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Biomedical subjects

N Koide

Publications and source records attributed to N Koide.

At least 109 records · Page 6Linked to original sources

Immunohistochemical study of proteoglycans in D-galactosamine-induced acute liver injury in rats.

In this study, we carried out an immunohistochemical investigation of time-dependent alterations in the distribution of proteoglycans, and the proliferation profiles of hepatocytes and fat-storing cells (FSCs) in the livers of rats intoxicated with D-galactosamine (GalN). The proliferative cells were analyzed by proliferative cell nuclear antigen (PCNA) staining. In untreated rats, heparan sulfate, dermatan sulfate, and chondroitin/chondroitin sulfate were detected within the portal spaces and the central veins, and, with the exception of chondroitin, also within the reticular fibers. After administration of GalN, the number of PCNA-positive cells (FSCs and hepatocytes) and FSCs increased, reaching maximal on the 2nd and 3rd days, respectively. Heparan sulfate showed complicated changes. Dermatan sulfate decreased in portal spaces from the 2nd to the 3rd day, and in reticular fibers from 12 h to the 6th day. Chondroitin/chondroitin sulfate staining was observed from 2 h to the 6th day in the sinusoidal endothelia, which suggests that the sinusoidal endothelia may produce chondroitin/chondroitin sulfate transiently during liver damage as part of the mechanism of regeneration. Heparan sulfate and chondroitin/chondroitin sulfate were detected in necrotic regions, but dermatan sulfate was not. These observations suggest that heparan sulfate and chondroitin/chondroitin sulfate are involved in cell proliferation or morphogenesis and that the dermatan sulfate plays a role in the differentiation or functional maintenance of cells in liver regeneration.

Alanine Transaminase↗

Enzyme-linked immunoassay for midkine, and its application to evaluation of midkine levels in developing mouse brain and sera from patients with hepatocellular carcinomas.

Midkine (MK) is a growth factor that promotes neurite outgrowth and survival of neurons, and enhances the plasminogen activator in endothelial cells. A highly sensitive enzyme-linked immunoassay for MK was developed, involving affinity-purified anti-MK antibodies, their biotinylated form, and avidin-beta-galactosidase. The amount of bound avidin-beta-galactosidase was determined using a fluorogenic substrate, 4-methylumbelliferyl-beta-D-galactoside. This method allowed the detection of human and mouse MK in the range of 50 pg-10 ng. Pleiotrophin, which is related to MK in its amino acid sequence, did not show any cross reactivity. Employing this method, the MK levels in the developing mouse brain were determined. The MK level was 2 micrograms/g of wet tissue on the 12th day of gestation, and then steadily decreased during embryogenesis and postnatal development to 30 ng/g two months after birth. The assay method can also be applied to serum samples. Although the MK levels in the sera of normal human subjects were low or undetectable, 0.6-8 ng/ml of MK was detected in samples in the majority of cases of hepatocellular carcinomas.

Animals↗

Lipopolysaccharide induces apoptotic cell death of B memory cells and regulates B cell memory in antigen-nonspecific manner.

Lipopolysaccharide (LPS) was administered into sheep red blood cells (SRBC)-primed mice, and the effect of LPS on SRBC-specific memory cells was investigated. Spleen cells from SRBC-primed mice which were injected with LPS exhibited much lower in vitro secondary plaque-forming cells (PFC) responses to SRBC than those from untreated SRBC-primed mice. The in vitro anti-SRBC response of the spleen cells to LPS was also reduced. The combination experiments of B cells and T cells from SRBC-primed mice which were injected with or without LPS demonstrated that the reduction of immune responses to SRBC after administration of LPS was caused by the defect of SRBC-specific B memory cells, but not T memory cells. B cell type rosette-forming cells (RFC) for SRBC markedly decreased after injection of LPS, while PFC as antibody-forming cells did not increase subsequently. Therefore, the reduction of RFC was not due to their differentiation into PFC. The lymphoid follicles in the spleens from mice injected with LPS were stained positively by in situ nick end labeling specific for fragmented DNA. A large percentage of Ig+ spleen cells from SRBC-primed mice which were injected with LPS was also stained positively. The injection of glucocorticoids into SRBC-primed mice induced similar reduction of B memory cells. It was suggested that LPS might induce apoptosis of B memory cells and regulate B cell memory in antigen-nonspecific manner.

Animals↗

Differential release of smooth-type lipopolysaccharide from Pseudomonas aeruginosa treated with carbapenem antibiotics and its relation to production of tumor necrosis factor alpha and nitric oxide.

