy of this region in mammals. Here, we examined comparative anatomy to understand the anatomical formation of the duodenojejunal flexure in mammals. MATERIALS AND METHODS: The areas around the duonenojejunal flexure were ob-served in mouse, rat, dog, pig, and human, and the anatomical structures around the duodenojejunal junction in the animals were compared with those in human. RESULTS: The superior and inferior duodenal folds, and the superior and inferior duodenal fossae were identified in all examined humans. In pig, the structures were not clearly identified because the duodenum strongly adhered to the retroperitoneum and to the mesocolon. In mouse, rat, and dog, only the plica duodenocolica, which is regarded as the animal counterpart of the superior duo-denal fold in human, was identified, and other folds or fossae were not observed, probably because the duodenum was not fixed to the parietal peritoneum in those animals. Transection of the plica duodenocolica could return the normally rotated intestine back to the state of non-rotation in rat. CONCLUSIONS: This study showed the anatomical similarities and dissimilarities of the duodenojejunal flexure among the mammals. Anatomical knowledge of the area is useful for duodenal and pancreatic surgeries, and for animal studies about the duodenum. (Folia Morphol 2018; 77, 2: 286-292).


Subject(s)
Duodenum/anatomy & histology , Jejunum/anatomy & histology , Anatomy, Comparative , Animals , Dogs , Humans , Rats , Species Specificity , Swine
2.
Am J Transplant ; 15(8): 2085-95, 2015 Aug.
Article in English | MEDLINE | ID: mdl-25908212

ABSTRACT

Islet autotransplantation following total pancreatectomy differs from allograft transplantation with respect to the requirement of biliary reconstruction. Although it is known that careful consideration should be given to postoperative cholestatic liver injury after biliary reconstruction, its direct effects on transplanted islets have not been completely elucidated. In this study, we developed a murine model of postoperative cholestatic liver injury after biliary reconstruction with islet autotransplantation that involved syngeneic intraportal islet transplantation into chemically induced diabetic mice and common bile duct ligation. We assessed the viability and function of the transplanted islets. The impaired viability of transplanted islets and increased blood glucose levels indicated restoration of the diabetic state after common bile duct ligation in this murine model. Furthermore, impaired islet viability and function occurred earlier in the transplanted islets than in the surrounding liver tissues, which was consistent with the faster and higher expression of oxidative stress markers in the transplanted islets. Transplanted islets may be more vulnerable to oxidative stress caused by cholestatic liver injury than the surrounding liver tissue. Therefore, patients should be intensively managed after total pancreatectomy with islet autotransplantation to preserve viability and function of the transplanted islets.


Subject(s)
Biliary Tract/physiopathology , Cholestasis/prevention & control , Islets of Langerhans/physiology , Animals , Male , Mice , Mice, Inbred BALB C , Oxidative Stress
3.
Am J Transplant ; 15(6): 1531-42, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25846610

ABSTRACT

Recent basic and clinical studies have assessed the use of highly sensitive imaging modalities for visualizing transplanted islets. We investigated the utility of enhanced ultrasonography, combined with fluorescent acoustic liposome nano/microbubbles (FALs), for evaluating angiogenesis and the endocrine function of transplanted islets. BALB/c mice were classified into three groups: Diabetic mice that underwent syngeneic islet transplantation into the subrenal capsule and achieved normoglycemia (Tx group); those that failed to achieve normoglycemia (Tx-DM group); and those not receiving any treatment (DM group). Mice were examined by FAL-enhanced high frequency ultrasonography. The echogenicity of the islets increased rapidly within the first minute after injection of FALs and remained at a higher level in the Tx group, while small increases were observed in the other two groups. In histological assessments, fluorescently stained erythrocytes could be seen in and around the transplanted islets, indicating that the transplanted islets were enhanced by infusion of FALs via vessel networks between the engrafted islets and tissue. Furthermore, the echogenicity correlated significantly with endocrine parameters, including blood glucose (BG), serum insulin, and the BG change in the glucose tolerance test. In conclusion, the echogenicity of the islets under FAS-enhanced ultrasonosonography correlated with the endocrine status of transplanted islets.


