AMO—1脂质体对缺血性心律失常大鼠的治疗研究(1)
[摘要] 目的 为治疗大鼠缺血性心律失常，构建抗体修饰的脂质体，递送AMO-1到缺血心肌。 方法 采用薄膜分散法制备脂质体，并分析脂质体的粒径、电位、包封率和释放率，用流式细胞仪评价心肌细胞对脂质体的摄取，小动物活体成像评价脂质体在大鼠体内的靶向性，通过细胞毒性实验确定给药浓度，采用心电图评价cT-A-LIP抗心律失常作用。 结果 脂质体的粒径均小于120 nm，电位大于-3.8 mV，包封率为(63.0±5.7)%，24 h累积释放率为(32.6±0.7)%。流式细胞仪结果显示，心肌细胞对A-LIP、cT-A-LIP都有一定的摄取率，但有一定差异。小动物活体成像结果显示，cT-A-LIP组药物靶向心脏缺血部位。细胞毒性实验结果显示，脂质体中0.5 μmol/L的AMO-1浓度是cT-A-LIP和A-LIP的最大有效安全剂量。心电图结果显示，给药24 h后ST段明显下降并接近于正常水平。 结论 抗cTnI抗体修饰AMO-1脂质体能够靶向到大鼠的缺血心肌，并且能够缓解缺血性心率失常。
[关键词] 缺血性心律失常；脂质体；AMO-1；抗cTnI抗体
[中图分类号] R943 [文献标识码] A [文章编号] 1673-7210(2016)07(b)-0008-04
[Abstract] Objective To construct an antibody modified liposome to deliver AMO-1 to ischemic myocardium for arrhythmia therapy in rats. Methods The liposomes were prepared by the thin film hydration process. The particle size， zeta potential， encapsulation efficiency and release rate of liposomes were analyzed. The uptake of liposomes by primary myocardial cells was evaluation by flow cytometry. The targeting distribution of liposomes was monitored by the in vivo imaging. The concentration of the drug was determined by the cytotoxicity test. The anti-arrhythmic effects of cT-A-LIP were evaluated by electrocardiograms (ECG). Results The size of liposomes was less than 120 nm， the potential was higher than -3.8 mV， the encapsulation efficiency was (63.0±5.7)%， and the cumulative release rate after 24 hours was (32.6±0.7)%. The results of flow cytometry showed that cT-A-LIP and A-LIP were internalized by primary myocardial cells to some extent， and there were significant differences. The data from the in vivo imaging showed that the drug was accumulated to the foci in the heart. The results of cell toxicity test showed that 0.5 μmol/L AMO-1 concentration in the liposome was the largest safe and effective dose of cT-A-LIP and A-LIP. ECG results showed that the ST segment was obviously decreased and close to the normal level at 24 hours after drug administration. Conclusion The liposomes modified with anti-cTnI antibody as AMO-1 carriers are able to target ischemic myocardium in rats and relieve ischemic arrhythmias.
[Key words] Ischemic arrhythmias； Liposomes； AMO-1； Anti-cTnI antibody
心肌梗死引起的缺血性心律失常是危害人类健康的重要疾病之一，具有较高的死亡率，然而传统的药物对缺血性心律失常的治疗效果并不明显[1-2]。2007年Yang等[3]发现miR-1是治疗缺血性心律失常的靶点。当心脏中miR-1表达上调时心律失常的发生率增加，而将miR-1特异性反义寡核苷酸(AMO-1)局部导入缺血心肌时，心律失常的发生率明显降低[4]。
在纳米载体中脂质体因具有良好的细胞亲和性、靶向性、缓释性和可以提高药物稳定性等优点，成为最有前景的基因药物载体[5-7]。因此，本研究选用脂质体作为药物的载体。心肌肌钙蛋白(cTnI)作为一种特异性心肌标志物，是心肌损伤的标志蛋白，具有高度特异性和敏感性，是临床上早期判断心肌梗死的重要依据[8-10]。心肌缺血患者心肌间质4～48 h cTnI浓度高于正常，这种缺血性心肌cTnI的特异性高表达为本研究提供了构建靶向制剂的特异靶点[11-13]。本研究通过单克隆抗体技术制备心肌特异性抗cTnI单克隆抗体，再将该抗体共价偶联到脂质体表面，利用抗体介导将载药脂质体富集到病理心肌组织中，以实现抗体介导靶向缺血心肌、治疗缺血性心律失常的目的。 (杨伟丽 刘肖莹 于辉)
[关键词] 缺血性心律失常；脂质体；AMO-1；抗cTnI抗体
[中图分类号] R943 [文献标识码] A [文章编号] 1673-7210(2016)07(b)-0008-04
[Abstract] Objective To construct an antibody modified liposome to deliver AMO-1 to ischemic myocardium for arrhythmia therapy in rats. Methods The liposomes were prepared by the thin film hydration process. The particle size， zeta potential， encapsulation efficiency and release rate of liposomes were analyzed. The uptake of liposomes by primary myocardial cells was evaluation by flow cytometry. The targeting distribution of liposomes was monitored by the in vivo imaging. The concentration of the drug was determined by the cytotoxicity test. The anti-arrhythmic effects of cT-A-LIP were evaluated by electrocardiograms (ECG). Results The size of liposomes was less than 120 nm， the potential was higher than -3.8 mV， the encapsulation efficiency was (63.0±5.7)%， and the cumulative release rate after 24 hours was (32.6±0.7)%. The results of flow cytometry showed that cT-A-LIP and A-LIP were internalized by primary myocardial cells to some extent， and there were significant differences. The data from the in vivo imaging showed that the drug was accumulated to the foci in the heart. The results of cell toxicity test showed that 0.5 μmol/L AMO-1 concentration in the liposome was the largest safe and effective dose of cT-A-LIP and A-LIP. ECG results showed that the ST segment was obviously decreased and close to the normal level at 24 hours after drug administration. Conclusion The liposomes modified with anti-cTnI antibody as AMO-1 carriers are able to target ischemic myocardium in rats and relieve ischemic arrhythmias.
[Key words] Ischemic arrhythmias； Liposomes； AMO-1； Anti-cTnI antibody
心肌梗死引起的缺血性心律失常是危害人类健康的重要疾病之一，具有较高的死亡率，然而传统的药物对缺血性心律失常的治疗效果并不明显[1-2]。2007年Yang等[3]发现miR-1是治疗缺血性心律失常的靶点。当心脏中miR-1表达上调时心律失常的发生率增加，而将miR-1特异性反义寡核苷酸(AMO-1)局部导入缺血心肌时，心律失常的发生率明显降低[4]。
在纳米载体中脂质体因具有良好的细胞亲和性、靶向性、缓释性和可以提高药物稳定性等优点，成为最有前景的基因药物载体[5-7]。因此，本研究选用脂质体作为药物的载体。心肌肌钙蛋白(cTnI)作为一种特异性心肌标志物，是心肌损伤的标志蛋白，具有高度特异性和敏感性，是临床上早期判断心肌梗死的重要依据[8-10]。心肌缺血患者心肌间质4～48 h cTnI浓度高于正常，这种缺血性心肌cTnI的特异性高表达为本研究提供了构建靶向制剂的特异靶点[11-13]。本研究通过单克隆抗体技术制备心肌特异性抗cTnI单克隆抗体，再将该抗体共价偶联到脂质体表面，利用抗体介导将载药脂质体富集到病理心肌组织中，以实现抗体介导靶向缺血心肌、治疗缺血性心律失常的目的。 (杨伟丽 刘肖莹 于辉)