Endotoxin release from Pseudomonas aeruginosa treated with cell wall-active carbapenem antibiotics and its effect on the production of tumor necrosis factor alpha and nitric oxide were examined. Treatment of bacteria with imipenem induced much lower levels of endotoxin release than treatment with meropenem. The endotoxin released was demonstrated to be of the smooth type and O-specific polysaccharide-rich. The exposure of the filtrates of P. aeruginosa treated with imipenem to physiologically relevant cells caused low-level production of tumor necrosis factor alpha and nitric oxide, while similar treatment with meropenem induced high levels of production.

Carbapenems↗

Augmented immunological activities of recombinant lipopolysaccharide possessing the mannose homopolymer as the O-specific polysaccharide.

Recombinant lipopolysaccharide possessing the mannose homopolymer as the O-specific polysaccharide was manufactured genetically by transforming Escherichia coli K-12 with various rfb genes capable of synthesizing the mannose homopolymer. Recombinant lipopolysaccharide exhibited levels of anticomplement activity, adjuvant activity, and regional lymph node-enlarging activity much higher than those exhibited by the original rough-type lipopolysaccharide from E. coli K-12 or lipopolysaccharide possessing the heteropolysaccharide from E. coli O111. Immunological activities of recombinant lipopolysaccharide were as strong as those of wild-type lipopolysaccharide possessing the mannose homopolymer. Characteristic activities of wild-type lipolysaccharide possessing the mannose homopolymer were exhibited by recombinant lipopolysaccharide. The abilities of lipopolysaccharide to activate B cells polyclonally and to produce cytokines did not seem to be related to the presence of the mannose homopolymer. Therefore, it was suggested that the mannose homopolymer in the O-specific polysaccharide might exclusively enhance anticomplement activity, adjuvant activity, and regional lymph node-enlarging activity among various lipid A activities.

Adjuvants, Immunologic↗

Apoptotic cell death in the response of D-galactosamine-sensitized mice to lipopolysaccharide as an experimental endotoxic shock model.

The apoptotic cell death induced in D-galactosamine-sensitized mice by administration of lipopolysaccharide was characterized. Administration of lipopolysaccharide caused apoptotic cell death in livers of D-galactosamine-sensitized mice. Apoptotic cells were also detected in the kidney, thymus, spleen, and lymph node. Severe hepatic apoptosis in D-galactosamine-sensitized mice was reproduced by transfer of the sera from mice injected with D-galactosamine and lipopolysaccharide. The hepatocyte apoptosis induced by lipopolysaccharide was completely prevented by an anti-tumor necrosis factor alpha antibody but not by an anti-gamma interferon antibody. Administration of recombinant tumor necrosis factor into D-galactosamine-sensitized mice also caused hepatocyte apoptosis. Lipopolysaccharide-induced hepatocyte apoptosis in D-galactosamine-sensitized mice did not seem to be mediated by Fas antigen. It was suggested that lipopolysaccharide- induced hepatic injury and failure in D-galactosamine-sensitized mice was due to the apoptotic cell death of hepatocytes caused by tumor necrosis factor alpha released in the circulation.

Animals↗

Transient accumulation of perisinusoidal chondroitin sulfate proteoglycans during liver regeneration and development.

After partial hepatectomy, the liver is capable of complete restoration of normal hepatic size, architecture, and function (regeneration). To study roles of the extracellular matrix in regeneration, the temporal and spatial sequences of deposition of several components, including collagen I, III, and IV, fibronectin, laminin, heparan sulfate proteoglycan (perlecan), and chondroitin sulfate proteoglycans were characterized by light microscopic immunohistochemistry in rat liver after 70% partial hepatectomy. Consistent with previous reports, there was a brisk mitosis of hepatocytes after the partial hepatectomy. Of the extracellular matrix components studied, 1B5 epitope generated by chondroitinase ABC digestion on chondroitin sulfate proteoglycans exhibited the most dramatic changes; the epitope was detectable as early as 1.5 hr after partial hepatectomy and its immunoreactivity reached a maximum at 24 hr, then declined gradually. This transient expression of the 1B5 epitope was also detected in neonatal rat liver during development. By Western blotting, the 1B5 epitope was found on two forms of the core protein of chondroitin sulfate proteoglycans with apparent molecular masses of 163 KD and 152 KD, which were also regulated in the same temporal manner.

Animals↗

Cytochrome P450s of isolated rat hepatocytes in spheroid and monolayer cultures.

