Human Cord Cell Hematopoiesis in Three-Dimensional Nonwoven Fibrous Matrices: In Vitro Simulation of the Marrow Microenvironment

Abstract

Current hematopoietic culture systems mainly utilize two-dimensional devices with limited ability to promote self-renewal of early progenitors. In vivo-like three-dimensional (3-D) culture environments might be conducive to regulating stem cell proliferation and differentiation similar to in vivo hematopoiesis. The few 3-D cultures reported in the literature either produced few progenitors or provided little information about microenvironment. In this study, we constructed a 3-D hematopoietic microenvironment composed of nonwoven matrix and human cord blood (CB) cells to simulate the marrow microenvironment and expand cord progenitors. Nonwoven polyethylene terephthalate (PET) fabric with defined microstructure was used as the 3-D scaffold and the PET surface was modified by hydrolysis to improve cell adhesion. Different cell organizations were formed in a 3-D matrix in a developmental manner, from individual cells and cells bridging between fibers to large cell aggregates. Both stromal and hematopoietic cells were distributed spatially within the scaffold. Compared to two-dimensional (2-D) CD34(+) cell culture, 3-D culture produced 30-100% higher total cells and progenitors without exogenous cytokines. With thrombopoietin and flt-3/flk-2 ligand, it supported two- to three-fold higher total cell number (62.1- vs. 24.6-fold), CD34(+) cell number (6.8- vs. 2.8-fold) and colony-forming unit (CFU) number for 7-9 weeks (n = 6), indicating a hematopoiesis pathway that promoted progenitor production. Culture in 3-D nonwoven matrices enhanced cell-cell and cell-matrix interactions and allowed 3-D distribution of stromal and hematopoietic cells. The formation of cell aggregates and higher progenitor content indicated that the spatial microenvironment in 3-D culture played an important role in promoting hematopoiesis. This 3-D culture system can be used as an in vitro model to study stem cell or progenitor behavior, and to achieve sustained progenitor expansion.

The effect of nanofibre surface amine density and conjugate structure on the adhesion and proliferation of human haematopoietic progenitor cells

“…Previous studies indicate that in addition to the signalling molecules supplemented in the medium, substrate surface properties including topographical, mechanical and biochemical properties can significantly influence the proliferation and differentiation of CD34 þ cells [12][13][14][15][16][17][18]. For example, Li et al [12] designed *Author for correspondence (hmao@jhu.edu).…”

Milieu-Adopted in vitro and in vivo Differentiation of Mesenchymal Tissues Derived from Different Adult Human CD34-Negative Progenitor Cell Clones

“…Although most of the multipotent stem cells are routinely isolated from the bone marrow [Pittenger et al, 1999], there is also evidence of circulating CD34-negative, CD45-negative and CD105-positive stem cells in the peripheral blood . Differentiation of stem cells can be induced and directed by various in vitro measures, like the application of appropriate growth factors and cytokines or the co-cultivation of stem cells on tissue-inductive scaffolds [Mackay et al, 1998;Bianchi et al, 2001;Li et al, 2001]. Previous experiments on bone formation and vascularized tissues from mesenchymal stem cells were performed with a number of cultured cells consisting of a heterogeneous cell population without evidence of clonality [Dennis et al, 2002;Harris and Cooper, 2004].…”

Artificial Hematopoietic Stem Cell Niches-Dimensionality Matters

“…With thrombopoietin and flt-3/flk-2 ligand a two-to three-fold higher total cell number was demonstrated compared to 2D culture [86]. 3D-PET scaffolds were also used when surface modified with fibronectin.…”