Immunohistochemical and molecular biological study on GSA-II binding epithelial cells of alimentary tract

消化道GSA-II结合上皮细胞的免疫组织化学和分子生物学研究

• 批准号: 04670022
• 负责人: SUGANUMA Tatsuo
• 金额: $ 1.28万
• 依托单位: Miyazaki Medical College
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04670022/
• 关键词: Mucus Glycoprotein; lectin; GSA-II; Alimentary Tract; mucous neck cell; Core-protein; Antibody; immunohistochemistry; ラット; 胃粘膜; アフィニティー精製; TFMS'; core蛋白質

Gastric mucus is thought to protect the gastric mucosa from mechanical trauma, desiccation, pathogenic microorganisms, acid and proteases. We purified Griffonia simplicifolia agglutinin (GSA)-II-binding mucus glycoprotein(GMG)from rat gastric mucosa by solubilization in a guanidine-containing buffer, gel permeation chromatography, Ricinus communis agglutinin(RCA)-I-affinity chromatography and GSA-II-affinity chromatography. Rat GMG showed high molecular weight on a Sephacryl S-1000 column, and a single band in 0.5%agarose-2% polyacrylamide composite gels and blots. A protein of about 60kDa was contained in the GMG preparation. GMG was deglycosylated with trifluoromethanesulfonic acid treatment. An antibody was rraised against deglycosylated GMG(deGMG). The antibody recognized deGMG, GMG, periodic acid-treated deGMG, and O-glycanase-digested dGMG, but did not react to trypsin-digested deGMG.These results that the antibody recognizes the peptide backbone of GMG.In imminohistochemistry, the mucous gel layr of the stomach luminal surface was stained with the antibody. The antibody recognized not only gastric mucous neck cells and pyloric gland cells but also gastric surface mucous cells, mucous cells in the duodenal gland, and goblet cells in the small intestine and colon. these indicate that GMG is a component of rat gastric mucus, and that the antibody recignizes mucous-secreting cells in rat stomach and intestine.

胃粘液被认为可以保护胃粘膜免受机械损伤、干燥、病原微生物、酸和蛋白酶的伤害。从大鼠胃粘膜中分离纯化了单叶灰树花凝集素(GSA)-II结合粘液糖蛋白(GMG)，通过凝胶渗透层析、蓖麻凝集素(RCA)-I-亲和层析和GSA-II-亲和层析从大鼠胃粘膜中纯化了GMG。大鼠GMG在Sephacryl S-1000柱上显示高相对分子质量，在0.5%琼脂糖凝胶-2%聚丙烯酰胺复合凝胶和斑点上显示单一条带。GMG制剂中含有约60 kDa的蛋白质。用三氟甲磺酸对GMG进行脱糖化处理。制备了抗脱糖化GMG(DeGMG)抗体。该抗体可识别deGMG、GMG、高碘酸处理的deGMG和O-葡聚糖消化的dGMG，但对胰酶消化的deGMG不起反应。结果表明，该抗体识别GMG的肽骨架。该抗体不仅能识别胃粘液颈细胞和幽门腺细胞，还能识别胃表面粘液细胞、十二指肠腺体粘液细胞以及小肠和结肠的杯状细胞。提示GMG是大鼠胃粘液的一种成分，该抗体可与大鼠胃和肠的粘液分泌细胞结合。

项目成果

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Oinuma, T.、Kawano, J.、Suganuma, T.：“非洲爪蟾发育过程中细胞分化和迁移的溴脱氧尿苷免疫组织化学”269. 205-212 (1992)。

Kawano J,Ide S., Oinuma T,Suganuma T: "A protein -specific monoclonal antivody to rat liver beta 1-4 galatosylransferase and its cpploication to immunohistochemistry." J.Histochem.Cytechem.(in press).

Kawano J、Ide S.、Oinuma T、Suganuma T：“针对大鼠肝脏 β 1-4 半乳糖基转移酶的蛋白质特异性单克隆抗体及其在免疫组织化学中的应用。”

Kawano J,Oinuma T,Nakayama T,Suganuma T: "Characterization of beta 1-4 galactosyltransferase purified from rat liver microsomes" J.Biochem.111. 568-572 (1992)

Kawano J、Oinuma T、Nakayama T、Suganuma T：“从大鼠肝微粒体中纯化的 β 1-4 半乳糖基转移酶的表征”J.Biochem.111。

Yamanari H,Suganuma T,Iwamura T,Kitamura N,Taniguchi S,Setoguchi T: "Extracellular matrix components regulating glandular differentiation and the formation of basal lamina of a human pancreatic cancer cell line in vitro." Exp.Cell Res.(in press).

Yamanari H、Suganuma T、Iwamura T、Kitamura N、Taniguchi S、Setoguchi T：“细胞外基质成分在体外调节人胰腺癌细胞系的腺体分化和基底层的形成。”

Torihara K., suganuma T., Ide S., Morimitsu T.: "Anionic sites in blood capillaries of the mouse cochlear duct." Hear.Res.(in press).

Torihara K.、suganuma T.、Ide S.、Morimitsu T.：“小鼠耳蜗管毛细血管中的阴离子位点。”