Falk Cardiovascular Research Centre, Stanford School of Medicine, USA.
Regen Med. 2010 Mar;5(2):231-44. doi: 10.2217/rme.09.83.
This study aim to enhance endothelial differentiation of human embryonic stem cells (hESCs) by transduction of an adenovirus (Ad) vector expressing hVEGF(165) gene (Ad-hVEGF(165) ). Purified hESC-derived CD133(+) endothelial progenitors were transplanted into a rat myocardial infarct model to assess their ability to contribute to heart regeneration.
Optimal transduction efficiency with high cell viability was achieved by exposing differentiating hESCs to viral particles at a ratio of 1:500 for 4 h on three consecutive days.
Reverse transcription-PCR analysis showed positive upregulation of VEGF, Ang-1, Flt-1, Tie-2, CD34, CD31, CD133 and Flk-1 gene expression in Ad-hVEGF(165) -transduced cells. Additionally, flow cytometric analysis of CD133, a cell surface marker, revealed an approximately fivefold increase of CD133 marker expression in Ad-hVEGF(165)-transduced cells compared with the nontransduced control. Within a rat myocardial infarct model, transplanted CD133(+) endothelial progenitor cells survived and participated, both actively and passively, in the regeneration of the infarcted myocardium, as seen by an approximately threefold increase in mature blood vessel density (13.62 +/- 1.56 vs 5.11 +/- 1.23; p < 0.01), as well as significantly reduced infarct size (28% +/- 8.2% vs 76% +/- 5.6%; p < 0.01) in the transplanted group compared with the culture medium-injected control. There was significant improvement in heart function 6 weeks post-transplantation, as confirmed by regional blood-flow analysis (1.72 +/- 0.612 ml/min/g vs 0.8 +/- 0.256 ml/min/g; p < 0.05), as well as echocardiography assessment of left ventricular ejection fraction (60.855% +/- 7.7% vs 38.22 +/- 8.6%; p < 0.05) and fractional shortening (38.63% +/- 9.3% vs 25.2% +/- 7.11%; p < 0.05).
hESC-derived CD133(+) endothelial progenitor cells can be utilized to regenerate the infarcted heart.
本研究旨在通过转导表达人 VEGF(165) 基因的腺病毒（Ad）载体增强人胚胎干细胞（hESC）的内皮分化。纯化的 hESC 衍生的 CD133(+)内皮祖细胞被移植到大鼠心肌梗死模型中，以评估其促进心脏再生的能力。
通过将分化的 hESC 暴露于病毒颗粒，比例为 1:500，连续 3 天，每天 4 小时，实现了具有高细胞活力的最佳转导效率。
逆转录-PCR 分析显示，Ad-hVEGF(165)转导的细胞中 VEGF、Ang-1、Flt-1、Tie-2、CD34、CD31、CD133 和 Flk-1 基因表达呈阳性上调。此外，通过流式细胞术分析 CD133 这一细胞表面标志物，与未转导的对照组相比，Ad-hVEGF(165)转导的细胞中 CD133 标志物的表达增加了约 5 倍。在大鼠心肌梗死模型中，移植的 CD133(+)内皮祖细胞存活下来，并积极和被动地参与梗死心肌的再生，成熟血管密度增加了约 3 倍（13.62±1.56 对 5.11±1.23；p<0.01），移植组的梗死面积明显减小（28%±8.2%对 76%±5.6%；p<0.01）。与对照组相比，移植后 6 周时心脏功能明显改善，通过区域性血流分析得到证实（1.72±0.612 ml/min/g 对 0.8±0.256 ml/min/g；p<0.05），以及超声心动图评估左心室射血分数（60.855%±7.7%对 38.22%±8.6%；p<0.05）和缩短分数（38.63%±9.3%对 25.2%±7.11%；p<0.05）。
hESC 衍生的 CD133(+)内皮祖细胞可用于再生梗死的心脏。
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