Subject(s)
Contrast Media , Diabetes Mellitus, Experimental/surgery , Islets of Langerhans Transplantation/diagnostic imaging , Islets of Langerhans/diagnostic imaging , Microbubbles , Ultrasonography/methods , Animals , Blood Glucose/metabolism , Diabetes Mellitus, Experimental/chemically induced , Disease Models, Animal , Insulin/blood , Islets of Langerhans/blood supply , Islets of Langerhans/physiology , Mice , Mice, Inbred BALB C , Neovascularization, Physiologic/physiology , Streptozocin/adverse effects , Treatment Outcome
4.
Proc Math Phys Eng Sci ; 478(2260): 20220073, 2022 Apr.
Article in English | MEDLINE | ID: mdl-35510221

ABSTRACT

We introduce the concept of a handlebody decomposition of a three-manifold, a generalization of a Heegaard splitting, or a trisection. We show that two handlebody decompositions of a closed orientable three-manifold are stably equivalent. As an application to materials science, we consider a mathematical model of polycontinuous patterns and discuss a topological study of microphase separation of a block copolymer melt.

5.
Front Rehabil Sci ; 3: 1019089, 2022.
Article in English | MEDLINE | ID: mdl-36569638

ABSTRACT

Background: As a type of welfare technology, care robotics is now widely seen as a potential aide to rehabilitation, increasing independence and enhancing the wellbeing of people with disabilities and older adults. Research into and development of care robots have both been vigorously promoted in North America, Europe and Asia, and the competition for technological advancement in robotics is becoming fierce. AI ethics and policy guidelines are being established. However, there are still differences in attitudes and perceptions, as well as national policies regarding this type of welfare technology. Moreover, despite the anticipated usefulness, it is believed that progress has been slow in the diffusion of care robots. Purpose: In order to explore how public discourses support technological innovation, such as care robots, while preparing society for potential risks and impact, we sought to ascertain whether public discourse on care robots varies from region to region. For example, what are the hopes and promises associated with care robots and what are the concerns? Methods: To address these questions, this article explored how care robots have been portrayed in five major broadsheet newspapers in five jurisdictions in Asia and Europe (France, Great Britain, Hong Kong SAR, Ireland and Japan). We obtained 545 articles for the period between January 2001 and September 2020, more than half of which originated in Japan. A thematic analysis was conducted of these articles written in four languages (Chinese, English, French and Japanese). Results: Positive and negative narratives were teased out, alongside other key prominent themes identified, such as Japan as the land of robots, the pandemic, and the impact of robots on the economy. As the number of robot-related articles grew from the year 2012 onwards, narratives became more nuanced in European newspapers, but not in Asian ones. Furthermore, recent articles began to address the social and relational impact of care robots, while providing concrete examples of improvements in the quality of life for users. Further careful examination will be necessary in the future in order to establish the impact of robotics use in rehabilitation for people with disabilities, older adults, their carers and society at large.

6.
Thorac Cardiovasc Surg ; 59(7): 416-20, 2011 Oct.
Article in English | MEDLINE | ID: mdl-21692022

ABSTRACT

BACKGROUND: The internal thoracic artery (ITA) is a useful graft for coronary artery bypass grafting. Skeletonization, a technique that uses an ultrasonic scalpel, is increasingly used. However, the cost of an ultrasonic scalpel is extremely high. The purpose of this study was to determine whether a new electrosurgical cautery device (ForceTriad™) is as effective as an ultrasonic scalpel. METHODS: Bilateral ITAs were harvested from eight pigs using the skeletonizing technique. The ITA on one side was harvested with an ultrasonic scalpel and on the other side using the ForceTriad™. Macroscopic and histological examinations were performed in sixteen ITAs. RESULTS: No significant differences in the time required for harvesting were observed. The macroscopic findings revealed no significant change in any of the samples. The histological findings showed that the degree of thermal injury was similar. The normal structure was maintained in all samples. The ForceTriad™ costs US$ 226.82 less per patient than the ultrasonic scalpel. CONCLUSION: The new electrosurgical cautery device ForceTriad™ was less expensive, but it was equally effective. It appears that skeletonization performed with the new device is equivalent to that performed with an ultrasonic scalpel.


