Biomedical subjects

M Miyake

Publications and source records attributed to M Miyake.

At least 55 records · Page 3Linked to original sources

PubMed2000

Novel proteinaceous toxins from the box jellyfish (sea wasp) Carybdea rastoni.

During summer and autumn, the box jellyfish (sea wasp) Carybdea rastoni is one of the most bothersome stinging pests to swimmers and bathers on the Japanese coast. Two labile but potent hemolytic toxins from the tentacles of Carybdea rastoni were isolated in their active forms using newly developed purification methods. The molecular masses of the isolated C. rastoni toxin-A and toxin-B (CrTX-A and CrTX-B) are 43 and 46 kDa, respectively, as calculated from SDS-PAGE. In the present study, we sequenced the full-length cDNA (1600 bp), which encodes both CrTX-A and CrTX-B. The deduced 450 amino acid sequence of the CrTXs, showed no significant homology with any known protein. This report presents the first complete sequence of a proteinaceous jellyfish toxin. Furthermore, it was revealed that CrTX-A was primarily localized in the nematocyst, whereas CrTX-B was detected only in the tentacle. Because the nematocyst is the organ responsible for the cnidarian sting, the remainder of the study focused on the toxicity of CrTX-A. We found that CrTX-A was fatally toxic to mice at 20 microg/kg (i.v.) and crayfish at 5 microg/kg (i.p.). Subcutaneously injected CrTX-A (0.1 microg) caused inflammation of mouse skin. These results showed that CrTX-A is responsible for the cutaneous inflammation observed in humans stung by C. rastoni.

Animals↗

PubMed2000

Binding of avian ovomucoid to shiga-like toxin type 1 and its utilization for receptor analog affinity chromatography.

Development of a simple and efficient purification procedure for Shiga-like toxin I (Stx1) was attempted. Since it has been suggested that pigeon egg white ovomucoid carries a P1 antigenic determinant, we examined its ability to bind Stx1. The ovomucoid glycoprotein fraction (GPro) was prepared from pigeon egg white by acetone precipitation, and a portion of the GPro was treated with pronase to obtain the glycopeptide fraction (GPep). When both GPro and GPep were coupled to CNBr-activated Sepharose 4B and subjected to affinity chromatography, Stx1 specifically bound to both columns. The Stx1 eluted with a buffer containing 4.5 M MgCl2 was shown to be highly purified to homogeneity by polyacrylamide gel electrophoresis under denatured condition; only two protein bands with molecular weights of 32,000 and 8000, which correspond to the A and the B subunits of Stx1, respectively, were recognized. The purified toxin showed cytotoxicity on Vero cells with a specific activity of approximately 6 x 10(8) CD50/mg protein; almost 100% of the activity was recovered from Escherichia coli cell lysate. We propose that the utilization of avian ovmucoid for the affinity chromatography provides a potentially simple, convenient, and widely available method to purify Shiga-like toxins.

Animals↗

PubMed2000

Successful implantation of in vitro-matured, electro-activated oocytes in the pig.

In the present study, we derived parthenogenetic porcine fetuses from in vitro-matured oocytes following a simple activation process in order to evaluate their developmental limitations in-vivo. Follicular oocytes collected from gilts at local slaughterhouses were matured for 48 h. They were subjected to a single square pulse of direct current for 100 microsec at 1,500 V/cm and then treated with 5 microg/mL cytochalasin B for 4 h to obtain activated diploid oocytes. The diploids were cultured in modified Whitten's medium until transfer. Diploids which had cleaved to the 2- and 3- to 4-cell stages were transferred to oviducts of recipients. Live and/or dead parthenogenetic fetuses were recovered in 6 of 8 trials at 17, 18, 19, 24 and 29 d post activation. The total proportion of fetuses to transferred diploids was 31.3% (62/198). When fetuses were recovered at 19 d post activation, the proportion of development into fetuses was 71% (15/21). Our results, however, suggest that periods of gestation longer than 19 d resulted in a decrease of these proportions to 45% (18/40) at 24 d and to 18% (7/40) at 29 d. The hearts were beating in nearly all of the fetuses recovered at 19, 24 and 29 d post activation. Thus, parthenogenetic porcine diploids developed to at least the stage of limb-bud formation beyond the early heart-beating stage. Abnormalities were also externally visible on some fetuses. Formation of cyst-like structures in the heart and liver, and insufficient development of the head region and acephali were observed in some cases.

