核干细胞因子增强间充质干细胞心肌保护作用

目的 研究核干细胞因子（nucleostemin，NS）过表达对脂肪来源间充质干细胞（ADSC）移植后的生存和心肌保护作用的影响。 方法 分离培养小鼠ADSC，分别转染NS腺病毒（ADSC-NS）和对照腺病毒（ADSC-Con），CM-DiI标记ADSC。制作小鼠心肌梗死（MI）模型，随机分为以下4组:①Sham组（与MI相同操作但不结扎冠脉），②MI+vehicle组（冠脉结扎后，在左心室前壁、侧壁和后壁立即心肌点注射总量20 μl PBS），③MI + ADSC-Con组（心肌点注射20 μl含2 × 105新鲜ADSC-Con的悬液），④MI + ADSC-NS组（心肌点注射20 μl含2 × 105的新鲜ADSC-NS的悬液）。MI 3 d后，免疫荧光和流式检测ADSC存活率，TUNEL法检测心肌细胞凋亡。MI 4周后，超声心动图检测心脏结构和功能，MASSON-三色染色测定MI面积，实时定量PCR检测心房钠尿肽（ANP）和脑钠尿肽（BNP）表达水平。在细胞水平将ADSC分为以下各组:①对照腺病毒（ADSC-Con）组，②NS腺病毒（ADSC-NS）组，③对照腺病毒 + 对照（ADSC-Con + Con）组，④NS腺病毒+对照（ADSC-NS + Con）组，⑤对照腺病毒+双氧水（ADSC-Con + H₂O₂）组，⑥ NS腺病毒+双氧水（ADSC-NS + H₂O₂）组。检测NS过表达对ADSC增殖、迁移和抗凋亡作用的影响。 结果 MI 3 d后，与ADSC-Con相比，ADSC-NS显著增加ADSC移植后在心肌组织中的存活率（P < 0.05），显著降低心肌细胞凋亡率（P < 0.05）。MI 4周后，ADSC-NS移植显著增加左心室射血分数，降低左心室内径，降低MI面积和ANP、BNP的mRNA表达。在细胞水平，NS过表达促进ADSC增殖和抗凋亡作用，对ADSC的迁移无显著影响。 结论 NS过表达通过增加ADSC的增殖和抗凋亡能力，增强ADSC移植对心脏的保护作用。

Nucleostemin enhances therapeutic efficacy of mesenchymal stromal cells in treatment of acute myocardial infarction

AIM To investigate the effects of nucleostemin (NS) on the survival and cardioprotective effects of adipose tissue-derived mesenchymal stromal cells (ADSC). METHODS ADSC obtained from C57BL/6J mice were transfected with adenovirus harboring NS (ADSC-NS) or control (ADSC-Con). Mice were subjected to myocardial infarction (MI) or sham operations. Immediately after coronary artery occlusion, MI mice were intramyocardially injected with ADSC-NS (2 × 105 fresh ADSC-NS per mouse) or ADSC-Con (2 × 105 fresh ADSC-Con per mouse), both of which were stained with CM-DiI dye before transplantation. The survival of ADSC and cardiomyocyte apoptosis were determined 3 days after MI. Cardiac functions, fibrosis, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) mRNA expressions were evaluated 4 weeks after MI. In the in vitro studies, cultured ADSC were subjected to the following six groups: (1) ADSC-Con, (2) ADSC-NS, (3) ADSC-Con + Control, (4) ADSC-NS+Control, (5) ADSC-Con + H₂O₂, (6) ADSC-NS + H₂O₂. ADSC proliferation, migration and anti-apoptosis were investigated. RESULTS Both immunostaining and flow cytometric analyses showed that ADSC-NS significantly increased (P < 0.05) ADSC survival in myocardial peri-infarcted area 3 days after injection as compared to ADSC-Con. In addition, ADSC-NS treatment significantly (P < 0.05) reduced cardiomyocyte apoptosis. Echocardiographic studies showed that ADSC-NS, not ADSC-Con, significantly improved the left ventricular ejection fraction (LVEF) 4 weeks after MI. ADSC-NS transplantation significantly mitigated MI-induced fibrosis. In vitro studies showed that NS over-expression significantly increased ADSC growth and reduced H₂O₂-induced ADSC apoptosis. CONCLUSION NS over-expression increases ADSC survival and cardioprotection.