Valeo Michele, Marie Sébastien, Rémy Murielle, Menguy Tiphaine, Le Coz Cédric, Molinari Michael, Feuillie Cécile, Granier Fabien, Durrieu Marie-Christine
Université de Bordeaux, CNRS, Bordeaux INP, CBMN, UMR 5248, F-33600 Pessac, France.
FGHI, Montarnaud, France.
J Mater Chem B. 2024 Dec 11;12(48):12508-12522. doi: 10.1039/d4tb01578a.
Dendrigrafts are multivalent macromolecules with less ordered topology and higher branching than dendrimers. Exhibiting a high density of terminal amines, poly-L-lysine dendrigrafts of the fifth generation (DGL G5) allow hydrogel formation with tailorable crosslinking density and surface modification. This work presents DGL G5 as multifunctional crosslinkers in biomimetic PEG hydrogels to favour the osteogenic differentiation of human mesenchymal stem cells (hMSCs). DGL G5 reaction with dicarboxylic-acid PEG chains yielded amide networks of variable stiffness, measured at the macro and surface nanoscale. Oscillatory rheometry and compression afforded consistent values of Young's modulus, increasing from 8 to more than 30 kPa and correlating with DGL G5 concentration. At the surface level, AFM measurements showed the same tendency but higher values, from approximately 15 to more than 100 kPa, respectively. To promote cell adhesion and differentiation, the hydrogels were functionalised with a GRGDSPC peptide and a biomimetic of the bone morphogenetic protein 2 (BMP-2), ensuring the same grafting concentrations (between 2.15 ± 0.54 and 2.28 ± 0.23 pmols mm) but different hydrogel stiffness. 6 h after seeding on functionalised hydrogels in serum-less media, hMSC showed nascent adhesions on the stiffer gels and greater spreading than on glass controls with serum. After two weeks in osteogenic media, hMSC seeded on the stiffer gels showed greater spreading, more polygonal morphologies and increased levels of osteopontin, an osteoblast marker, compared to controls, which peaked on 22 kPa-gels. Together, these results demonstrate that DGL G5-PEG hydrogel bioactivity can influence the adhesion, spreading and early commitment of hMSCs.
树枝状嫁接物是一种多价大分子，其拓扑结构比树枝状聚合物更无序，分支更多。第五代聚-L-赖氨酸树枝状嫁接物（DGL G5）具有高密度的末端胺基，能够形成具有可定制交联密度和表面修饰的水凝胶。这项工作展示了DGL G5作为仿生聚乙二醇水凝胶中的多功能交联剂，有利于人间充质干细胞（hMSC）的成骨分化。DGL G5与二羧酸聚乙二醇链反应产生了可变刚度的酰胺网络，在宏观和表面纳米尺度上进行了测量。振荡流变学和压缩实验得到了一致的杨氏模量值，从8 kPa增加到超过30 kPa，并与DGL G5浓度相关。在表面水平上，原子力显微镜测量显示出相同的趋势，但值更高，分别从约15 kPa到超过100 kPa。为了促进细胞粘附和分化，水凝胶用GRGDSPC肽和骨形态发生蛋白2（BMP-2）的仿生物进行了功能化，确保相同的接枝浓度（在2.15±0.54和2.28±0.23 pmols/mm之间）但不同的水凝胶刚度。在无血清培养基中接种到功能化水凝胶上6小时后，hMSC在较硬的凝胶上显示出新生粘附，并且比在有血清的玻璃对照上有更大的铺展。在成骨培养基中培养两周后，接种在较硬凝胶上的hMSC与对照相比显示出更大的铺展、更多的多边形形态以及成骨细胞标志物骨桥蛋白水平的增加，在22 kPa凝胶上达到峰值。总之，这些结果表明DGL G5-PEG水凝胶的生物活性可以影响hMSC的粘附、铺展和早期定向。
