Activin A directs striatal projection neuron differentiation of human pluripotent stem cells

Abstract

The efficient generation of striatal neurons from human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) is fundamental for realising their promise in disease modelling, pharmaceutical drug screening and cell therapy for Huntington's disease. GABAergic medium-sized spiny neurons (MSNs) are the principal projection neurons of the striatum and specifically degenerate in the early phase of Huntington's disease. Here we report that activin A induces lateral ganglionic eminence (LGE) characteristics in nascent neural progenitors derived from hESCs and hiPSCs in a sonic hedgehog-independent manner. Correct specification of striatal phenotype was further demonstrated by the induction of the striatal transcription factors CTIP2, GSX2 and FOXP2. Crucially, these human LGE progenitors readily differentiate into postmitotic neurons expressing the striatal projection neuron signature marker DARPP32, both in culture and following transplantation in the adult striatum in a rat model of Huntington's disease. Activin-induced neurons also exhibit appropriate striatal-like electrophysiology in vitro. Together, our findings demonstrate a novel route for efficient differentiation of GABAergic striatal MSNs from human pluripotent stem cells.

Transplantation in HD: Are We Transplanting the Right Cells?

“…The numbers of DARPP-32-positive neurons yielded in vitro ranged from 20 to 50%, depending on the PSC line being assessed, and expression of striatal markers calbindin and CTIP2 was also reported. Although no functional effect was seen in vivo after transplantation of these cells, the striatal neuronal differentiation reported was considerably better than in previous studies [75]. Most graft-derived cells expressed GABA, and DARPP-32-positive neurons comprised 49% of total graft-derived cells.…”

“…Specifically, addition of both noggin and SB431542 (a BMP inhibitor and Activin/Nodal inhibitor, respectively) was shown to increase the yield of cells expressing the neural markers Pax6, Foxg1 and Sox1, whilst expression of the pluripotent marker Oct4 decreased [72]. This method is now widely used as the first stage in the generation of neural cells from PSCs and has successfully been applied to the initial stages of striatal differentiation protocols [73][74][75] (discussed below, and see Table 1). …”

“…A novel striatal conversion protocol, still utilising dual SMAD inhibition for the initial neural induction phase, but introducing Activin A in the patterning stage and reporting the redundance of SHH, resulted in DARPP-32-positive neurons after 36-40 days in vitro [75]. The numbers of DARPP-32-positive neurons yielded in vitro ranged from 20 to 50%, depending on the PSC line being assessed, and expression of striatal markers calbindin and CTIP2 was also reported.…”

“…Other striatal phenotypes observed were calbindin, FoxP2, dopamine receptor D2 and substance P. The proliferative marker, Ki-67, was detected at 4-week post-transplantation, but was absent from grafts at 16-week post-transplantation. Furthermore, the largest graft comprised approximately 500,000 cells (determined by assessment of human nuclei immunostaining), at 16-week post-transplantation, following engraftment of 400,000 cells per graft [75].…”

Stem Cells in Neuroendocrinology

“…Clinical trials are being planned in the US, Japan and Sweden, which have led to the formation of G-Force PD, a global effort to coordinate hPSC-based cell therapy efforts in PD (Barker et al 2015). Another neurodegenerative disease being targeted is Huntington's disease where several protocols have been published to generate authentic, striatal medium spiny neurons and where there is some initial evidence of efficacy in preclinical models (Arber et al 2015;Delli Carri et al 2013;Ma et al 2012). Finally, several promising strategies are under development using glial cells.…”