摘要
目的分析Claudin-7基因敲除后小鼠表型及Claudin-7潜在的抑癌功能。方法将小鼠分为2组,Claudin-7基因敲除组及野生型组,每组各15只,分别采用PCR鉴定小鼠基因型,HE染色观察Claudin-7基因敲除后各脏器的组织病理学变化;免疫共沉淀(CO-IP)检测Claudin-7与intergrinα2相互作用;蛋白印迹技术检测Claudin-7在小鼠不同脏器及C-fos、C-jun、Cox-2蛋白在小鼠肠道中表达,同时检测Claudin-7在人不同分化大肠癌及肝转移组织中的表达情况,分正常人大肠组织、高分化大肠癌组织、低分化大肠癌组织及大肠癌肝转移组织4组,每组20例患者。结果 Claudin-7基因敲除小鼠自出生后第4 d左右停止生长,7 d左右死亡;HE染色以肠道病理变化最为显著,表现为黏膜上皮细胞严重脱落、空泡形成、并有炎性反应细胞浸润,其他脏器病理变化显示除有轻微的中性粒细胞浸润外未见明显的组织损伤;Claudin-7在肠道、胃、肺、膀胱、皮肤、胸腺、肾脏脏器中均有表达,小肠和大肠广泛强表达;Claudin-7和intergrinα2免疫共沉淀,C-fos、C-jun及Cox-2蛋白表达在Claudin-7基因敲除小鼠肠道中显著高表达(P<0.001);Claudin-7蛋白表达随大肠癌的分化降低而降低,在大肠癌肝转移组织中表达显著降低(P<0.001),差异有统计学意义。结论 Claudin-7基因敲除可诱导严重的肠道炎性反应,Claudin-7可能为潜在的抑癌基因。
Objective To investigate the phenotype of Claudin-7 knockout mice （KO） and potential anti-tumor function of Claudin-7 gene. Methods DNA extracted from mouse tails were used to determine mouse genotype by PCR methods, and Claudin-7 protein expression in various tissues was detected by Western blotting, and pathomorphological changes in intestine were observed by HE staining. Interaction between protein Claudin-7 and intergrin c^2 were detected by co-immunoprecipitation method. Claudin-7 protein expression in each organ and protein C-los, C-jun, COX-2 expression in intestine of WT and KO mouse were measured by Western blotting, as well as Claudin-7 expression at different degrees of differentiation of colon cancer tissues. Results Mouse failing to grow could be found 4 days after birth for Claudin-7 knockout mice （KO） , which would die within about 7 days after birth. Claudin-7 could be detected strongly in various tissues such as intestine, stomach, lung, bladder, skin, thymus and kidney, but ubiquitously expressed along the whole intestine including small and large intestine. Moreover, serious intestinal mucosal disruption including severe epithelial cell loss, vacuolation and infiltrated with inflammatory cells such as neutrophil granulocyte could be viewed by light micrographs with H＆E-stained, but there were no obvious changes in other tissues except light inflammatory cells infiltration. C-fos, C-jun and Cox-2 protein expression was upregulated significantly in intestine of Claudin-7 knockout mouse as compared with that in normal mouse （ WT）, as well as Claudin-7 protein expression downregulated in poorly differentiated and liver metastasis tissues of colonic carcinoma. Importantly, Claudin-7 and intergrin c^2 could precipitate together by co-immunoprecipitation （Co-IP） method. Conclusion Mouse intestinal inflammation model was modulated by Claudin-7 knockout successfully, and may be a potential tumor suppressor gene.
出处
《首都医科大学学报》
CAS
2012年第5期569-574,共6页
Journal of Capital Medical University
基金
首都医科大学基础-临床科研合作基金(12JL15)
首都医科大学附属北京世纪坛医院院基金(2012-C13)~~