Animals↗

PubMed2000

Heterologous expression of clostridial hydrogenase in the Cyanobacterium synechococcus PCC7942.

The Clostridium pasteurianum hydrogenase I has been expressed in the cyanobacterium Synechococcus PCC7942. The Shine-Dalgarno sequence of the structural gene encoding hydrogenase I from C. pasteurianum was changed to that of the cat (chloramphenicol acetyltransferase) gene. The hydrogenase gene was cloned downstream of a strong promoter, isolated from Synechococcus PCC7942, with the cat gene as a reporter gene. Expression of clostridial hydrogenase was confirmed by Western and Northern blot analyses in Synechococcus and Escherichia coli, whereas in vivo/in vitro measurements and activity staining of soluble proteins separated on non-denaturing polyacrylamide gels revealed functional expression of hydrogenase only in cyanobacterial cells. The changed Shine-Dalgarno sequence appeared to be essential for the functional expression of clostridial hydrogenase in Synechococcus, but had no influence on the expression and activity of clostridial hydrogenase expressed in E. coli.

Blotting, Northern↗

PubMed2000

Biochemical characterization of sialoprotein "anti-agglutinin" purified from boar epididymal and seminal plasma.

Sialoprotein "anti-agglutinin," previously shown to inhibit sperm head-to-head agglutination, is found in both boar epididymal and seminal plasma. The present report characterizes anti-agglutinin by mass spectrometry, by N-terminal amino acid sequence analysis, and by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blotting techniques to assess phosphate content of the molecule. Anti-agglutinin had the SDS-PAGE mobility of approximately 25 kDa. By electrospray ionization-mass spectrometry, however, mass spectra of anti-agglutinin were characterized by two major peaks (19,379-19,382 Da and 19,395-19,397 Da) and several minor peaks. Mass spectrometry of tryptic peptide fragments of deglycosylated anti-agglutinin and amino acid sequence analysis revealed that the protein has a unique peptide-mass fingerprinting of fragments (12,668 Da, 5,209 Da, 1,226 Da, and 1,168 Da) and a novel N-terminal amino acid sequence (KTDDY AISGA KEEEF YDYME ELYAV), respectively. Additionally Western blot techniques, using commercially available monoclonal antibodies, were used to detect presence of phosphothreonine and phosphoserine substituents, but two different monoclonal antibodies did not detect phosphotyrosine. Moreover, treatment with two different alkaline phosphotases converted the molecule, as assessed by SDS-PAGE and detection by silver stain, from the parent form of about 25 kDa to forms of approximately 19 kDa (similar to that assigned by mass spectrometry) and/or 15 kDa. Original antiserum generated toward, and reacting with native anti-agglutinin, reacted only with 19 kDa form. These results are consistent with the conclusion that the native anti-agglutinin may be a novel protein that is phosphorylated at serine and/or threonine residues.

Agglutinins↗

PubMed2000

Citrus limonoids obacunone and limonin inhibit azoxymethane-induced colon carcinogenesis in rats.

Obacunone and limonin are bitter limonoids in citrus. Their modifying effects on the development of aberrant crypt foci (ACF), the activity of detoxification enzymes, glutathione S-transferase (GST) and quinone reductase (QR), and cell proliferation activity were investigated in male F344 rats treated with azoxymethane (AOM). Obacunone and limonin were administered in the diet, during the initiation (for 4 weeks) or postinitiation phase (for 4 weeks) of AOM-induced tumorigenesis. Feeding of obacunone and limonin (0.02% or 0.05%) caused significant reduction (55-65% by "initiation" feeding and 28-42% by "postinitiation" feeding) in the yield of ACF. The ability to reduce the proliferating cell nuclear antigen-labeling index in crypts and correlated well with the prevention of ACF. In a subsequent long-term experiment (38 weeks), in which rats were initiated with AOM and fed 0.05% obacunone or 0.05% limonin during the initiation or post-initiation phase, both compounds in diet caused significant reduction (65%-92% inhibition) in the incidence of colonic adenocarcinoma. Thus, citrus bitter limonoids obacunone and limonin possess chemopreventive effects on chemically induced rat colon carcinogenesis.

