Dopamine neurons derived from human ES cells efficiently engraft in animal models of Parkinson’s disease

Abstract

SUMMARY Human pluripotent stem cells (hPSCs) are a promising source of cells for applications in regenerative medicine. Directed differentiation of hPSCs into specialized cells such as spinal motoneurons1 or midbrain dopamine (DA) neurons2 has been achieved. However, the effective use of hPSCs for cell therapy has lagged behind. While mouse PSC-derived DA neurons have shown efficacy in models of Parkinson’s disease (PD)3, 4, DA neurons from human PSCs generally display poor in vivo performance5. There are also considerable safety concerns for hPSCs related to their potential for teratoma formation or neural overgrowth6, 7 Here we present a novel floor plate-based strategy for the derivation of human DA neurons that efficiently engraft in vivo, suggesting that past failures were due to incomplete specification rather than a specific vulnerability of the cells. Midbrain floor plate precursors are derived from hPSCs in 11 days following exposure to small molecule activators of sonic hedgehog (SHH) and canonical WNT signaling. Engraftable midbrain DA neurons are obtained by day 25 and can be maintained in vitro for several months. Extensive molecular profiling, biochemical and electrophysiological data define developmental progression and confirm identity of hPSC-derived midbrain DA neurons. In vivo survival and function is demonstrated in PD models using three host species. Long-term engraftment in 6-OHDA-lesioned mice and rats demonstrates robust survival of midbrain DA neurons, complete restoration of amphetamine-induced rotation behavior and improvements in tests of forelimb use and akinesia. Finally, scalability is demonstrated by transplantation into Parkinsonian monkeys. Excellent DA neuron survival, function and lack of neural overgrowth in the three animal models indicate promise for the development of cell based therapies in PD.

Using iPSC‐derived human DA neurons from opioid‐dependent subjects to study dopamine dynamics

“…2011) as detailed above. DA neurons differentiated from this protocol express FOXA2, LMX1A, TH (tyrosine hydroxylase), Nurr1, VMAT2, and DAT (Sheng et al.…”

“…2011). Indeed, our DA neuronal cultures expressed FOXA2, a unique property of the midbrain dopaminergic neurons as well as Nurr1, Ptx3, DAT, VMAT2, and the DA autoreceptor Drd2.…”

“…2011). Briefly, midbrain floor‐plate precursors were derived from iPSCs without feeder cells after 11 days of exposure to small molecule activators of sonic hedgehog (SHH) and canonical WNT signaling.…”

“…The precursor cells were then further matured into dopaminergic neurons in neurobasal/B27 medium supplemented with ascorbic acid, BDNF, GDNF, TGF β 3, cAMP, and DAPT as described previously (Kriks et al. 2011). Cells were passaged onto poly‐ l ‐ornithine/laminin/fibronectin‐coated plates on days in vitro (DIV) 18 and cultured in the neuronal differentiation media until DIV28 before analyzed for various assays.…”

Induction of pluripotency by defined factors

“…Transplantation of these differentiated cells is expected to result in functional recovery in patients. In fact, the transplantation of differentiated cells derived from iPS cells has already been shown to mitigate the disease phenotypes in mouse models of sickle cell anemia, 77) platelet deficiency, 78) Parkinson's disease 79) and spinal cord injury. 80),81) iPS cells can be generated from patients' own somatic cells.…”