2型糖尿病大鼠后肢缺血模型的建立及评价

目的建立2型糖尿病大鼠后肢缺血模型并进行评价，为后续的干预实验提供研究平台。方法将15只SD大鼠随机分为正常对照组、糖尿病组及糖尿病后肢缺血组，每组5只。糖尿病组及糖尿病后肢缺血组的10只大鼠均给予高脂饮食喂养4周后，腹腔注射链脲佐菌素（STZ，40mg／kg）以建立2型糖尿病模型。糖尿病后肢缺血组大鼠建模成功后行双侧髂总动脉结扎术以建立后肢缺血模型，正常对照组和糖尿病组大鼠仅分离髂总动脉，不予结扎。2周后对3组大鼠股动脉的起始段行彩色多普勒超声检查，以检测股动脉的血流峰值速度和血流加速时间；取缺血部位的小腿三头肌及大腿股四头肌组织，分别行HE染色及免疫组化SP染色，以观察3组大鼠肌细胞的营养状况及血管再生情况。结果后肢缺血模型建模2周后，正常对照组、糖尿病组和糖尿病后肢缺血组大鼠的血流峰值速度分别为（22．49±3．02）cm／s、（17．36±2．60）cm／s和（11．23±1．26）cm／s，血流加速时间分别为（0．080±0．009）S、（0．120±0．009）S和（0．160±0．020）s，糖尿病后肢缺血组大鼠的股动脉血流峰值速度小于正常对照组和糖尿病组（P〈0．05），而血流加速时间较长（P〈0．05）。HE染色结果显示：糖尿病后肢缺血组大鼠小腿三头肌的结构破坏，有大量炎症细胞浸润，肌肉损伤程度重于正常对照组和糖尿病组。免疫组化sP染色结果显示：糖尿病后肢缺血组大鼠大腿股四头肌的毛细血管密度[（1．40±0．55）个／HPF]小于正常对照组[（6．80±0．84）个／HPF]及糖尿病组[（4．60±0．55）个／HPF]，差异均有统计学意义伊〈O．05）。结论对SD大鼠给予高脂饮食联合小剂量STZ注射可以成功诱导2型糖尿病模型，在此模型基础上结扎髂总动脉可以成功制备糖尿病后肢缺血模型。

Establishment and Evaluation of Ischemic Hind Limb Model in Type 2 Diabetic Rats

Objective To establish and evaluate the hind limb ischemia model in type 2 diabetic rats, and to provide a platform for subsequent intervention experiment. Methods Fifteen SD rats were randomly divided into 3 groups： normal control group, diabetes group, and diabetic ischemia group, each group enrolled 5 rats. The 10 rats of later 2 groups were fed with high fat diet for 4 weeks, and then received intraperitoneal injection of streptozotocin （40 mg/kg） to establish type 2 diabetic model. Three days later, the rats of diabetic ischemia group underwent ligation of the bilateral common iliac artery to establish the hind limb ischemia model, but iliac artery of rats in other 2 groups didn＇t received ligation. Ultrasound and color Doppler flow imaging detection was used to determine the peak velocity and acceleration time of femoral artery in rats of 3 groups in 2 weeks after operation, and triceps tissues and quadriceps tissues were collected to perform HE staining and SP staining for the observation on status of nutrition and vascular regeneration respectively. Results The peak velocity of femoral artery in rats of normal control group, diabetes group, and diabetic ischemia group were （22.49_±3.02） cm/s, （17. 36±2. 60） cm/s, and （11.23±1.26） cm/s, the acceleration time were （0. 080±0. 009） s, （0. 120±0. 009） s, and （0. 160±0. 020） s, the arteriolar density measured by immunohistochemistry SP method were 6. 80_±0. 84/HPF, 4. 60±0. 55/HPF, and 1.40±0. 55/HPF respectively. The peak velocity of femoral artery and arteriolar density of rats in diabetic ischemia group were both lower （P 〈 0. 05）, but acceleration time was longer （P〈 0. 05）. Results of HE staining showed that structure of triceps tissues was damaged, with infiltration of lots of inflamm- atory cells, which was worse than normal control group and diabetes group. Conclusion Method of high-fat diet in combination with small dose of streptozotocin can induce type 2 diabetic rat model, and hind limb ischemia model can be successfully established by ligation of common iliac artery on this model.