Biomedical subjects

H Munakata

Publications and source records attributed to H Munakata.

At least 73 records · Page 4Linked to original sources

PubMed1985

Enzymatic sulfation of exogenous high molecular weight glycopeptides by microsomal fraction of the rabbit uterine endometrium.

Incorporation of radioactive sulfate into exogenous glycopeptides and glycoproteins from adenosine 3'-phosphate 5'-phospho[35S]sulfate was studied with the microsomal fraction of the uterine endometrium of rabbits. A high molecular weight (Mr greater than 750,000) glycopeptide fraction from rat adenocarcinoma was found to be active as substrate, while several other glycoproteins were not. The rate of incorporation of sulfate was almost proportional to the concentration of glycopeptide substrate, the quantity of microsomal fraction, and the length of the incubation period. Cellulose acetate membrane electrophoresis demonstrated that radioactivity was incorporated into the glycopeptide fraction. The radioactive glycopeptide was excluded from a Sephadex G-50 column, but the 35S radioactivity and oligosaccharides were found in the retarded fractions after treatment with alkali in the absence of sodium borohydride. These observations indicated the presence of an enzyme which catalyzes the transfer of sulfate residue from adenosine 3'-phosphate 5'-phosphosulfate into the carbohydrate units attached to the polypeptide via O-glycosidic linkage. The sulfotransferase activity was measured with the microsomal fractions from the animals which had been treated with female sex hormones. As a result, it was found that estrogen enhances the activity and that progesterone suppresses the effect of estrogen.

Adenocarcinoma↗

PubMed1985

Effects of female hormones on the activity of 3'-phosphoadenylylsulphate: desulphated heparan sulphate sulphotransferase in the endometrium of rabbit uterus.

Sulphotransferase activity in the microsomal fraction of the uterine endometrium of rabbits was measured with heparan sulphate, N-desulphated heparan sulphate and N-, O-desulphated heparan sulphate as exogenous substrates in the presence of 3'-phosphoadenylyl[35S]sulphate. Of the three acceptors, N-, O-desulphated heparan sulphate was most effectively radiolabelled. When N-, O-desulphated heparan sulphate was used as an acceptor, it was found that estrogen enhanced the sulphotransferase activity and that progesterone suppressed the effect of estrogen.

Animals↗

PubMed1985

Characterization of blood group active glycopeptides derived from porcine kidneys.

Sialoglycopeptide fractions were prepared from the pronase digest of porcine kidneys by DEAE-Sephadex A-25 column chromatography and gel-filtration through Sephadex G-100. Their chemical compositions and large molecular size suggested that these glycopeptides were derived from mucin-type glycoprotein(s). The results of the beta-elimination reaction indicated that they have the O-glycosidic linkages between N-acetylgalactosamine and serine/threonine. The glycopeptides exhibited blood group A and H activities. The present study revealed that the porcine kidney contains the blood group antigens of glycoprotein nature.

ABO Blood-Group System↗

PubMed1985

Sulfated glycopeptides, containing desmosine and isodesmosine, isolated from porcine aorta.

Intima-media of porcine thoracic aorta was digested with pronase, after extraction of saline-soluble matters and fat. A crude sulfated glycopeptide fraction (CSGP) was precipitated with 90% (v/v) ethanol from the 80% ethanol-soluble fraction of the trichloroacetic acid (7%)-soluble fraction of the pronase digest. CSGP was fractionated by DEAE-Sephadex A-25 (Cl- form) column chromatography. Of the resulting 9 fractions, 4 major fractions were further purified by gel filtration on Sephadex G-50, followed by affinity chromatography on concanavalin (Con) A-Sepharose 4B to a homogeneous state in electrophoresis on cellulose acetate membrane. All the purified fractions contained glucosamine, galactose, mannose, and sialic acid as the major sugars, and desmosine and isodesmosine as the unique constituents. The fractions with affinity for Con A (S1 and S2) contained much more mannose than those without affinity for this lectin (S3 and S4). The latter contained sulfate. The predominant amino acids in the former were glycine, aspartic acid (and/or asparagine), and serine, while those in the latter were glycine, proline, and alanine.

