摘要目的 探讨细胞黏附的重要分子P选择素糖蛋白配体1(PSGL-1)在内皮祖细胞过表达对内皮祖细胞黏附能力以及促新生血管形成的影响.方法 将人PSGL-1 cDNA亚克隆至穿梭质粒,包装扩增病毒.用所产病毒转染大鼠内皮祖细胞,用逆转录聚合酶链反应(RT-PCR)和Western blot等方法检测PSGL-1在内皮祖细胞中的表达,用黏附实验检测PSGL-1的功能.将过表达PSGL-1的内皮祖细胞(实验组)移植裸鼠下肢缺血模型,同时设置对照组(转染空载体的内皮祖细胞移植组)和空白组(注射PBS组),用免疫组织化学方法检测缺血部位新生毛细血管的形成,用激光多普勒检测缺血部位的血流恢复情况.结果 PSGL-1基因重组腺病毒载体构建成功,病毒滴度可达3.1×1011 pfu/ml.该重组腺病毒转染内皮祖细胞后,PSGL-1表达显著增加,内皮祖细胞的黏附活性增高;细胞移植至裸鼠缺血下肢后,缺血部位实验组新生毛细血管数量为[(41.0±2.2)个/高倍视野]较对照组[(21.0±2.5)个/高倍视野]及空白组[(10.0±1.6)个/高倍视野]显著增加(P＜0.01);缺血部位的血流恢复也明显增加[实验组(119.1%±7.0%)、对照组(93.3%±3.0%)、空白组(76.3%±12.0%),P＜0.01].结论 转染PSGL-1可增加内皮祖细胞的黏附活性,移植高表达PSGL-1的内皮祖细胞能促进缺血组织新生血管形成,为改善移植内皮祖细胞治疗缺血性疾病提供了实验依据.

abstractsObjective To explore whether overexpression of P-selectin glycoprotein ligand-1 (PSGL-1) promotes the adhesive ability of endothelial progenitor cells and functionally facilitates neovascularization in mouse model of hindlimb ischemia. Methods Rat endothelial progenitor cells were transfected with recombinant adenovirus vector encoding human PSGL-1. The mRNA and protein expression levels of PSGL-1 were measured by RT-PCR and Western blot, respectively. The effect of overexpression of PSGL-1 in endothelial progenitor cells was analyzed by adherence assay. Histological examination of skeletal muscle sections retrieved from the mouse ischemic hindlimbs was performed, and the hindlimb blood flow was measured by laser Doppler flow meter. Results Adenovirus vector expressing of PSGL-1 gene was successfully constructed with high titer of 3.1 × 1011 pfu/ml. After transfection, PSGL-1 gene was highly expressed in the transfected endothelial progenitor cells. In vitro assay showed that overexpression of PSGL-1 enhanced the adhesive properties of endothelial progenitor cells. When the transfected endothelial progenitor cells were transplanted into the ischemic hindlimb of nude mice, the number of new capillary vessels was (41.0 ±2.2)/HPF compared to that of (21.0 ±2.5) /HPF in the negative control group and (10.0 ±1.6)/HPF in the blank control group (P＜0.01). Furthermore, the blood flow was increased in the experimental group (119.1% ±7.0%), whereas in the negative control group, it was (93.3% ±3.0%)and in the. blank control group it was (76.3%±12.0%) , P＜0.01. Conclusions Overexpression of PSGL-1 enhances the adhesive and angiogenic properties of endothelial progenitor cells. The approach may provide an effective therapeutic strategy to improve the efficiency of cell-based proangiogenic therapy.