LDL particle size by gradient-gel electrophoresis cannot be estimated by LDL-cholesterol/apolipoprotein B ratios.
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Apolipoproteins B↗
Biomedical subjects
D Kobayashi
Publications and source records attributed to D Kobayashi.
A sensitive assay of tumor necrosis factor alpha in sera from Duchenne muscular dystrophy patients.
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Adolescent↗
Screening of cationic compounds as an absorption enhancer for nasal drug delivery.
Several cationic compounds were screened as potential nasal absorption enhancers to increase intranasal absorption of a model drug, fluorescein isothiocyanate labeled dextran (MW 4.4 kDa, FD-4), without nasal membrane damage in rats. Their effects were compared with those of classical enhancers. Various cationic compounds (poly-L-arginines with different molecular weights (MW 8.9, 45.5 and 92.0 kDa, poly-L-Arg (10), (50) and (100), respectively), L-arginine (L-Arg), L-lysine (L-Lys), and cetylpyridinium chloride (CPCL) were evaluated. Of the cationic compounds, poly-L-Arg and CPCL greatly enhanced the intranasal absorption of FD-4, as did chitosan, a cationic polysaccharide which has been reported to show a great effect on the transnasal delivery of peptide and protein drugs. The enhancing intensity by poly-L-Arg was dependent on its molecular weight. Rank order of the enhancing ratio, calculated from the AUC ratio for the enhancer treatment against the untreatment, was 0.5% poly-L-Arg (100) congruent with0.5% sodium dodecylsulfate congruent with0.5% CPCL?0.5% poly-L-Arg (50)?0.5% sodium deoxycholate congruent with0.5% sodium taurodihydrofusidate?0.5% polyoxyethylene-9-lauryl ether congruent with0.5% lysophosphatidylcholine?0.5% chitosan congruent with0.5% poly-L-Arg (10)>/=10% L-Arg congruent with10% L-Lys?0.5% sodium glycocholate congruent with0.5% sodium taurocholate congruent with0.5% EDTA. Only the poly-L-Args represented almost the same degree of hemolysis of cationic compounds compared with pH 7.0 phosphate buffered saline in the rat erythrocyte lysis experiment. The enhancing ratio by classical enhancers correlated with leaching of protein, phospholipids and LDH from isolated rabbit nasal mucosa. CPCL also fell on the regression lines between the enhancing ratio and their degree of leaching from classical enhancers. In contrast, the enhancing intensities by poly-L-Arg (10), (50) and (100) were greatly shifted from the regression line: the amount of leaching was markedly low in spite of a great enhancement of FD-4 absorption. These findings suggest that of the assessed enhancers only the poly-L-Args enhance the transnasal delivery of high molecular substances without severe damage to the nasal mucosal membrane. Poly-L-Arg is therefore a promising candidate having a good balance between enhancing activity and safety for nasal peptide and protein delivery.
Absorption↗
Medaka (Oryzias latipes) FTZ-F1 potentially regulates the transcription of P-450 aromatase in ovarian follicles: cDNA cloning and functional characterization.
Our previous findings suggest the activity of cytochrome P-450 aromatase (P-450arom), the enzyme which converts testosterone to estradiol-17beta, in the ovarian follicle of medaka (Oryzias latipes) is regulated at the transcriptional level. In this study, we cloned a cDNA encoding a FTZ-F1-like protein (mdFtz-F1) from ovarian follicles of medaka. In vitro translated mdFTZ-F1, and nuclear extract from medaka ovarian follicles, formed complexes with oligonucleotide probes containing putative orphan nuclear receptor binding motifs, which are present in the promoter region of the medaka P-450arom gene. The expression pattern of mdFtz-F1 transcripts during oogenesis coincides with that of P-450arom transcripts. Transfection assays further suggest a potential transcriptional regulatory activity of mdFTZ-F1 upon the medaka P-450arom promoter. Taken together, these results suggest a potential role of mdFTZ-F1 in the transcriptional regulation of P-450arom in the ovarian follicle of medaka.
Amino Acid Sequence↗
The hsp operons are repressed by the hrc37 of the hsp70 operon in Staphylococcus aureus.
