Stem Cell-Derived CPECs

CPEC derivation from human pluripotent stem cells

Based on developmental principles we defined previously (e.g., Currle et al., 2005), our lab published and patented a method to derive choroid plexus epithelial cells (CPECs) from mouse and human embryonic stem (ES) cells. This derivation is a two-step process – pluripotent stem cells (PSCs) are first differentiated into early-stage neuroepithelial progenitors, then exposed to bone morphogenic protein 4 (BMP4) to induce CPEC differentiation (Watanabe et al., 2012). The method has now been simplified and further optimized for more scalable production using human ES or induced pluripotent stem cells (iPSCs).

Immunostaining for CPEC markers OTX2 and TTR at the edge of a stem cell-derived CPEC island.

Human developmental modeling

Cell differentiations using human PSCs can be invaluable experimental models of human development. Given how little is known about human CPEC development, a variety of studies are being carried out on the derived CPECs. These include single cell RNA sequencing (scRNA-seq) and novel bioinformatic analyses in collaboration with the Genomic High-Throughput Facility (GHTF) and NSF-Simons Center for Multiscale Cell Fate Research. Interesting or unexpected in silico findings are tested or validated in various ways, often using separate CPEC derivations or perinatal ChP tissues from the autopsy service of the Department of Pathology & Laboratory Medicine.

Clustering analysis to identify cell types present at one stage of CPEC drivation.

Human disease modeling

In addition to normal development, patient-derived or genetically-engineered iPSCs can be unique and powerful tools for modeling human diseases. Current diseases of interest include mitochondrial disorders and Alzheimer’s Disease (AD). CPECs have the highest mitochondrial volume of any CNS cell, and in almost all postmortem subjects, occasional CPECs will accumulate excessive mitochondria with age. One current mitochondrial study involves iPSCs from a toddler who died with Leigh’s Syndrome, a novel mitochondrial mutation, devastating neuropathology, and striking mitochondrial accumulation in all CPECs. For the common sporadic form of AD, the APOE gene is, by far, the strongest genetic risk factor, and CPECs express particularly high levels of APOE. Together with the iPSC Core of the Alzheimer’s Disease Research Center (ADRC) and CRISPR Core of the Stem Cell Research Center (SCRC), we are using isogenic and non-isogenic APOE iPSC lines to derive human CPECs and study APOE-dependent functions of relevance to AD.

Drug Screens

A third regenerative medicine application (in addition to developmental and disease modeling) is drug screening, and PSC technologies enable direct screening of human cells. Human CPEC drug screens are particularly compelling when one considers the straightforward nature of drug delivery to the ChP and CPECs in vivo given their rich supply of fenestrated capillaries and disproportionate share of cerebral blood flow. In collaboration with the Conrad Prebys Center for Chemical Genomics, we received NIH funding to develop high-throughput and high-content CPEC screens. The goal is to identify pharmacologic tools and therapeutic leads for defining CPEC functions (e.g. secretion, uptake, and detoxification) that could rectify disease states. For example, drugs that reduce CSF production by CPECs could treat hydrocephalus, whereas drugs that increase CSF or CPEC secretion of anti-amyloid proteins could protect the brain from AD.

Cell-Based Therapies

Transplant studies by others implicate the ChP in neuroregeneration and repair, and our previous work highlighted the potential for derived CPECs to engraft host ChP after intraventricular injection (Watanabe et al., 2012). These and other observations underscore the promise of CPEC-based therapies – e.g. using genetically engineered CPECs (perhaps from a patient’s own iPSCs) to produce and secrete therapeutic proteins directly into the CSF and correct patient-specific deficiencies that lead to neurologic disease. Such an approach would circumvent the blood-brain and blood-CSF barriers that can render peripheral medications incapable of reaching their CNS targets. Stereotactic procedures and anesthesia systems for mice have been developed by our lab in the SCRC vivarium to facilitate this work.