Enhancing the Therapeutic Capacity of the MSC Secretome Using Engineered Biomaterials

Mesenchymal stromal cells (MSCs) are under broad investigation to catalyze tissue regeneration and treat numerous diseases, and cell-based approaches are a promising alternative to high concentrations of recombinant growth factors. Despite robust lineage-specific differentiation potential in vitro, MSC function in vivo is largely attributed to their potent secretome composed of a complex mixture of reparative growth factors (GFs). GF secretion is markedly increased when MSCs are formed into spheroids. However, MSC spheroids alone are insufficient to induce robust bone regeneration compared to BMP-2 delivery, often delivered in supraphysiological concentrations. We hypothesize that the regenerative potential of MSC spheroids could be enhanced through 1) synergistic coupling with a complementary cell secretome and 2) engineering GF-sequestering materials for entrapment. Considering the established synergistic relationship between muscle and bone, we first hypothesized that the MSC secretome could enhance the bioactivity of myokines secreted by myoblasts by increasing the concentration and diversity of endogenous GFs present for bone regeneration. We found that the osteogenic potential and GF concentration of conditioned media increased when exposed to myoblasts. When delivered into a rat critical-sized femoral defect using an alginate hydrogel, we observed increased bone formation in defects treated with conditioned media compared to the delivery of MSCs alone. As expected and in agreement with other studies, the majority of GF in our platform eluted within 24 hours, limiting the therapeutic effect of the scaffolds and motivating the critical need for engineered scaffolds that retain bioactive components from the system. We next modified alginate hydrogels with sulfate groups to sequester heparin-binding GFs secreted from MSCs to enhance the potency and availability of the MSC secretome. We confirmed that sulfated alginate hydrogels sequestered a mixture of endogenous biomolecules secreted by entrapped MSCs, thereby prolonging the therapeutic effect of MSC spheroids for tissue regeneration. We further confirmed these growth factors remain bioactive, both indirectly by stimulating limited endothelial cell tubulogenesis from conditioned media and directly by measuring enhanced myoblast infiltration. We further investigated this platform in vivo by implanting it in a rat muscle crush injury. We found the combination of spheroids and sulfated alginate limited early fibrotic response and stimulated neuromuscular junction formation. This platform has the potential to significantly enhance current cellular tissue engineering approaches as well as become a model for investigating growth factor sequestration for other applications