The heat-shock proteins are coded for the polycistronic operons hsp70 and hsp60 in Staphylococcus aureus. The hsp70 operon is comprised of five genes, hrc37, hsp20, hsp70, hsp40 and orf35, and the hsp60 is comprised of two genes, hsp10 and hsp60. The hsp70 operon transcribed five different sizes of mRNA from three promoters: P1, the most active promoter, transcribed 6.0 and 3.6 kb mRNAs; P2 transcribed a single 1.8 kb mRNA; and P3 transcribed 4.2 and 2.4 kb mRNAs. The hsp60 operon transcribed a single 2.1 kb transcript from only one promoter, P1. Both operons had a common structure of inverted repeat element (CIRCE, Controlling Inverted Repeat of Chaperon Expression) at the promoter region. All of the transcripts were heat (46 C) inducible. One of the unidentified genes, hrc37, was characterized. The disruptant of the hrc37 in the hsp70 operon enhanced the transcription of both operons at 37 C (derepression). Complementation of the disruptant with the cloned hrc37 plasmid recovered the repression of the transcription of both operons at 37 C. The product of hrc37, Hrc37, was found to bind to the CIRCE element. These findings indicated that Hrc37 from the hsp70 operon repressed the transcription of both the hsp70 and hsp60 operons by binding to the CIRCE element located at the promoter region.
Base Sequence↗
Possibility for discriminating between two representative non two-state thermal unfolding models of proteins by DSC.
Possible differences between two representative non two-state thermal unfolding mechanisms of protein are discussed concerning differential scanning calorimetry. Numerical simulations showed that, by DSC measurement, it is hard to discriminate between the independent model, which assumes independent unfolding domains in a protein, and the sequential model, which assumes intermediate(s) between native and denatured states, especially when values of molecular weight, denaturation enthalpy, and difference in denaturation temperature of each denaturation process are large. DSC curve analysis of Aspergillus niger glucoamylase based on these two models gave essentially the same thermodynamic parameters.
Aspergillus niger↗
Systemic lupus erythematosus with a giant rectal ulcer and perforation.
A 41-year-old man with systemic lupus erythematosus (SLE) who developed pelvic inflammation due to perforation of a giant rectal ulcer is described. The patient presented with persistent diarrhea, abdominal pain and fever without development of disease activity of SLE. Endoscopic and radiological examinations revealed a perforated giant ulcer on the posterior wall at the rectum below the peritoneal evagination. The ulcerated area was decreased after a colostomy was performed at the transverse colon to preserve anal function. The patient is currently being monitored on an outpatient basis. It should be noted that life-threatening complications such as perforated ulcer of the intestinal tract could occur without SLE disease activity.
Adult↗
Antibody mimicking an anti-E antibody that binds to patient's E negative red cells.
We have reported a rare case of auto anti-E antibody with specificity mimicking alloantibody with E specificity. A patient whose red cells typed as R1R1 and who had a positive direct antiglobulin test was admitted to our hospital. After standard serologic testing was performed, flow cytometry, Western blot analysis and differential allogenic adsorption test were used to verify whether antibody binds to the patient's red cells and normal red cells. A high titer anti-E antibody was detected transiently from the patient's serum and eluate. An indirect antiglobulin test using red cells treated by cysteine-activated papain and dithiothreitol (ZZAP) and chloroquine showed that both the patient's serum and eluate bound an apparent anti-E antibody to E negative patient's red cells. Fluorescence activated cells sorter (FACS) and Western blot analysis verified that the patient's red cells lacked E antigen. Further, it was clarified that the antibody does not bind to any E negative normal red cells by differential allogenic adsorption test. These results provide evidence that an antibody mimicking an E alloantibody can bind to patient's own E negative red cells but not to allogenic E negative red cells.
Antibodies, Anti-Idiotypic↗
[The control survey in CD marker analysis of leukemic cells].
Analysis of cell surface antigens is thought to be more objective compared to classification by the morphology. However, it has been indicated that the evaluated data are different each other between institutes. Therefore, we performed a control survey of CD marker analysis in leukemia cells using flow cytometry in six commercial laboratories. The expression of CD2, 4, 5, 7, 8, 10, 13, 14, 19, 20, 33, 34, 38, and 71, HLA-DR in tumor cells from megakaryoblastic leukemia, lymphoblastic crisis of CML, and ALL patients, were examined. There were large differences in the results of CD marker analysis among the six commercial laboratories. One possible reason for this result is the differences in gating condition and monoclonal antibodies used in each laboratory. However, there were the differences in the results even when the same gating condition and antibodies were used. The reasons for the difference in transport or treatment conditions for samples remain a matter of considerable controversy. In the present condition, it was suggested that the exchange of information about the origin of tumor cells between the physician in charge and examination experts is thought to be the best way to analyze proper cell surface antigen expression. Further, no national survey has been carried out, but the establishment of a standard assay is preferred to obtain proper data for the patients and medical parties concerned.
