超声靶向微泡破坏介导Ang1基因转染改善犬心肌梗死后左心室同步性的研究

Ultrasound targeted microbubbles destruction-mediated the Angl gene transfection improves left ventricularmechanical synchrony during the progression of myocardial infarction in canines

目的利用超声靶向微泡破坏（UTMD）介导血管生成素1（Ang1）基因转染犬梗死心肌，促进血管新生改善心肌运动同步性，以逆转左室重构治疗心肌梗死。方法21只杂种犬随机平分为3组，每组7只：①对照组（未经处理的健康犬）；②心梗组（心梗造模但未经治疗）；③UTMD组（心梗造模＋UTMD）。1个月后行常规超声测量心脏大小和收缩功能；二维斑点追踪成像（2I）_STI）技术检测左室心肌运动纵向、径向和环向应变达峰时间标准差（Tls—SD、Trs—SD和TCS—SD）和最大差值（T1s—Dif、Trs—Dif和Tes—Dif）作为评价左室运动同步性的指标；心肌组织取材免疫组化CD31和a—SMA分别定位毛细血管和微小动脉；Westernblotting检测Ang1蛋白和钙离子运作关键蛋白肌浆网Ca2＋-ATP酶2a蛋白（SERCA2a）和受磷蛋白（PLB）表达量。比较各组上述指标以评价uTMD介导Angl基因转染改善心梗后左室同步性的治疗效果。结果①三组间常规超声参数比较，心梗组较对照组左室舒张末期内径（LVEDD）、收缩末期内径（LVEsD）、g／e’增大，左室射血分数（LVEF）、e’和E／A减小，uTMD组较心梗组有所恢复，但仍低于对照组（P均〈0．05）；②左室同步性参数比较，Tls—SD、Tls—Dif和Trs—SD在三组间差异均有统计学意义（P均〈0．05），与对照组相比，心梗组不同步程度增加，UTMD组不同步程度降低，而TrsDif、Tcs—SD和Tes—Dif在三组问差异无统计学意义（P〉0．05）；③CD31和a—SMA组化结果显示心梗组较对照组血管密度均明显减低，UTMD组新生血管较心梗组增加（P均〈0．05）；④Westernblotting显示Angl蛋白在UTMD组较对照组和心梗组增加。SERCA2a蛋白在心梗组较对照组明显减低，UTMD后增加，PLB与之相反，差异均有统计学意义（P均〈0．115）。结论经UTMD介导Angl基因转染可以促进梗死心肌周边血管新生、逆转左室重构、改善左室同步性。该心肌运动同步性可能与钙离子运作关键蛋白SERCA2a和PLB表达量相关。

Objective To evaluate the left ventricular synchrony after myocardial infarction （MI） by ultrasound targeted microbubbles destruction （UTMD）-mediated angiogenin 1 （Angl） gene transfection in canine. Methods Twenty-one dogs were divided into three groups （ n = 7 in each group） ：（1）control group （healthy dogs）; MI group （MI dogs without treatment）; UTMD group （MI dogs with UTMD treatment）. One month later, the size and systolic function of heart were measured by echocardiography. The synchronization parameters derived from two dimensional-speckle tracking imaging（2D-STI） included the standard deviation and maximum difference of time to peak strain for all left ventrieular segments （Tls- SD, Trs-SD, Tcs-SD, Tls-Dif, Trs-Dif and Tcs-Dif）. CD31 and ^-SMA were applied for quantifying capillary and arteriolar density. The Angl, SERCA2a and PLB protein were detected by Western blotting. Results One month later, the conventional ultrasonic parameters were compared among three groups, theLVEDD,LVESD and E/e＇ increased and LVEF, e＇ and E/A reduced in MI group than those in control group, all of them partially recovered in UTMD group than those in MI group, but were still lower than those in control group （ P〈0.05） ; （R）The left ventricular synchrony parameters of Tls-SD, Tls Dif and Trs- SD showed significant differences among the three groups（ P 〈0.05）, the degree of dyssynchrony increased in MI group than control group, they were lower in UTMD group than those in MI group. The Tcs-SD, Tcs- Dif and Trs-Dif showed no significant difference among three groups （P 〉 0.05 ）; The immunohistochemistry showed the higher blood vessel density in UTMD group than that in MI group（ P 〈 0.05） ;The relative quantity of Angl was significantly higher in UTMD group. The relative quantity of SERCA2a protein was lower in MI group than that in control group, increased in UTMD group, the trend of PLB was contrary to it. The differences were statistically significant （all P〈0.05）. Conclusions The UTMD-mediated Angl gene transfection can promote angiogenesis after MI, reverse left ventricular remodeling and improve left ventricular synchrony. The myocardial synchrony may be related to the expression of calcium ions key protein SERCA2a and PLB.