Animals↗

PubMed2000

Standard radial bone mineral density and physical factors in ordinary Japanese women.

To prevent osteoporosis, which is expected to increase in incidence in this rapidly aging society, in recent years bone mineral density (BMD) has frequently been measured as a predisposition index. However, these measurements are made on different sites with different apparatus, and the results are independently studied by different institutions. In our present investigation, to establish the standard radius BMD as determined by dual-energy X-ray absorptiometry (DXA), we carried out a general population survey in 29 municipalities and prefectures on 11,252 locally residing females aged 15 to 83 years (mean, 35.61 +/- 12.85 years). Their YAM (young adult mean) BMD was estimated at 0.664 +/- 0.054 g/cm2, which was almost the same as the figure given in the 1996 version of the diagnostic criteria for primary osteoporosis. We further studied the relationships of BMD to age and physical factors known to be influential to BMD. It was found that BMD was correlated negatively to age and positively to body mass index (BMI). The average values we obtained for age and physique groups appeared to have provided reliable indices for the primary prevention of osteoporosis.

Adolescent↗

PubMed2000

A long-term study of implanted artificial hydroxyapatite particles surrounding the carotid artery in adult dogs.

In this long-term study, we implanted HAP into adult dogs using a silicone chamber attached to the carotid artery to clarify tissue reaction to HAP implantation over a long period. We designed chambers and both hemispheres of the chambers were filled with HAP particles, and were placed around both carotid arteries of seven adult dogs. The implants were removed after 150, 300, 380 days, and histological and ultrastructual examination was undertaken. We observed bone-like tissue which was formed where HAP particles were implanted. Immunohistochemical findings showed that osteocalcin and osteonectin were as positive in the bone-like tissue as in normal bone. This study suggests that biological factors from the arterial wall might play an important role in new bone-like tissue forming, and that HAP has a strong osteoconductive ability even at heterogeneous sites.

Animals↗

PubMed2000

In vivo EPR dosimetry of accidental exposures to radiation: experimental results indicating the feasibility of practical use in human subjects.

Low frequency electron paramagnetic resonance (EPR) provides the potential advantage of making accurate and sensitive measurements of absorbed radiation dose in teeth in situ, i.e. without removing the teeth from the potential victim. The potential limiting factors for making such measurements are: (1) whether low frequency EPR is sufficiently sensitive to detect radiation-induced signal in human teeth; (2) whether sufficient sensitivity can be maintained under in vivo conditions. In this manuscript, we summarize results indicating that this approach is feasible. Using 1.2 GHz EPR spectroscopy, we found that the lower limit for these measurements in isolated human teeth is 0.2 Gy or lower. Measurements of radiation-induced EPR signals in the teeth of living rats were achieved with sufficient sensitivity to indicate that, when taking into consideration the larger mass of human teeth, similar measurements in human teeth in situ would provide sensitivity in the dose range for potential accidental exposures. We estimate that the current lower limit for detecting radiation doses in human teeth in situ (in vivo) is 0.5-1.0 Gy; this would be sufficient for determining if a person has been exposed to potentially life threatening doses of ionizing radiation. The limiting factor for sensitivity appears to be background signals rather than signal/noise, and there are feasible means to overcome this problem and further increase sensitivity. The additional instrumental developments required to make an effective in vivo EPR dosimetric spectrometer for the measurements in teeth in human subjects in situ, seem quite achievable.

Animals↗

PubMed2000

A novel natural killer cell line (KHYG-1) from a patient with aggressive natural killer cell leukemia carrying a p53 point mutation.

