论文下载：自体骨髓干细胞移植治疗心肌梗死的临床研究

自体骨髓干细胞移植治疗心肌梗死的临床研究

http://www.100md.com 2007年2月1日杨承健张郁青徐欣胡世文曹佳宁张月军黄伟金

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     [摘要]目的研究自体骨髓干细胞经冠脉移植对心肌梗死患者心功能的影响及其安全性。方法 6例冠心病伴心肌梗死患者，男5例、女1例，平均年龄62岁(51~75岁)，体外分离培养自体骨髓干细胞2~3周，开通梗死相关“罪犯”血管，通过冠脉转运将骨髓干细胞植入患者的心肌梗死区，移植的骨髓间充质干细胞数为(0.9~3.2)×106个，骨髓单个核细胞数为(1.6~5.3)×106个。术前和术后6个月分别行99mTcMIBI心肌灌注显像、二维超声心动图、心电图和动态心电图检查。结果 6例患者二维超声心动图检查示左室射血分数(LVEF)较术前明显增加[(52.33±8.70)% vs (42.92±12.60)%,P<0.05]，左室舒张期内径(LVDd)较术前明显减少[(54.33±4.84) vs (61.67±6.83),P<0.05]；6例99mTcMIBI检查显示梗死部位心肌灌注明显改善。术中及术后随访6~18个月无心律失常和其他合并症发生。结论进行自体骨髓干细胞经冠脉移植治疗冠心病、心肌梗死具有抑制左室重构、改善心脏功能的作用。

    [关键词]骨髓干细胞;细胞移植;心肌梗死

    Clinical research for treatment of myocardial infarction with autologous bone marrow stem cell transplantation

    YANG Chengjian1,ZHANG Yuqing1,XU Xin1,HU Shiwen1,CAO Jianing1,ZHANG Yuejun1,HUANG Wei1,JIN Yan1,ZHANG Fumin2,YANG Zhijian2

    (1 Department of Cardiology,the Second Hospital of Wuxi,Affiliated to Nanjing Medical University,Wuxi 214000,China;

    2 The First Affiliated Hospital,Nanjing Medical University,Nanjing 210029,China)

    Abstract:Objective To evaluate the efficacy and safety of transplantation of autologous bone marrow mesenchymal stem cells(BMSCs) and mononuclear cells(BMMNCs) for treating myocardial infarction.MethodsSix patients suffered from coronary heart disease with myocardial infarction were enrolled.In 6 patients BMSCs+BMMNCs were transplanted immediately following PCI.Autologous BMSCs+BMMNCs were isolated and cultured for 2~3 weeks.BMSCs [(0.9~3.2)×106 ] and BMMNCs [(1.6~5.3)×106] were transplanted through the intracoroary way.Cardiac functions were determined by 2Dechocardiography，technetium99m methoxyisobutylisonitrile (99mTc MIBI) and ECG Holter monitor before and 6 months after the procedure,respectively.Results Left ventricular ejection fraction(LVEF) significantly increased [(52.33±8.70)% vs (42.92±12.60)%, P<0.05]and left ventricular diastolic diameter(LVDd) reduced [(54.33±4.84) vs (61.67±6.83), P<0.05].Neither obvious arrhythmia nor other complication was observed during 6~18 months follow up in all 6 patients.Conclusions The preliminary study has shown that in the patients suffered from coronary heart disease complicated with myocardiac infarction,transplantation of BMSCs+BMMNCs can improve cardiac function without significant complications and arrhythmia during 6~18 months followup.

    Key words:bone marrow stem cells；cell transplantation; myocardial infarction (Modern Medical Journal,2007,35: 710)

    干细胞移植为冠心病提供了一个崭新的治疗途径。大量动物实验证实，干细胞移植可以改善心功能[18]。自体骨髓干细胞移植因来源丰富，没有免疫排斥作用，因而备受临床医师重视[910]。我们与南京医科大学第一附属医院合作，经医院伦理学委员会批准并获得患者知情同意后，于2003年6月起对冠心病、心肌梗死患者进行自体骨髓干细胞经冠脉移植治疗冠心病，现将已随访6~18个月6例患者的结果报告如下。

    1 对象和方法

    1.1 病例选择6例冠心病、心肌梗死患者，男5例、女1例，平均年龄62岁(51~75岁)；心功能Ⅱ~Ⅳ级，其中Ⅱ级1例，Ⅲ级2例，Ⅳ级3例;前壁心肌梗死5例，下壁心肌梗死1例。

