Toward transgene-free induced pluripotent stem cells: lessons from transdifferentiation studies.

Abstract Regenerative medicine has received much attention over the years due to its clinical and commercial potential. The excitement around regenerative medicine waxes and wanes as new discoveries add to its foundation but are not immediately clinically applicable. The recent discovery of induced pluripotent stem cells has lead to a sustained effort from many research groups to develop clinically relevant regenerative medicine therapies. A major focus of cellular reprogramming is to generate safe cellular products through the use of proteins or small molecules instead of transgenes. The successful reprogramming of somatic nuclei to generate pluripotential cells capable of embryo development was pioneered over 50 years ago by Briggs and King and followed by Gurdon in the early 1960s. The success of these studies, the cloning of Dolly, and more current studies involving adult stem cells and transdifferentiation provide us with a large repository of potential candidate molecules and experimental systems that will assist in the generation of safe, transgene-free pluripotential cells.

The effect of umbilical cord blood cells on outcomes after experimental traumatic spinal cord injury.

STUDY DESIGN: A cytokine expression profile of umbilical cord blood (UCB) derived multipotential stem cells (MPSC) was produced. We then transplanted MPSCs into a rat model of spinal cord injury (SCI) and assessed neurologic function as well as spinal cord histology. OBJECTIVE: To determine if MPSCs transplanted into a rat model of acute SCI would lead to a beneficial neurologic effect. SUMMARY OF BACKGROUND DATA: Conditioned medium from UCB contains factors that could promote healing of endogenous neural tissues. Previously, our laboratory has demonstrated that UCB hematopoietic cells can develop into MPSCs capable of differentiating into multiple cell types including oligodendrocyte-like cells. METHODS: We cultured MPSCs from UCB cells using fibroblast growth factor 4, stem cell factor and fms-like tyrosine kinase receptor-3 ligand supplemented serum-free medium. Using a cytokine antibody array, we produced a cytokines expression profile of MPSCs. We then transplanted MPSCs into an immunosuppressed rat model of SCI and assessed neurologic function weekly for 6 weeks by the Basso, Beattie, and Bresnahan locomotor test. The spinal cords were examined histologically and lesion areas quantified. RESULTS: We detected elevated levels of cytokines and growth factors with known neuroprotective, angiogenic, and anti-inflammatory effects in the MPSC conditioned media. The SCI rats treated with MPSCs showed a significant improvement in Basso, Beattie, and Bresnahan scores after 6 weeks compared with the group that received vehicle only. Immunohistochemistry revealed transplanted human cells were present in the injured spinal cord after 1 week, but were no longer present by 6 weeks. There was a trend for the lesion size in treated rats to be smaller than that of the control group. CONCLUSION: We conclude that UCB MPSCs improve neurologic function of rats with acute SCI, possibly by the release of factors that reduce secondary injury.

Neural progenitors, neurons and oligodendrocytes from human umbilical cord blood cells in a serum-free, feeder-free cell culture.

We have previously demonstrated that lineage negative cells (Lin(neg)) from umbilical cord blood (UCB) develop into multipotent cells capable of differentiation into bone, muscle, endothelial and neural cells. The objective of this study was to determine the optimal conditions required for Lin(neg) UCB cells to differentiate into neuronal cells and oligodendrocytes. We demonstrate that early neural stage markers (nestin, neurofilament, A2B5 and Sox2) are expressed in Lin(neg) cells cultured in FGF4, SCF, Flt3-ligand reprogramming culture media followed by the early macroglial cell marker O4. Early stage oligodendrocyte markers CNPase, GalC, Olig2 and the late-stage marker MOSP are observed, as is the Schwann cell marker PMP22. In summary, Lin(neg) UCB cells, when appropriately cultured, are able to exhibit characteristics of neuronal and macroglial cells that can specifically differentiate into oligodendrocytes and Schwann cells and express proteins associated with myelin production after in vitro differentiation.