心房钠尿肽合成细胞在大鼠胃的超微结构定位

Ultrastructural localization of atrial natriuretic peptide-synthesizing cells in rat stomach

背景：心房钠尿肽不仅存在于心脏且分布于全身许多部位，心房钠尿肽合成细胞在大鼠胃中形态学定位及在胃黏膜不同分区的分布特点仍处于研究阶段。目的：对心房钠尿肽合成细胞在大鼠胃中进行超微结构定位，观察其在胃不同区域的分布及形态。设计：重复测量实验。单位：承德医学院。材料：实验于2004-10／2007-07在承德医学院中药研究所免疫学实验室（省级重点实验室）完成，选用18只成年Wistar大鼠，体质量175-300g，雌雄不拘，由承德医学院实验动物科提供。实验过程中对动物的处置符合动物伦理学标准。实验用心房钠尿肽抗体及血清为美国Santa Cruz Biotechnology公司产品，胶体金标记抗体液（胶体金颗粒的直径为15nm，华美生物工程公司产品），CMTAS多功能真彩色病理图象分析系统由北京航空航天大学研制开发，日本产JEM-1200EX透射电镜。方法：大鼠经麻醉后，剪下胃及右心房（为心房钠尿肽抗体免疫组织化学染色时的阳性对照），沿大鼠胃贲门区、胃底区、幽门区的方向纵行取材。①将右心房作为阳性对照与胃组织在同一条件下进行免疫组织化学染色，观察心房钠尿肽在胃组织中不同部位的分布特点，确定心房钠尿肽合成细胞所在部位。②免疫电镜胶体金标记法对胃黏膜心房钠尿肽合成细胞的超微结构定位，采用CMTAS多功能真彩色病理图象分析系统对心房钠尿肽合成细胞细胞在不同的分布区域（贲门区、幽门区及胃底区）面密度的百分比。主要观察指标：胃心房钠尿肽合成细胞形态、所在部位及胃的不同分区面密度百分比。结果：①心房钠尿肽合成细胞形态及部位：阳性对照大鼠心房肌的心房钠尿肽合成细胞呈现阳性表达。大鼠胃中心房钠尿肽合成细胞也呈阳性表达，主要位于胃黏膜的下1／3。心房钠尿肽合成细胞的形态为圆形、锥体形和烧瓶型。阴性染色的部位为黏膜表层。黏膜下层和肌层。②心房钠尿肽合成细胞面密度的百分比：胃贲门区面密度的百分比最大，各面密度的百分比分布顺序是贲门区〉幽门区〉胃底区。结论：心房钠尿肽合成细胞主要位于胃黏膜的下1／3；大鼠胃不同分区心房钠尿肽合成细胞的面密度的百分比不相同，以胃贲门区最大。

BACKGROUND： Atrial natriuretic peptide （ANP）-synthesizing cells distribute in all over the body. However, very little is known about the morphological localization and the distribution characteristics of ANP-synthesizing cells in different regions of rat stomach. OBJECTIVE： To investigate the ultrastructural localization and distribution characteristics of ANP-synthesizing cells in rat stomach. DESIGN： Repeated measurement experiment. SETTING： Chengde Medical Collage, Chengde, Hebei Province, China. MATERIALS： This study was performed at the Laboratory for Institute of Chinese Materia Medica Immunology （provincial key laboratory）, Chengde Medical College between October 2004 and July 2007. Eighteen adult Wistar rats of either gender, weighing 175-300 g, were provided by the Laboratory Animal Center of Chengde Medical College. The disposal of animals was conducted in accordance with ethical guidelines for the use and care of animals. ANP antibody and serum （Santa Cruz Biotechnology, Inc, USA）, Protein A-15 nm colloidal gold labeled （Sino-American Biotechnology Company, China）, CMIAS image analysis system （Beijing University of Aeronautics and Astronautics, China） and JEM-1200EX transmission electron microscope （Japan） were used in the study. METHODS： After gastric and right atrial tissues （as a positive control in immunohistochemical staining for ANP-synthesizing cells） were freshly excised from anesthetized Wistar rats, the specimens were longitudinally harvested along rat gastric cardiac region, gastric fundic mucosa and gastric pyloric region. Gastric tissue was performed immunohistochemical staining （for positive control） together with fight atrial tissue, to observe the distribution characteristics of ANP-synthesizing cells in different regions of rat gastric tissue, and to localize ANP-synthesizing cells. The ultrastructural localization of ANP-synthesizing cells in the rat gastric mucosa was performed by postembedding immunoelectron microscopy. The area-density percentage of ANP-synthesizing cells in the different regions of rat stomach （gastric cardiac region, pyloric region and fundic region） was calculated with a CMIAS image analysis system. MAIN OUTCOME MEASURES： Morphological appearance and localization of ANP-synthesizing cells as well as the different area-density percentage in different regions of rat stomach. RESULTS： As a positive control, ANP-synthesizing cells showed intense positive expression in atrial myocytes and cytoplasm and in gastric mucosa （mainly within the lower third of rat gastric mucosa）. The morphological appearance of the individual ANP-synthesizing cell was variable, exhibiting round, pyramidal and flask shapes. Negative staining for ANP-synthesizing cells was detected in the lamina propria, submucosa, and smooth muscle. ANP-synthesizing cells existed in different regions of rat stomach and its area-density was the largest in the pyloric region. The distribution order of ANP-syuthesizing cells in area-density was gastric cardiac region 〉 gastric pyloric region 〉 gastric fundic mucosa in mucosa layer. CONCLUSION： ANP-synthesizing cells mainly exist within the lower third of the rat gastric mucosa. The percentage of area-density of ANP-synthesizing cells was variable in different regions of rat gastric mucosa, and its percentage of area-density was the largest in the gastric cardiac region.