    1.2 方法

    1.2.1 骨髓干细胞分离、培养 0.5%碘伏常规消毒，1%利多卡因局麻，无菌条件下从患者髂后上嵴穿刺抽取骨髓20~40ml，采用Ficoll密度梯度离心法进行骨髓干细胞分离后接种于含20%胎牛血清的IMDM细胞培养液中，37℃、5%CO2孵箱中进行培养；待细胞密度达到70%左右时进行传代培养，体外培养2~3周；移植前收集细胞，悬浮于10ml含肝素、无胎牛血清的IMDM细胞培养液中。骨髓干细胞分离前和干细胞培养结束时均分别行病原学检测。

    1.2.2 术前评价术前1周采用99mTcMIBI心肌灌注显像、二维超声心动图及24h动态心电图评价患者心肌灌注、室壁运动、心功能及心律失常情况。

    1.2.3 骨髓干细胞移植 6例患者均在梗死相关血管经皮冠状动脉介入治疗(PCI)术后进行干细胞移植。移植的骨髓间充质干细胞数为(0.9~3.2)×106个，骨髓单个核细胞数为(1.6~5.3)×106个。采用over the wire 导管将预先准备好的10ml含肝素细胞悬液分3或4次注入梗死相关动脉。每次移植细胞时间3min，间隔3min后进行第2次移植。移植过程中每次扩张球囊3min，严密监测患者有无胸痛情况、心电图及腔内压力改变。6例移植过程均顺利。

    1.2.4 术后随访术后在心脏病监护病房(CCU)监护24~48h，常规监护血压、心电图；术后6、12、24h抽血测定肌钙蛋白I(cTnI)。术后每个月电话随访患者1次，每3个月记录心电图1次，术后6个月随访二维超声心动图、99mTcMIBI心肌灌注显像和24h动态心电图。

    1.3 统计学处理数据以±s表示,组内比较采用配对t检验,P<0.05为差异有统计学意义。

    2 结果

    2.1 6例患者手术前后二维超声心动图检查结果6例患者随访均超过6个月(6~18个月)，心功能从Ⅱ~Ⅳ级提高至Ⅰ~Ⅱ级(I级3例、II级3例)，左室射血分数(LVEF)较术前明显增加[(52.33±8.70)% vs (42.92±12.60)%，P<0.05]，左室舒张末期内径(LVDd)较术前明显减少[(54.33±4.84)mm vs (61.67±6.83)mm，P<0.05](图1)。

    图1 干细胞移植患者术前、术后6个月二维超声心动图心功能对比(左图为术前,右图为术后) 略

    2.2 99mTcMIBI心肌灌注显像术后6个月99mTcMIBI心肌灌注显像示6例患者上述靶部位心肌灌注均得到不同程度的改善(图2)。

    2.3 其他6例患者骨髓干细胞分离培养前后病原学培养均阴性。细胞移植后6、12、24h血cTnI水平均在实验室正常范围，提示细胞移植未引起冠状循环微栓塞及心肌细胞坏死。6例患者术中及术后6~18个月随访中无严重心律失常及其他合并症。

    图2 术前、术后6个月心肌灌注显像对比(第1、3行为术前，第2、4行为术后)略

    3 讨论

    研究证实，移植心肌细胞、骨骼肌卫星细胞、平滑肌细胞和骨髓干细胞[1119]都可以改善心肌梗死后的心功能。前期急性心肌梗死动物实验证实[2025]，移植的自体骨髓干细胞可以分化为带横纹的肌细胞，表达肌球蛋白重链β和连接蛋白Connexin 43，增加梗死区周边毛细血管密度，改善99mTcMIBI心肌灌注显像。我们共进行了6例自体骨髓干细胞心脏移植，并对其进行了随访，6例均在PCI后进行细胞移植。6~18个月的随访结果令人鼓舞：6例患者LVEF提高、LVDd缩小，移植区心肌灌注得到改善，提示移植的细胞在心肌微环境中有增加梗死区新生血管作用，改善了心肌灌注及心功能。晚近研究[2627]认为，尽管骨髓干细胞心肌移植后能否转化为心肌细胞尚存争论，但动物和临床研究[2830]均证实骨髓干细胞移植能够改善心功能，其机制可能为骨髓干细胞移植有新生血管和抑制左室重构作用。我们的临床研究也证实了在心肌灌注显像改善的同时心功能得到不同程度的提高。本研究6例患者随访6~18个月均无心悸感觉，心电图及24h动态心电图未发现严重心律失常及其他不良反应。我们的研究结果与国外的结果[31]相似：自体骨髓干细胞移植并不增加心律失常的发生，相反随着心功能的改善，心律失常的发生有所减少；如果这些乐观的结果能为日后严密的临床试验所证实，自体骨髓干细胞移植将为冠心病病人提供一种崭新的治疗手段。

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    1.南京医科大学附属无锡第二医院心内科，江苏无锡 214000；

    2.南京医科大学第一附属医院，江苏南京 21002920060826

    [作者简介]李江津(1972-)，男，湖北宜昌人，主治医师。

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