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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,40 mg/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(P<0.05)。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。
Objective To establish and evaluate the model of hindlimb ischemia in type 2 diabetic rats and provide a research platform for subsequent intervention experiments. Methods Fifteen SD rats were randomly divided into normal control group, diabetic group and diabetic hindlimb ischemia group, with 5 rats in each group. Diabetic rats and diabetic hind limb ischemic rats were fed with a high-fat diet for 4 weeks and then intraperitoneally injected with streptozotocin (STZ, 40 mg / kg) to establish a type 2 diabetic model. Rats with diabetic hindlimb ischemia model were successfully established bilateral ligation of common iliac artery to establish hindlimb ischemia model, normal control group and diabetic rats only isolated common iliac artery, not ligation. Two weeks later, the color Doppler echocardiography was performed on the initial section of the femoral artery of the three groups to detect the peak velocity of blood flow and the acceleration of blood flow in the femoral artery. The triceps and thigh quadriceps The biceps femoris muscle tissues were stained with HE and SP immunohistochemically to observe the nutritional status and angiogenesis of the three groups. Results After 2 weeks of modeling, the peak velocity of blood flow in the control group, diabetic group and diabetic hind limb ischemic group were (22.49 ± 3.02) cm / s and (17.36 ± 2.60) cm / s, respectively (11.23 ± 1.26) cm / s, and the acceleration time of blood flow were (0.080 ± 0.009) s, (0.120 ± 0.009) s and (0.160 ± 0.020) s respectively. The peak velocity of femoral artery in diabetic hindlimb ischemia group Less than the normal control group and diabetic group (P <0.05), while the blood flow acceleration time was longer (P <0.05). The results of HE staining showed that the structures of triceps brachii of diabetic hindlimb ischemia group were damaged, with a large number of infiltration of inflammatory cells, the degree of muscle damage was heavier than that of normal control group and diabetic group. Immunohistochemical SP staining showed that the capillary density of the thigh quadriceps [(1.40 ± 0.55) / HPF] in diabetic hindlimb ischemia group was smaller than that in the normal control group [(6.80 ± 0.84) / HPF) and diabetes Group 〔(4.60 ± 0.55)〕 / HPF〕, the differences were statistically significant (P <0.05). Conclusion High fat diet combined with low dose STZ injection in SD rats can successfully induce type 2 diabetes mellitus. Based on this model, ligation of common iliac artery can successfully prepare diabetic hindlimb ischemia model.