Nestin阳性细胞在小鼠心肌组织中的表达及分布特征

Expression and Distribution Characteristics of Nestin-positive Cells in Myocardial Tissue of Mouse

【目的】了解Nestin在小鼠正常和损伤心脏组织中的表达及分布特征,探讨Nestin能否作为心肌干细胞的标志物。【方法】通过倒置荧光显微镜观察、RT-PCR和实时荧光定量PCR检测,分析Nestin在胚胎期13.5 d、出生后1 d、7 d、1月、3月的Nestin-GFP转基因小鼠心脏组织中的表达情况;建立小鼠心肌梗死模型,观察损伤后心肌组织中Nestin的表达情况;利用免疫荧光染色法检测小鼠心脏组织中Nestin与多种干细胞标记物的共表达关系。【结果】胚胎13.5 d Nestin表达主要集中于大脑、脊髓、视网膜等部位,但在心脏组织中也可观察到Nestin的绿色荧光,小鼠出生后Nestin的表达随着小鼠年龄的增加而逐渐减少,并且RT-PCR、Q-PCR的结果显示Nestin表达同样呈递减趋势。在梗死部位的心肌组织中可见Nestin阳性的细胞较正常组织显著增多。可见Sca-1、c-kit、Isl-1、Nkx2.5在心肌组织中广泛表达,但是未与Nestin共表达,而Nestin在正常与损伤组织均可与musashi-1、vimentin共表达。【结论】Nestin与心脏发育成熟有密切的关系,可能为神经外胚层来源的一类心肌干细胞。

[Objective] The main aim of this study was to systematically evaluate the expression patterns of the Nestin in the developing or damaged adult heart issue, and probe into whether Nestin can be as a marker of cardiac stem cell. [Methods] Nestin expression was assessed in the embryonic 13.5 d and postnatal 1 d, 7 d, 1 month, 3 months old Nestin-GFP transgenic mouse heart tissue by fluorescence microscopy, real-time quantitative PCR and RT-PCR. Myocardial infaction model was established by ligation of left anterior descending coronary in adult Nestin-GFP mice and the Nestin expression was observed in the myocardium at 7d after injury. Then, the correlation between Nestin and other stem cell markers expression in mouse heart tissue were determined by immunofluoresent assay. [Results] In embryonic 13.5 d, the Nestin mainly expressed in the brain, spinal cord, and the retina, and also can be observed in the heart tissue. After the mouse was born, Nestin expression is gradually reduced with growth, and that was also confirmed by the RT-PCR and Q-PCR analysis. Nestin-positive cells increased significantly in myocardial infarction area compared to the normal tissue. Sca-1, c-kit, Isl-1, and Nkx2.5 were widely expressed in heart tissue, but not co-expressed with Nestin. However, in normal and injured tissue, Nestin was co-expressed with vimentin and musashi-1, neural cell marker. [Conclusion] These results indicate that Nestin expression is highly correlated with cardiac development, and the Nestin-positive myocardial cell might be arise from neural linage cells, which suggest that such cells play an important role in the growth and maintenance of the cardiogenesis and regeneration.