小鼠胆囊酪氨酸激酶受体的表达及其意义

Expression of c-kit in CD1 mouse gallbladder and its significance

目的研究酪氨酸激酶受体（c-kit）在CD1小鼠胆囊的表达及其意义。方法采用免疫组化方法，应用激光扫描共聚焦显微镜检测CD1小鼠胆囊组织切片和急性分离单细胞的c-kit免疫反应阳性细胞的表达；RT-PCR法检测胆囊c-kit mRNA的表达；Western blot检测胆囊c-kit蛋白的表达。结果免疫荧光激光扫描共聚焦显微镜检测结果显示CD1小鼠胆囊从黏膜层到肌层均有c-kit免疫反应阳性细胞存在，其呈类似网络状分布和点和（或）丝状分布。急性分离胆囊单细胞检测结果显示c-kit免疫反应阳性细胞大致可区分为两种类型，一种为分支状，大多数细胞呈现该形态；一种为类梭形，少见分支，类似平滑肌细胞，部分细胞呈现此形态。而典型呈梭形的平滑肌细胞则无荧光显现，呈c-kit免疫反应阴性。RT-PCR法检测结果表明胆囊c-kit mRNA表达阳性。Western印迹显示c-kit蛋白表达阳性。结论c-kit在CD1小鼠胆囊表达阳性，即CD1小鼠胆囊存在Cajal间质细胞（ICC）。ICC可能与胆囊运动有关，有可能为胆囊起搏细胞。

Objective To study the expression of c-kit in CD1 mouse gallbladder and explore its significance. Methods Gallbladder tissue samples obtained from CD1 mouse were prepared for immunohistochemical investigations using the specific ICC marker c-kit antibody. Enzymatically dispersed cells were identified as ICC using anti-c-kit. Confocal microscopy was used to detect the expression of c-kit. Reverse transcription （RT）-PCR and western blot were used to confirm the expression of mRNA specific for the c-kit gene and production of c-kit protein in the CD1 mouse gallbladder. Results Anti-c-kit labeling demonstrated that ICC was present throughout the wall of the CD1 mouse gallbladder, from the mucosal/suhepithelium to the muscle regions, and indicated the similar network of ICC in the gallbladder as that in the small intestine of the same CD1 mice. Besides the network of ICC, c- kit labeling had a punctuate nature compared with the filamentous distribution. Using anti-c-kit, enzymatic dissociation of cells from tissue pieces containing spindle-shaped smooth muscle cells and branched cells, and the latter were usually immunopositive for c-kit, whereas the former, only a few showed fluorescent staining. And the dispersed spindle-shaped, smooth muscle cells showed no fluorescent staining. Meanwhile, RT-PCR indicated that the CD1 mouse gallbladder expressed mRNA specific for the c-kit gene and Western blot analysis showed the evidence of production of c-kit protein in the CD1 mouse gallbladder. Conclusion ICC are found in the CD1 mouse gallbladder, which is likely to lead to the development of motility study of gallbladder.