Adolescent↗
Detection of human serum tumor necrosis factor-alpha in healthy donors, using a highly sensitive immuno-PCR assay.
BACKGROUND: Tumor necrosis factor-alpha (TNFalpha) is an important mediator of inflammatory and autoimmune diseases. Analysis of its pathophysiologic roles has been difficult because low concentrations of TNFalpha, including those in healthy controls, cannot be measured by existing methods. METHODS: We developed a sensitive immuno-PCR assay for the detection of TNFalpha in human serum. The DNA label was generated by PCR amplification using biotinylated primer and was bound with streptavidin to the biotinylated third antibody. TNFalpha sandwiched by antibodies was detected by amplification of the DNA label using PCR. RESULTS: The limit of detection of the assay was 0. 001 ng/L, an approximately 5 x 10(4)-fold improvement compared with a conventional ELISA. The mean serum TNFalpha concentration (+/- SD) in healthy donors was 0.021 +/- 0.044 ng/L in men (n = 29) and 0.033 +/- 0.065 ng/L in women (n = 25). CONCLUSION: This method may be useful for analyzing the significance of TNFalpha concentration in various diseases.
Adult↗
Differentiation induction by a tumor-necrosis-factor mutant 471 in human myelogenous leukemic cells via tumor-necrosis-factor receptor-p55.
The present study examined differentiation-inducing activity by various tumor-necrosis-factor(TNF) mutants against the human leukemic cell lines HL-60 and U-937. Mutant TNF 471, from which 7 N-terminal amino acids of native TNF were deleted and Pro8, Ser9 and Asp10 were replaced by Arg, Lys and Arg, possessed the highest activity among the TNF mutants, and its activity was 120-fold that of native TNF. The various biological activities of TNF were signaled through 2 distinct receptors, p55 and p75. Although cytotoxicity was reported to involve mainly p55, this differentiation-inducing activity was not well understood. The fact that the affinity of TNF 471 was higher to p55 and lower to p75 than that of native TNF by a binding competition assay suggested that the differentiation-inducing activity was also signaled through p55. To verify this hypothesis, the human myelogenous leukemic cell line, KG-1, which scarcely expresses either receptor and does not differentiate with TNF, was transduced with the p55 or p75 gene. Subsequently p55 transfectants manifested a greater ability to differentiate; however, p75 transfectants did not differ from parental cells or from mock-transfectants. Further, the differentiation of p55 transfectants induced by TNF was reduced by the inhibitor of protein-kinase-C (PKC), staurosporine. These results indicate that the differentiation-inducing activity was signaled through the TNF receptor, p55, via PKC and that the excellent ability of TNF 471 to induce differentiation was related to its high affinity for p55.
Antigens, CD↗
Suppression of intracellular resistance factors by adriamycin augments heat-induced apoptosis via interleukin-1beta-converting enzyme activation in pancreatic carcinoma cells.
Combination of heat and various anticancer drugs can exert a synergistic antitumor effect in vitro and in vivo, though the mechanism is not clear. We have previously shown that endogenous tumor necrosis factor (enTNF) acts as an intracellular resistance factor to inhibit the cytotoxic effect of heat by scavenging oxygen-free radicals via the induction of manganous superoxide dismutase (MnSOD). Consequently, we examined whether the suppression of these resistance factors by combining anticancer drugs and heat causes an augmentation of heat-induced cytotoxicity. The human pancreatic carcinoma cell line, PANC-1, constitutively expresses appreciable amounts of enTNF and also demonstrates heat resistance. After treatment of these cells for 15 hr with adriamycin (ADM), the expression of enTNF was decreased by 43%, and MnSOD activity was suppressed by 55%. The cytotoxic effects of the treatment of PANC-1 cells with ADM followed by heat were greater than the sum of those observed with the agents administrated individually. Heat-induced apoptosis was also augmented by pretreatment with ADM. Furthermore, the interleukin-1beta-converting enzyme inhibitor, Ac-YVAD-CMK, reversed the augmented cytotoxicity. Our results indicate that suppression of intracellular resistance factors such as enTNF and MnSOD plays an important role in apoptosis seen after heat and ADM combined therapy.
Apoptosis↗
BMP-induced osteogenesis on the surface of hydroxyapatite with geometrically feasible and nonfeasible structures: topology of osteogenesis.