Amino Acids↗

PubMed1985

An application of the high-iron diamine staining for detection of sulfated glycoproteins (glycopeptides) in electrophoresis on cellulose acetate membrane.

The high-iron diamine staining (HID), which has been used in histochemistry to stain sulfated glycoconjugates (SGC), was tested for detectability of authentic acidic substances (chondroitin sulfates A plus C, dermatan sulfate, heparan sulfate, chondroitin, hyaluronic acid, alpha 1-acid glycoprotein and ribonucleic acid) in electrophoresis on cellulose acetate membrane (Separax). The results showed that only SGC were detectable by the HID, although all the above substances were stained with alcian blue. The glycoconjugate preparations obtained from the liver, kidney, lung, small intestine, colon, stomach, brain and spleen of rats were examined by two-dimensional electrophoresis on Separax. The new spots (or bands), besides those of sulfated glycosaminoglycans, were detected by the HID on the electrophoretograms of all the samples except for the kidney one. The substances giving the new spots (or bands) were indicated to be sulfated glycopeptides (SGP) by crude heparinase digestion of a representative sample. The present results revealed that the HID was applicable for detection of SGP in electrophoresis on cellulose acetate membrane. Also, it is a novel finding that the liver and spleen contain SGP.

Animals↗

PubMed1985

Glycosaminoglycans from renal brush border membranes of rabbits.

Brush border membranes were prepared from the rabbit kidney cortex by sucrose density gradient centrifugation and digested with pronase. A glycosaminoglycan fraction was isolated from the digest by ion-exchange chromatography. Electrophoretogram on cellulose acetate membrane (Separax) of the glycosaminoglycan fraction indicated the presence of two components, which were identified as chondroitin sulfate A/C and dermatan sulfate on the basis of their mobility in Separax electrophoresis and sensitivity to chondroitinase ABC. Neither hyaluronic acid nor heparan sulfate was detected. The results of high-performance liquid chromatography of the pyridylamino derivatives of unsaturated disaccharides in the digests with chondroitinase AC and ABC of the glycosaminoglycan fraction indicated that the ratio of chondroitin 4-sulfate, chondroitin 6-sulfate and dermatan sulfate was 5.5 : 1.0 : 9.1.

Animals↗

PubMed1985

Distribution of fibronectin and other connective tissue components in human placenta.

Human placenta specimens obtained at term were investigated for distribution of fibronectin, collagens and glycosaminoglycans. When examined by the immunofluorescence staining technique with anti-plasma fibronectin antiserum, fibronectin was shown to be present around the fetal blood vessels and in the stroma of placental villi. The distribution of type IV collagen also was examined with specific antiserum. It was found that its distribution was similar to that of fibronectin. Conventional alcian blue staining indicated that the placental villi contained only small amounts of glycosaminoglycans. These data suggest that fibronectin and type IV collagen play important roles in tissue organization of the placental villi.

Chorionic Villi↗

PubMed1985

Unusual low affinity of human liver fibronectin for gelatin.

Fibronectin extracted from the human livers was shown to have very low affinity for gelatin unlike plasma fibronectin. It was demonstrated that this unusual property of liver fibronectin was not due to its proteolytic fragmentation.

Chromatography, Affinity↗

PubMed1985

Alteration in glycosidases from well-differentiated colorectal adenocarcinoma of rat.

The activities of six glycosidases in a rat colorectal adenocarcinoma were measured and compared with those of normal colonic mucosa. The specific activities of beta-galactosidase (EC 3.2.1.23) and beta-glucuronidase (EC 3.2.1.31) in the adenocarcinoma were similar to those of the corresponding ones in the normal mucosa, whereas those of beta-N-acetylglucosaminidase (EC 3.2.1.30), alpha-L-fucosidase (EC 3.2.1.51), alpha-galactosidase (EC 3.2.1.22) and beta-glucosidase (EC 3.2.1.21) were reduced in the former as compared with those in the latter. In the case of alpha-L-fucosidase, two forms were newly detected in the tumor. The relative abundance of three forms of beta-N-acetylglucosaminidase was quite different between the adenocarcinoma and the normal mucosa, and the level of the intermediate form in the tumor was markedly reduced. However, thermostability and Km values of two forms A and B in the tumor were not different from those of the corresponding ones in the normal tissue.

