Gelatin methacryloyl-alginate core-shell microcapsules as efficient delivery platforms for prevascularized microtissues in endodontic regeneration.

Combined injectable cell-laden microspheres and angiogenesis approaches are promising for functional vascularized endodontic regeneration. However, advanced microsphere designs and production techniques that benefit practical applications are rarely developed. Herein, gelatin methacryloyl (GelMA)-alginate core-shell microcapsules were fabricated to co-encapsulate human dental pulp stem cells (hDPSCs) and human umbilical vein endothelial cells (HUVECs) based on a coaxial electrostatic microdroplet technique. This technique enables high-throughput production, convenient collection, and minimal material waste. The average diameter of core-shell microcapsules was ∼359 µm, and that of GelMA cores was ∼278 µm. There were higher proliferation rates for hDPSCs and HUVECs co-encapsulated in the GelMA cores than for hDPSCs or HUVECs monoculture group. HUVECs assembled to form 3D capillary-like networks in co-culture microcapsules. Moreover, HUVECs promoted the osteo/odontogenic differentiation of hDPSCs in microcapsules. After 14 days of cultivation, prevascularized microtissues formed in microcapsules that contained abundant deposited extracellular matrix (ECM); no microcapsule aggregation occurred. In vivo studies confirmed that better microvessel formation and pulp-like tissue regeneration occurred in the co-culture group than in hDPSCs group. Thus, an effective platform for prevascularization microtissue preparation was proposed and showed great promise in endodontic regeneration and tissue engineering applications. STATEMENT OF SIGNIFICANCE: Cell-laden microspheres combined with the proangiogenesis approach are promising in endodontic regeneration. We proposed GelMA-alginate core-shell microcapsules generated via the coaxial electrostatic microdroplet (CEM) method, which utilizes a double-lumen needle to allow for core-shell structures to form. The microcapsules were used for co-culturing hDPSCs and HUVECs to harvest large amounts of prevascularized microtissues, which further showed improved vascularization and pulp-like tissue regeneration in vivo. This CEM method and the microcapsule system have advantages of high-throughput generation, convenient collection, and avoid aggregation during long-term culturing. We proposed a high-effective platform for mass production of prevascularized microtissues, which exhibit great promise in the clinical transformation of endodontic regeneration and other applications in regenerative medicine.