Cellular Neurobiology

The Stem Cell groups at the Stanley Center for Psychiatric Research are developing stem cell–derived neuronal models and using genome engineering in human pluripotent stem cells (hPSCs) to investigate in vitro phenotypes associated with genetic variants underlying psychiatric disease. The Stem Cell groups are focused on the following projects:

Human Cellular Models
Stem Cell Genome Engineering
Human Brain Organoid Models
Neuronal Spheroids for Screening and Therapeutic Target Studies
The Brain Interaction Network
A Human Stem Cell Genome Project
Massively Mosaic Experimental Systems

Human Cellular Models

The Nehme lab aspires to identify the cellular phenotypes linked to human genetic variants associated with neuropsychiatric disorders. Through a large collaborative effort with the McCarroll and Eggan labs, we have established the Stanley Center Stem Cell Resource, a rich resource of curated and whole-genome sequenced human pluripotent stem cell (hPSC) lines.  We have assembled, sequenced and annotated stem cell lines from hundreds of patients with a spectrum of psychiatric conditions as well as from ancestrally matched controls. To achieve a more equitable representation of diverse genetic backgrounds in functional studies, we are also leading a large effort to reprogram iPSC lines from diverse genetic backgrounds. Projects leveraging this resource aim to generate and integrate datasets from multiple phenotypic measurements in disease relevant cell-types, including transcriptional, morphological and functional, using arrayed and pooled approaches we and colleagues have developed (Nehme et al, 2018Mitchell et al., bioRxiv, 2020Fan et al., 2018). 

Stem Cell Genome Engineering

The Lindy Barrett lab developed a series of CRISPR-Cas9–based genome engineering pipelines in hPSCs (see Stem Cell Reports and Scientific Reports) to enable functional studies of genes and variants identified through whole-exome and whole-genome sequencing of psychiatric cohorts. Cellular resources generated through these efforts are utilized in research projects throughout the Stanley Center community, including editing genes in different backgrounds for human brain organoid models. The group now leverages these and other resources to interrogate convergent molecular and cellular mechanisms across neurodevelopmental diseases.

Human Brain Organoid Models

Developing improved cellular systems that model complex human genetic traits, while accounting for non-disease-relevant variation in genetic background, is an important goal in the Paola Arlotta lab. Drawing from foundational work on cortical neuronal development and circuitry in the mouse cortex, the group has optimized a protocol, published in Nature, that supports long-term development of self-organizing whole brain organoids derived from human stem cells. These human organoids generate a large diversity of cell classes from distinct regions of the brain and display robust neuronal networks, presence of synaptic spines, and light sensitivity. Human brain organoids present promises and challenges for gaining insight into human neurodevelopmental and neuropsychiatric pathologies.

Neuronal Spheroids for Screening and Therapeutic Target Studies

Developing screening assays relevant to psychiatric disease using human neurons is the focus of the Lee Rubin lab. The group recently reported a robust spheroid culture protocol in Stem Cell Reports, noteworthy for large-scale production of hPSC-derived neurons with mature features. The group now seeks to “close the gap between spheroid and organoids” with continued technical improvements to increase human brain-like mature features, in order to eventually implement the method in their wide studies of psychiatric disease mechanisms. In collaboration with Beth Stevens and the Stanley Center Therapeutics group, some of these Stanley Center–wide efforts include following up on the Nature paper linking the human leukocyte antigen (HLA) locus, a schizophrenia GWAS hit, with mechanisms of synaptic pruning downstream of the complement 4 (C4) protein.

The Brain Interaction Network

In collaboration with the Eggan lab at the Stanley Center, the Lage lab has teased out cell-type-specific protein interaction networks that help understand how common and rare genetic variation can affect whole neuronal processes. By combining genetic information from different neuropsychiatric disorders with proteomics, the team has built a robust pipeline, published in Nature Communications, that spans the generation of experimental data, data quality control and analysis, and integration with a variety of publicly available genetic datasets to maximize results interpretation. Ongoing efforts aim to further extend this research to cell types that are most strongly implicated in neuropsychiatric disease.

A Human Stem Cell Genome Project

The ability of hPSCs to renew indefinitely enables limitless propagation in culture, but also raises concerns about the long-term genomic integrity of hPSCs. To gain a holistic view of the genetic landscape of hPSCs, Kevin Eggan and team performed whole exome sequencing (WES) and WGS of 114 publically available human embryonic stem cell lines, acquired and banked at the Stanley Center. The report on stem cell exomes, published in Nature, showed that hPSCs in culture can acquire and expand cancer-associated mutations that would not have been detected by methods currently used for quality control (such as karyotyping and SNP genotyping). These findings point to the importance of sequence-based approaches for genetic screening of hPSCs and their differentiated derivatives, both in experimental models of disease and cell replacement therapies, across the stem cell field.

Massively Mosaic Experimental Systems

In collaboration with Steve McCarroll, the Eggan lab is harnessing WGS data to generate unique fingerprints — composed of genetic variants used as an intrinsic barcode — of large numbers of hPSC lines. This bioinformatics framework serves as the foundation for development of mosaic experimental systems, in which large numbers of hPSC lines are co-cultured in the same dish and analyzed using Drop-seq, and then have their donor identity inferred after the experiment. Efforts underway aim to develop Drop-seq to measure gene expression levels at a cellular level. These approaches enable scaling of stem cell disease models in a way that is internally controlled, pointing towards applications for mapping eQTLs as a function of psychiatric disease.