Subject(s)
Electrocoagulation/instrumentation , Electrosurgery/instrumentation , Mammary Arteries/surgery , Tissue and Organ Harvesting/instrumentation , Ultrasonic Surgical Procedures/instrumentation , Animals , Cost-Benefit Analysis , Electrocoagulation/adverse effects , Electrocoagulation/economics , Electrosurgery/adverse effects , Electrosurgery/economics , Equipment Design , Mammary Arteries/pathology , Swine , Time Factors , Tissue and Organ Harvesting/adverse effects , Tissue and Organ Harvesting/economics , Ultrasonic Surgical Procedures/adverse effects , Ultrasonic Surgical Procedures/economics
7.
Transplant Proc ; 41(1): 346-9, 2009.
Article in English | MEDLINE | ID: mdl-19249553

ABSTRACT

BACKGROUND: It is important to determine the efficacy of intraportal (IP) islet transplantation in comparison with other transplant sites. In this study, we sought to determine the optimal number of islets to achieve normoglycemia following transplantation into the liver versus the kidney using a mouse model. METHODS: Streptozotocin-induced diabetic mice (Balb/C) were transplanted with syngeneic islets via the IP versus renal subcapsular (SC) routes. The transplanted islet numbers were 0 to 800 (n = 3-5). We assessed the correlation between parameters and islet numbers, comparing IP versus SC groups. The parameters were: (1) percentage of normoglycemia; (2) postoperative days to normoglycemia; (3) mean blood glucose levels at various points from pretransplantation to the end of the study (postoperative day 28); (4) mean serum insulin; and (5) area under the curve of blood glucose levels after glucose injection. RESULTS: Two hundred islets yielded normoglycemia in renal subcapsular grafts, while 800 islets were the minimum required for normoglycemia with IP transplantation. The transplant efficacy in SC transplantation was 2 to 5 times greater than that of IP transplantation. The days to normoglycemia were significantly different between IP versus renal SC islets (13.25 +/- 4.38 days vs 4.50 +/- 0.81 days; P = .007). CONCLUSION: The efficacy of islet transplantation in murine diabetic models was significantly greater under the kidney capsule. Clinical islet transplantation could benefit from trials of alternative transplant sites.


Subject(s)
Diabetes Mellitus, Experimental/surgery , Islets of Langerhans Transplantation/methods , Animals , Blood Glucose/metabolism , Diabetes Mellitus, Experimental/blood , Female , Kidney , Mice , Mice, Inbred BALB C , Portal System , Postoperative Period , Transplantation, Isogeneic
8.
J Dent Res ; 98(9): 1011-1019, 2019 08.
Article in English | MEDLINE | ID: mdl-31329042

ABSTRACT

Ral small GTPases, consisting of RalA and RalB, are members of the Ras family. Their activity is upregulated by RalGEFs. Since several RalGEFs are downstream effectors of Ras, Ral is activated by the oncogenic mutant Ras. Ral is negatively regulated by RalGAP complexes that consist of a catalytic α1 or α2 subunit and its common partner ß subunit and similarly regulate the activity of RalA as well as RalB in vitro. Ral plays an important role in the formation and progression of pancreatic and lung cancers. However, the involvement of Ral in oral squamous cell carcinoma (OSCC) is unclear. In this study, we investigated OSCC by focusing on Ral. OSCC cell lines with high Ral activation exhibited higher motility. We showed that knockdown of RalGAPß increased the activation level of RalA and promoted the migration and invasion of HSC-2 OSCC cells in vitro. In contrast, overexpression of wild-type RalGAPα2 in TSU OSCC cells attenuated the activation level of RalA and inhibited cell migration and invasion. Real-time quantitative polymerase chain reaction analysis of samples from patients with OSCC showed that RalGAPα2 was downregulated in oral cancer tissues as compared with normal epithelia. Among patients with OSCC, those with a lower expression of RalGAPα2 showed a worse overall survival rate. A comparison of DNA methylation and histone modifications of the RalGAPα2 gene in OSCC cell lines suggested that crosstalk among DNA methylation, histone H4Ac, and H3K27me2 was involved in the downregulation of RalGAPα2. Thus, activation of Ral GTPase by downregulation of RalGAP expression via a potential epigenetic mechanism may enhance OSCC progression.


