Measurement of rat gastric mucus secretion using ex-vivo gastric chamber system

为了评估从大鼠胃粘膜分泌到胃腔的胃粘液，我们研究了Takeuchi等人开发的离体胃腔系统。为了应用该系统测量分泌到流出液中的粘蛋白，我们必须开发一种高灵敏度的方法来确定胃粘蛋白含量。最后采用高效液相色谱法测定了粘蛋白分子中主要糖基组分氨基己糖的含量。通过使用该方法结合包括非粘液糖蛋白的低分子量含己糖胺材料的去除处理，可以实现测量分泌到流出液中的粘蛋白的时间过程变化。本研究应用卡巴胆碱对大鼠胃粘膜粘液分泌的影响，结果表明卡巴胆碱对胃粘膜粘液分泌有促进作用，并试图获得特异性抗胃粘蛋白的单克隆抗体，用于免疫组化法测定粘蛋白含量。我们建立了几个单克隆抗体，这是具体的碳水化合物部分的粘蛋白存在于特定区域的大鼠胃粘膜。目前，我们正试图建立一种测定胃粘膜表面粘液细胞、胃体腺粘液细胞、胃窦腺粘液细胞等特定部位粘液含量的方法。

For the assessment of gastric mucus secreted from rat gastric mucosa into gastric lumen, we examined an ex-vivo gastric chamber system which had been developed by Takeuchi et al. To apply this system for the measurement of mucin secreted into the effluent solution, we had to develop a highly sensitive method to determine gastric mucin content. Finally we adopted an HPLC method to determine the content of hexosamine which constitutes the major carbohydarate component of mucin molecule. By using this method coupled with the removal treatment of low molecular weight hexosamine-containing materials including non-mucinous glycoproteins, it could be achieved to measure the time course changes of mucin secreted into the effluent solution. This method was applied to estimate the effect of carbachol administration on the mucus secretion in the rat gastric mucosa, and obtained a result that this drug to be an accelerator of mucus secretion into the gastric lumen.In the course of this study, we tried to obtain some monoclonal antibodies(MAbs) which are specific for the gastric mucin, and utilize them to the immunohistochemical method to determine the mucin content. We established several MAbs which are specific for the carbohydrate moieties of mucins present in the specific region of rat gastric mucosa. At present, we are trying to establish a method to assess the content of mucin originated from a specific site of gastric mucosa such as surface mucous cells, corpus gland mucous cells, antral gland mucous cells.