The maintenance of cytochrome P450s (P450s) and NADPH-cytochrome P450 reductase (P450 reductase) in the monolayer and spheroid cultures of hepatocytes from male rats was examined. The content of total P450 in monolayer culture decreased to almost none after 144 hr, whereas the level in spheroid culture remained within 6-13% of initial values during an incubation period of 144-192 hr. P450 2C11, a major P450 in male rat, in monolayer cells rapidly decreased in 144 hr, while the level in spheroid cells after 144 hr and 192 hr maintained 25% and 15%, respectively, of initial level. On the other hand, P450 2A1 and P450 2E1 in both monolayer and spheroid cells rapidly decreased. P450 reductase in both cells showed a gradual decline reaching a level of 43-44% of the initial level at 96 hr, and remained within 16-17% of the initial value during an incubation period of 192 hr. These results indicate that P450 2C11 in spheroid cells maintained more stable than in the monolayer cells, and that P450 reductase in both cultures declined only moderately, compared with P450s.

Animals↗

Role of tumor necrosis factor-alpha and glucocorticoid on lipopolysaccharide (LPS)-induced apoptosis of thymocytes.

Administration of bacterial lipopolysaccharide (LPS) into mice markedly induced the apoptosis of CD4+8+ thymocytes. The injection of anti-tumor necrosis factor (TNF)-alpha antibody or RU38486, a glucocorticoid receptor antagonist, into mice definitely inhibited LPS-induced apoptosis of thymocytes. Addition of the sera 1 h after injection of LPS into in vitro cultures of thymocytes caused thymocyte apoptosis. It was also prevented by either anti-TNF-alpha antibody or RU38486. Further, recombinant TNF-alpha and hydrocortisone collaborated in induction of the thymocyte apoptosis in vitro. The in vivo phenomenon of LPS-induced apoptosis of thymocytes was reproducible by the in vitro experimental system. It was therefore suggested that both TNF-alpha and glucocorticoid participate and collaborate as effector molecules in LPS-induced apoptosis of thymocytes.

Animals↗

Binding of mannose-binding protein to Klebsiella O3 lipopolysaccharide possessing the mannose homopolysaccharide as the O-specific polysaccharide and its relation to complement activation.

Lipopolysaccharide from Klebsiella pneumoniae O3, which possesses the mannose homopolysaccharide as the O-specific polysaccharide, exhibits an extraordinarily high ability to activate the human complement system. We isolated the mannose-binding protein with a Klebsiella O3 lipopolysaccharide affinity column. The protein isolated had a molecular mass of much higher than 200 kDa, and it consisted of subunits with an apparent molecular mass of 32 kDa. The NH2-terminal sequence of the 32-kDa subunits was completely consistent with a part of the amino acid sequence of human serum mannose-binding protein. In immunoblotting, an anti-mannose-binding protein monoclonal antibody was definitely reactive with the isolated protein with the higher molecular mass. The protein isolated was bound exclusively to lipopolysaccharides possessing the mannose homopolysaccharide, not to lipopolysaccharide possessing the heteropolysaccharides. Klebsiella O3 lipopolysaccharide did not exhibit a high anticomplement activity in the serum from which the mannose-binding protein was depleted. It was concluded that the serum factor that bound to Klebsiella O3 lipopolysaccharide may be mannose-binding protein and that it may play a crucial role in the strong complement activation by Klebsiella O3 lipopolysaccharide.

Amino Acid Sequence↗

Gene expression of liver-specific proteins in hepatocyte spheroids in primary culture.

Adult rat hepatocytes assemble to form multicellular spheroids under non-adherent environments such as immobilized chondroitin sulfate-proteoglycan in primary culture. Previously, we demonstrated that hepatocyte spheroids exhibited various differentiated structures as observed in the liver tissue. It was also shown that hepatocyte growth was highly suppressed and several differentiated functions, including albumin production and gluconeogenesis, were well preserved in spheroids. To investigate the differentiated functions of cultured hepatocytes in relation to cell morphology, we compared the expression of the albumin and transferrin genes in spheroids with those in monolayers by Northern blot analysis. Production of these proteins in the culture medium was simultaneously examined by ELISA. Gene expression and protein production of both albumin and transferrin were better preserved in spheroids. We also examined changes in the expression of liver-specific genes in response to IL-6. Reduced mRNA levels of both albumin and transferrin was only found in spheroids and no change was observed in monolayers. These results suggest that the regulation of tissue-specific gene expression is better preserved in spheroids, in which hepatocytes are in close contact with each other.

