芪参益气滴丸对肾性高血压大鼠心肌钙调节蛋白激酶Ⅱ的影响

Effects of qishenyiqi gutta pills on calcium/calmodulin dependent protein kinase Ⅱ in rats with renalhypertension

目的观察芪参益气滴丸对肾性高血压大鼠左室肥厚及心肌钙调节蛋白激酶Ⅱ（CAMKU）表达的影响，探讨芪参益气滴丸对心肌肥厚干预的机制。方法将50只Wistar大鼠随机数字表法分为5组（n=10），分别为假手术组、对照组、芪参益气滴丸大剂量干预组（大剂量组）、芪参益气滴丸小剂量干预组（小剂量组）、缬沙坦干预组（缬沙坦组）。其中对照组、大剂量组、小剂量组、缬沙坦组行两肾一夹术建立肾血管性高血压大鼠左室肥厚模型。术后第5周起给缬沙坦组灌胃缬沙坦30mg·k-1·d-1；大、小剂量组分别灌胃芪参益气滴丸250、125mg·kg-1·d-1，假手术组和对照组给予相同剂量蒸馏水灌胃，每周检测各组尾动脉收缩压（SBP），持续8周。第12周末终止实验，处死大鼠。取左心室组织检测左心室重量指数（LVMI）、VG染色测量心肌组织胶原容积积分（CVF），ELISA法检测超氧化物歧化酶（SOD）和活性氧自由基（ROS）、用免疫组化和免疫印迹法（Western印迹）检测钙调蛋白激酶II（CAMKⅡ）的表达。结果（1）大、小剂量组及对照组的血压均显著高于假手术组和缬沙坦组[（167．66±11．48）、（166．72±13．51）、（174．34±14．52）mmHg比（119．57±6．30）、（131．80±12．49）mmHg，P〈0．01]。大、小剂量组之间对血压影响差异无统计学意义。（2）与假手术组相比，大、小剂量组及缬沙坦组LVMI为（1．98±0．16）、（2．09±0．14）、（1．97±0．17）g/kg比（1．74±0．17）g／kg，CVF为0．94％±0．22％、2．53％±0．61％、0．81％±0．20％比0．45％±0．13％，显著性升高（P〈0．01，P〈0．05）；但与对照组相比，大、小剂量组及缬沙坦组LVMI为（1．98±0．16）、（2．09±0．14）、（1．97±0．17）g/kg比（2．28±0．28）g／kg，CVF为0．94％4±0．22％、2．53％±0．61％、0．81％±0．20％比4．73％±1．04％，显著性降低（P〈0．01，P〈0．05）。（3）大、小剂量组、缬沙坦组、对照组的CAMKⅡ均显著高于假手术组（65．9％、95．3％、84．8％、160．1％比67．7％），但大、小剂量组、缬沙坦组的CAMKⅡ均显著低于对照组（65．9％、95．3％、84．8％比160．1％），而大、小剂量组与缬沙坦组CAMKⅡ无显著差异。结论芪参益气滴丸对肾性高血压大鼠左室肥厚有延缓作用，其机制可能与其具有抗氧化活性及抑制心肌CAMKⅡ表达有关。

Objective To explore the effects of qishenyiqi gutta pills on myocardial hypertrophy of left ventricle and calcium/calmodulin dependent protein kinase Ⅱ （CAMK Ⅱ ） in rats with renal hypertension and elucidate its intervention mechanism for myocardial hypertrophy. Methods A total of 50 Wistar rats were randomly divided into 5 groups of sham-operation, control, high-dose qishenyiqi gutta pills, low-dose qishenyiqi gutta pills and valsartan （n = 10 each）. The rat model of myocardial hypertrophy with renal hypertension was established by the 2-kidney 1-clip （2KIC） method. The experimental animals were divided into control, high-dose, low-dose and valsartan groups. At Week 5 postoperation, valsartan group received an oral dose of valsartan （30 mg · kg-1 · d-1）, high-dose and low-dose groups took qishenyiqi gutta pills （250 and 125 mg · kg-1 ·d-1 ） while sham-operation and control groups had the same dose ofnormal saline solution. Tail arterial pressure was detected weekly and continued for 8 weeks. At the end of Week 12, the animals were sacrificed to harvest myocardial tissue of left ventricle for detecting left ventricular mass index （LVMI）. The collagen volume fraction （CVF） of myocardium was examined by Van Gieson staining, the activities of superoxide dismutase （SOD） and reactive oxygen species （ROS） were detected by enzyme-linked immunosorbent assay （ELISA） and the expression of CAMKⅡ was detected by immunohistochemistry and Western blot. Results （ 1 ） Blood pressures were significantly higher in high- dose, low-dose and control groups than those in sham-operation and valsartan groups （ （ 167.66± 11.48 ） , （166.72±13.51）, （174.34±14.52） vs（119.57±6.30）, （131.80±12.49）mmHg, P〈0.01）. The changes of blood pressure had no significant difference between high-dose and low-dose groups. （2） LVMI and CVF increased significantly in high-dose, low-dose and valsartan groups versus sham-operation group （LVMI： （1.98±0.16）, （2.09 ±0.14）, （1.97 ±0.17） vs （1.74 ±0.17） g/kg; CVF： 0.94% ±0.22%, 2.53% ±0.61%, 0.81% ±0.20% vs 0.45% ±0. 13%）（P〈0.01, P〈0.05）, but decreased significantly versus control group （ LVMI： （ 1.98 ± 0. 16 ）, （ 2. 09 ± 0. 14 ）, （ 1.97 ± 0. 17 ） vs （ 2. 28 ± 0.28） g/kg; CVF： 0.94% ±0.22%, 2.53% ±0.61%, 0.81% ±0.20% vs 4.73% ± 1.04%） （P〈 0. 01, P 〈 0. 05 ）. （3） The expression of CAMK Ⅱ was significantly higher in high-dose, low-dose, valsartan and control groups than that in sham-operation group （65.9%, 95.3%, 84. 8%, 160. 1% vs 67.7% ）. And it was significantly lower in high-dose, low-dose and valsartan groups than that in control group （65.9% , 95.3% , 84. 8% vs 160. 1% ）. There was no statistical difference among high-dose, low- dose and valsartan groups. Conclusions Qishenyiqi gutta pills may retard myocardial hypertrophy of left ventricle in rats with renal hypertension. And the mechanism is probably be correlated with its antioxidant activity and inhibited expression of myocardial CAMK Ⅱ.