Dynamics of gastric mucous gel layer in vivo and in vitro using high pressure freezing method

冷冻固定是目前公认的最好的初始固定步骤，不仅可以保存生物标本的精细结构，还可以保留其抗原性。为了阐明胃壁细胞的功能转化，我们建立了一种新的体外实验模型，命名为离体胃黏膜。本研究用组胺刺激或西咪替丁抑制壁细胞的酸分泌，并对标本进行深低温冷冻(光镜观察)或高压冷冻(电镜观察)。结果表明，腺体细胞的组织结构在分离后至少2小时内保存完好，与新鲜切除的大鼠胃粘膜组织结构相似。H+/K+-ATPase免疫组织化学染色显示，在组胺刺激下，该酶从细胞质移位到顶膜。在西咪替丁处理的粘膜中，大多数壁细胞在形态上处于静息状态，在细胞质中显示大量的管泡。相反，组胺刺激的壁细胞显示出发达的细胞内小管和长的微绒毛。胃腺的功能形态动力学还需要进一步的研究。

Cryofixation is currently accepted as the best initial fixation step to preserve not only the fine structure but also the antigenicity of biological specimens. To elucidate the functional transformation of gastric parietal cells, we have newly developed an in vitro experimental model, named the isolated gastric mucosa. In this study, acid secretion of the parietal cell was stimulated with histamine or inhibited with cimetidine, and the samples were cryofixed by plunge freezing for light microscopy or high-pressure freezing for electron microscopy. As a result, the organization of glandular cells was well-preserved and quite similar to freshly excised rat gastric mucosa for at least 2 h after isolation. Immunohistochemistry of H+/K+-ATPase demonstrated a translocation of the enzyme from the cytoplasm to the apical membrane associated with histamine-stimulation. In cimetidine-treated mucosa, most of the parietal cells were morphologically in the resting state, showing numerous tubulovesicles in their cytoplasm. In contrast, histamine-stimulated parietal cells exhibited well-developed intracellular canaliculi lined with long microvilli.Further study should be required to clarify the functional morphological dynamics of gastric gland.