Subject(s)
Carcinoma, Squamous Cell/genetics , GTPase-Activating Proteins/genetics , Mouth Neoplasms/genetics , ral GTP-Binding Proteins/genetics , Cell Line, Tumor , DNA Methylation , Disease Progression , Down-Regulation , Epigenesis, Genetic , Gene Knockdown Techniques , Histones , Humans
10.
Ophthalmology ; 114(12): 2138-41, 2007 Dec.
Article in English | MEDLINE | ID: mdl-18054632

ABSTRACT

PURPOSE: To evaluate the changes in regular and irregular corneal astigmatism after 25-gauge transconjunctival sutureless vitrectomy and 20-gauge standard vitrectomy. DESIGN: Prospective observational comparative case series. PARTICIPANTS: Thirty-two eyes of 32 patients undergoing 25-gauge transconjunctival sutureless vitrectomy and 25 eyes of 24 patients undergoing 20-gauge standard vitrectomy. METHODS: Corneal topography was obtained preoperatively and at 2 weeks and 1 month postoperatively. MAIN OUTCOME MEASURES: The dioptric data of the central 3-mm zone of the cornea were decomposed using Fourier harmonic analysis into spherical power, regular astigmatism, asymmetry, and higher-order irregularity. RESULTS: None of the 4 Fourier indices changed throughout the observation period in the 25-gauge group. In the 20-gauge group, regular astigmatism, asymmetry, and higher-order irregularity were increased significantly at 2 weeks after vitrectomy (P<0.05, Wilcoxon signed-ranks test) and returned to preoperative levels by 1 month. The spherical power in the 20-gauge group did not change after surgery. For regular astigmatism, asymmetry, and higher-order irregularity, the 20-gauge group showed significantly greater surgically induced changes than the 25-gauge group (P<0.05, Mann-Whitney U test). CONCLUSIONS: Twenty-five-gauge transconjunctival sutureless vitrectomy does not induce significant changes in corneal topography and exerts little influence on the optical quality of the cornea.


Subject(s)
Astigmatism/etiology , Cornea/pathology , Corneal Topography , Minimally Invasive Surgical Procedures/adverse effects , Suture Techniques/adverse effects , Vitrectomy/adverse effects , Astigmatism/diagnosis , Conjunctiva , Female , Fourier Analysis , Humans , Male , Middle Aged , Prospective Studies , Retinal Diseases/surgery
11.
Water Sci Technol ; 55(3): 167-75, 2007.
Article in English | MEDLINE | ID: mdl-17410853

ABSTRACT

Lake Biwa is the largest lake in Japan, and water quality in the lake is heterogeneous. Therefore, it is important for water quality management that spatial distribution of water quality in the lake should be clearly understood. The objectives of this study are to show a methodology and to develop a simulation system to calculate COD distribution in Lake Biwa taking internal COD production into consideration. This study also aims to examine transition of COD in the lake using the simulation system. In the simulation system, runoff loads of COD from the Lake Biwa basin are calculated by Macro Model for each tributary. The external COD concentration in 233 inshore meshes of the Lake Biwa water surface was calculated using the runoff loads. The internal COD was calculated using relationships among limiting nutrients, chlorophyll-a and COD. Then, the spatial distribution of water quality in Lake Biwa was calculated both for the external and internal COD by spline technique. Simulations using the system were implemented for 1986-1998, and a clear difference in characteristics between a drought year and a flood year was shown. In the result, it was shown that the simulation system developed here was available to calculate COD distribution in Lake Biwa, and that it had the possibility to explain the recent phenomenon of COD increase in the lake.