Albumins↗

A family with hereditary serum cholinesterase deficiency.

A family with serum cholinesterase (SChE) deficiency is reported. A 64-year-old woman was admitted for the excision of colon adenoma; her laboratory data revealed a markedly decreased level of SChE. SChE genes of the patient and her family members were amplified by the polymerase chain reaction (PCR) and analyzed by direct sequencing. The patient's SChE gene had a homozygous frame shift mutation, in which an extra adenine was inserted in codon 315 (ACC-->AACC), resulting in the appearance of a new stop codon in codon 322. The family study disclosed that her brother and sister had the same frame shift mutations in homozygote and heterozygote, respectively.

Base Sequence↗

Increase of fucosylated serum cholinesterase in relation to high risk groups for hepatocellular carcinomas.

Serum cholinesterase (ChE) (E.C. 3.1.1.8) is a glycoprotein which has 36 potential sites of asparagine-N-linked sugar chains. The structures of oligosaccharides released from ChE on hydrazinolysis were studied by serial lectin affinity column chromatography, exoglycosidase digestion, and methylation analysis. Seventy-three % of the sugar chains occurred as biantennary oligosaccharides and the remainder as C-2 and C-2,4/C-2,6 branched tri- and tetraantennary oligosaccharides. Several percentages of the Lewis X antigenic determinant and fucosylated mannose core were linked to them, and their sialic acid residues were linked to nonreducing terminal galactose residues at the C-3 and C-6 positions. Aleuria aurantia lectin-reactive ChE with the Lewis X antigenic determinant increased in hepatocellular carcinomas and liver cirrhosis compared with chronic hepatitis; on the other hand, Aleuria aurantia lectin-reactive ChE did not change significantly after transcatheter arterial embolization and was not related to the serum levels of alpha-fetoprotein and carcinoembryonic antigen in patients with hepatocellular carcinomas. Accordingly, the analysis of Aleuria aurantia lectin-reactive ChE is clinically useful for differentiating liver cirrhosis from chronic hepatitis and to identify high risk groups for hepatocellular carcinomas, i.e., cirrhotic patients in Child's A grade.

Biomarkers↗

Host factors required for internal initiation of translation on poliovirus RNA.

Translation of poliovirus RNA is initiated by entry of ribosomes into the nucleotide sequence (internal ribosomal entry site; IRES) within the 5'-untranslated region (5'-UTR). Efficiency of this translation initiation in rabbit reticulocyte lysates (RRL) was very low and was greatly enhanced by addition of the ribosomal salt-wash fraction (RSW) prepared from HeLa cells. This stimulating activity in the RSW was partially purified by gel-filtration column chromatography and its molecular weight was estimated to be higher than 240,000. Several proteins that bind specifically to the poliovirus IRES were detected in the active fraction. Among those, a 57 kDa protein, recognized by antibodies against polypyrimidide tract-binding protein (PTB), was found. In addition, La protein (52 kDa) which is a human antigen recognized by antibodies from patients with autoimmune disorders was also detected. Further purification on a hydroxylapatite column resulted in considerable loss of the stimulatory activity, accompanied by a reduction of the apparent molecular weight of active component(s). These results suggest that fully active HeLa cell stimulatory factors for the translation initiation on poliovirus RNA function in RRL as a large complex consisted of several components including PTB and La protein.

Acrylic Resins↗

[Lugol staining for esophageal carcinoma and influence of radiotherapy].

For evaluating the endoscopic staining of the esophageal carcinoma with lugol solution, 50 patients who underwent esophagectomy for carcinoma were subjected to this study. Among the 50 patients, 21 were radiated before surgery and 29 were not radiated. The findings of the lugol staining were compared between endoscopic staining and staining on removed specimens. Non-staining area demonstrated by endoscopic procedure almost agreed with that by the procedure on removed specimen in non-radiation group, but both areas of 28.6% cases disagreed in radiation group. On the second step, the extent of non-staining area demonstrated by the procedure of removed specimen was compared with histological extent of carcinoma. The non-staining area on the removed specimen was more extended than histological extent of carcinoma; 10.3% in the non-radiation group and 71.4% in the radiation group. As one of the causes of the large non-corresponding rate in the radiation group, radiation esophagitis was demonstrated. It can be finally concluded that the reliability of endoscopic lugol staining is reduced by preoperative irradiation.

Esophageal Neoplasms↗