We present the establishment of a natural killer (NK) leukemia cell line, designated KHYG-1, from the blood of a patient with aggressive NK leukemia, which both possessed the same p53 point mutation. The immunophenotype of the primary leukemia cells was CD2+, surface CD3-, cytoplasmic CD3epsilon+, CD7+, CD8alphaalpha+, CD16+, CD56+, CD57+ and HLA-DR+. A new cell line (KHYG-1) was established by culturing peripheral leukemia cells with 100 units of recombinant interleukin (IL)-2. The KHYG-1 cells showed LGL morphology with a large nucleus, coarse chromatin, conspicuous nucleoli, and abundant basophilic cytoplasm with many azurophilic granules. The immunophenotype of KHYG-1 cells was CD1-, CD2+, surface CD3-, cytoplasmic CD3epsilon+, CD7+, CD8alphaalpha+, CD16-, CD25-, CD33+, CD34-, CD56+, CD57-, CD122+, CD132+, and TdT-. Southern blot analysis of these cells revealed a normal germline configuration for the beta, delta, and gamma chains of the T cell receptor and the immunoglobulin heavy-chain genes. Moreover, the KHYG-1 cells displayed NK cell activity and IL-2-dependent proliferation in vitro, suggesting that they are of NK cell origin. Epstein-Barr virus (EBV) DNA was not detected in KHYG-1 cells by Southern blot analysis with a terminal repeat probe from an EBV genome. A point mutation in exon 7 of the p53 gene was detected in the KHYG-1 cells by PCR/SSCP analysis, and direct sequencing revealed the conversion of C to T at nucleotide 877 in codon 248. The primary leukemia cells also carried the same point mutation. Although the precise role of the p53 point mutation in leukemogenesis remains to be clarified, the establishment of an NK leukemia cell line with a p53 point mutation could be valuable in the study of leukemogenesis.

Antigens, CD↗

PubMed2000

Quantitative trait loci (QTL) analysis in a Meishan x Göttingen cross population.

In order to locate the genetic regions in the swine genome that are responsible for economically important traits, a resource population has been constructed by mating two female Meishan pigs with a male Göttingen miniature pig. In subsequent generations, 265 F2 offspring were produced from two F1 males and 19 F1 females. The F2 offspring were scored for eight traits including growth rate, teat number, vertebra number and backfat thickness, and genotyped for 318 genetic markers spanning the swine genome. Least-square analysis revealed quantitative trait loci (QTL) effects for vertebra number on chromosomes 1 and 2; for teat number on chromosomes 1 and 7; for birth weight on chromosome 1; for average daily gain between 4 and 13 weeks of age on chromosomes 9 and 10; for backfat thickness on chromosome 7; and for backskin thickness on chromosome 3.

Animals↗

PubMed2000

Enhancement of poly-adenosine diphosphate-ribosylation in human hepatocellular carcinoma.

BACKGROUND: Poly-adenosine diphosphate (ADP)-ribosylation, catalysed by poly(ADP-ribose) polymerase (PARP), is a post-translational modification of nuclear proteins and is involved in a wide range of biological processes including DNA repair, cell proliferation and malignant transformation. Alteration of this reaction in human hepatocellular carcinoma (HCC) is of interest, but has not yet been explored. The aim of this study was to evaluate poly-ADP-ribosylation and to compare the expression of PARP in HCC and adjacent non-tumour tissues. METHODS: Tumorous and adjacent non-tumorous tissues were obtained from five consecutive patients with HCC during surgery for tumour resection. Tissue homogenates were subjected to ADP-ribosylation with [32P]-nicotinamide adenine dinucleotide. The ADP-ribosylated proteins were separated by sodium dodecylsulfate-polyacrylamide gel electrophoresis, followed by autoradiography. Expression of PARP was also evaluated by western blotting. RESULTS: Several proteins were ADP-ribosylated in human HCC tissues. Notably, the radiolabelling of a 116-kDa protein was remarkably greater than that in adjacent non-tumorous tissues (86.5 +/- 35.2 arbitrary units by densitometry vs 12.2 +/- 9.9, mean +/- SD, n = 5, P < 0.02). The radiolabelling of the 116-kDa protein was decreased in the presence of PARP inhibitors in a concentration-dependent manner. Immunoblot analyses revealed that the radiolabelled protein was PARP and that its expression was significantly greater in HCC than in adjacent non-tumorous tissues (333 +/- 204% of non-tumorous tissue, P < 0.05). CONCLUSIONS: We found that poly-ADP-ribosylation and PARP expression were significantly increased in human HCC compared with those in adjacent non-tumorous tissues in surgically obtained specimens.

