Angelo DePalma earned his Ph.D. in organic chemistry from Stony Brook University and was previously senior scientist at Schering-Plough. He has written extensively on biotechnology, biomanufacturing, medical devices, pharmaceutical commerce, laboratory instrumentation, and advanced materials.
One often hears that environment or niche is critical for optimizing stem cell utilization. This was demonstrated again in late 2014 when Doo Yeon Kim and Rudolf Tanzi at Harvard University succeeded in developing the first in vitro model for Alzheimer’s disease (AD) that incorporated two hallmarks of the disorder: amyloid plaques and neurofibrillary tangles [1]. Mouse models express one but not the other.
Kim and Tanzi had previously attempted and failed to develop an AD model from a two-dimensional culture of ReNcells, a human neuronal stem cell line from EMD Millipore. The cells had been modified to over-express the amyloid precursor protein and presenelin-1, which underlies early-onset familial AD. When the investigators switched to a three-dimensional ReNcell culture using Corning’s Matrigel® matrix, the model expressed both plaques and tangles.
Tanzi noted at the time that the new culturing system, which could be applied to other neurodegenerative diseases, could revolutionize drug discovery and basic AD research. Testing drugs in animal models possessing either plaques or tangles, but not both, is expensive and time-consuming. “With our three-dimensional model that recapitulates both plaques and tangles, we now can screen hundreds of thousands of drugs in a matter of months, without using animals, in a system that is considerably more relevant to the events occurring in the brains of Alzheimer's patients,” he said in a statement.
“Cells behave differently and have different phenotypes when grown in three dimensions,” says Nick Asbrock, stem cell and molecular biology product manager, EMD Millipore. “3D cell cultures are more physiologically relevant and more closely resemble in vivo tissues. This unique 3D Alzheimer’s application developed at Harvard has sparked renewed interest in ReNcell lines and highlights their utility in future neurological disease modeling.”
Asbrock notes the potential advantages of using induced pluripotent stem cells (iPSCs) rather than tissue-specific adult neuronal stem cells like ReNcell lines, which are derived from fetal tissue, isolated and immortalized in culture. “iPSCs are easier to isolate from patients [and] grow in large numbers and do not have ethical issues associated with their derivation. Once in hand, experimenters can differentiate them into neural progenitor cells using growth factors or small-molecule stimuli. It is also possible to use CRISPR or other gene-editing tools to knock in specific genes to push cells towards a specific neuronal or glial lineage.”
Flexibility of iPSCs
The advent of iPSCs made possible the more rigorous study of human neural stem cells (NSCs) for basic research, drug discovery and cell therapy. Before 2010, most studies used mouse and rat NSCs, which behaved differently from human NSCs but were critical for the development of methods underlying today’s human NSC experimentation.
The other major sources of NSCs were cells derived from human embryonic stem cells (ESCs). “At the time, NSCs were expensive and not scalable or expandable,” says Mohan Vemuri, R&D leader for cell biology at Thermo Fisher Scientific. “The isolation method was tedious and manual. If you began with one million pluripotent stem cells, you were lucky to get one million NSCs.” Another disadvantage is that pluripotent NSCs came with a fixed genetic fate from which two neuron types, at most, were accessible.
Today, iPSCs may be grown in suspension culture and differentiated into NSCs in about one week. These cells may in turn be expanded to a stable bank of billions of cells, enabling efficient drug screening or disease modeling.
Generating NSCs from iPSCs is also of great interest for personalized medicine settings, where investigators might wish to screen drugs for a particular patient.
“Labs still purchase NSCs commercially, particularly if time is critical or they can afford it,” Vemuri continues. “But if they’re interested from a patient-specific perspective, or wish to observe the entire process, they will generate the cells themselves.”
Migrating from murine cells
Sam Lloyd-Burton, senior product marketing manager at STEMCELL Technologies, concurs that neuroscientists who traditionally used rodent models are switching to NSCs derived from iPSCs, but that mouse models will not leave the stage any time soon. “We see from posters and presentations that iPSC-based neurological modeling has moved from proof of principle to the application phase, with protocols being established for many neurodevelopmental and neurodegenerative diseases, and many interesting phenotypes being reported. The increased physiological relevance of these models complements traditional rodent models and improves the chances of positive funding and publication outcomes.”
STEMCELL Technologies specializes in media and accessory products for neural stem/progenitor cell research. For example, the company’s STEMdiff™ Neural System is optimized for derivation, expansion and downstream differentiation of ESC- and iPSC-derived neural progenitor cells.
There has also been increased interest in more complex models of cortical development, for example the cerebral organoid model. With increased recognition of the vital and multifaceted roles of glial cells in development and disease, more researchers are trying to differentiate iPS/ES cells into astrocytes and oligodendrocytes. “The more complex models involve cultures of neurons and glia together,” Lloyd-Burton says.
Additionally, labs collaborating with Arnold Kriegstein formerly at the Broad Institute, MIT and now at University of California, San Francisco, have been working on a population of neural stem cells called outer radial glia, which exist during human but not rodent brain development. These studies shed light on human cortical evolution, and provide further evidence for why rodent models are sometimes not predictive of human disease and/or developmental mechanisms.
Reference
[1] Choi, SH, et al., “A three-dimensional human neural cell culture model of Alzheimer's disease,” Nature, 515:274-278, 2014. [PMID:25307057]
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