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目的:观察参附注射液对老年大鼠肢体缺血再灌注(hind limb ischemia/reperfusion,I/R)损伤后肾功能变化、血红素加氧酶-1(Heme-oxygenase 1,HO-1)表达的影响,并对其保护机制作一初步探讨。方法:64只雄性大鼠,随机分为4组,分别为:假手术对照组、肢体缺血再灌注组、参附干预组、参附+锌原卟啉干预组。假手术对照组:大鼠麻醉后仅分离不夹闭股动脉,分离血管前10 min以7.5ml/kg腹腔注射生理盐水;肢体缺血再灌注组:夹闭股动脉前10 min以7.5ml/kg腹腔注射生理盐水,夹闭股动脉缺血3 h,再灌注4 h;参附干预组:夹闭股动脉前10 min以中等实验剂量7.5ml/kg(常用实验剂量有效范围5-20ml/kg)腹腔注射参附注射液,夹闭股动脉缺血3 h,再灌注4 h;参附+锌原卟啉干预组:术前30min腹腔注射锌原卟啉Ⅸ5mg/kg,余同参附干预组。再灌注完毕后取材,测定肾组织丙二醛(Malondialdehyde,MDA)含量、超氧化物歧化酶(Superoxide Dismutase,SOD)活性;采用免疫组化法测定肾组织HO-1的表达和定量;光镜下观察肾脏病理学改变;取外周静脉血测血清肌酐(Cr)、尿素氮(BUN)含量。结果:与假手术组比较,各肢体缺血再灌注造模组MDA含量均升高;肢体缺血再灌注组、参附+锌原卟啉干预组SOD含量降低;各组血清肌酐(Cr)、尿素氮(BUN)含量升高,肾组织损伤明显(P<0.05或0.01)。与肢体缺血再灌注组比较,参附+锌原卟啉干预组各项指标未见显著差异(P>0.05),参附干预组MDA含量降低,SOD含量升高(P<0.05);血清肌酐(Cr)、尿素氮(BUN)含量降低,肾组织损伤明显减轻(P<0.05)。与参附干预组比较,参附+锌原卟啉干预组MDA含量升高,SOD含量降低(P<0.05);血清肌酐(Cr)、尿素氮(BUN)含量升高,肾组织损伤加重(P<0.05)。与假手术组比较,肢体缺血再灌注组、参附干预组、参附+锌原卟啉干预组肾组织HO-1表达升高,(P<0.05);与肢体缺血再灌注组比较,参附干预组、参附+锌原卟啉干预组肾组织HO-1表达升高(P<0.05)。与参附干预组比较,参附+锌原卟啉干预组HO-1表达未见差异(P>0.05)。结论:肢体缺血再灌注可造成肾脏功能损伤,给予常规剂量7.5ml/kg参附注射液预处理可以减轻肾脏损害程度,这种保护作用可能与参附注射液预处理上调HO-1蛋白在肾组织中的表达,抑制氧自由基生成有关。
Objective: To observe the effect of Shenfu injection on the changes of renal function, heme-oxygenase-1 (HO-1) in aged rats after hind limb ischemia / reperfusion (I / R) The impact of expression, and its protection mechanism to make a preliminary discussion. Methods: Sixty-four male rats were randomly divided into 4 groups: sham operation control group, limb ischemia-reperfusion group, Shenfu intervention group and Shenfu + Zinc protoporphyrin intervention group. Sham-operated control group: rats were anesthetized only isolated without femoral artery separation, 10 min before the separation of blood vessels to 7.5ml / kg intraperitoneal injection of saline; limb ischemia-reperfusion group: clamping the femoral artery before 10 min to 7.5ml / kg intraperitoneal injection of saline, occlusion of the femoral artery ischemia 3 h, reperfusion 4 h; Shenfu intervention group: 10 minutes before clamping the femoral artery to a moderate experimental dose 7.5ml / kg (commonly used experimental dose range 5-20ml / kg) intraperitoneal injection of Shen Fu injection, occlusion of femoral artery ischemia 3 h, reperfusion 4 h; Shen Fu + zinc protoporphyrin intervention group: 30 minutes before surgery, intraperitoneal injection of zinc protoporphyrin Ⅸ 5mg / kg, Intervention group. After reperfusion, the contents of malondialdehyde (MDA) and superoxide dismutase (SOD) in renal tissue were determined. The expression and quantitation of HO-1 in renal tissue were detected by immunohistochemistry. The changes of renal pathology were observed. Serum creatinine (Cr) and blood urea nitrogen (BUN) were measured by peripheral venous blood. Results: Compared with the sham operation group, the content of MDA in each model of ischemia-reperfusion injury was increased. The content of SOD in limb ischemia-reperfusion group and Shenfu + , The content of BUN increased, and the damage of kidney tissue was obvious (P <0.05 or 0.01). Compared with limb ischemia / reperfusion group, the parameters of Shenfu + ZP did not show significant difference (P> 0.05), the content of MDA decreased and the content of SOD increased in Shenfu intervention group (P <0.05) Creatinine (Cr), blood urea nitrogen (BUN) decreased, renal tissue injury significantly reduced (P <0.05). Compared with Shenfu group, the content of MDA and the content of SOD in Shenfu + Zinc Protoporphyrin intervention group were decreased (P <0.05), while the content of serum creatinine (Cr) and urea nitrogen (BUN) P <0.05). Compared with sham group, the expression of HO-1 in limb ischemia-reperfusion group, Shen-Fu intervention group and Shen-Fu + Zinc protoporphyrin group increased (P <0.05), compared with sham-operated group , Shenfu intervention group and Shenfu + Zn protoporphyrin intervention group, the expression of HO-1 in renal tissue increased (P <0.05). Compared with Shenfu group, there was no difference in the expression of HO-1 between Shenfu and ZP group (P> 0.05). Conclusion: Limb ischemia-reperfusion can cause renal dysfunction. Pretreatment with Shenfu Injection 7.5ml / kg can reduce the degree of renal damage. This protective effect may be associated with the increase of HO-1 protein Kidney tissue expression, inhibition of oxygen free radicals generated.