Subject(s)
Environmental Monitoring/methods , Fresh Water/analysis , Water Pollution/analysis , Geographic Information Systems , Geography , Japan , Models, Theoretical , Water Movements
12.
Bone Marrow Transplant ; 37(6): 601-5, 2006 Mar.
Article in English | MEDLINE | ID: mdl-16444280

ABSTRACT

Eight children underwent reduced-intensity stem cell transplantation (RIST) from an HLA-matched sibling. They received a fludarabine-melphalan based preparative regimen. Stem cell source was bone marrow, and GVHD prophylaxis consisted of cyclosporine A alone. Acute GVHD grade II-IV and grade III-IV were observed in four (50%) and three (37.5%), respectively, out of these eight patients. This incidence was significantly higher than that after conventional bone marrow transplantation, without severe tissue damage, in the same setting of stem cell source and GVHD prophylaxis. Although the number of patients is small, our results suggest that incidence of acute GVHD after RIST for children is significant. It should be remembered that RIST for children does not seem to be an easy transplant procedure from the viewpoint of acute GVHD, although RIST is less toxic.


Subject(s)
Bone Marrow Transplantation/methods , Graft vs Host Disease/epidemiology , Histocompatibility Testing , Leukemia/therapy , Lymphoma/therapy , Neoplasms/therapy , Stem Cell Transplantation/methods , Adolescent , Cell Count , Child , Child, Preschool , Graft vs Host Disease/physiopathology , Humans , Incidence , Infant , Japan , Male , Retrospective Studies
13.
Waste Manag ; 26(7): 752-7, 2006.
Article in English | MEDLINE | ID: mdl-16650753

ABSTRACT

Nuclear waste repositories need highly durable cementitious materials to function for over thousands of years while resisting leaching and degradation. The durability of cementitious material can be effectively improved by reducing permeability and by changing cement hydrates to a less soluble matrix. This paper describes the properties of carbonated new cementitious materials containing belite-rich cement and gamma-2CaO.SiO2 as main components. In addition, the long-term leaching properties are investigated and compared with ordinary Portland cement by using a predictive leaching model.


Subject(s)
Carbon/chemistry , Construction Materials , Minerals/chemistry
14.
Water Sci Technol ; 53(2): 23-32, 2006.
Article in English | MEDLINE | ID: mdl-16594320

ABSTRACT

Many strategies for water quality conservation in Lake Biwa are being carried out mainly by reducing runoff pollutant loads into the lake. But influence of the runoff load reduction on the water quality in Lake Biwa has not been clarified enough so far. This study is aimed at discussing methodology to estimate water quality distribution in Lake Biwa using runoff pollutant loads from its basin. The runoff loads from the basin are calculated by Macro Model with GIS database of the Lake Biwa basin, and the water quality distribution in the lake is estimated by the spline technique with the calculated runoff loads. As a result, it has been proved that the methodology has enough reproducibility to estimate the water quality distribution in Lake Biwa and is available to examine the water quality in the lake.


Subject(s)
Water Pollutants, Chemical , Conservation of Natural Resources , Environment , Environmental Monitoring , Hazardous Substances , Japan , Rain , Rivers , Waste Disposal, Fluid , Water , Water Pollutants , Water Supply
15.
Biochim Biophys Acta ; 1437(1): 71-9, 1999 Jan 29.
Article in English | MEDLINE | ID: mdl-9931444

ABSTRACT

Apolipoprotein B100 (apoB) is a large secretory protein that forms very low density lipoprotein in liver. An in vitro degradation assay was developed using rabbit reticulocyte (RR) lysate in order to investigate the mechanism of intracellular degradation of newly synthesized apoB by the ubiquitin-proteasome pathway. [3H]apoB, isolated from [3H]leucine pulsed/chased Hep G2 cells, was degraded 51% when incubated for 2 h at 37 degreesC in an assay mixture that included RR lysate (source of the ubiquitin conjugation system and proteasome) and an exogenous ATP regenerating system. ApoB degradation was ATP-dependent and degradation fragments were not observed suggesting that the very large apoB molecule was extensively degraded. ApoB degradation was decreased to 50% when potent proteasome inhibitors, clasto-lactacystin beta-lactone (10 microM) or MG-132 (50 microM), were added to the reaction mixture, but was not affected by the cysteine protease inhibitor, E-64, or the serine protease inhibitor, phenylmethylsulfonyl fluoride. ApoB degradation was inhibited by the mutant ubiquitin protein K48R and by ubiquitin aldehyde, an inhibitor of ubiquitin-protein isopeptidases. During incubation ubiquitination of apoB increased even as apoB was being degraded. These results suggest that in vitro degradation of apoB, a large secretory protein that is normally found in the endoplasmic reticulum (ER) lumen or associated with the ER membrane, was proteasome-dependent and involved both ubiquitination and deubiquitination steps.


