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N Usuda

Publications and source records attributed to N Usuda.

88 records · Page 5Linked to original sources
PubMed1988

Immunohistochemical studies on the localization of fibronectin in human thyroid neoplastic tissues.

Fibronectin (FN) has been considered to be involved in the malignant transformation of cells. It was thus of interest to morphologically study the cell surface distribution of FN in various types of thyroid neoplasms employing immunohistochemical techniques. Immunohistochemical staining using the specific anti-human FN antibody was performed by the avidin-biotin-peroxidase complex procedure. Compared to the staining pattern of normal thyroid tissues (control), differences in staining in malignant thyroid lesions and even in fetal and embryonal adenomas were evident, often on the lateral and basal aspects of the cell membrane. Colloid adenomas, however, showed FN staining on the basement membrane, basically similar to the controls. Moreover, in thyroid carcinoma, different staining patterns relating to each of the histological types were observed. The present investigation demonstrates the heterogeneity of the thyroid neoplasms in terms of FN binding.

Adenocarcinoma↗
PubMed1988

Immunocytochemical localization of liver-specific proteins in pancreatic hepatocytes of rat.

Hepatocytes are induced in the pancreas of rats maintained first on a copper-deficient diet for 8 weeks and then on normal rat chow. These cells are morphologically identical to parenchymal cells of the liver. These hepatocytes contain two liver-specific proteins: carbamyl phosphate synthetase I, a mitochondrial matrix protein that participates in the conversion of ammonia to carbamyl phosphate; and urate oxidase, an enzyme that catalyzes the oxidation of uric acid to allantoin. In addition, we also present evidence indicating that dietary administration of ciprofibrate induces peroxisomal beta-oxidation pathway enzymes, while the levels of catalase are unaltered in pancreatic hepatocytes. These observations along with the previously published results further establish the identity of pancreatic hepatocytes to parenchymal cells of liver and clearly indicate that transdifferentiation of pancreatic cells to hepatocytes is associated with activation of several liver-specific genes.

Animals↗
PubMed1988

Immunohistochemical studies on local antitumor effects of streptococcal immunopotentiator, OK-432, in human solid malignant tumors.

Immunocytochemical techniques were used to clarify the local inhibitory effects of a streptococcal immunopotentiator, OK-432, against solid malignant tumor growth. Natural killer (NK) cells and fibronectin were chosen as immunostaining markers to demonstrate the antitumor effects. Immunocytochemical staining was performed by the avidin-biotin-peroxidase complex method. These investigations demonstrated that (1) local administration of OK-432 seems to promote a marked induction of NK cells and fibroblasts around or entering into the cancerous lesions and (2) the cancer cell-killing effect of NK cells and the fibronectin-enriched stromal reaction augmented by the injection of OK-432 suggest at least the possibility of protection against neoplastic growth with invasion and the spread of distant or nodal metastases of solid carcinomas.

Adenocarcinoma↗
PubMed1988

Tissue specificity and species differences in the distribution of urate oxidase in peroxisomes.

The localization of urate oxidase in different tissues of rat and in the livers of selected mammalian species was investigated by immunoblot analysis and protein A-gold immunoelectron microscopy. Urate oxidase was purified from rat liver and used as an antigen to generate polyclonal antibodies in the rabbit. The antibodies were found to be monospecific by immunodiffusion and immunoblot analyses. By immunoblot analysis, urate oxidase was detected in the livers of rat, two strains of mice, hamster, dog, cat, and cow, but not in the Cynomolgus monkey and human liver. Urate oxidase was not detected by immunoblot method in rat kidney, jejunal mucosa, adrenal gland, testis, and pancreas. The subcellular localization of urate oxidase was ascertained by the protein A-gold immunocytochemical staining of the Lowicryl K4M embedded tissues. Urate oxidase was localized exclusively in the crystalloid core of the peroxisome in hepatic parenchymal cells of rat, mouse, hamster, dog, cat, and cow. The limiting membrane and the matrix of hepatic peroxisomes in these species were negative for the staining. The marginal plates of feline, canine, and bovine hepatic peroxisomes were also negative for urate oxidase. This enzyme was also not detected within the peroxisomes of human and monkey livers by the immunocytochemical technique. Peroxisomes (microperoxisomes) in extrahepatic rat tissues did not stain positively for urate oxidase by the protein A-gold immunocytochemical method, although they were positive for catalase. Fatty acyl-CoA oxidase was present in peroxisomes of jejunal mucosa, Leydig cells of test-is and pancreas but not in adrenal gland. Administration of a hepatic peroxisome proliferator, ciprofibrate or Wy-14643, failed to induce urate oxidase in rat liver. These results indicate that urate oxidase is a liver specific protein in rat and its localization within the liver peroxisomes of six mammals, excluding man and a nonhuman primate, and that its localization is limited exclusively to the crystalloid core. Unlike fatty acyl-CoA oxidase, urate oxidase does not appear to be inducible significantly by peroxisome proliferator treatment in the rat liver.

