Core-shell microsphere structure stimulation: Effects on cell behavior and collagen synthesis for regenerative applications.

The unique structure of core-shell microspheres offers novel strategies for manipulating the sequence of cell contact, as the regenerative process is time-dependent and relies on stimulation provided by the implant materials. In this study, 6 types of core-shell microspheres with hydrophilic core/hydrophobic shell and hydrophobic core/hydrophilic shell configurations were prepared (i.e. PCL@AG, PCL@CS, and PCL@Alg, AG@PCL, CS@PCL, Alg@PCL). Among these, PCL@AG microspheres with an average particle size of 26.97 μm, exhibited high core-shell formation probability of 88.21%, and showed a prolonged degradation rate of 13.12% during 60-day test. Evaluation using in vitro co-culture systems revealed that all microspheres no significant short-term cytotoxicity (live cell ratio ranging from 95.33% to 98.86%). Microspheres with a hydrophobic PCL shell and a hydrophilic natural polymer core (PCL@AG, PCL@CS, PCL@Alg) significantly outperformed their inverse counterparts in promoting cell viability (relative viability 106.55%-156.59%) and migration (PCL@AG, highest wound closure 63.13%) by providing initial stimulation. The group treated by PCL@AG showed the most pronounced effects in promoting the synthesis of type I collagen (34.38 ng/mL) and type III collagen (125.65 ng/mL) as well as COX-2 expression (110.12 pg/mL), while maintaining a relatively low IL-6 level (28.51 pg/mL). The RT-qPCR analysis further confirmed that the PCL-shell/hydrophilic-core structure significantly upregulated the expression of COL1A1 and COL3A1. Dynamic observation of fibroblasts co-cultured with PCL@AG microspheres on a microfluidic chip revealed that the microspheres guided fibroblasts to migrate from the 2D plane onto the microsphere surface, gradually forming a 3D encapsulation that mimics the function of an extracellular matrix scaffold. In summary, core-shell microspheres combined with sequential mechanical stimulation can regulate fibroblast cell behavior. The findings verified the need for a PCL shell to provide essential stimulation to drive appropriate cell proliferation at an early stage, while the hydrophilic core, particularly one with gelling properties at room temperature, was able to maintain continuous stimulation for collagen synthesis and remodeling at a low inflammatory level.