ADP-Ribosylation Factors↗

PubMed2000

p16 protein expression is associated with a poor prognosis in squamous cell carcinoma of the lung.

An immunohistochemical analysis for p16 protein was performed in 171 patients with non-small-cell lung cancer (NSCLC). Sixty-two carcinomas (36.3%) were classified as p16-negative. p16-negative tumours in squamous cell carcinomas (SCCs) were significantly more than those in adenocarcinomas (P = 0.039). There was no significant difference in survival according to tumour p16 status in patients with NSCLCs or in patients with adenocarcinomas. In contrast, of patients with SCCs, the 5-year survival rate of patients with p16-negative tumours was significantly lower than those with p16-positive tumours (P = 0.001). Especially, the survival of patients with p16-negative tumours was significantly worse than that of patients with p16-positive tumours in the early stage of the SCC, e.g. stage I (P = 0.005). Multivariate analysis showed that p16 status and nodal status were significant prognostic factors for the survival of patients with SCCs of the lung (P = 0.024 and P = 0.008 respectively). In conclusion, our study showed that alteration of p16 was one of the significant factors of a poor prognosis in SCCs of the lung, and that p16 might play an important role in some SCCs of the lung due to its high prevalence and prognostic value.

Adenocarcinoma↗

PubMed2000

Role of cyclic adenosine 3',5'-monophosphate and serum albumin in head-to-head agglutination of boar spermatozoa.

It has previously been shown that when boar spermatozoa are incubated in a modified Krebs-Ringer bicarbonate (mKRB), head-to-head agglutination occurs in many cells. The aim of the present study was to investigate the effects of cyclic adenosine 3',5'-monophosphate (cAMP) and serum albumin on sperm agglutination and to discuss a possible mechanism for sperm agglutination. Spermatozoa were collected from four mature boars, washed and incubated in mKRB. After a 1-h incubation, a sample of each sperm suspension was smeared gently on a separate glass slide, dried and stained in a phosphate-buffered solution of Giemsa to assess the percentage of head-to-head agglutinated cells in each suspension. In the samples incubated in mKRB, approximately 50% of the spermatozoa were agglutinated with one another at the acrosome. However, the percentages of head-to-head agglutinated spermatozoa were greatly reduced by a lack of calcium chloride in mKRB, but were recovered by the addition of dibutyryl cAMP (dbcAMP, a cAMP analogue) in a dose-dependent manner between 1 and 1000 microM. Addition of 3-isobutyl-1-methylxanthine (IBMX, 100 and 500 microM) instead of dbcAMP also significantly increased the percentages of head-to-head agglutinated spermatozoa. Moreover, the effects of adding dbcAMP were attenuated by treatment with Rp-adenosine 3',5'-cyclic monophosphorothioate triethylamine salt (0.25-1.0 mM, a cAMP antagonist) or H-89 (5 microM, a protein kinase-A inhibitor), but were enhanced by treatment with okadaic acid (500 nM) and calyculinA (500 nM) (inhibitors of protein serine/threonine phosphatase). In sperm samples incubated in mKRB containing 0.1% polyvinyl alcohol (mKRB-P) or mKRB-P lacking calcium chloride and supplemented with 1 mM dbcAMP, a lack of bovine serum albumin (BSA) resulted in a significant decrease in the percentages of head-to-head agglutinated spermatozoa. Addition of porcine serum albumin (PSA, 1-4 mg mL(-1)) or methyl-beta-cyclodextrin (MBC, 5-10 mg mL(-1)) instead of BSA was as effective as BSA (4 mg mL(-1)) in enhancing sperm agglutination. However, the effects of BSA (4 mg mL(-1)) or MBC (5 mg mL(-1)) were reduced by pre-mixing these reagents with cholesterol 3-sulfate (a cholesterol analogue, 5 microg mL(-1) for BSA and 375 microg mL(-1) for MBC). In addition, a protein 'anti-agglutinin' inhibiting sperm agglutination, was extracted from spermatozoa incubated with serum albumin or MBC and detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting techniques. The obtained Western blots revealed that sperm-bound anti-agglutinin was detected less in the samples incubated with either BSA (4 mg mL(-1)) or MBC (5-10 mg mL(-1)), compared with control samples. Moreover, pre-mixing MBC (5 mg mL(-1)) with cholesterol 3-sulfate (375 microg mL(-1)) reduced this reagent's effects on the loss of sperm-bound anti-agglutinin. Additionally, the assay of sperm agglutination and a chlortetracycline staining assay revealed that the percentages of head-to-head agglutinated spermatozoa were positively correlated with those of spermatozoa classified into B pattern (capacitated spermatozoa). These results are consistent with the following suggestions: (i) an adenylyl cyclase-cAMP-protein kinase system mediates a signalling pathway leading to head-to-head agglutination; and (ii) loss of anti-agglutinin from the spermatozoa may be modulated by changes in the plasma membrane induced by actions of serum albumin or MBC contained in a medium.