Animals↗
PubMed1987

Purification, properties, and immunocytochemical localization of human liver peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase.

A molecular understanding of genetic disease in which peroxisomal functions are impaired depends on analysis of the structure of normal and mutant enzymes of peroxisomes. We report experiments describing the isolation, characterization, and immunocytochemical localization of enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme (PBE) of the peroxisomal fatty acid beta-oxidation system from normal human liver and compared it with that of rat liver enzyme. The human enzyme, purified approximately equal to 2300-fold by ion-exchange chromatography, is homogeneous as judged by NaDodSO4/PAGE. This PBE is localized exclusively in the matrix of peroxisomes in liver cells by the protein A/gold immunocytochemical method. The human PBE is similar to rat enzyme in size (Mr, approximately equal to 79,000), isoelectric point (pI, 9.8), pH optima, molecular structure as observed by rotary shadowing, and peptide pattern on NaDodSO4/PAGE after proteolytic digestion with Staphylococcus aureus V8 protease. The human and rat enzymes differed in their immunological properties by having partial identity with each other; this is reflected in their slightly dissimilar composition of the amino acids aspartic acid, threonine, glutamic acid, tyrosine, and glycine. COOH-terminal amino acid were similar for both the enzymes: -Gly-Ser-Leu-Ile-COOH. These results suggest that the human and rat liver PBE may be different in their amino acid sequences at their antigenic sites.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1987

Absence of gamma-glutamyl transpeptidase activity in neoplastic lesions induced in the liver of male F-344 rats by di-(2-ethylhexyl)phthalate, a peroxisome proliferator.

Male F-344 rats were fed a diet containing 2% di-(2-ethylhexyl)phthalate (DEHP) for 95 weeks. Liver nodules and/or hepatocellular carcinomas (HCC) developed in 6/10 rats fed DEHP and none were found in controls (P less than 0.005 by chi 2 test). All the nodules and HCC were negative for gamma-glutamyl transpeptidase. In the non-tumorous portions of liver, the hepatocytes contained an increased number of peroxisomes and extensive accumulation of lipofuscin. By immunocytochemical analysis, the liver peroxisomes in rats treated chronically with DEHP had visually detectable decrease in the H2O2-degrading catalase and increase in H2O2-producing fatty acyl-CoA oxidase. These results show that higher dietary level of DEHP, which causes substantially greater degree of peroxisome proliferation than the 1.2% dietary level used in the National Toxicology Program bioassay (1982, Publication no. NTP-80-37, Tech. Report Series No. 217), can induce liver tumors in male rats.

Animals↗
PubMed1986

Purification of membrane polypeptides of rat liver peroxisomes.

Peroxisomes were obtained by sucrose density gradient centrifugation from the livers of di(2-ethylhexyl)phthalate-fed rats, and the membranes were prepared by carbonate extraction (Fujiki, Y., Fowler, S., Shio, H., Hubbard, A.L., & Lazarow, P.B. (1982) J. Cell Biol. 93, 103-110). The integrated membrane polypeptides were solubilized with sodium dodecyl sulfate, and purified by repeated polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Separation of 70 and 68 kDa polypeptides was not attempted in the present study because of their close migration in polyacrylamide gel electrophoresis. Other polypeptides with apparent molecular masses of 41, 27, 26, and 22 kDa were purified to near homogeneity. Antibodies were raised against these purified preparations. The 68 kDa polypeptide is suggested to be produced by the proteolytic modification of 70 kDa polypeptide, since the former increased concomitantly with decrease of the latter when the liver homogenate was incubated, and this change was prevented in the presence of leupeptin during the incubation. The 41 kDa polypeptide was a minor component. The 70 and 68 kDa polypeptides and 41 kDa polypeptide and their antibodies were cross-reactive, but the relation of these polypeptides was not clear. The 27 and 26 kDa polypeptides seemed to be another species of membrane polypeptides, although the relationship of these two polypeptides remains to be clarified. The 22 kDa polypeptide is not related to other membrane polypeptides. The results of immunoblot analysis of subcellular fractions of the liver and an electron microscopic immunocytochemical study to locate the antigenic sites with protein A-gold complex suggest that all of these polypeptides are localized on peroxisomal membranes. On proliferation of rat liver peroxisomes by administration of di(2-ethylhexyl)phthalate, a peroxisome proliferator, all of these polypeptides were markedly increased.

