Cord blood-hematopoietic stem cell expansion in 3D fibrin scaffolds with stromal support

Abstract

Expansion of multipotent, undifferentiated and proliferating cord blood (CB)-hematopoietic stem cells (HSC) in vitro is limited and insufficient. Bone marrow (BM) engineering in vitro allows mimicking the main components of the hematopoietic niche compared to conventional expansion strategies. In this study, four different 3D biomaterial scaffolds (PCL, PLGA, fibrin and collagen) were tested for freshly isolated cord blood (CB)-CD34(+) cell expansion in presence of (i) efficient exogenous cytokine supplementation and (ii) umbilical cord (UC)-mesenchymal stem cells (MSC). Cell morphology, growth and proliferation were analyzed in vitro as well as multi-organ engraftment and multilineage differentiation in a murine transplantation model. All scaffolds, except 3D PLGA meshes, supported CB-CD34(+) cell expansion, which was additionally stimulated by UC-MSC support. CB-CD34(+) cells cultured on human-derived 3D fibrin scaffolds with UC-MSC support i) reached the highest overall growth (5 × 10(8)-fold expansion of total nuclear cells after fourteen days and 3 × 10(7)-fold expansion of CD34(+) cells after seven days, p < 0.001), ii) maintained a more primitive immunophenotype for more cell divisions, iii) exhibited superior morphological, migratory and adhesive properties, and iv) showed the significantly highest numbers of engraftment and multilineage differentiation (CD45, CD34, CD13, CD3 and CD19) in BM, spleen and peripheral blood in long-term transplanted NSG mice compared to the other 3D biomaterial scaffolds. Thus, the 3D fibrin scaffold based BM-mimicry strategy reveals optimal requirements for translation into clinical protocols for CB expansion and transplantation.