Adenocarcinoma↗

PubMed1985

Glycosaminoglycans in brush border membranes from rabbit small intestine.

A brush border membrane fraction was prepared from mucosae of rabbit small intestine by differential centrifugation. The membrane fraction was digested with pronase and a glycosaminoglycan fraction was isolated from the digest by fractionation with cetylpyridinium chloride. Individual glycosaminoglycans were quantitated by specific mucopolysaccharide-degrading enzymes and nitrous acid. The glycosaminoglycan composition was then calculated to be as follows: Hyaluronic acid, 34%; chondroitin sulfate A/C, 16%; dermatan sulfate, 11%; heparan sulfate, 39%.

Animals↗

PubMed1985

Sulfated glycopeptides from middle ear effusions of secretory otitis media.

The middle ear effusion specimens were obtained by myringotomy and aspiration from 4 children of 4-7 years old, who had been diagnosed as patients with secretory otitis media on the basis of conductive hearing loss and tympanogram. In cases 1 and 2, their ear fluids were macroscopically serous, while those of cases 3 and 4 were mucous. These ear fluids were digested with pronase and the digests were analyzed by cellulose acetate membrane electrophoresis with alcian blue and high-iron-diamine stainings. All samples were found to contain glycopeptides possibly derived from sulfated mucin-type glycoproteins with small amounts of glycosaminoglycans. The glycoconjugates from cases 3 and 4 were further examined after hyaluronidase and chondroitinase ABC treatments, followed by heparitinase digestion. The resultant glycopeptide fractions appeared to be electrophoretically homogeneous and their chemical compositions suggested that they were typical mucin-type glycopeptides. Furthermore, they contained sulfates. The data suggest that in secretory otitis media, one of the major components of middle ear effusions is sulfated mucin-type glycoprotein.

Amino Acids↗

PubMed1985

Evidence for the occurrence of lactosaminoglycan in porcine kidney.

Weakly acidic glycopeptides were obtained from he pronase digest of porcine kidney. The glycopeptides were digested with endo-beta-galactosidase. An oligosaccharide fraction was separated from the digest by gel-filtration through Sephadex G-50. The fraction contained galactose and glucosamine in an equimolar ratio.

Amino Sugars↗

PubMed1985

N-sulfation of heparan sulfate with microsomal fraction of endometrium of rabbit uterus.

Heparan sulfate was prepared from the pronase digest of the porcine kidney and modified chemically to yield N-desulfated, N-, O-desulfated, and N- O-desulfated N-acetylated heparan sulfates. Heparan sulfate and its modifications thus obtained were used as substrates to measure the sulfotransferase activity in the microsomal fraction of the endometrium of the rabbit uterus. N-, O-Desulfated heparan sulfate was the best acceptor of sulfate from 3'-phosphoadenylyl [35S] sulfate and intact heparan sulfate was the poorest one. The products of enzymatic reaction were treated with nitrous acid and examined by gel-filtration and electrophoresis. The results indicated that sulfate was almost exclusively incorporated as N-sulfate into all the present substrates.

Animals↗

PubMed1985

Glycosaminoglycans of porcine uteri.

Glycosaminoglycan fractions were obtained from the pronase digests of the endometrium and myometrium of porcine uteri. Their glycosaminoglycan compositions were determined by the use of specific mucopolysaccharide-degrading enzymes and nitrous acid. It was found that the endometrium contained chondroitin sulfates, dermatan sulfate and heparan sulfate and a smaller amount of hyaluronic acid, whereas hyaluronic acid and dermatan sulfate were the major glycosaminoglycans in the myometrium.

Animals↗

PubMed1985

Changes of glycosaminoglycan composition of uterine myometrium of rabbit induced by female sex steroids.