1-Methyl-3-isobutylxanthine↗

PubMed2000

Early clearance of circulating hepatitis C virus enhanced by induction therapy with twice-a-day intravenous injection of IFN-beta.

To improve the long-term efficacy of interferon (IFN) for treatment of chronic hepatitis C virus (HCV) infection, we proposed induction therapy with twice-a-day IFN-beta injection. This study was intended to clarify the antiviral mechanism. Thirty patients were randomly assigned to two groups: group A (twice-a-day therapy) received 3 MU IFN-beta intravenously (i.v.) twice a day for 2 weeks; group B (once-a-day therapy) received 6 MU of IFN-beta daily. HCV RNA, IFN-beta, alanine aminotransferase (ALT), 2'5'-oligoadenylate synthetase (2'5'-AS) activity, and beta2-microglobulin in serum were compared between the two groups during the first 2 weeks of IFN therapy. The clearance rate of serum HCV RNA in group A (86.7%) was significantly higher than that in group B (13.3%) at day 3 (p = 0.0006). No accumulation of IFN-beta was shown in serum throughout the therapy. The ratio (day 3/day 1) of 2'5'-AS activity was significantly higher in group A. Multivariate analysis indicated twice-a-day IFN-beta injection therapy led to significantly early clearance of circulating HCV. Twice-a-day IFN-beta injection therapy could induce biologically enhanced antiviral activities and be an efficient induction therapy for eradication of HCV.

Adult↗

PubMed2000

Haemophilus influenzae infection and Guillain-Barré syndrome.

