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Biomedical subjects

M Maeno

Publications and source records attributed to M Maeno.

At least 55 records · Page 3Linked to original sources
PubMed1992

Characterization of mineral-binding 40-kDa glycoprotein extracted from young adult rabbit alveolar bone.

A forty-kilodalton (40-kDa) protein was extracted from alveolar bone of young adult rabbit with 0.5 M EDTA after extraction with 4 M GuHCl, and purified by gel-filtration, anion-exchange and hydroxyapatite columns using a high-pressure liquid chromatography system under denaturing conditions. The purified 40-kDa protein was not susceptible to bacterial collagenase and thrombin, but was cleaved by cyanogen bromide. The protein was stained blue with Stains-all. Among various lectins, concanavalin A and lentil lectin agglutinin bound to this protein, but peanut agglutinin, Ricinus communis agglutinin, phytohemagglutinin-E and wheatgerm lectin agglutinin did not. Lectin binding assays showed that the protein is a glycoprotein containing large amounts of mannose and/or glucose residues, but is not a fragment of proteoglycan. The amino acid composition of the protein shows a characteristically high content of acidic amino acids. Therefore, the mineral-binding 40-kDa glycoprotein is considered to be osteonectin/secreted protein acidic and rich in cysteine (SPARC), in terms of similarities to bovine and porcine osteonectins with regard to molecular weight and contents of glycoses and amino acids.

Alveolar Process↗
PubMed1991

In vitro transcription of the hepatitis B virus gene by nuclear extracts of human hepatoma cells.

In vitro transcription of hepatitis B virus DNA (HBV DNA) was studied using nuclear extracts of human hepatoma cell lines. RNA polymerase II-dependent run-off transcription of pre-S mRNA under the control of pre-S1 promoter was observed in nuclear extracts obtained from HepG2 and PLC/PRF/5 cells, and the efficiencies in these extracts were significantly higher than those in nuclear extracts of non-liver cells such as HeLa, Molt-4, and Ehrlich. Analysis of run-off transcripts by the pre-S1 promoter, using deletion mutants of HBV DNA as templates and synthetic oligonucleotides as competitors, showed that hepatocyte nuclear factor 1 was necessary for initiation of in vitro transcription of pre-S mRNA. The run-off transcript of pregenome RNA was also detected and its initiation site was determined. Nuclear extracts of not only hepatoma cells but non-liver cells were active in transcription of pregenome RNA in vitro. However, run-off transcripts of S mRNA and X mRNA were not observed in this system. These results suggest that there were some differences between the mechanisms of HBV DNA transcription in vitro and in vivo. This in vitro transcription system will be useful for clarifying the mechanism regulating transcription of HBV DNA since the biochemical and functional characteristics of the nuclear factors can readily be analyzed.

Base Sequence↗
PubMed1991

Biochemical and immunocytochemical characterization of mineral binding proteoglycans in rat bone.

We examined biochemically and immunocytochemically the type and distribution of mineral binding proteoglycans (PGs) in rat mid-shaft subperiosteal bone using three monoclonal antibodies (MAb 1-B-5, 9-A-2, and 3-B-3) which specifically recognize unsulfated chondroitin, chondroitin 4-sulfate (C4-S) and dermatan sulfate (DS), and chondroitin 6-sulfate. Bone proteins were extracted from fresh specimens with a three-step technique: 4 M guanidine HCl (GdnCl), aqueous EDTA without GdnCl (E-extract), followed by GdnCl. Western blot analysis of SDS-polyacrylamide gel electrophoresis revealed that E-extract after chondroitinase ABC digestion reacted strongly with MAb 9-A-2 but not with MAb 1-B-5 or 3-B-3. After adehyde fixation, ethanolic trimethylammonium EDTA was used as a demineralizing agent for light and electron immunocytochemistry. This provided good retention of water-soluble PGs in the specimens. After chondroitinase ABC pre-treatment of tissue sections, MAb 9-A-2 specifically stained C4-S and/or DS in the walls of osteocyte lacunae and bone canaliculi in the mineralized matrix as well as in the unmineralized matrix such as pre-bone, vascular canals, and pericellular matrix surrounding osteocytes; the remainder of the mineralized matrix lacked staining. These results indicate that mineral binding PGs contain C4-S and/or DS and are exclusively localized in the walls of the bone lacuna and canaliculus.