Animals↗
PubMed1986

Immunocytochemical localization of D-amino acid oxidase in the central clear matrix of rat kidney peroxisomes.

Light and electron microscopic localizations of D-amino acid oxidase (DAO) in rat kidney was investigated using immunoenzyme and protein A-gold techniques. The enzyme was purified from rat kidney homogenate and its antibody was raised in rabbits. By Ouchterlony double-diffusion analysis and immunoblot analysis with anti-(rat kidney DAO) immunoglobulin, the antibody was confirmed to be monospecific. The tissue sections (200 micron thick) of fixed rat kidney were embedded in Epon or Lowicryl K4M. Semi-thin sections were stained for DAO by the immunoenzyme technique after removal of epoxy resin for LM, and ultra-thin sections of Lowicryl-embedded material were labeled for DAO by the protein A-gold technique for EM. By LM, fine cytoplasmic granules of proximal tubule were stained exclusively. Among three segments of proximal tubules, and S2 and S3 segments were heavily stained but the S1 segment only weakly so. By EM, gold particles indicating the antigenic sites for DAO were exclusively confined to peroxisomes. Within peroxisomes, the gold particles were localized in the central clear matrix but not in the peripheral tubular substructures. The results indicate that D-amino acid oxidase in rat kidney is present exclusively in peroxisomes in the proximal tubule and that within peroxisomes it is found only in central clear matrix and not in the peripheral tubular substructures.

Animals↗
PubMed1986

[Distribution pattern of fibronectin in human lung cancer tissue].

The distribution pattern of fibronectin in human lung cancer tissues has been studied using the indirect immunoperoxidase technique. In adenocarcinoma fibronectin is observed in the basal side of cancer cells and tumor stroma. In squamous cell carcinoma, it is seen in the basal side of cancer cells, surface of cancer cells and tumor stroma. The difference between adenocarcinoma and squamous cell carcinoma in invasion and metastasis was investigated to have some relation to the distribution pattern of fibronectin.

Adenocarcinoma↗
PubMed1985

The localization of a new antithrombotic agent, cilostazol, in CHO-K1 cells as demonstrated by autoradiography.

Cilostazol (6-[4-(1-cyclohexyl-1H-tetrazol-5-yl)butoxy]-3,4-dihydro-2(1H)- qui nolinone, OPC-13013), a new antithrombotic agent, has a potent inhibitory action on platelet aggregation. The cellular and subcellular distribution of this compound was investigated. Autoradiographic studies using tritiated cilostazol showed that this compound penetrates the cell membrane of Chinese hamster ovary cells, clone K1 (CHO-K1 cells) and is distributed mainly in the cytoplasma. These results suggest that this compound exhibits its pharmacological action through specific inhibition of cytoplasmic cyclic AMP phosphodiesterase.

Animals↗
PubMed1983

Immunocytochemical demonstration of retinol-binding protein in the lysosomes of the proximal tubules of the human kidney.

The localization of plasma retinol-binding protein (RBP) in the human kidney was determined by two immunocytochemical techniques, the PAP method and the protein A-gold technique. By using the affinity purified antibody against RBP obtained from the urine of the patients with cadmium poisoning (Itai-Itai disease), the immunoreactive substances were located by light microscopy in the proximal tubules of the human kidney. By immuno-electron microscopy, the stained organelles were identified as lysosomes in both S1 and S2 segments of the tubules. These data suggested that the reabsorption of low molecular weight plasma proteins like RBP can occur in the two segments. We inferred a similarity between the S1 and S2 segments concerning the reabsorption of RBP.

Histocytochemistry↗
PubMed1983

Clinical application of the enzyme immunoassay for pancreatic oncofetal antigen.

