干细胞白血病基因药物对糖尿病膀胱病豚鼠膀胱Cajal间质细胞表面c-kit蛋白表达的影响  

Effects of Stem Cell Leukemia Gene Drug on C-kit Protein Expression in Bladder ICCs of DCP Guinea Pigs

作　　者：李朋 徐浩 申茂磊 赵国立 钱彪[1] 王勤章[1] LI Peng;XU Hao;SHEN Maolei;ZHAO Guoli;QIAN Biao;WANG Qinzhang(Department of Urology Surgery,the First Affiliated Hospital of Medical College,Shihezi University,Shihezi 832000,China;People's Liberation Army 69235 Troops Health Center,Kuitun 833200,China)

机构地区：[1]石河子大学医学院第一附属医院泌尿外科,新疆石河子832000 [2]解放军69235部队卫生队,新疆奎屯833200

出　　处：《医药导报》2020年第10期1335-1339,共5页Herald of Medicine

基　　金：国家自然科学基金资助项目(81360120)。

摘　　要：目的研究干细胞白血病(SCL)基因药物对糖尿病膀胱病(DCP)豚鼠受损膀胱Cajal样间质细胞(ICCs)表面c-kit蛋白表达的影响。方法选取60只健康豚鼠单次腹腔注射1%链脲佐菌素(STZ)200 mg·kg-1构建豚鼠糖尿病膀胱病模型。按随机数字表法将27只糖尿病膀胱病豚鼠随机分为3组:实验组、阳性对照组和空白对照组,每组9只。实验组单次尿道灌注SCL基因重组慢病毒药物(滴度2×107 U)0.2 mL,阳性对照组单次尿道灌注空慢病毒液(不含SCL基因)0.2 mL,空白对照组单次尿道灌注磷酸盐缓冲液(PBS)0.2 mL。每组每次各选取3只豚鼠于灌注后第7,14,28天麻醉后取其膀胱,提取培养DCP豚鼠膀胱ICCs,于倒置显微镜下观察其细胞形态,将用绿色荧光抗体标记的ICCs置于激光共聚焦显微镜下检测ICCs细胞荧光强度,采用Western blotting检测ICCs表面c-kit蛋白的表达。结果SCL基因重组慢病毒在14 d时转染效果最佳,激光共聚焦显微镜下观察到实验组ICCs荧光强度和c-kit蛋白表达较其他两组明显增强(P<0.05),与空白对照组比较,阳性对照组ICCs荧光强度和c-kit蛋白表达差异无统计学意义(P>0.05)。结论慢病毒为载体将SCL基因药物转染入DCP膀胱后能使高糖环境下ICCs膜表面c-kit蛋白表达增强,可能有利于DCP膀胱受损的ICCs功能的恢复。Objective To investigate the effect of stem cell leukemia(SCL)gene therapy on the expression of c-kit protein on the surface of Cajal like stromal cells(ICCs)in the injured bladder of guinea pigs with diabetic bladder disease(DCP).Methods Sixty healthy guinea pigs were administrated with 1%streptozotocin(200 mg·kg-1)in a single intraperitoneal injection to construct diabetic bladder disease model.Twenty-seven guinea pigs with diabetic bladder disease were randomly divided into three groups according to the random number table method:experimental group,positive control group and blank control group,with 9 guinea pigs in each group.SCL gene recombinant lentivirus solution(0.2 mL)with a titer of 2×107 U was injected through the urethra in the experimental group.The positive control group was perfused with 0.2 mL of empty lentivirus carrying SCL gene through the urethra,and the blank control group was perfused with 0.2 mL phosphate buffer(PBS).Three guinea pigs were selected from each group,and the bladder was taken after anesthesia on the 7th,14th,and 28th day after perfusion.ICCs cells in guinea pig DCP bladder were extracted by enzyme digestion.The morphology of ICCs was observed under an inverted microscope.Green fluorescent antibody was used.The green fluorescent labeled ICCs were detected by laser confocal microscopy and the fluorescence intensity of ICCs was determined.Western blot was used to detect the expression of c-kit protein on the surface of ICCs.Results The recombinant lentivirus was successfully transfected into guinea pig DCP bladder,and the transfection effect was the best at 14 days.The fluorescence intensity and c-kit protein expression of ICCs in the experimental group were significantly enhanced compared with the other two groups(P<0.05).No significant difference in fluorescence intensity and c-kit protein expression between the positive control group and the blank control group was found(P>0.05).Conclusion Transfection of SCL gene into DCP bladder can enhance the expression of c-kit protein

关 键 词：干细胞白血病基因药物 糖尿病膀胱病 CAJAL间质细胞 C-KIT蛋白 

分 类 号：R966[医药卫生—药理学]