G-CSF动员骨髓干细胞向缺血-再灌注肾脏归巢并促进肾脏修复的研究  

Mobilization of bone marrow stem cells by G-CSF to accelerate renal regeneration after ischemia- reperfusion injury in mice

作　　者：谢晓强[1] 吕碧锋[1] 章振保[2] 孔德领[3] 李宗金[4] 徐勇[5] 

机构地区：[1]厦门市第二医院急诊部,福建省厦门361021 [2]厦门市第二医院泌尿外科,福建省厦门361021 [3]南开大学生物活性材料教育部重点实验室 [4]南开大学医学院 [5]天津市泌尿外科研究所

出　　处：《中华急诊医学杂志》2012年第5期491-496,共6页Chinese Journal of Emergency Medicine

基　　金：基金项目：国家重大科学研究计划973项目（2011CB964903）;国家自然科学基金（31071308）

摘　　要：目的观察粒细胞集落刺激因子（G-CSF）是否可以有效动员骨髓干细胞进入外周血并向缺血一再灌注损伤肾脏归巢，提高肾脏修复的效能。方法6周龄全身表达绿色荧光蛋白（GFP）的C57BL／6J转基因小鼠提供骨髓，6—8周龄同种无荧光标记的C57BL／6J小鼠20只作为骨髓受体。骨髓移植前，受体小鼠接受致死剂量的吖放射线137Cs照射，骨髓重建情况经流式细胞仪检测确认。骨髓重建完毕后所有小鼠均接受单侧肾脏缺血一再灌注损伤。实验动物分组：（1）对照组（n=10），术前3d至术后4d，皮下注射生理盐水，0．2ml／d。（2）动员组（n=10），术前3d至术后dd，皮下注射人重组粒细胞集落刺激因子200μg/（kg·d）。干细胞动员效果及向肾脏归巢情况经流式细胞仪检测鉴定。损伤1周后取肾脏标本行HE染色，评估肾小管损伤程度。损伤4周后通过组织切片免疫荧光组织化学方法观察并计数血管内皮细胞数。实验数据中计量资料以均数±标准差（石±s）表示，两组计量资料之间比较采用独立样本t检验。结果G—CSF动员后，分别为Sca-1、c-Kit、CD29、CD34阳性的四群干细胞占外周血非红系细胞的比例均高于对照组（P〈0．05）。损伤1周后，G-CSF动员组的缺血侧肾脏中，骨髓来源并且分别表达Sca-1、c-Kit、CD34的干细胞的比例均高于对照组（P〈0．05）；肾小管损伤程度评分分值低于对照组（P〈0．05）。损伤4周后动员组的。肾脏中CD31阳性的内皮细胞数目高于对照组（P〈0．05）。结论（1）G-CSF可以有效动员骨髓于细胞进入外周血并向缺血肾脏归巢；（2）G—CSF动员可以提高肾脏修复的效能，减轻肾脏组织病理学损伤程度。Objective To investigate mobilization of the bone-marrow-derived stem cell （BMSC） into peripheral blood by granulocyte-colony stimulating factor （G-CSF） to accelerate the renal regeneration. Methods Six-week-old transgenic C57BL/6J mice labeled with green fluorescent protein （GFP） as bone marrow donors and C57BL/6 mice without fluorescence label as recipients （ n = 20 ） of bone marrow transplantation were used. All recipients received lethal dose of 8. 5 Gy total body γ-ray irradiation with 137 Cs before bone marrow transplantation, and the transplantation of bone marrow mononuclear ceils 2 × 10^5 by retrobulbar injection was done two hours later after irradiation. Bone marrow reconstruction after transplantation was proved by flow cytometry five weeks after transplantation. Six weeks after the bone marrow reconstruction completed, left renal pedicles of all mice were cross-clasped for 30 minutes followed by reperfusion to establish the animal model of ischemia-reperfusion injury. Mice were divided into twogroups： （ 1 ） Saline control group （ n = 10）, saline O. 2 ml/day was injected subcutaneously into chimeric mice from 3 days before to 4 days after operation; （2） G-CSF mobilization group （n = 10）, chimeric mice were injected subcutanously with recombinant human G-CSF, 200μg/kg/day, once a day from three days before surgery for a week. On the 1 st day after mobilization, the percentage of stem cell in non-erythroid cells of peripheral blood was detected by using flow cytometry. One week after ischemia, the homing of BMSC to kidney was identified by flow cytometory. Renal tissue sections were stained with Hemotoxylin and Eosin staining method for pathological study, and the degree of renal tubular injury was analyzed by semi- quantitative method of Vyacheslav. Four weeks after ischemia, the differences in degree of renal regeneration between the two groups by analysis the numbers of vascular endothelial cells in the kidney. Results After G-CSF mobilization, the percentage of s

关 键 词：骨髓干细胞 急性肾脏损伤 缺血-再灌注损伤 粒细胞集落刺激因子 修复 

分 类 号：R692[医药卫生—泌尿科学]