Animals↗
PubMed1989

Characterization of multiple forms of small collagenous apatite-binding proteins in bone.

A number of small (Mrs 25-28 kDa) collagenous apatite-binding (SCAB) proteins that stain blue with 'Stains-All' have been isolated from fetal porcine bone by sequential extractions with 4M GuHCl (G1), followed by 0.5M EDTA (E), and again with 4M GuHCl (G2). Following purification under dissociative conditions, two types of SCAB proteins both with approximately one-third of their structure being collagenous, were identified in the EDTA extract. One type, which appears to be a novel protein, was revealed in two forms (SCABs 1 and 2, Mrs 25 and 28 kDa) that were recognized by a monoclonal antibody (MBP-322). The second type, SCAB 3, was also present in two forms; one form (SCAB 3a) having a lower affinity for hydroxyapatite than the other (SCAB 3b). These proteins were resistant to CNBr and displayed the chemical and immunochemical properties of the alpha 1 pN-propeptide of type I collagen. A third form of the propeptide (G2-28K) was a prominent component of the second 4M GuHCl extract. The chromatographic properties of serum alpha 1 (I) pN-propeptide were similar to SCAB 3a, indicating that SCAB 3b and G2-28K are post-translationally modified forms of the propeptide produced by bone cells. These propeptides may provide a link between the hydroxyapatite and collagen fibrils, and also have the potential to suppress collagen synthesis during bone resorption.

Animals↗
PubMed1988

Identification of small collagenous proteins with properties of procollagen alpha 1 (I) pN-propeptide in fetal porcine calvarial bone.

Several small collagenous apatite binding (SCAB) proteins have been extracted from the mineralized matrix of fetal porcine calvarial bone. One protein (SCAB 3), released on demineralization of bone with 0.5 M EDTA, appears to represent the alpha 1 pN-propeptide that is normally released during proteolytic processing of type I procollagen. The 28 Kd protein, which stains blue with "Stains-all", is reduced to a 19 Kd fragment by bacterial collagenase digestion, but is not susceptible to cyanogen bromide. The amino acid composition, blocked amino-terminus and immunological properties are all consistent with properties of alpha 1 (I) pN-propeptide. Fractionation on hydroxylapatite in the presence of urea has revealed a nonbinding (SCAB 3a) and a binding (SCAB 3b) form. Extraction of the demineralized matrix of bone with 4 M GuHCl revealed a third form (G2-28) which was similar to SCAB 3a on hydroxylapatite chromatography but showed differences on FPLC "Mono Q" resin. The occurrence of these different forms of pN-propeptide in bone may be of significance in collagen fibril-associated hydroxylapatite formation and in the regulation of osteoblastic function during bone resorption.

Amino Acids↗
PubMed1988

Differential effects of transforming growth factor-beta on the synthesis of extracellular matrix proteins by normal fetal rat calvarial bone cell populations.

To determine the effects of transforming growth factor-beta (TGF-beta) on the different cell types that exist in bone, cell populations (I-IV), progressively enriched in osteoblastic cells relative to fibroblastic cells, were prepared from fetal rat calvaria using timed collagenase digestions. TGF-beta did not induce anchorage-independent growth of these cells, nor was anchorage-dependent growth stimulated in most populations studied, despite a two- to threefold increase in the synthesis of cellular proteins. In all populations the synthesis of secreted proteins increased 2-3.5-fold. In particular, collagen, fibronectin, and plasminogen activator inhibitor synthesis was stimulated. However, different degrees of stimulation of individual proteins were observed both within and between cell populations. A marked preferential stimulation of plasminogen activator inhibitor was observed in each population, together with a slight preferential stimulation of collagen; the effect on collagen expression being directed primarily at type I collagen. In contrast, the synthesis of SPARC (secreted protein acidic rich in cysteine/osteonectin was stimulated approximately two-fold by TGF-beta, but only in fibroblastic populations. Collectively, these results demonstrate that TGF-beta stimulates matrix production by bone cells and, through differential effects on individual matrix components, may also influence the nature of the matrix formed by different bone cell populations. In the presence of TGF-beta, osteoblastic cells lost their polygonal morphology and alkaline phosphatase activity was decreased, reflecting a suppression of osteoblastic features. The differential effects of TGF-beta on bone cell populations are likely to be important in bone remodeling and fracture repair.