Subject(s)
Apolipoproteins B/metabolism , Peptide Hydrolases/metabolism , Reticulocytes/metabolism , Ubiquitins/metabolism , Adenosine Triphosphate/metabolism , Animals , Apolipoprotein B-100 , Apolipoproteins B/chemistry , Apolipoproteins B/isolation & purification , Cell Line , Immunoblotting , Protease Inhibitors/pharmacology , Rabbits , Time Factors , Tritium , Ubiquitins/analysis
16.
Biochim Biophys Acta ; 1067(2): 159-65, 1991 Aug 26.
Article in English | MEDLINE | ID: mdl-1878370

ABSTRACT

To examine the effects of lipid peroxidation on the protein conformation in the porcine intestinal brush-border membranes, a fluorogenic thiol reagent, N-(1-pyrene)maleimide (NPM) was employed. By treatment of NPM-labeled membranes with 100 microM ascorbic acid/10 microM Fe2+ in the presence of various concentrations of tert-butyl hydroperoxide (t-BuOOH), the fluorescence intensity of the complex decreased with the formation of conjugated diene, depending on the hydroperoxide concentration. The temperature dependence profile of the fluorescence intensity of NPM-labeled control membranes showed a thermal transition of the NPM fluorescence at 27-28 degrees C. The transition phenomenon of the NPM fluorescence in the membranes around this temperature disappeared by treatment of the labeled membranes with 100 microM ascorbic acid/10 microM Fe2+/0.6 mM t-BuOOH. The difference in response of the fluorescence characteristics of the bound NPM for temperature variation between the control and peroxidized membranes was also observed in the quenching efficiency with acrylamide. Measurement of the fluorescence polarization revealed that the harmonic mean of the rotational relaxation times of the bound NPM molecules to the membrane proteins increased from 1.96 to 4.93 microseconds by lipid peroxidation of the membranes. This indicates that the movement of the region containing NPM-labeled SH groups in the membrane proteins is restricted by lipid peroxidation. Treatment of NPM-labeled peroxidized membranes with sodium dodecyl sulfate (SDS) resulted in a restoration of the intensity of the NPM fluorescence to the level of the control ones. In addition, the temperature dependence profile of the fluorescence intensity of NPM-labeled peroxidized membranes in the presence of SDS also showed an appearance of a transition phenomenon around 30 degrees C. The result of SDS-polyacrylamide gel electrophoresis of the peroxidized membranes revealed that high-molecular-weight aggregates of the membrane proteins were not formed by lipid peroxidation. On the basis of these results, changes in the environmental properties around NPM-labeled SH groups in the membrane proteins by lipid peroxidation are discussed.


Subject(s)
Intestines/drug effects , Maleimides/toxicity , Microvilli/drug effects , Animals , Electrophoresis, Polyacrylamide Gel , Fluorescence Polarization , Fluorescent Dyes , Lipid Peroxidation , Membrane Lipids/metabolism , Membrane Proteins/metabolism , Swine , Temperature
17.
Biochim Biophys Acta ; 1042(2): 210-6, 1990 Feb 06.
Article in English | MEDLINE | ID: mdl-2302423

ABSTRACT

The uptake of LDL and acetylated LDL and the ability of cholesteryl ester accumulation by cells of a human monocytic cell line, U937, has been characterized by flow cytometric assay using a fluorescent probe, DiI, and by high-performance liquid chromatography (HPLC). The increase of mean fluorescence intensity of U937 incubated with DiI-labeled lipoproteins demonstrates that this cell line could incorporate DiI-AcLDL, as well as DiI-labeled LDL. Competition and saturation studies indicate that the manner of taking up DiI-AcLDL is receptor-mediated. While differentiated U937 incubated with 16 nM phorbol myristate acetate for 24 h took up little DiI-AcLDL, HPLC analysis confirmed that intracellular free and esterified cholesterols significantly increase in the U937 cells incubated with AcLDL or LDL. The ability of mouse peritoneal macrophage to abundantly accumulate at least five kinds of cholesteryl ester were also shown in this analysis. In contrast, in U937 cells, free fatty acids are incorporated into various substances rather than into cholesteryl esters (as revealed by HPLC analysis), so that the cholesterol in AcLDL taken up by U937 cells is not synthesized into cholesteryl esters to any great extent.


