Babaei Mahrokh, Nasernejad Bahram, Sharifikolouei Elham, Shokrgozar Mohammad Ali, Bonakdar Shahin
Department of Chemical Engineering, Amirkabir University of Technology (Tehran Polytechnic), Tehran 15875-4413, Iran.
Department of Applied Science and Technology, Politecnico di Torino, Turin 10129, Italy.
ACS Omega. 2022 Jul 21;7(30):26353-26367. doi: 10.1021/acsomega.2c02206. eCollection 2022 Aug 2.
Physical and chemical parameters that mimic the physiological niche of the human body have an influence on stem cell fate by creating directional signals to cells. Micro/nano cell-patterned polydimethylsiloxane (PDMS) substrates, due to their ability to mimic the physiological niche, have been widely used in surface modification. Integration of other factors such as the biochemical coating on the surface can achieve more similar microenvironmental conditions and promote stem cell differentiation to the target cell line. Herein, we investigated the effect of physical topography, chemical functionalization by acid bone lysate (ABL) nanocoating, and the combined functionalization of the bone proteins' nanocoated surface and the topographically modified surface. We prepared four distinguishing surfaces: plain PDMS, physically modified PDMS by 3D cell topography patterning, chemically modified PDMS with bone protein nanocoating, and chemically modified nano 3D cell-imprinted PDMS by bone proteins (ABL). Characterization of extracted ABL was carried out by Bradford staining and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis, followed by the MTT assay for evaluation of cell viability on ABL-coated PDMS. Moreover, field emission scanning electron microscopy and profilometry were used for the determination of optimal coating thickness, and the appropriate coating concentration was identified and used in the study. The binding and retention of ABL to PDMS were confirmed by Fourier transform infrared spectroscopy and bicinchoninic acid assay. Sessile drop static water contact angle measurements on substrates showed that the combined chemical functionalization and nano 3D cell-imprinting on the PDMS surface improved surface wettability by 66% compared to plain PDMS. The results of ALP measurement, alizarin red S staining, immunofluorescence staining, and real-time PCR showed that the nano 3D cell-imprinted PDMS surface functionalized by extracted bone proteins, ABL, is able to guide the fate of adipose derived stem cellss toward osteogenic differentiation. Eventually, chemical modification of the cell-imprinted PDMS substrate by bone protein extraction not only improved the cell adhesion and proliferation but also contributed to the topographical effect itself and caused a significant synergistic influence on the process of osteogenic differentiation.
模拟人体生理微环境的物理和化学参数通过为细胞创造定向信号来影响干细胞的命运。微/纳米细胞图案化的聚二甲基硅氧烷（PDMS）底物由于能够模拟生理微环境，已被广泛用于表面修饰。整合其他因素，如表面的生化涂层，可以实现更相似的微环境条件，并促进干细胞向目标细胞系分化。在此，我们研究了物理形貌、酸性骨裂解物（ABL）纳米涂层的化学功能化以及骨蛋白纳米涂层表面与形貌修饰表面的联合功能化的影响。我们制备了四种不同的表面：普通PDMS、通过3D细胞形貌图案化进行物理修饰的PDMS、用骨蛋白纳米涂层进行化学修饰的PDMS以及用骨蛋白（ABL）进行化学修饰的纳米3D细胞印迹PDMS。通过Bradford染色和十二烷基硫酸钠聚丙烯酰胺凝胶电泳分析对提取的ABL进行表征，随后进行MTT测定以评估ABL包被的PDMS上的细胞活力。此外，使用场发射扫描电子显微镜和轮廓仪来确定最佳涂层厚度，并确定合适的涂层浓度并用于该研究。通过傅里叶变换红外光谱和二喹啉甲酸测定法确认了ABL与PDMS的结合和保留。对底物进行的静滴静态水接触角测量表明，与普通PDMS相比，PDMS表面的化学功能化与纳米3D细胞印迹相结合使表面润湿性提高了66%。碱性磷酸酶测量、茜素红S染色、免疫荧光染色和实时PCR的结果表明，用提取的骨蛋白ABL功能化的纳米3D细胞印迹PDMS表面能够引导脂肪来源干细胞的命运向成骨分化。最终，通过骨蛋白提取对细胞印迹的PDMS底物进行化学修饰不仅改善了细胞粘附和增殖，还对形貌效应本身有贡献，并对成骨分化过程产生了显著的协同影响。
