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

(1)胃底腺的粘液颈细胞(MNC)在系统发育上被认为最早出现在两栖动物中。我们研究了非洲爪哇胃底腺中单核细胞的来源和分化。用单叶灰树花凝集素(GSA)-II凝集素组织化学方法，在成虫胃底腺中特异性地检测到单核细胞。在变态幼虫胃的形态发生过程中，GSA-II反应细胞随机出现在发育不良的胃底腺的不同部位，然后迅速定位于颈部。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MNCs和SMC中间细胞和MNCs和OPC中间细胞两种类型。MNC和OPC的中间细胞也见于成人。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di…更多的患者对GSA-II无反应。这些观察结果表明，GSA-II在研究MNC及其前体细胞的分化方面是一个有用的标记，而不受物种分化的影响。(2)我们用溴脱氧尿嘧啶核苷(BrdU)免疫组织化学方法研究了成虫及其幼虫胃底腺的细胞分化和迁移。在成人的胃底腺中，BrdU的累积标记显示在SMCs和MNC之间有一个增殖细胞区，位于胃底腺的颈部。MNCs的标记指数在5周后迅速增加，OPC的标记指数在7周后出现延迟性增加，与MNCs标记指数的下降相一致。在幼虫未成熟的胃底腺中，标记的增殖细胞随机分布在发育中的胃粘膜中。在变态期间，未成熟上皮细胞的标记指数在第63期最高。63期幼虫注射BrdU后，在变态期间标记的上皮细胞没有明显的丢失。此外，在给药后第7天，SMCs/生殖细胞、MNCs和OPC等上皮细胞之间的标记指数也没有显著差异。在成人胃粘膜中，生殖细胞可能直接向上分化为SMC。大多数OPC似乎在向下迁移过程中出现了分化。生殖细胞经过一个阶段的跨国公司，最终成为OPC。在幼虫胃区组织发育期，未分化细胞可在原位增殖分化为SMCs、MNCs和OPC。他们可能是在和65岁阶段迁徙的。成人胃粘膜的组织发生在66期完成。(3)我们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，我们建立了GSA-II结合糖蛋白的酶联凝集素分析(ELLA)检测体系。大鼠胃粘膜糖蛋白在盐酸胍缓冲液中增溶，然后应用于Sephacryl S-1000柱。将含有GSA-II结合糖蛋白的部分应用于RCA-I-琼脂糖柱。保留在RCA-I柱中的样品通过GSA-IIagarose柱进行亲和纯化。纯化的GSA-II结合糖蛋白很可能可以开发出特异性抗体，并阐明这一项目。较少

(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