不同年龄鼠骨髓干细胞移植治疗心梗后心衰的实验研究

目的：通过对比观察不同年龄鼠骨髓干细胞在体外的生长状态和移入受损心肌后对心肌梗死大鼠心功能的影响,说明年龄对骨髓干细胞的增殖能力和对移植效果的影响。方法：分别分离培养3日龄,1月龄,6月龄,12月龄Wister雄鼠骨髓干细胞,观察细胞生长状态,描记生长曲线。将60只大鼠用结扎冠状动脉前降支的方法制成心肌梗死模型后两周,随机分为三组：Ⅰ组：3日龄组（n=20只）：给予3日龄的5-溴脱氧尿嘧啶（Brdu）标记的鼠骨髓干细胞;Ⅱ组：6月龄组（n=20只）：给予6月龄的Brdu标记的鼠骨髓干细胞;Ⅲ组：对照组（n=20只）：给予培养液。各组均于治疗前及治疗后4周进行心脏超声检查评价心功能改善情况。处死动物取出心脏,进行直接测量后取左心室作石蜡切片,行苏木素-伊红（HE）染色及免疫组化检测,鉴定植入的细胞和心肌、毛细血管再生情况。结果：不同年龄鼠骨髓干细胞培养结果示3日龄及1月龄组增殖能力均高于6月龄及12月龄组,但3日龄与1月龄组之间无统计学差异,12月龄组在原代培养后期即死亡。治疗前超声心动图显示Ⅰ、Ⅱ、Ⅲ组心功能无显著差异（P〉0.05）,治疗之后四周超声心动图检查结果示：Ⅰ、Ⅱ组LVEF、FS、IVST、LVPW和LVESD较治疗前均有明显提高,且显著高于Ⅲ组;Ⅰ组LVEF、FS、IVST、LVPWs和LVESD又明显高于Ⅱ组;Ⅰ、Ⅱ、Ⅲ组LVEDD与治疗前比较无统计学意义。心脏直测结果显示Ⅰ、Ⅱ组的Wh、Wh、Wb、L、I度均较第Ⅲ组有所增加,其中Ⅰ组又明显高于Ⅱ组。免疫组化结果显示：Ⅰ、Ⅱ组于梗死周边均发现Brdu（＋）细胞存在,心肌特异性抗体阳性,且Ⅰ组阳性率明显高于Ⅱ组。毛细血管密度检测结果显示Ⅰ组毛细血管增生情况明显优于Ⅱ组。结论：不同年龄鼠骨髓干细胞的体外增殖能力不同,年龄越小,增殖能力越强。骨髓干细胞移植可以改善心肌梗死后大鼠的心脏功能,增加梗死区毛细血管密度,抑制心室重构,且年龄与移植效果成反比。

Comparative Treatment on Heart Failure of Acute Myocardial Infaction Following Rat Bone Marrow Stem Cell Transplantation of Different Ages

Objective： To study treatment of heart failure of acute myocardial infarction with rat bone marrow stem cell transplantation of different ages and observe cardiac functional alteration,regenerative cadiocyte and capillary.Methods： Separate and culture the MSCs of different ages,observe the living characteristic of the cell,and draw the growth curve.Sixty male rats were made into myocardial infarction model by occlusion of left coronary artery.They were randomly divided into 3 groups： 3-day bone marrow cells transplanting group（Ⅰ,n=20）;6-month group（Ⅱ,n=20） and control group（Ⅲ,n=20）.Each group was measured by ultrasonic cardiography（UCG） before treatment.Mononuclear bone marrow cell was marked with 5-bromodeoxyuridine（Brdu）.All rats survived evaluate improveing of cardiac function after four weeks by treated.To sacrifice aminals and take out hearts,LV remodeling indexes were assessed by dissection and pathological examinations were done for judgement of implanted cells,regenerative cadiocyte and capillary.Results： It turned out that 3-day-old and 1-month-old group had better multiplication capacity than 6-month-old group and 12-month-old group;but there was no significant difference between the 3-day-old group and 1-month-old group.The indexes of UCG showed there was no difference among all the groups before treatment,but after treatment,EF,FS,IVSTd,IVSTs,LVPWs and LVPWd and LVESD of Ⅰ,Ⅱgroups were obviously larger than Ⅲ group.Ⅰgroups was better than Ⅱgroups which explained myocardial contraction force and improve cardiac function after treatment.Ⅰ,Ⅱ,Ⅲ groups LVEDD have no evidently improvement.LV indexed showed： Wh,Wh/Wb,L,I of Ⅰ,Ⅱ groups were bigger than Ⅲ groups and Ⅰ group is better than Ⅱgroup.The pathologic results： through immunohistochemical staining showed positive cells marked by Brdu can be seen at infracted myocardium nearby in Ⅰand Ⅱ groups,but myosin heavy chain（MHC） is negative.Imformation of regenerative capillary showed Ⅰgroup was better than Ⅱ groups.Conclusion： Bone marrow mononuclear cells transplantation can improve myocardial infarction in rat heart function,and the younger the donor bone marrow mononuclear cells on cardiac function can improve the situation much better than older donor bone marrow mononuclear cells.