Pancreatic oncofetal antigen (POA) purified by us has been detected in sera of patients with carcinoma of the pancreas by the micro-Ouchterlony method. In an attempt to improve the sensitivity and clinical usefulness of the serum POA assay, we established an enzyme immunoassay for POA and reported the results with this method. In this study, we investigated serum POA levels in pancreatic cancer and other diseases. The tissue localization of this POA in the pancreas was also studied. For the establishment of the enzyme immunoassay, an anti-POA-F(ab')2 fragment prepared from absorbed antiserum was conjugated with beta-D-galactosidase. The solid-phase "sandwich" principle was used. The normal upper limit of the serum POA level was defined as 400 units/ml. Among 60 patients with pancreatic cancer, 44 had elevated levels (73.3%). Of 22 cases with chronic pancreatitis, 4 had elevated levels (18.2%). In malignant diseases other than pancreatic cancer, elevated levels of serum POA were seen in 17.6% to 48.5% of the patients, most of whom had only slightly elevated levels. These results indicate that enzyme immunoassay for POA is clinically useful for making a diagnosis of pancreatic cancer. Immunoperoxidase staining showed POA to be found at the apical surface of ductular cells in fetal pancreas, at the luminal surface of glandular structures in pancreatic cancer tissue, and also at the luminal surface of the small duct in normal pancreas. Thus it is suggested that a high level of serum POA in patients with pancreatic cancer is derived from pancreatic cancer tissue.

Adult↗
PubMedDate not supplied

Assessment of proliferative activity of glandular cells in hyperfunctioning parathyroid gland using flow cytometric and immunohistochemical methods.

The proliferative activity of epithelial cells in hyperfunctioning parathyroid glands was estimated by flow cytometric and immunohistochemical procedures. A total of 30 parathyroid glands, 29 hyperfunctioning glands, and 1 normal gland were studied. The pathology of the 29 glands was determined to be hyperplasia in 19 and adenoma in 10. The S-phase cell population was expressed in terms of the S-phase fraction (%SPF) calculated from the histogram by DNA flow cytometry and in terms of the BrdU immunostaining labeling index (BrdU LI). Cells in all stages of the cell cycle were studied by Ki-67 immunostaining and expressed in terms of the labeling index (Ki-67 LI). Both BrdU LI and Ki-67 LI values were low, ranging from 0% to 0.58% and from 0.21% to 2.62%, respectively. The BrdU LIs were lower than the Ki-67 LIs, ranging from one-sixth to one-twelfth of the values depending on the disease. There were significant correlations between the two indices (p < 0.001). The %SPF determined by flow cytometry was consistently higher than both the BrdU LI and the Ki-67 LI. This discrepancy cannot be explained precisely, and further improvements are required for the flow cytometric analysis of %SPF. The cell cycle study by BrdU and Ki-67 immunohistochemistry suggested that the glandular cells of the hyperfunctioning parathyroid were characterized by low proliferative activity. No evidence of rapid cell turnover rate assumed from the flow cytometric study could be observed in the hyperfunctioning parathyroid gland.

Adenoma↗
PubMedDate not supplied

Study on RNA synthesis in the retina and retinal pigment epithelium of mice by light microscopic radioautography.

With the aim of determining the distribution of the incorporation of 3H-uridine in both retina and retinal pigment epithelium (RPE), the mouse eyes at embryonic day 9.5 (E 9.5), E 12.5, E 14.5, E 16.5, E 18.5 of gestational ages, and postnatal day 1 (P 1), P 3, P 7, P 14 were analyzed by light microscopic radioautography. Small pieces of the ocular tissues were labelled with 3H-uridine in vitro and light microscopic radioautographs were prepared. The average grain numbers per cell of the respective regions of tissues were calculated. In the retina, the grain numbers increased gradually from E 9.5 to P 1 and reached the maximal value at P 1, and then decreased until P 14. However, the grain numbers were more in the vitreal portion than those in the scleral portion at E 16.5 and then became more in the scleral portion from E 18.5 to P 14. It is considered that the ganglion and bipolar cells finish the RNA synthesis earlier, while the photoreceptor cells do it later during the fetal and postnatal development. In the RPE, the grain numbers gradually increased from E 12.5 to P 7 and then decreased until P 14. Considering the same ages, the grain numbers increased in the following order, anterior, equatorial and posterior regions during embryonic stages, but decreased in the same order after birth. Therefore, it is suggested that the activity of RNA synthesis in PE cells is higher in the posterior region than in the anterior region during embryonic stages. But the activity ascends generally and becomes relatively higher in the anterior region, after birth. Comparing the retina and RPE, it was noted that the grain numbers in the RPE were more important than in the retina and that the maximal value was at P 1 in the retina, while it was at P 7 in the RPE. From these results, it can be concluded that the RNA synthesis ceases earlier in the retina than in the RPE.

Aging↗