冠状静脉窦逆行灌注血管内皮生长因子对兔心肌梗死心室肌细胞L型钙电流的影响

Effects of retrograde coronary sinus perfusion with vascular endothelial growth factor on L-type calcium ion current in rabbit myocardial infarction model

目的研究经冠状静脉窦逆行灌注血管内皮生长因子(vascular endothelial growth factor,VEGF)对兔心肌梗死模型心室肌细胞L型钙离子电流(L-type calcium ion current,Ica-L)的影响,探讨VEGF抗心律失常的细胞学离子机制。方法 45只新西兰大耳大白兔按电脑随机数字法分为3组(每组15只),胸骨正中切开,暴露心脏,Ⅰ组(假手术对照组)冠状静脉窦插管备取静脉血;II组(0.9%氯化钠溶液对照组)结扎左心室支建立心肌梗死模型,冠状静脉窦插管逆灌0.9%氯化钠溶液;III组(VEGF逆灌治疗组)冠状静脉窦插管逆行灌注VEGF蛋白。全程监测心电,记录恶性心律失常发生情况,酶解法分离缺血部位单个心室肌细胞,采用全细胞膜片钳技术记录Ica-L,于术前及术后120 min取冠状静脉窦血检测VEGF、一氧化氮(nitric oxide,NO)、肌钙蛋白T(cardiotroponin T,TnT)浓度。结果术前3组动物VEGF、NO、TnT浓度比较,差异无统计学意义(P>0.05)。Ⅱ组心律失常发生率及电流密度峰值(mV)明显高于Ⅰ组及Ⅲ组,差异有统计学意义(P<0.05)。术后120 min,Ⅱ、Ⅲ组TnT浓度均显著高于Ⅰ组[(4.93±0.57)μg/L vs.(0.72±0.21)μg/L,P<0.05;(2.26±0.49)μg/L vs.(0.72±0.21)μg/L,P<0.05];但Ⅲ组低于Ⅱ组,差异有统计学意义[(2.26±0.49)μg/L vs.(4.93±0.57)μg/L,P<0.05]。Ⅲ组NO[(108.14±14.57)μmol/L vs.(62.98±10.29)μmol/L,P<0.05;(108.14±14.57)μmol/L vs.(67.28±13.45)μmol/L,P<0.05]、VEGF[(834.65±49.23)ng/L vs.(112.34±30.18)ng/L,P<0.05;(834.65±49.23)ng/L vs.(123.45±27.98)ng/L,P<0.05]浓度明显高于Ⅰ、Ⅱ组,差异有统计学意义。结论经冠状静脉窦逆行灌注VEGF能使缺血区心室肌细胞明显增加的Ica-L得到逆转,减少恶性心律失常发生率,其机制可能与增加NO水平有关。

Objectives To investigate effects of retrograde coronary sinus perfusion with vascular endothelial growth factor （VEGF） on L-type calcium ion current（Ica-L）in rabbit myocardial infarction model, so as to explore the basis of VEGF treatment for antiarrhythmia. Methods Forty-five rabbits were randomly divided into three groups. Thorax of rabbits were opened to expose the hearts. Models＇ coronary sinus were only puntured in group I （sham-operated control group） for blood sample taken and no interventions were done. Group Ⅱ （0.9% sodium chloride solution control group） was clamped the main stem of left ventricular coronary, and 0.9% sodium chloride solution was retrograde perfused through coronary sinus. Group Ⅲ （VEGF treatment group）was treated with VEGF. The incidences of malignant arrhythmia were recorded. Single ventricular myocyte were isolated from the epicardial zone of the infarcted region, and whole cell patch clamp technique was used to record Ica-L. Concentrations of VEGF, nitric oxide （NO） and cardiotroponin T （TnT） before operation and 120 min after operation were tested. Results There were no significant difference in concentrations of VEGF, NO and TnT among the three groups preoperative （P〉0.05）. Incidences of malignant arrhythmia and peak of current density （mV） in group Ⅱ were higher than those in group I and group Ⅲ （P〈0.05 or 0.01）. Concentration of TnT was significantly higher in group II and IU than that in group I 120 min after operation [（4.93±0.57）pLg/L vs. （0.72±0.21）μg/L,P 〈0.05; （2.26 ±0.49）/μg/L vs. （0.72 0.21）μg/L,P 〈0.05],and concentration of TnT in group llI was lower than that in group Ⅱ [ （2.26±0.49）μg/L vs. （4.93±0.57）μg/L, P〈0.05 ]. Concentrations of VEGF [（834.65±49.23）ng/L vs. （112.34±30.18） ng/L,P〈0.05; （834.65±49.23） ng/L vs. （ 123.45±27.98） ng/L, P〈0.05 ] and NO [ （ 108.14±14.57） μmol/L vs. （62.98±10.29） p, mol/L, P〈0.05 ; （ 108.14±14.57 ） μmol/L vs. （ 67.28±13.45 ） μmol/L, P〈0.05 ] in group llI were significantly higher than those in group I and I1.