Performance evaluation of a cerebrovascular stent coated with bone marrow mesenchymal stem cells

doi:10.3969/j.issn.2095-4344.2014.34.016

Abstract

Abstract:

BACKGROUND: Generally, the stent surface modification, especially seeding cells, may accelerate or cause stent endothelium, and cause restenosis for prevention of in-stent thrombosis.

OBJECTIVE: To develop the optimal conditions for vascular stents coated with bone marrow mesenchymal stem cells.

METHODS: Cerebrovascular stent was co-cultured with passage 3 bone marrow mesenchymal stem cells from rats at 1×10⁶, 1×10⁷, 1×10⁸, and 1×10⁹/L. Cells on the stents were examined with transmission electron microscopy after 48 hours. A total of 160 male Sprague-Dawley rats were enrolled, among which, 20 rats were as normal control group, and the remaining 140 were used for producing models of ischemic stroke that were randomly sub-divided into seven groups at 8 weeks after modeling: stainless steel stent implanted group, polymer stent group, and different concentrations of cell stent composite groups. After 8 weeks of implantation, the expression of vascular endothelial growth factor in these cells was examined by western blot assay. Rat platelet activation in different groups was determined by flow cytometry.

RESULTS AND CONCLUSION: Implanted stem cells were able to grow adherently on the stainless steel stent wall. When the planting cell concentration was 1×10⁷ cells/L, the cells and organelles were morphologically normal and covered the stent surface well. These coated cells also expressed vascular endothelial growth factor, suggesting that they functioned as endothelial cells, and they also significantly lowered platelet activation. When co-cultured with 1×10⁷/L bone marrow mesenchymal stem cells, the stent was covered well with endothelial-like cells and had significant lower platelet activation in vivo.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

全文链接：

Key words: stainless steel, stem cells, blood platelets

CLC Number:

R318

Tang Ying, Liu Li-na, Wang Qing-min, Liu Chang, Yang Yi-li. Performance evaluation of a cerebrovascular stent coated with bone marrow mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(34): 5503-5508.

Figures/Tables 2

2.1 骨髓间充质干细胞的分离培养与鉴定骨髓间充质干细胞在24 h首次换液后即可见大量贴壁细胞，部分伸出伪足呈短棒状。细胞消化后呈圆形，接种2.0-3.0 h后即贴壁伸展，逐渐恢复为长梭形。传代后的细胞增殖速度较快，不再以集落的方式生长，而是呈均匀分布的纺锤形细胞平均生长，5-7 d即可传代1次，且细胞形态单一均匀，融合后呈典型的极性，漩涡状生长，见图1。流式细胞仪检测体外传代培养第1代间充质干细胞表面标记CD105、CD45、CD44、CD29的表达，结果显示CD44、CD29呈阳性表达，CD105，CD45呈阴性表达，提示分离培养的细胞具有间充质干细胞的性质，见图2。 2.2 电镜观察支架内细胞形态变化电镜结果显示，当细胞浓度为1×106 L-1时，支架内细胞间间隙过大，细胞之间的相互作用弱，暴露的非细胞覆盖面积过大，见图3A。当细胞浓度为1×107 L-1时，细胞间连接紧密，细胞形态及细胞器结构完整，具有一定的生理功能，见图3B。当细胞浓度≥1×108 L-1时，细胞密度过大，细胞产生接触抑制效应，细胞器内产生大量的高密度物质，细胞生理功能受损，见图3C，D。 2.3 支架植入实验中各组血管内皮生长因子的表达蛋白免疫印迹技术检测发现，细胞种植浓度为1×106，1×107，1×108，1×109 L-1时，血管内皮生长因子灰度比值分别为0.104 6±0.034 0，0.312 6±0.053 0，0.201 1±0.025 0，0.101 7± 0.019 0，具体见图4。实验提示当种植浓度为1×107 L-1时，细胞生长良好，在血管内环境的诱导下，具有良好的内皮化趋势。 2.4 支架植入实验中各组血小板活化检测结果流式检测发现，正常对照组、模型组、钢制支架组、高分子材料洗脱支架组、1×106，1×107，1×108，1×109 L-1细胞支架组血小板活化程度分别为(3.31±1.05)%，(72.39±7.26)%，(75.31±5.39)%，(74.88±8.61)%，(39.56±4.54)%，(14.25± 5.17)%，(22.33±5.81)%，(41.14±6.26)%。与正常对照组比较，其余各组中血小板活化程度显著增高(P < 0.01)；与模型组比较，钢制支架组和高分子洗脱支架组血小板活化程度具有上升趋势，但差异无显著性意义，细胞种植支架组血小板活化程度均有下降趋势(P < 0.01)，其中1×107 L-1组下降最为明显，见图5。 "

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