Bone morphogenetic protein (BMP) is known to require a suitable carrier to induce ectopic bone formation in vivo. Hydroxyapatite ceramics have been reported to be effective in some forms but ineffective in others as a carrier of BMP-induced bone formation. In this study we compare three geometrically different forms of hydroxyapatite to examine their functions as carriers of BMP-induced bone formation. A fraction containing all the active BMPs (BMP cocktail) was partially purified from a 4M guanidine extract from bovine bone by a three-step chromatographic procedure. The BMP cocktail was combined with each of three forms of hydroxyapatite--solid particles (SPHAP), porous particles (PPHAP), and coral-replicated porous tablets (coral-HAP)--and implanted subcutaneously into rats. Both the PPHAP and coral-HAP systems induced osteogenesis 2 weeks after implantation, as evidenced by morphological and biochemical observations. Details of the osteogenetic process were followed by double-fluorescence labeling in the coral-HAP system to confirm bone formation on the surface of hydroxyapatite. However, there was no evidence of osteogenesis or chondrogenesis in the SPHAP system. The results indicate that the geometry of the interconnected porous structure in PPHAP and coral-HAP create spaces for vasculature that lead to osteogenesis while the smooth structure and close contact of particles in SPHAP inhibit vascular formation and proliferation of mesenchymal cells, preventing bone and cartilage formation. It was concluded that the geometrical structure in hydroxyapatite ceramics that induces vasculature is crucial as a carrier for BMP-induced bone formation.
Alkaline Phosphatase↗
In vivo characteristics of injectable poly(DL-lactic acid) microspheres for long-acting drug delivery.
Poly(DL-lactic acid) (PLA) microspheres containing testosterone (T) were prepared by the solvent evaporation process to evaluate their physical properties such as size distribution, shape, drug content, in vivo controlled drug release, pharmacological influences on the prostate gland in castrated rats, and histopathological findings of tissues surrounding the implants. The in vivo release of T from PLA microspheres containing 30 mg of drug obtained with chloroform was continued over a 6-week period. This effect is attributed to high dispersibility of T in the device when obtained with chloroform. Both serum drug levels and prostate gland weight recovery suggested the effects of a long-acting drug delivery system. The histopathological findings showed that the devices used were completely degraded 10 weeks after injection.
Animals↗
Preparation and characterization of oil-in-water type poly (D,L-lactic acid) microspheres containing testosterone enanthate.
Poly (D,L-lactic acid) (PLA) microspheres containing testosterone enanthate (ET) were prepared by using an oil-in-water (O/W) emulsion technique. The size distribution of the microspheres obtained could be explained by a log-normal distribution, and as a result, it was found that ET fully incorporates into microspheres even when the drug is loaded at up to 50%. On the other hand, the dissolution behavior of ET from microspheres was strongly dependent on particle size, suggesting that dissolution of the drug from microspheres can be easily controlled by controlling the preparative conditions.
Calorimetry, Differential Scanning↗
[A gastric cancer showing an exclusive metastatic nest at the gastroenemius muscle].
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Adenocarcinoma, Papillary↗
Quantitative reverse transcription-PCR assay of the RNA component of human telomerase using the TaqMan fluorogenic detection system.
We established the validity of a quantitative reverse transcription (RT)-PCR assay for the RNA component of human telomerase (hTR), using the TaqMan fluorogenic detection system. Using this assay, we quantified hTR expression in two human pancreatic cancer cell lines, ASPC-1 and MIAPaCa-2. Our results indicated that hTR expression in MIAPaCa-2 was 1.99-fold higher than that in ASPC-1 cells. This TaqMan RT-PCR assay appears to be useful in determining the amount of hTR in clinical specimens.
Cell Line↗
Avirulence gene D of Pseudomonas syringae pv. tomato may have undergone horizontal gene transfer.
Avirulence gene D (avrD) is carried on the B-plasmid of the plant pathogen Pseudomonas syringae pv. tomato with plasmid-borne avrD homologs widely distributed among the Pseudomonads. We now report sequences in the soft rot pathogen Erwinia carotovora that cross-hybridize to avrD suggesting a conserved function beyond avirulence. Alternatively, avrD may have been transferred horizontally among species: (i) DNA linked to avrD shows evidence of class II transpositions and contains a novel IS3-related insertion sequence, and (ii) short sequences linked to avrD are similar to pathogenicity genes from a variety of unrelated pathogens. We have also identified the gene cluster that controls B-plasmid stability.
Bacterial Proteins↗