Amirikia Mehdi, Shariatzadeh Seyed Mohammad Ali, Jorsaraei Seyed Gholam Ali, Soleimani Mehranjani Malek
Department of Biology, Faculty of Science, Arak University, Arak, Iran.
Department of Biology, Faculty of Science, Arak University, Arak, Iran.
Tissue Cell. 2017 Dec;49(6):657-663. doi: 10.1016/j.tice.2017.09.002. Epub 2017 Sep 20.
Cell behaviours such as proliferation and attachment can be affected by the length of pre-incubation period of the scaffolds in the culture medium for long term. The aim of this study was to investigate the long term pre-incubation of 3D silk fibroin scaffolds in complete culture medium on cell attachment and proliferation. After the preparation of silk fibroin scaffolds by the technique of freeze drying, the scaffolds were pre-incubated in complete culture medium for 2 d, 6 d or 10 d before apical papilla stem cells (SCAP) seeding. Modifications of the scaffold surface and wettability were examined by FE-SEM and water contact angle, respectively. Results showed a decrease both in roughness and water contact angle as pre-incubation time increases. DNA measurement after 18h and 10 d cell seeding showed a significant increase of DNA concentration which represents better attachment and proliferation with pre-incubation time increase. Qualitative examination, live&dead assay or H&E staining method after 30h and 10 d cell seeding respectively, indicated that pre-incubation of scaffolds has time dependent effect on cell proliferation and attachment. This suggests that improvement of cell attachment and proliferation may be mediated by differences in the amount of wettability (decreased water contact angle) after exposure of scaffold to culture medium for long term which, in turn, causes more protein adsorption in the surface of silk fibroin scaffold (decreased roughness).
细胞行为，如增殖和附着，可能会受到支架在培养基中长时间预孵育期长度的影响。本研究的目的是调查三维丝素蛋白支架在完全培养基中的长期预孵育对细胞附着和增殖的影响。通过冷冻干燥技术制备丝素蛋白支架后，在接种根尖乳头干细胞（SCAP）之前，将支架在完全培养基中预孵育2天、6天或10天。分别通过场发射扫描电子显微镜（FE-SEM）和水接触角检测支架表面和润湿性的变化。结果表明，随着预孵育时间的增加，粗糙度和水接触角均降低。在接种细胞18小时和10天后进行的DNA测量显示，DNA浓度显著增加，这表明随着预孵育时间的增加，细胞附着和增殖情况更好。分别在接种细胞30小时和10天后进行的定性检查、活死细胞检测或苏木精-伊红（H&E）染色法表明，支架的预孵育对细胞增殖和附着具有时间依赖性影响。这表明，细胞附着和增殖的改善可能是由支架长期暴露于培养基后润湿性的差异（水接触角降低）介导的，进而导致丝素蛋白支架表面更多的蛋白质吸附（粗糙度降低）。