Glycosaminoglycan fractions were separately prepared from the myometrium of control, estrogen-treated, and estrogen-progesterone-treated rabbits after digestion with pronase. The glycosaminoglycan composition of each glycosaminoglycan fraction was determined by digestion with specific mucopolysaccharidases and nitrous acid. It was found that foregoing hormones showed no effects on the total glycosaminoglycan content, expressed as mg per g of the delipidated dry tissue. These hormones, however, markedly affected the glycosaminoglycan composition. As compared with the control, the estrogen-treatment decreased the contents of hyaluronic acid and dermatan sulfate, but increased those of chondroitin 4- and 6-sulfates. The effect of estrogen on the content of hyaluronic acid was strikingly enhanced by subsequent treatment with progesterone, and those of chondroitin 4- and 6-sulfates were also enhanced. On the other hand, progesterone abolished effect on the content of dermatan sulfate. Although no effect was observed on the content of heparan sulfate by the estrogen treatment, subsequent treatment with progesterone decreased the content of this glycosaminoglycan.

Animals↗

PubMed1985

[Intra-aortic PGE1 infusion combined with anti-malignant chemotherapy in the treatment of advanced neuroblastoma].

Neuroblastoma is one of the malignant childhood tumors which have a dismal prognosis, but is known to occasionally mature into benign histology spontaneously. Two cases with either stage III or IV neuroblastoma, whose tumors had not been extirpated at the initial operation, were treated by intra-aortic PGE1 infusion combined with oral papaverine and conventional anti-malignant chemotherapy. This therapeutic modality was effective in one case, where the tumor decreased in size and was removed in a second exploratory operation. The histology of the extirpated tumor was benign ganglioneuroma. In this case, however, distant bone metastases were found after the second operation. The role of PGE1 in the mechanism of bone metastasis remains to be determined.

Administration, Oral↗

PubMed1984

Further characterization of porcine plasma fibronectin which contains fucosylated carbohydrate chains.

Porcine plasma fibronectin and its functional four fragments produced by cathepsin B digestion were examined for biological, immunochemical and biochemical properties. Native fibronectin, 150-kDa and 130-kDa fragments exhibited similar cell attachment-promoting activity to each other. In an Ouchterlony double immunodiffusion system, these three polypeptides formed a precipitin line with anti-fibronectin antiserum, while the 50-kDa and 30-kDa fragments did not. The 150-kDa and 130-kDa fragments contained free sulfhydryl(s). The glycopeptide fractions were prepared by pronase digestion of porcine and human plasma fibronectin, and radiolabeled with [14C]acetic anhydride. The results of affinity chromatography with concanavalin A and lentil lectin immobilized on agarose indicated that the porcine glycopeptide fraction was different from the human fraction in that a larger part (58%) of the former was bound to lentil lectin. About 90% of this lentil lectin-reactive glycopeptides lost this reactivity upon alpha-L-fucosidase digestion. The glycopeptide fractions were also prepared from three carbohydrate-containing domains. Less than 30% of the radioactivity of the glycopeptide fractions of 150-kDa and 130-kDa fragments was retained on the lentil lectin-agarose, while about 90% of that from the 50-kDa fragment was retained. These results indicate that porcine plasma fibronectin has characteristics very similar to those of human plasma fibronectin and others, but is unique in that it contains fucosylated carbohydrate chains which unevenly distribute through functional domains.

Animals↗

PubMed1984

Biochemical and immunochemical comparison of fibronectin and polynectin from porcine plasma.

Polynectin, a glycoprotein of porcine blood plasma (also known as glycine-rich gelatin-binding protein (Isemura, M., Sato, N., and Yosizawa, Z. (1982) J. Biol. Chem. 257, 14854-14857] is similar to fibronectin with respect to binding characteristics to affinity gels. However, no immunoprecipitation reaction was observed either between polynectin and anti-fibronectin antiserum, or between fibronectin and anti-polynectin antiserum prepared in the present experiments. No cross-reaction was found between fibronectin and polynectin by enzyme-linked immunosorbent assays. Immunohistochemical studies revealed the presence in abundance of polynectin in intestinal and gastric glands. It was found that the pattern of distribution of polynectin was entirely different from that of fibronectin. Polynectin, in contrast to fibronectin, exhibited no cell attachment-promoting effects on BHK-21, a baby hamster kidney cell line, and a cell line of human embryonic lung fibroblasts. In addition, changes in binding characteristics to affinity gels after reduction and alkylation differed between polynectin and fibronectin. Although both proteins appear to have similar carbohydrate chains in view of the similar reactivities with lectins, the results of the present experiments demonstrated that polynectin is entirely different from fibronectin.

Alkylation↗