In vitro culture and characterization of gene targeted mouse endothelium
In vitro culture and characterization of gene targeted mouse endothelium
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DOI：
10.1046/j.1365-201x.2001.00901.x
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发表时间：
2001-09-01
期刊：
影响因子：
--
通讯作者：
Bullard, DC
中科院分区：
文献类型：
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作者：
Kevil, CG;Bullard, DC
内皮细胞在维持心血管稳态中发挥着至关重要的作用。尽管许多心血管疾病涉及内皮细胞功能障碍，但所涉及的具体细胞和分子机制尚不清楚。我们试图建立一种从基因靶向小鼠中分离内皮细胞的可重复方法，以专门检查内皮病理生理机制。原代主动脉内皮细胞培养物是从野生型和细胞间粘附分子-1 (ICAM-1) 缺陷型小鼠中建立的。通过荧光激活细胞分选法分离小鼠主动脉内皮细胞 (MAEC) 通常会产生纯净、均质的原代培养物。野生型和 ICAM-1 缺陷型内皮细胞形态相似，两种培养物均显示鹅卵石形态和 Dil-Ac-LDL 染色。与野生型MAEC相比，单核细胞对ICAM-1缺陷的主动脉内皮细胞的粘附减少了86%。还使用 YN-1（一种 ICAM-1 阻断抗体）测定单核细胞粘附。 YN-1使单核细胞与野生型主动脉内皮细胞的粘附降低25%，而YN-1并没有进一步降低单核细胞与ICAM-1缺陷的MAEC的粘附。这些数据表明，基因靶向内皮细胞培养物是识别涉及内皮细胞生理学和功能障碍的特定细胞和分子机制的有效手段。
Endothelial cells play a crucial role in maintaining cardiovascular homeostasis. Although many cardiovascular disorders involve endothelial cell dysfunction, the specific cellular and molecular mechanisms involved are not well known. We sought to establish a reproducible method of endothelial cell isolation from gene targeted mice to specifically examine endothelial pathophysiological mechanisms. Primary aortic endothelial cell cultures were established from wild type and intercellular adhesion molecule-1 (ICAM-1) deficient mice. Isolation of mouse aortic endothelial cells (MAEC) by fluorescent activated cell sorting routinely resulted in pure, homogenous, primary cultures. Wild type and ICAM-1 deficient endothelial cell morphology was similar, with both cultures showing cobblestone morphology and Dil-Ac-LDL staining. Monocyte adhesion to ICAM-1 deficient aortic endothelial cells was decreased by 86% as compared with wild type MAEC. Monocyte adhesion was also determined using YN-1, an ICAM-1 blocking antibody. YN-1 decreased monocyte adhesion to wild type aortic endothelial cells by 25%, whereas YN-1 did not further decrease monocyte adhesion to ICAM-1 deficient MAEC. These data demonstrate that gene targeted endothelial cell cultures are an effective means of identifying specific cellular and molecular mechanisms involved in endothelial cell physiology and dysfunction.