A surface-tethered model to assess size-specific effects of hyaluronan (HA) on endothelial cells
A surface-tethered model to assess size-specific effects of hyaluronan (HA) on endothelial cells
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DOI：
10.1016/j.biomaterials.2006.09.030
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发表时间：
2007-02-01
期刊：
影响因子：
14
通讯作者：
Ramamurthi, Anand
中科院分区：
文献类型：
--
作者：
Ibrahim, Samir;Joddar, Binata;Ramamurthi, Anand
含有长链（MW > 1 x 10（6）Da）透明质酸（HA）（一种结缔组织GAG）的交联gets（hylans）显示出优异的血管植入生物相容性，但与血管内皮细胞（EC）的相互作用较差。以往的研究表明，在原位破碎的HA紫外光生物激活hylan凝胶，并引起增强EC的反应。由于片段化的HA可能是促炎性的，因此重要的是确定HA片段在hylan表面上的最佳尺寸分布，这将募集和支持正常功能的EC并限制进一步的反应。相关研究表明，HA的外源性模型不一定复制细胞对HA支架的反应。由于支架不能单独基于片段化的HA创建，我们试图通过创建HA系留的培养表面来确定EC对限定片段大小的HA底物的大小特异性反应。使用碳二亚胺反应将HA（1000、200、20 kDa）和低聚物混合物栓系到氨基硅烷（APTMS）处理的玻璃表面上。MALDI-TOF显示HA聚合物含有HA 4- 8聚体，其中4 mer为75 +/-0.4%w/w。免疫荧光，SEM，AFM和XPS分析显示均匀的胺和HA表面。胺s-SDTB分析和HA荧光团辅助碳水化合物电泳（FACE）分别显示表面密度为9 +/- 3个胺基/nm（2）和0.57 +/- 0.44 μ g/cm（2）。HA/HA片段/寡聚体在无血清培养基中孵育超过21天是稳定的。EC在这些表面上的增殖有限，这可能是光滑表面形貌、高阴离子性以及在4聚体的情况下不与主要HA细胞表面受体（CD 44）相互作用的影响。这项工作是重要的，因为它允许测试细胞对由单一大小的HA片段组成的底物的反应，这些HA片段本身不能交联成凝胶。我们实验室的未来工作将使用该模型来评估其他HA寡聚体大小对EC行为的影响。(c)2006爱思唯尔有限公司保留所有权利。
Crosslinked gets (hylans) containing long-chain (MW > 1 x 10(6) Da) hyaluroman (HA), a connective tissue GAG, show exceptional biocompatibility for vascular implantation but poorly interact with vascular endothelial cells (ECs). Previous studies showed in situ fragmentation of HA by UV light to bioactivate hylan gels and elicit enhanced EC responses. Since fragmented HA can be proinflammatory, it is important to define an optimal size distribution of HA fragments on the hylan surface that will recruit and support normally functional ECs and limit ulterior responses. Related studies have shown that exogenous models of HA do not necessarily replicate cell responses to HA scaffolds. Since scaffolds cannot be created based on fragmented HA alone, we sought to determine size-specific responses of ECs to HA substrates of defined fragment sizes by creation of HA-tethered culture surfaces. HA (1000, 200, 20 kDa) and an oligomer mixture were tethered onto an aminosilane (APTMS)-treated glass surfaces using a carbodiimide reaction. MALDI-TOF showed the HA digests to contain HA 4-8mers with a 75 +/- 0.4% w/w of 4mers. Immuno-fluorescence, SEM, AFM and XPS analysis revealed homogeneous amine and HA surfaces. An amine s-SDTB assay and HA fluorophore-assisted carbohydrate electrophoresis (FACE) indicated surface densities of 9 +/- 3 amine groups/nm(2) and 0.57 +/- 0.44 mu g/cm(2), respectively. HA/HA fragments/oligomers were stable over 21 days of incubation in serum-free culture media. EC proliferation on these surfaces resulted was limited, a possible effect of smooth surface topography, high anionicity, and in case of 4mers, non-interaction with primary HA cell-surface receptors (CD44). This work is significant in that it allows testing of cell responses to substrates composed of single-sized fragments of HA that cannot by themselves be cross-linked into a get. Future work in our lab will use this model to assess the effects of other HA oligomer sizes on EC behavior. (c) 2006 Elsevier Ltd. All rights reserved.