A new 3D human brain tissue platform developed by MIT researchers is the first to integrate all major brain cell types, including neurons, glial cells and the vasculature into a single culture. Grown from individual donors' induced pluripotent stem cells, these models—dubbed Multicellular Integrated Brains (miBrains)—replicate key features and functions of human brain tissue, are readily customizable through gene editing, and can be produced in quantities that support large-scale research.
Although each unit is smaller than a dime, miBrains may be worth a great deal to researchers and drug developers who need more complex living lab models to better understand brain biology and treat diseases.
"The miBrain is the only in vitro system that contains all six major cell types that are present in the human brain," said Li-Huei Tsai , Picower Professor, director of The Picower Professor of Learning and Memory, and senior author of the study describing miBrains, published DATE in the Proceedings of the National Academy of Sciences.
"In their first application, miBrains enabled us to discover how one of the most common genetic markers for Alzheimer's disease alters cells' interactions to produce pathology," she added.
Tsai's co-senior authors are Robert Langer , David H. Koch (1962) Institute Professor, and Joel Blanchard, associate professor in the Icahn School of Medicine at Mt. Sinai in New York and a former Tsai Laboratory postdoc. The study is led by Alice Stanton, former postdoc in the Langer and Tsai labs and now assistant professor at Harvard Medical School and Massachusetts General Hospital, and Adele Bubnys, a former Tsai lab postdoc and current senior scientist at Arbor Biotechnologies.
Benefits from two kinds of models
The more closely a model recapitulates the brain's complexity, the better suited it is for extrapolating how human biology works and how potential therapies may affect patients. In the brain, neurons interact with each other and with various helper cells, all of which are arranged in a three-dimensional tissue environment that includes blood vessels and other components. All of these interactions are necessary for health and any of them can contribute to disease.
Simple cultures of just one or a few cell types can be created in quantity relatively easily and quickly, but they cannot tell researchers about the myriad interactions that are essential to understanding health or disease. Animal models embody the brain's complexity, but can be difficult and expensive to maintain, slow to yield results, and different enough from humans to yield occasionally divergent results.
miBrains combine advantages from each type of model, retaining much of the accessibility and speed of lab-cultured cell lines while allowing researchers to obtain results that more closely reflect the complex biology of human brain tissue. Moreover, they are derived from individual patients, making them personalized to an individual's genome. In the model, the six cell types self-assemble into functioning units, including blood vessels, immune defenses, and nerve signal conduction, among other features. Researchers ensured that miBrains also possess a blood-brain-barrier capable of gatekeeping which substances may enter the brain, including most traditional drugs.
"The miBrain is very exciting as a scientific achievement," said Langer. "Recent trends toward minimizing the use of animal models in drug development could make systems like this one increasingly important tools for discovering and developing new human drug targets."
Two ideal blends for functional brain models
Designing a model integrating so many cell types presented challenges that required many years to overcome. Among the most crucial was identifying a substrate able to provide physical structure for cells and support their viability. The research team drew inspiration from the environment that surrounds cells in natural tissue, the extracellular matrix (ECM). The miBrain's hydrogel-based "neuromatrix" mimics the brain's ECM with a custom blend of polysaccharides, proteoglycans, and basement membrane that provide a scaffold for all the brain's major cell types while promoting the development of functional neurons.
A second blend would also prove critical: the proportion of cells that would result in functional neurovascular units. The actual ratios of cell types have been a matter of debate for the last several decades, with even the more advanced methodologies providing only rough brushstrokes for guidance, for example 45-75% for oligodendroglia of all cells or 19-40% for astrocytes.
The researchers developed the six cell types from patient-donated induced pluripotent stem cells, verifying that each cultured cell type closely recreated naturally-occurring brain cells. Then, the team experimentally iterated until they hit on a balance of cell types that resulted in functional, properly structured neurovascular units. This laborious process would turn out to be an advantageous feature of miBrains: because cell types are cultured separately, they can each be genetically edited so that the resulting model is tailored to replicate specific health and disease states.
"Its highly modular design sets the miBrain apart, offering precise control over cellular inputs, genetic backgrounds, and sensors—useful features for applications such as disease modeling and drug testing," said Stanton.
Alzheimer's discovery using miBrain
To test miBrain's capabilities, the researchers embarked on a study of the gene variant APOE4, which is the strongest genetic predictor for the development of Alzheimer's disease. Although one brain cell type, astrocytes, are known to be a primary producer of the APOE protein, the role that astrocytes carrying the APOE4 variant play in disease pathology is poorly understood.
miBrains were well-suited to the task for two reasons. First of all, they integrate astrocytes with the brain's other cell types, so that their natural interactions with other cells can be mimicked. Second, because the platform allowed the team to integrate cell types individually, APOE4 astrocytes could be studied in cultures where all other cell types carried APOE3, a gene variant that does not increase Alzheimer's risk. This enabled the researchers to isolate the contribution APOE4 astrocytes make to pathology.
In one experiment, the researchers examined APOE4 astrocytes cultured alone, vs. ones in APOE4 miBrains. They found that only in the miBrains did the astrocytes express many measures of immune reactivity associated with Alzheimer's disease, suggesting the multicellular environment contributes to that state.
The researchers also tracked the Alzheimer's-associated proteins amyloid and phosphorylated tau, and found all-APOE4 miBrains accumulated them, whereas all-APOE3 miBrains did not, as expected. However, in APOE3 miBrains with APOE4 astrocytes, they found that APOE4 miBrains still exhibited amyloid and tau accumulation.
Then the team dug deeper into how APOE4 astrocytes' interactions with other cell types might lead to their contribution to disease pathology. Prior studies have implicated molecular cross-talk with the brain's microglia immune cells. Notably, when the researchers cultured APOE4 miBrains without microglia, their production of phosphorylated tau was significantly reduced. When the researchers dosed APOE4 miBrains with culture media from astrocytes and microglia combined, phosphorylated tau increased, whereas when they dosed them with media from cultures of astrocytes or microglia alone, the tau production did not increase. The results therefore provided new evidence that molecular cross-talk between microglia and astrocytes is indeed required for phosphorylated tau pathology.
In the future, the research team plans to add new features to miBrains to more closely model characteristics of working brains, such as leveraging microfluidics to add flow through blood vessels or single cell RNA sequencing methods to improve profiling of neurons.
Researchers expect that miBrains could advance research discoveries and treatment modalities for Alzheimer's disease and beyond.
"Given its sophistication and modularity, there are limitless future directions," said Stanton. "Among them, we would like to harness it to gain new insights into disease targets, advanced readouts of therapeutic efficacy, and optimization of drug delivery vehicles."
"I'm most excited by the possibility to create individualized miBrains for different individuals," added Tsai. "This promises to pave the way for developing personalized medicine."
Funding for the study came from the BT Charitable Foundation, Freedom Together Foundation, the Robert A. and Renee E. Belfer Family, Lester A. Gimpelson, Eduardo Eurnekian, Kathleen and Miguel Octavio, David B. Emmes, the Halis Family, The Picower Institute for Learning and Memory, and an anonymous donor.
