Functional nanofiber scaffolds with different spacers modulate adhesion and expansion of cryopreserved umbilical cord blood hematopoietic stem/progenitor cells

Abstract

Objective-Nanofiber scaffolds with amino groups conjugated to fiber surface through different spacers (ethylene, butylenes, and hexylene groups, respectively) were prepared and the effect of spacer length on adhesion and expansion of umbilical cord blood hematopoietic stem/progenitor cells (HSPCs) was investigated.Materials and Methods-Electrospun polymer nanofiber scaffolds were functionalized with poly (acrylic acid) grafting, followed by conjugation of amino groups with different spacers. HSPCs were expanded on aminated scaffolds for 10 days. Cell proliferation, surface marker expression, clonogenic potential, and nonobese diabetic (NOD)/severe combined immunodeficient (SCID) repopulation potential of the expanded cells were evaluated following expansion culture.Results-Aminated nanofiber scaffolds with ethylene and butylene spacers showed high-expansion efficiencies (773-and 805-fold expansion of total cells, 200-and 235-fold expansion of CD34 + CD45 + cells, respectively). HSPC proliferation on aminated scaffold with hexylene spacer was significantly lower (210-fold expansion of total cells and 86-fold expansion of CD34 + CD45 + cells), but maintained the highest CD34 + CD45 + cell fraction (41.1%). Colony-forming unit granulocyte-erythrocyte-monocyte-megakaryocyte and long-term culture-initiating cell maintenance was similar for HSPCs expanded on all three aminated nanofiber scaffolds; nevertheless, the NOD/SCID mice engraftment potential of HSPCs expanded on aminoethyl and aminobutyl conjugated nanofibers was significantly higher than that on aminohexyl conjugated nanofibers.Conclusion-This study demonstrated that aminated nanofibers are superior substrates for ex vivo HSPC expansion, which was correlated with the enhanced HSPC adhesion to these aminated nanofibers. The spacer, through which amino groups were conjugated to nanofiber surface, affected the expansion outcome. Our results highlighted the importance of scaffold topography and cellsubstrate interaction to regulating HSPC proliferation and self-renewal in cytokine-supplemented expansion.Offprint requests to: Hai-Quan Mao, Ph.D., Department of Materials Science and Engineering, Johns Hopkins University, 102 Maryland Hall, 3400 N. Charles Street, Baltimore, MD 21218; E-mail: hmao@jhu.edu. In conventional ex vivo expansion culture, HSPCs are generally regarded as suspension cells and numerous protocols implement HSPC suspension cultures in flasks or bags in the presence of various combinations of early acting cytokines [3][4][5][6][7]. The importance of topographical and substrate-immobilized biochemical cues has not been taken into consideration in these culture systems. Suspension culture conditions differ drastically from conditions provided by the natural bone marrow microenvironment, where a complex network of stromal cells and extracellular matrix (ECM) serves as a stem cell niche that regulates HSPC functions such as homing, self-renewal, proliferation, and fate choice [2,8]. A growing body of evidence has suggested the importance of...

Hematopoiesis in Regenerated Bone Marrow Within Hydroxyapatite Scaffold

“…It is particularly important that complex BM components are histologically regenerated in these scaffolds. Several attempts using combinations of cytokines (10), stromal cells (11), osteoblasts (13), nanofiber scaffolds (14), and 3D scaffolds (15) have been made to support, regulate, and expand HSPCs. Although recent studies have suggested a functional classification of BM niches into two kinds, the endsteal (4) and the vascular (8), it seems likely that native BM in vivo is more complex and has longer term functions than those currently identified because actual BM is composed of many types of cells.…”

“…Moreover, some degree of efficacy has been demonstrated for fibronectin, nanofiber scaffolds (14), and 3D scaffolds (15) in the expansion of HSPCs. However, these attempts could not maintain HSPC properties in the long term.…”

A synthetic nanofibrillar matrix promotes in vitro hepatic differentiation of embryonic stem cells and induced pluripotent stem cells

“…NF scaffolds have been shown to recapitulate the structural features of stem cell niche (Lim and Mao, 2009) and have been used for the ex vivo expansion of various types of stem cells such as murine ES cells (Hashemi et al, 2011;Nur-EKamal et al, 2006) and human tissue stem cells. In addition, the ECM was found to deposit as an extensive scaffold on the basal surface of the cells attached to NFs (Shih et al, 2006;Chua et al, 2007;Ma et al, 2008). Importantly, the sNF system has been reported to enhance not only the differentiation of murine ES cells into neural lineages (Lim et al, 2010;Purcell et al, 2012;Xie et al, 2009), but also differentiation from human MSCs into hepatoblasts (Ghaedi et al, 2012;Kazemnejad et al, 2009).…”

Strategies for the chemical and biological functionalization of scaffolds for cardiac tissue engineering: a review

“…In one study, diamino-poly(ethylene glycol) (di-NH2-PEG) was used as a linker molecule to add functional amine groups to previously exposed carboxyl groups, by soaking PLLA nanofibres in NaOH solution [178]. Using a similar strategy, carboxyl functional groups were grafted on polyethersulfone by using a linker molecule (polyacrylic acid) introduced on the polymer surface by photo polymerization [179]. Moreover, polyacrylic acid was used as linker molecule to covalently bond collagen in the form of lamellae or sheets on PMMA and other polymers [180].…”

“…Moreover, polyacrylic acid was used as linker molecule to covalently bond collagen in the form of lamellae or sheets on PMMA and other polymers [180]. It is very important to consider the properties of the linker molecules before using them, as several studies have demonstrated that linker molecule properties influence the interaction between cells and the immobilized biomolecules, such as cell adhesion ligands [179,181,182].…”