Center for Biomaterial Development and Berlin-Brandenburg Centre for Regenerative Therapies (BCRT), Institute of Polymer Research, Helmholtz-Zentrum Geesthacht, Teltow, Germany.
Clin Hemorheol Microcirc. 2012;52(2-4):391-401. doi: 10.3233/CH-2012-1613.
As shown in several studies, various properties of biomaterials such as stiffness, surface roughness, chemical composition or the amount of functional groups at the surface can influence adhesion, viability, proliferation and functionalities of cells. The aim of this work was to explore whether a cell-selective effect could be achieved for acrylonitrile-based copolymers containing different contents of positively charged functional groups, which were introduced by incorporation of methacrylic acid-2-aminoethylester hydrochloride (AEMA) units. The p(AN-co-AEMA) copolymers were synthesized by suspension polymerization in water and processed into disk shaped test specimen via a sintering process to ensure the absence of organic solvents in the copolymers. Copolymers with an AEMA content of 1.4, 1.6, and 4.4 mol-% were investigated according to their cell-selective capacity, which should support the adhesion, viability and proliferation of keratinocytes, while the adherence of fibroblasts should rather be disabled. The test samples were seeded with primary human keratinocytes and primary human dermal fibroblasts in mono- as well as in co-cultures. Tissue culture plate polystyrene (TCP) was used to control the physiologic growth of the cells. Density and viability of attached and non-adherent cells were analyzed by live/dead staining, lactate dehydrogenase (LDH) assay and flow cytometry with DAPI staining. For the assured discrimination of adherent cell types in coculture a keratin/vimentin-staining was performed. On copolymers with 4.4 mol-% AEMA adherent keratinocytes in monoculture and cocultured keratinocytes and fibroblasts showed a higher viability, a lower impairment of cell membranes and higher densities of viable cells compared to both other copolymers. For adherent fibroblasts these parameters did not differ between the copolymers and an increasing ratio of keratinocytes to fibroblasts in cocultures were found with increasing AEMA content. The results showed that keratinocytes and fibroblasts can be influenced by copolymers with different contents of positively charged functional groups. Since the tendency of a better adherence and viability of keratinocytes with increasing amounts of positively charged functional groups was shown, the potential enhancement by further increase of the amount of positively charged functional groups shall be tested in a future study.
如图所示，生物材料的各种性质，如硬度、表面粗糙度、化学成分或表面官能团的数量，都可以影响细胞的黏附、活力、增殖和功能。本工作旨在探索是否可以通过引入带正电荷的官能团来实现丙烯腈基共聚物的细胞选择性，该共聚物是通过在水相中悬浮聚合并通过烧结工艺加工成圆盘状测试样品来制备的，以确保共聚物中不存在有机溶剂。研究了含有 1.4、1.6 和 4.4 mol-%AEMA 单元的 p(AN-co-AEMA)共聚物，根据其细胞选择性，这些共聚物应该支持角质形成细胞的黏附、活力和增殖，而纤维母细胞的黏附应该受到抑制。测试样品与原代人角质形成细胞和原代人真皮成纤维细胞一起在单培养和共培养中接种。组织培养板聚苯乙烯（TCP）用于控制细胞的生理生长。通过活/死染色、乳酸脱氢酶（LDH）测定和 DAPI 染色的流式细胞术分析贴壁和非贴壁细胞的密度和活力。为了确保在共培养中区分黏附细胞类型，进行了角蛋白/波形蛋白染色。在含有 4.4 mol-%AEMA 的共聚物上，与其他两种共聚物相比，在单培养和共培养中的角质形成细胞具有更高的活力、更低的细胞膜损伤和更高的活细胞密度。对于黏附的成纤维细胞，这些参数在共聚物之间没有差异，并且随着共培养中角质形成细胞与成纤维细胞比例的增加，发现共聚物中带正电荷的官能团的含量增加。结果表明，不同带正电荷官能团含量的共聚物可以影响角质形成细胞和纤维母细胞。由于显示出随着带正电荷官能团数量的增加，角质形成细胞具有更好的黏附性和活力的趋势，因此应在未来的研究中测试进一步增加带正电荷官能团数量的潜在增强效果。