Subject(s)
Cholesterol Esters/metabolism , Lipoproteins, LDL/metabolism , Monocytes/metabolism , Acetylation , Animals , Cell Line , Cholesterol/metabolism , Chromatography, High Pressure Liquid , Flow Cytometry , Humans , Mice , Mice, Inbred ICR , Peritoneal Cavity/cytology
18.
Biochim Biophys Acta ; 1042(3): 352-8, 1990 Feb 23.
Article in English | MEDLINE | ID: mdl-2306486

ABSTRACT

Dormant lymphocytes are known to show little LDL receptor (LDL-R) activities. The present study was designed to determine whether or not LDL-R activities of lymphocytes from normal subjects were high enough to be measured by flow cytometry after the cells had been stimulated with recombinant interleukin-2 (IL-2) and anti-CD3 monoclonal antibody (mAb). IL-2 or anti-CD3 mAb individually provokes proliferation of lymphocytes in a serum-free medium. Proliferation rate was accelerated when the two reagents were used in combination. Stimulated cells cultured for 5 days expressed more than 85% CD3 positive, less than 0.5% CD14 positive, and less than 1.5% CD20 positive. The LDL-R activities of the cells were examined by the uptake of a fluorescence probe, DiI-labeled LDL (DiI-LDL) and analyzed by flow cytometry. Stimulated cells showed increased uptake of DiI-LDL and 84 +/- 9% were positive, whereas only 3.0 +/- 2.5% of the cells without stimulation were positive (P less than 0.001). Under the same conditions stimulated lymphocytes from a homozygous familial hypercholesterolemia (FH) patient showed little LDL-R activities; 14% of the cells were positive. Displacement assays reveal that the uptake of LDL by these cells is occurring by way of its specific pathway. These data imply the lymphocytes stimulated with the reagents used in the study might be used for detecting defects in LDL-R, perhaps defects in other genomic systems as well.


Subject(s)
Antibodies, Monoclonal/pharmacology , Interleukin-2/pharmacology , Lymphocytes/metabolism , Receptors, LDL/metabolism , Cell Division/drug effects , Cells, Cultured , Female , Flow Cytometry , Fluorescent Dyes , Humans , Hyperlipoproteinemia Type II/blood , Lymphocyte Activation/drug effects , Lymphocytes/drug effects , Male , Receptors, LDL/drug effects , Recombinant Proteins/pharmacology , Time Factors
19.
Biochim Biophys Acta ; 1550(2): 164-74, 2001 Dec 17.
Article in English | MEDLINE | ID: mdl-11755205

ABSTRACT

The AT motif-binding factor 1 (ATBF1)-A is a large transcription factor containing four homeodomains and 23 zinc finger motifs. It has a number of motifs involved in transcriptional regulation, and in addition, several motifs found in enzymes, such as ATPases and helicases. In this study, we examined whether ATPase activity is associated with the ATBF1-A molecule. A 263-amino acid segment of the ATBF1-A molecule, termed AHZ, which contains the ATPase A-motif, homeodomain IV and zinc finger 21, was expressed in Escherichia coli in the form of glutathione S-transferase fusion protein and analyzed for ATPase activity. We found that AHZ was able to hydrolyze ATP with K(m) 10.6 microM and K(cat) 0.055 min(-1) at 5 mM Mg(2+) and pH 7.75. AHZ retained bacterial DNA and removal of the DNA resulted in 70% decrease in ATPase activity. The addition of double- or single-stranded DNAs restored 70-75% ATPase activity and that of RNA restored 50-55% activity. Site-directed mutagenesis of the A-motif resulted in 34% reduction of ATPase activity with no significant loss of bound DNA. In contrast, mutation of homeodomain IV and zinc finger 21 resulted in 90 and 80% reduction of ATPase, respectively, with the loss of the ability to bind to DNA and RNA. These results show that ATBF1 has at least one enzyme activity in addition to regulation of DNA transcription. The ATPase activity associated with ATBF1-A is DNA/RNA-dependent and unique in that it requires both homeodomain and zinc finger motifs.