It has been reported recently that Haemophilus influenzae can elicit an axonal form of Guillain-Barré syndrome. To investigate the incidence and features of H. influenzae-related Guillain-Barré syndrome, anti-H. influenzae antibody titres were measured by enzyme-linked immunosorbent assay (ELISA) in 46 consecutive Japanese patients with Guillain-Barré syndrome, 49 normal controls, 24 patients with multiple sclerosis and 27 patients with amyotrophic lateral sclerosis (ALS). Whole bacteria of non-encapsulated (non-typable) H. influenzae isolated from one of the Guillain-Barré syndrome patients was the antigen used. Elevated anti-H. influenzae antibodies for two or three classes of IgG, IgM and IgA were found in six (13%) Guillain-Barré syndrome patients, but not in the normal controls and patients with multiple sclerosis or ALS. The incidence was significantly higher in patients with Guillain-Barré syndrome than in the normal controls (P = 0.01) and patients with multiple sclerosis or ALS (P = 0.009). Western blot analysis confirmed that the H. influenzae-positive patients' IgG recognized the lipopolysaccharides of H. influenzae. Guillain-Barré syndrome patients with anti-H. influenzae antibodies showed relatively uniform clinical and laboratory features: prodromal respiratory infection, less frequent cranial and sensory nerve involvement, pure motor axonal degeneration on electrophysiology, and positivity for IgG anti-GM1 antibodies. Although the features were similar to those in Guillain-Barré syndrome patients infected by Campylobacter jejuni, the recoveries seemed to be better in patients with H. influenzae-related Guillain-Barré syndrome. It is concluded that a form of Guillain-Barré syndrome occurs after respiratory infection by H. influenzae in the Japanese population. A particular strain of non-typable H. influenzae has a ganglioside GM1-like structure and elicits axonal Guillain-Barré syndrome similar to C. jejuni-related Guillain-Barré syndrome.

Adult↗

PubMed2000

Natural autoantibody to MUC1 is a prognostic indicator for non-small cell lung cancer.

A great deal of attention has been focused on the antitumor effects of anti-MUC1 humoral and cellular responses. We examined whether anti-MUC1 antibody is present in patients with lung cancer, and evaluated its prognostic value. Serum was obtained from 30 patients with nonresectable, non-small cell lung cancer (NSCLC) and 60 healthy volunteers. The presence of anti-MUC1 antibody was determined by enzyme-linked immunosorbent assay. The patients were observed for a median follow-up time of 54.0 mo. Overall survival was estimated by the Kaplan-Meier method. Multivariate analyses were performed using the Cox proportional hazards regression model. Anti-KL-6/MUC1 antibody levels of the patients were significantly lower than those of normal individuals (p < 0.001). One-year survival rate of patients with high concentrations of anti-KL-6/MUC1 antibody was significantly higher than that of patients with low levels of anti-KL-6/MUC1 antibody (90.9% versus 21.1%, p < 0.001). Anti-KL-6/MUC1 antibody status was most strongly correlated with mortality, followed by lymph node status and albumin levels, whereas sex, serum lactate dehydrogenase (LDH), and carcinoembryonic antigen (CEA) levels, and metastasis status did not correlate with mortality. These preliminary results indicate that the degree of decrease in antibody level may be associated with a patient's prognosis.

Adult↗

PubMed2000

Different recognition by peroxisome proliferator structures in rat peroxisomal induction: application of sandwich ELISA using monoclonal antibody against rat peroxisomes.

A novel assay for a peroxisomal beta-oxidation enzyme by sandwich ELISA using a monoclonal antibody (RPX-5) against purified rat liver peroxisomes was developed. Immunoblot analysis revealed that RPX-5 recognized a 78 Kd protein, which is a peroxisomal bifunctional enzyme (PBE) in the beta-oxidation pathway. Immunoprecipitation by RPX-5 and the resulting reduction of PBE activity were dependent on RPX-5 concentrations. Sandwich ELISA using RPX-5 could be used to assay PBE in the range of 30 to 2000 ng protein/ml. In rat hepatocyte cultures, the PBE amount by this assay correlated well with PBE activity, with correlation coefficients of 0.965. Studying the mechanisms of peroxisomal induction, patterns of peroxisomal induction were examined by co-treatment of rat hepatocytes with various peroxisome proliferators (PxPs). Treatment with clofibrate and bezafibrate resulted in neither an additive nor synergistic effect on PBE level. On the other hand, co-treatment with either bezafibrate-Wy-14,643 or clofibrate-MEHP(mono(2-ethylhexyl)phthalate) both resulted in an additive effect. From these results, it is suggested that PxPs of the fibrate group may exert their functions via a common process, and non-fibrate PxPs via a different process in hepatocytes. The cognition site for peroxisome proliferators, therefore, might not involve a single site for inducing peroxisomal enzymes.

3-Hydroxyacyl CoA Dehydrogenases↗