Animals↗
PubMed1987

Antigen reversion of glycogen phosphorylase isoenzyme in carcinoma and proliferative zone of intestinal metaplasia of the human stomach. An immunohistochemical study.

Specific antibodies to three purified glycogen phosphorylase isoenzymes, i.e., brain (or fetal), muscle, and liver types, were used to study the localization of these isoenzymes in carcinoma and the proliferative zone of intestinal metaplasia of the human stomach. Both the malignant cells of well-differentiated adenocarcinoma and the proliferative zone of some intestinal metaplasia of the stomach were stained when the antibrain-type phosphorylase isoenzyme antibody was used, but not when the other two types were used. The results suggest that brain-type phosphorylase in gastric carcinoma could be one example of fetal protein expression in cancer, and that the proliferative zone of some intestinal metaplasia having brain-type phosphorylase may histogenetically relate to well-differentiated adenocarcinoma.

Adenocarcinoma↗
PubMed1987

Characterization of a monoclonal antibody recognizing small collagenous proteins in fetal bone.

A monoclonal antibody (MBP-322) that recognizes two small collagenous, apatite-binding (SCAB) proteins associated with the mineral phase of fetal bone, has been prepared. The SCAB proteins, which are quantitatively extracted from bone with EDTA, have been shown by immunotransfer analyses to have MrS of 28K and 25K and both were selectively degraded by bacterial collagenase. Amino acid analysis of the collagenase-digested protein revealed a hypro:pro:gly ratio of approximately 0.5:1:0.7 for both proteins and indicated that one-third of the protein could have a collagen-like sequence. The SCAB proteins, unlike other collagens and collagen fragments tested bound quantitatively to hydroxylapatite in the presence of 4M guanidine hydrochloride and appear to be unique to bone. The antibody, however, was not specific for the SCAB proteins and showed comparable immunoreactivity against denatured alpha 1 chains of types I, II and III collagens and the alpha 2 chains of types I and V collagens but not type IV collagen nor native collagens I-V. The epitope was further localized to the CB6 fragment in the alpha 1(I) chain and the CB5 fragment of alpha 1(III) chain, and was present in both the TCA and TCB fragments of alpha 2(I). Despite the immunological reactivity, the properties of the SCAB proteins were not consistent with their being derived from known collagen types. Immunocytochemical staining of permeabilized bone cells with MPB-322 showed a perinuclear, punctate staining pattern in most cells with some cells showing specific nuclear staining. In non-permeabilized cells, the antibody stained various sized spherical particles, many of which were closely associated with the cell surface. Immunoblots of cell proteins revealed a number of immunoreactive proteins sensitive to collagenase digestion including two proteins with MrS similar to the SCAB proteins. The MBP-322 antibody appears useful for identifying sequence homology in various collagens, and for recognizing denatured collagen and specific collagen fragments in tissues, as well as being important for the further characterization of the SCAB proteins.

Amino Acids↗
PubMed1986

Immunohistochemical detection of glycogen phosphorylase isoenzymes in rat and human tissues.

Glycogen phosphorylase has at least three isoenzymes, i.e. muscle-, liver-, and brain-types. Antibodies have been raised against highly purified isoenzymes from rat muscle, liver and brain and found to react specifically to extracts from human muscle, liver and brain, respectively. Using these antibodies and the unlabelled antibody-enzyme method, each of the three isoenzymes has been localized in both rat and human tissues.

Animals↗
PubMed1985

Low mobility T cells in the peripheral blood of cancer patients: surface modification and mobility.

To elucidate how low-mobility T cells emerge in the peripheral blood of cancer patients, lymphocytes from normal individuals were treated with cancer patients' plasma, immunosuppressive (IS) substance with or without interleukin-2, or neuraminidase. A delay in cell mobility was induced by IS substance and neuraminidase treatments, which degrade the cell surface charge. Compared to the normal mobility pattern of lymphocytes, patterns induced by these treatments were characterized by a general shift of the fast cell peak to the left side. These patterns were different from those of cancer patients in terms of the inverse relationship between slow and fast peaks. These results indicate that, though humoral factors in the peripheral blood of cancer patients take a partial role, other mechanisms might operate to induce the characteristic mobility pattern of lymphocytes. One of the possibilities, the emerging of a special T cell subset recruited from the thymus, is discussed.

Cell Movement↗