Polyurethane scaffolds seeded with CD34+ cells maintain early stem cells whilst also facilitating prolonged egress of haematopoietic progenitors

Abstract

We describe a 3D erythroid culture system that utilises a porous polyurethane (PU) scaffold to mimic the compartmentalisation found in the bone marrow. PU scaffolds seeded with peripheral blood CD34+ cells exhibit a remarkable reproducibility of egress, with an increased output when directly compared to human bone scaffolds over 28 days. Immunofluorescence demonstrated the persistence of CD34+ cells within the scaffolds for the entirety of the culture. To characterise scaffold outputs, we designed a flow cytometry panel that utilises surface marker expression observed in standard 2D erythroid and megakaryocyte cultures. This showed that the egress population is comprised of haematopoietic progenitor cells (CD36+GPA−/low). Control cultures conducted in parallel but in the absence of a scaffold were also generally maintained for the longevity of the culture albeit with a higher level of cell death. The harvested scaffold egress can also be expanded and differentiated to the reticulocyte stage. In summary, PU scaffolds can behave as a subtractive compartmentalised culture system retaining and allowing maintenance of the seeded “CD34+ cell” population despite this population decreasing in amount as the culture progresses, whilst also facilitating egress of increasingly differentiated cells.

Differentiation of human induced pluripotent stem cells into erythroid cells

“…In this regard, lncRNAs might allow the generation of functional and mature RBCs from iPSCs [148,149]. Several groups have recently shown that 3D scaffolds such as poly (D, L-lactide-co-glycolide), polyurethane, collagen type I, and porous polyvinyl fluoride resin can mimic the bone marrow niche and improve maintenance of immature hematopoietic cells [150][151][152][153][154]. Although the in vitro maturation of iPSC-derived RBCs still presents several barriers, the cultured erythroid cells from iPSCs provide an important step toward fully defined and animal-free cultivation protocols that can be applied for transfusion medicine [67].…”

Towards manufactured red blood cells for the treatment of inherited anemia

“…45 Alternatively, better biomimicry of the stem cell niche to recapitulate conditions ex vivo has the potential to maintain HSPC stemness for longer periods and therefore increase yield; however, these technologies still require further development. [46][47][48][49][50][51][52] It will be very interesting to see if applications of these innovations can translate into higher yields for largescale mRBC production.…”

BrisSynBio Art-Science Dossier

“…One of the laboratory practices I observed includes growing stem cells within a series of three dimensional scaffold structures that serve as an analogue of bone marrow [25]. Produced from a polymer foam, these tiny synthetic specimens each measure 5 mm cubed.…”