兔特发性室性心动过速心室流出道HCN4蛋白表达的实验观察

Expression of HCN4 protein in ventricular outflow tract of rabbit with idiopathic ventricular tachycardia

目的探讨超极化激活的环核苷酸门控离子通道亚型4（HCN4）蛋白在室性心动过速（室速）中的作用。方法数字随机选择新西兰大白兔10只，应用HE染色及免疫组织化学检测中间神经纤维丝（NF-M），观察心室流出道组织是否存在浦肯野纤维。另选择新西兰大白兔40只平均分4组即对照组（SO）、室速组（VT）、室速＋艾司洛尔干预组（VT＋ESM）、室速＋伊伐布雷定干预组（VT＋IVA）。采用免疫组化方法检测心室流出道HCN4蛋白的表达，观察并记录各组诱发室速时所需电压的输出幅值、停止刺激后自发性室速发生的次数及持续时间。结果（1）兔心室流出道存在浦肯野纤维。（2）左、右心室流出道HCN4蛋白的表达VT组与s0组比较明显增多（左室：97．6±16．7与29．0±8．0，P〈0．01；右室：92．7±12．3与26．0±10．8，P〈0．01）；VT＋IVA组与VT组比较明显减少（左室：32．0±9．4与97．6±16．7，P〈0．01；右室：30．8±12．4与92．7±12．3，P〈0．01）。（3）VT＋ESM组和VT＋IVA组与VT组相比，在同等高频刺激的条件下诱发流出道室速所需的输出电压幅值增加（P〈0．01），停止刺激后自发性室速的发生次数减少（P〈0．01），自发性室速持续时间缩短（P〈0．01）。结论（1）心室流出道存在浦肯野纤维，且室速发生时HCN4蛋白表达上调；（2）艾司洛尔及伊伐布雷定可以预防及减少室速的发生，伊伐布雷定作为HCN通道的特异性通道抑制剂效果更强。

Objective To confirm the existence of purkinje fibers in rabbit outflow tract tissue and explore the role of Hyperpolarization-Activated Cyclic Nueleotide-Gated Channel 4 （HCN4） protein in idiopathic ventricular tachycardia. Methods A total of ten New Zealand white rabbits were randomly selected to observe whether there were pukinje fibers in outflow tract by the methods of HE staining and immunohistoehemieal detection of midsize neurofilament （NF-M）. Forty rabbits were randomly divided into four groups ： normal control group （ SO ） , ventricular tachycardia group （VT） , ventrlcular tachycardia ＋ esmolol intervention group （ VT ＋ ESM） and ventricular tachycardia ＋ ivabradine intervention group （ VT ＋ IVA）. Immunobistochemistry was used to detect the expression of HCN4 protein in vcntricular outflow tract ; the required output voltage amplitude was recorded when ventrlcular tachycardia was induced ; the times and duration of spontaneous ventricular tachycardia when stimulation stopped were also recorded for each group. Results （ 1 ） Purkinje fibers existed in the myocardial tissue of rabbit outflow tract. （2） HCN4 protein expression significantly increased in VT group compared with SO group （ left ventricular： 97.6 ±16. 7 vs 29.0 ±8.0, P 〈 0. 01 ; right ventricular： 92. 7 ±12. 3 vs 26.0 ± 10. 8, P 〈 0. 01 ）, the expression of HCN4 protein obviously reduced in VT ＋ IVA group compared with VT group （left ventricular： 32. 0± 9.4 vs 97.6 ± 16. 7, P 〈 0. 01 ; right ventricular ： 30. 8 ± 12. 4 vs 92. 7 ±12. 3, P 〈 0. 01 ）. （3）The output voltage amplitude required to induce the desired ventricular tachycardia in VT ＋ ESM group and VT ＋ IVA groupwere higher than the VT group, under the same high frequency stimulation （P 〈 0.01 ）; the times and duration of spontaneous ventricular tachycardia in VT ＋ ESM group and VT ＋ IVA group significantly reduced when the stimulation stopped, compared with the VT group （both P 〈 0. 01 ）. Conclusions （ 1 ） Purkinje fibers exist in ventricular outflow tract, which may be the histological origin of the ventricular tachycardia. （2） HCN4 protein is up-regulated in ventricular outflow tract when ventricular tachycardia occurs. （3） Esmolol and ivabradine can prevent and reduce the occurrence of ventricular tachycardia, and as the specific inhibitor of the HCN channel, the effect of ivabradine is more obvious.