BMP4 Sufficiency to Induce Choroid Plexus Epithelial Fate from Embryonic Stem Cell-Derived Neuroepithelial Progenitors

Abstract

Choroid plexus epithelial cells (CPECs) have essential developmental and homeostatic roles related to the cerebrospinal fluid (CSF) and blood-CSF barrier they produce. Accordingly, CPEC dysfunction has been implicated in many neurological disorders, such as Alzheimer’s disease, and transplant studies have provided proof-of-concept for CPEC-based therapies. However, such therapies have been hindered by the inability to expand or generate CPECs in culture. During development, CPECs differentiate from preneurogenic neuroepithelial cells and require Bone Morphogenetic Protein (BMP) signaling, but whether BMPs suffice for CPEC induction is unknown. Here we provide evidence for BMP4 sufficiency to induce CPEC fate from neural progenitors derived from mouse embryonic stem cells (ESCs). CPEC specification by BMP4 was restricted to an early time period after neural induction in culture, with peak CPEC competency correlating to neuroepithelial cells rather than radial glia. In addition to molecular, cellular, and ultrastructural criteria, derived CPECs (dCPECs) had functions that were indistinguishable from primary CPECs, including self-assembly into secretory vesicles and integration into endogenous choroid plexus epithelium following intraventricular injection. We then used BMP4 to generate dCPECs from human ESC-derived neuroepithelial cells. These findings demonstrate BMP4 sufficiency to instruct CPEC fate, expand the repertoire of stem cell-derived neural derivatives in culture, and herald dCPEC-based therapeutic applications aimed at the unique interface between blood, CSF, and brain governed by CPECs.

The Choroid Plexus and Cerebrospinal Fluid: Emerging Roles in Development, Disease, and Therapy

“…Genetic lineage analyses indicate that CPECs originate from the roof plate (Awatramani et al, 2003;Currle et al, 2005;Hunter and Dymecki, 2007), a well-known signaling center in early CNS development. Roof plate cells differentiate directly into CPECs at all four ChP sites, although in the telencephalon, most CPECs are induced indirectly by the roof plate (Bailey, 1915;Currle et al, 2005), and multiple studies implicate bone morphogenetic proteins as the instructive roof plate-derived molecules for CPEC fate (Hebert et al, 2002;Cheng et al, 2006;Fernandes et al, 2007;Watanabe et al, 2012). CPEC differentiation is evident before definitive neurogenesis or ependymal differentiation ensues, and in vivo and in vitro mouse studies indicate that CPEC competency is restricted to early-stage neuroepithelial cells in the telencephalon and roof plate regions (Thomas and Dziadek, 1993;Watanabe et al, 2012).…”

“…(Kim et al, 2010;Lehtinen et al, 2011) or the ciliary gene Arl13b (Higginbotham et al, 2013), they share a striking deficiency in IGF signaling receptors and decreased IGF signaling pathway activation on the apical surface of neural stem cells. Recent evidence indicates that IGF1 binding to the IGF1R on primary cilia of radial glial progenitor cells accelerates G 1 -S progression and ciliary resorption by a noncanonical G␤␥ signaling pathway (Yeh et al, 2013). Thus, the intrinsic regulation of apical morphology as well as ciliogenesis may be crucial for the capacity of apical progenitors to access and respond to CSF-distributed signals.…”

“…2). Efforts to assemble the critical mass of patients and experts necessary to advance cures of the disease have proved relatively unsuccessful, and treatment options and survival rates remain limited (Wolff et al, 2002;LafayCousin and Strother, 2009). Recent developments have innovated a cross-species genomics approach that allows mapping of the cells of origin of childhood brain tumors.…”

“…A stem cell-based approach for generating CPECs (patent pending) (Watanabe et al, 2012) that could address many of these limitations was recently described. CPECs are the workhorse cells that perform the major secretory, transport, detoxification, and barrier functions of the ChP.…”

“…2). In vivo, CPECs are long-lived, exhibiting little proliferation or turnover; accordingly, primary CPECs have been difficult to expand in culture to a significant degree (Watanabe et al, 2012). Using ES cells as starting material (which circumvents the expandability problem), the production of derived CPECs (dCPECs) follows important developmental principles, such as the ability of BMP4 to act as an instructive morphogen for CPEC fate (Cheng et al, 2006;Hu et al, 2008) and the restriction of CPEC potential to early-stage neuroepithelial cells rather than later-stage radial glia (Thomas and Dziadek, 1993).…”