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Histochemical and immunohistochemical study on GSA-II binding glycoprotein of alimentary tract

消化道GSA-II结合糖蛋白的组织化学和免疫组织化学研究

基本信息

批准号：
02670014
负责人：
SUGANUMA Tatsuo
金额：
$ 1.41万
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：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02670014/
关键词：
Glycoconjugate lectin GSA-II Alimentary tract Mucous neck cell Histochemistry Bromodeoxyuridine Ontogenesis 胃

项目摘要

(1) Mucous neck cells (MNCs) of the fundic gland are phylogenetically thought to have first appeared in amphibians. We studied the origin and differentiation of MNCs in fundic glands of xenopus laeves. By means of lectin histochemical methods using Griffonia simplicifolia agglutinin(GSA)-II, MNCs were detected specifically in fundic glands of adult X. laeves. During the morphogenetic period of the stomach in metamorphosing larvae, GSA-II reactive cells randomly appeared in various portions of the underdeveloped fundic glands and then rapidly localized in the neck portion. At this time, newly appearing mucous granules of MNC type were labeled by GSA-II-colloidal gold(CG). Two types of cells intermediate to MNCs and SMCs and intermediate to MNCs and OPCs were observed in the larval gastric region. Cellsintermediate to MNCs and OPCs were also found in adults. In these cells, mucous granules of MNC type were labeled by GSA-II-CG, but mucous granules of SMC type and zymogen-like granules di … More d not react to GSA-II. These observations suggest that GSA-II is a useful marker in studying the differentiation of MNCs and their precursors regardless of species differentiation.(2)We have examined cellular differentiation and migration in the fundic glands of adult X. laevis as well as its larvae using bromodeoxyuridine(BrdU)immunohistochemistry. In the adult fundic glands, cumulative labeling with BrdU revealed a proliferative cell zone between the SMCs and MNCS, in what is referred to as in the neck portion of the fundic gland. The labeling index of MNCs rapidly increased by week 5. The labeling index of OPCs showed the delayed-increase by week 7, coincident with the decrease in the labeling of MNCs. In the immature fundic glands of larvae, the labeled proliferating cells randomly distributed throughout the developing gastric mucosa. During the metamorphosing period, the labeling index of immature epithelial cells was the highest at stage 63. After the administration of BrdU to larvae at stage 63, there was no significant loss of the labeled epithelial cells during the metamorphosing period. Furthermore, there was no significant difference in the labeling indices among the epithelial cells such as SMCs/generative cells, MNCs, and OPCs at 7 days after administration. In adult gastric mucosa, generative cells might differentiate directly to SMCs in an upward direction. A majority of OPCs seemed to differentiate during a downward migration. Generative cells pass through a stage as MNCS, finally becoming OPCS. In histogenetic period of larval gastric region, undifferentiated cells might proliferate and differentiate to SMCS, MNCs and OPCs in situ. They may have migrated during stages 64 and 65. Histogenesis of adult gastric mucosa completed at stage 66.(3)We have purified GSA-II-binding glycoprotein from rat gastric mucosa. Before trying purification of this glycoprotein, we have established the assay system for GSA-II binding glycoprotein using the enzyme-linked lectin assay(ELLA)method. Glycoprotein from rat gastric mucosa was solubilized in the guanidineHCL buffer, then applied on to Sephacryl S-1000 column. The fraction containing GSA-II binding glycoprotein was applied to RCA-I-agarose column. The sample retained in RCA-I column was subsequently affinity-purified through GSA-IIagarose column. It is likely that purified GSA-II-binding glycoprotein allows to develop specific antibody and to shed light on this project. Less

(1)胃底腺的粘液颈细胞（MNCs）在遗传学上被认为最早出现在两栖动物中。本文研究了非洲爪蟾胃底腺中MNCs的起源和分化。应用GSA-Ⅱ凝集素组织化学方法，对黄脊灰蝶成虫胃底腺中的MNCs进行了特异性检测。莱维斯在变态幼虫的胃的形态发生期间，GSA-II反应细胞随机出现在不发达的胃底腺的各个部分，然后迅速定位在颈部。此时，用GSA-II-胶体金（CG）标记新出现的MNC型粘液颗粒。在幼虫胃区观察到两种类型的细胞，即介于MNC和SMC之间的细胞和介于MNC和OPCs之间的细胞。在成年人中也发现了MNCs和OPCs的中间细胞。在这些细胞中，MNC型粘液颗粒被GSA-II-CG标记，而SMC型粘液颗粒和酶原样颗粒被GSA-II-CG标记。 ...更多信息对GSA-II没有反应。这些观察结果表明，GSA-II是一个有用的标记，在研究分化的跨国公司及其前体，无论物种分化。(2)We已经研究了成年X的胃底腺中的细胞分化和迁移。用溴脱氧尿苷（BrdU）免疫组织化学方法对中国对虾及其幼虫进行了鉴定。在成人胃底腺中，累积标记BrdU揭示了SMC和MNC之间的增殖细胞区，即胃底腺的颈部。MNCs的标记指数在第5周迅速增加。OPCs的标记指数在第7周出现延迟性增加，与MNCs标记指数的减少一致。在未成熟的胃底腺中，标记的增殖细胞随机分布于整个发育中的胃粘膜。在变态期，未成熟上皮细胞的标记指数在63期最高。在第63期幼虫给药后，在变态期标记的上皮细胞没有明显的损失。此外，在给药后7天，上皮细胞如SMC/生殖细胞、MNC和OPC之间的标记指数没有显著差异。在成人胃粘膜中，生殖细胞可直接向上分化为平滑肌细胞。大多数OPC似乎在向下迁移过程中分化。生殖细胞经过一个阶段作为MNC，最终成为OPCS。在组织发生期，未分化细胞可原位增殖分化为SMCS、MNCs和OPCs。它们可能在第64和65阶段迁移。成人胃粘膜的组织发生在第66期完成。(3)We从大鼠胃粘膜中纯化了GSA-II结合糖蛋白。在尝试纯化该糖蛋白之前，我们使用酶联凝集素测定（ELLA）方法建立了GSA-II结合糖蛋白的测定系统。大鼠胃粘膜糖蛋白用盐酸胍缓冲液溶解后，上Sephacryl S-1000柱。将含有GSA-II结合糖蛋白的级分上样至RCA-I-琼脂糖柱。保留在RCA-Ⅰ柱上的样品随后通过GSA-Ⅱ琼脂糖柱进行亲和纯化。纯化的GSA-II结合糖蛋白可能允许开发特异性抗体并为该项目提供线索。少