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September 15, 2010 (Vol. 30, No. 16)

Expansion of Hematopoietic Stem Cells

Nanofiber-Based Ex Vivo Method Set Up to Help HSCs Realize Preclinical & Clinical Potential

  • Click Image To Enlarge +
    Figure 1. (A) Nanex fibers under scanning electron microscope (SEM): Typically the fiber diameter ranges from 300 nm to 800 nm. (B) 2,000X, (C) 5,000X, and (D) 10,000X SEM images of hematopoietic stem cells growing on top of the Nanex surface.

    Experimental and clinical transplantation procedures involving hematopoietic stem cells (HSCs) are challenging because of the sparse population of stem cells in adult tissues. In addition, preclinical studies to understand basic stem cell transplantation biology such as clonal selection and the integrity and kinetics of stem cells cannot be conducted using current stem cell isolation or expansion technology.

    For the past two decades investigators have undertaken HSC ex vivo expansion, which has led to multiple clinical studies. No significant clinical benefit has been reported, however. As a result, it is believed  that stem cell expansion technology is not optimum enough to maintain progenitor cell populations without differentiation.

    Increased knowledge of the hematopoietic stem cell niche, new methods to promote progenitor cell expansion without differentiation, and serum-free and animal protein free media for expanded adult HSCs may facilitate preclinical and clinical applications of stem cells.

    Arteriocyte’s Nanex™ Stem Cell Ex vivo Expansion System is for adult HSCs. Nanex, which was originally developed by Hai-Quan Mao at Johns Hopkins University, is a biofunctional nanofiber-based scaffold that partially mimics the bone marrow stem cell niche to provide efficient expansion of human bone marrow, peripheral blood, or umbilical cord blood derived HSCs.

    Nanex is prepared by electrospinning polyethersulfone fibers in nanoscale and then treating the surface with amine groups (Figure 1). The biofunctional surface together with its topographical structure facilitates the proliferation of HSCs while maintaining the stem cell phenotype in a 10-day serum-free and animal protein free culture (Figure 2).

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    Figure 2. Nanex-coated 6-well and 24-well plates for research applications

    Advantages of Nanex include that it is easily scalable to larger culture surfaces and that the cells on the Nanex surface are loosely attached, which makes the harvesting process easier, with high viability without enzymatic detachment.

    Arteriocyte has obtained promising results using Nanex-coated plates for ex vivo expansion of CD133+/CD34+ cells. An over 200-fold expansion of umbilical cord blood (UCB) derived HSCs was observed after 10 days of Nanex culture in a serum-free medium. More than 70% of the total cell population after 10-day expansion maintained the CD34+ phenotype.

    The Arteriocyte team has also demonstrated the differentiation potential of Nanex-expanded HSCs into smooth muscle and endothelial lineages, as well as reticulocytes. Nanex-expanded HSCs successfully reconstituted hematopoiesis in a NOD/SCID mouse vascular injury model at efficiencies higher than that of standard suspension culture.

    Further functional studies using Nanex-expanded HSCs in a rat LAD cardiac infarction model also showed a significant cardiac function improvement. This data  indicates that Nanex-expanded HSCs maintain higher angiogenic and vasculogenic potential.

    Additional studies showed that Nanex-expanded HSCs maintained self-renewal capability for at least 5 cycles (10 days each cycle), with a total of 10 million fold expansion. In addition, it has also been shown that Nanex-expanded HSCs can be genetically modified with various therapeutic genes and growth factors.

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