Subject(s)
Adenosine Triphosphatases/metabolism , Homeodomain Proteins/chemistry , Zinc Fingers , Adenosine Triphosphatases/chemistry , Adenosine Triphosphatases/genetics , Adenosine Triphosphate/chemistry , Amino Acid Sequence , Base Sequence , DNA/pharmacology , DNA, Bacterial/chemistry , Escherichia coli/genetics , Escherichia coli/metabolism , Homeodomain Proteins/genetics , Molecular Sequence Data , Mutagenesis, Site-Directed , Peptide Fragments/chemistry , Peptide Fragments/genetics , Peptide Fragments/metabolism , Plasmids , RNA/pharmacology , Recombinant Fusion Proteins/metabolism , Substrate Specificity
20.
Diabetes ; 45(8): 1037-43, 1996 Aug.
Article in English | MEDLINE | ID: mdl-8690149

ABSTRACT

Recent immunohistological studies using antibodies against advanced glycation end products (AGEs) have demonstrated the presence of AGEs in several tissues. By an enzyme-linked immunosorbent assay using the monoclonal anti-AGE antibody, the present study aimed to determine AGEs in pepsin-insoluble collagen (PIC) as well as in pepsin-soluble collagen (PSC) from the aortas of streptozotocin (STZ)-induced diabetic rats (at 4, 16, and 28 weeks after STZ injection) and those of age-matched control rats. Addition of EDTA to the immunoassay buffer has led us to successful determination of AGEs in the aortic PIC samples with following results: 1) in diabetic rats, there was a time-related increase in the AGE contents at 28 weeks (n = 9, 226.4 +/- 13.5 ng/mg collagen [mean +/- SE]), compared with that at 4 and 16 weeks (n = 6, 79.6 +/- 9.5 ng/mg collagen, and n = 8, 149.4 +/- 30.9 ng/mg collagen at 4 and 16 weeks, respectively; both P < 0.05, between 4 and 16 weeks and 28 weeks); 2) after 28 weeks of diabetes, the AGE contents in PIC of aortas were significantly higher in diabetic rats than in controls (n = 9, 226.4 +/- 13.5 ng/mg collagen vs. n = 8, 129.6 +/- 14.9 ng/mg collagen, P < 0.01, diabetic vs. control); and 3) the level of the AGE content was strongly correlated with the PIC/total collagen (TC) ratio (n = 45, r = 0.698, P = 0.0001). By treating the samples of PSC with alkaline solution, the AGE content of PSC was also determined. In the PSC fraction, the AGE levels in the diabetic rats tended to increase with time and to be higher than those of control rats at 28 weeks although these changes were not statistically significant (diabetic: n = 4, 19.4 +/- 9.7; n = 6, 22.3 +/- 6.2; n = 6, 39.6 +/- 10.8; control: n = 4, 19.7 +/- 9.8; n = 6, 22.9 +/- 7.3; n = 7, 30.7 +/- 7.2; at 4, 16, and 28 weeks, respectively). Compared with the AGE levels of PSC, those of PIC were about four to seven times and four to five times higher in diabetic and control rats, respectively (PIC versus PSC in diabetic or control rats, all P < 0.001, at 4, 16, and 28 weeks, respectively). These findings provide the first immunochemical evidence that AGE adducts are present in the materials extracted sequentially by pepsin and collagenase and that these adducts in PIC accumulated as a function of the increase in the aortic PIC/TC ratio.


Subject(s)
Collagen/metabolism , Diabetes Mellitus, Experimental/metabolism , Glycation End Products, Advanced/metabolism , Animals , Aorta/chemistry , Collagen/chemistry , Enzyme-Linked Immunosorbent Assay , Fructosamine , Hexosamines/metabolism , Male , Pepsin A/metabolism , Rats , Rats, Sprague-Dawley , Solubility
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