外加心脏局部稳恒磁场强化SPIO标记脂肪干细胞移植后心肌定植和心肌梗塞治疗作用

Stem cell implantation holds great promise of effective and feasible therapy for myocardial infarction. However, the majority of implanted stem cells are usually trapped and colonized in non-targeted organs such as the lungs, liver, spleen and kidney. As a result, most studies have only shown mild to moderate improvement in left ventricular ejection fraction after cardiac cell therapy. The low cardiac retention of implanted cells is an important reason for the limited therapeutic benefits. Thus, increase of cardiac retention of implanted cells appears a logical approach to improve therapeutic efficacy of cardiac cell therapy. ..Superparamagnetic iron oxide nanoparticles (SPIOs) are dextran-coated iron oxide particles with a diameter of 50-100 nm. Cell labeling is usually accomplished by the endocytosis of SPIO blinding with a transfection agent, such as protamine sulfate or poly-L-lysine. SPIO has been used for in vivo MRI tracking of delivered stem cells. Use of an external magnet has been shown to augment the delivery of SPIO-conjugated drug to a targeted region of the body. Moreover, a magnet near a mass of tumor seeded subcutaneously resulted in approximately three-fold increase in the extravasation of intravenously delivered SPIO-labeled monocytes into the surrounding tumor. Most studies have shown that low-intensity static magnetic field does not affect the viability, growth, proliferation, metabolism and structural morphology of stem cells. Therefore, we hypothesized that the combination of externally static magnetic field and SPIO labeling could significantly increase the cardiac retention of implanted cells. There is the positive relationship between cardiac retention of implanted stem cells and therapeutic benefits in hearts. Thus, development of techniques that can significantly increase cardiac retention of implanted cells is a logical approach to improve the efficacy of cardiac cell therapy. ..Micron-sized iron oxide particles (MPIOs) consist of an iron oxide core encased within an inert divinylbenzene polymer shell, having a diameter approaching or greater than 1μm. MPIO can be taken up by incubating cells with the particles directly. Previous studies showed that the differentiated function and proliferative capacity of mesenchymal and hematopoetic stem cells were not affected by labeling with MPIO. Cells labeled with MPIO generally contained iron levels of 100pg, which is approximately threefold higher than those of SPIO-labeled cells. The mass magnetization of MPIO is significantly greater than that of SPIO, which imply that the movement of MPIO can be readily obtained in the presence of static magnetic filed. Thus, we hypothesized that MPIO-labeled stem cells predisposed to colonize into the targeted hearts with the conrol of static magnetic field.

干细胞移植后多数细胞迁移定植于心脏外非靶向器官。心肌干细胞低定植比率严重限制其心肌梗塞治疗作用的发挥。超顺磁性氧化铁纳米粒子（SPIO）在转染试剂介导下可以安全高效地转运至细胞内。稳恒磁场能够靶向导引SPIO偶联药物或标记单核细胞浓聚于肿瘤组织内发挥抗癌作用。低强度稳恒磁场对干细胞生物学特性无不良影响。本课题调查外加心脏局部稳恒磁场能否导引SPIO标记干细胞心肌靶向定植，进而强化其心肌梗塞治疗作用。微米级超顺磁性氧化铁颗粒（MPIO）能够通过直接胞饮作用进入细胞内。MPIO氧化铁核心直径数十倍于SPIO，具有良好生物相容性、较大饱和磁化强度、标记后较高胞浆铁浓度等优点。本课题也调查MPIO标记干细胞在稳恒磁场导向作用下心肌靶向定植能力是否优于SPIO标记干细胞。希望建立一种简单、安全、高效的磁场导向技术强化干细胞心肌梗塞治疗效率，并探讨MPIO作为磁性载体介导磁场导向下干细胞心肌定植效率。

本项目主要研究目的如下：1）调查稳恒静磁场对脂肪干细胞生物学特性（活力、增殖、DNA遗传物质完整性、干细胞表面抗原表达、细胞因子分泌、脂肪及成骨诱导分化）的影响。2）调查外置心脏局部稳恒磁场能否强化SPIO磁性标记脂肪干细胞心肌定植，并强化干细胞移植后心肌梗塞治疗作用。3）调查慢性心肌梗塞首次灌注和延迟强化MRI模式与组织病理学改变的关系，揭示心肌梗塞MRI高度强化病理生理学机制。4）调查心肌梗塞后侧枝循环形成与心肌梗塞MRI造影剂增强模式的关系。5）调查细胞外小分子造影剂Gd-DTPA和血管内大分子造影剂P792慢性心肌梗塞MRI增强影像学模式的差异。.本项目主要研究发现如下：1）0.5T稳恒静磁场作用7d抑制脂肪干细胞活力、增殖、表面抗原表达、细胞因子分泌、干细胞表面抗原表达、脂肪和成骨诱导分化，但是并未影响DNA遗传物质完整性。2）移植脂肪干细胞能够分化为血管内皮细胞，整合进入新生血管，促进心肌梗塞后血管再生，抑制缺血性细胞凋亡。外置稳恒磁场（0.1T, 7d）对脂肪干细胞生物学特性无不良影响，能够显著增加移植脂肪干细胞心肌定植，促进心肌梗塞后左室收缩功能恢复。3）心肌梗塞后侧枝循环形成有效地输送造影剂进入梗塞心肌组织。梗塞组织内血管通透性只能允许小分子Gd-DTPA造影剂透过血管壁进入细胞外组织间隙，而大分子P792仍旧被限制在血管内空间。心肌细胞坏死、细胞结构消失、疏松胶原纤维网状结构形成均扩大了细胞外空间容积，增加Gd-DTPA分布系数，产生延迟高度信号强化。4）心肌梗塞后进行性侧枝循环恢复明显影响不同阶段（急性、亚急性、慢性）心肌梗塞Gd-DTPA延迟增强MRI模式。首次灌注和延迟强化MRI模式不仅取决于细胞膜破坏和扩大的细胞外间隙容积，也取决于侧枝循环建立的程度。5）慢性心肌梗塞特点为梗塞室壁局部收缩功能和厚度显著减低。小分子Gd-DTPA和大分子P792均不能鉴别慢性心肌梗塞与正常心肌，说明慢性梗塞心肌已经发展到类似正常心肌的侧枝循环。慢性心肌梗塞重构伴随有心肌细胞结构消失和疏松胶原纤维网络形成，这些病理改变扩大了Gd-DTPA细胞外组织间隙分布，产生延迟MRI高度增强模式。