3-D spheroids like this cardiac spheroid, which was grown in suspension from pluripotent stem cells, can produce enough mature cells to speed high-throughput drug discovery and solid-organ regenerative therapies.Credit: Thermo Fisher Scientific
When pluripotent stem cells (PSCs) were first isolated from human embryos in 1998, they promised a new age of regenerative medicine. These cell lines, the first lab-grown cells capable of differentiating into all the types of cells in the body, would produce new cells and tissues to replace those damaged by age or disease. The field’s prospects grew in 2006, when researchers reprogrammed mature human cells back to a pluripotent state to produce induced PSCs, thereby avoiding controversial embryonic sources.
The journey to the clinic, however, has often slowed to a crawl—particularly for therapies that could restore function to solid organs like the liver, kidney, pancreas and heart. Often that’s because too few PSCs are available to differentiate into the specific cell types needed for a treatment, especially when preparing cells for more than a single individual. A cardiac repair treatment, for instance, may require starting with upwards of a billion PSCs per patient. The standard 2-D cell-culture systems used today, which grow PSCs as monolayers on flat, rigid plastic or glass surfaces, can't economically produce enough cells for many important therapies.
From monolayers to spheroids
Modern 3-D cell-culture methods avoid many of the problems of monolayer growth. The cells can grow efficiently in a liquid suspension culture that is agitated (either shaken or stirred), achieving higher cell densities than cells grown in a monolayer. For example, in 2018 researchers from the Chinese Academy of Sciences reported a new 3-D system in Cell Death and Disease that could generate 1.5 billion viable PSCs for every 1.5 liters of culture medium—enough for high-throughput drug-discovery screening and most solid-organ regenerative therapies.
When grown properly in 3-D suspension culture, PSCs pack themselves into clusters called spheroids, where they encounter microenvironments like those they experience in the body. "Pluripotent stem cells like to be in contact with each other," says David Kuninger, director of R&D at Thermo Fisher Scientific, in Frederick, Maryland. PSCs grown this way remain pluripotent and genetically stable, he adds.
A new 3-D suspension culture medium developed by Thermo Fisher manages all these feats. PSCs cultured in StemScale PSC Suspension Medium form spheroids with a consistent size and uniform population of cells. “That’s incredibly important when you want to do a targeted differentiation," Kuninger says. And it helps affordably cultivate between 1 and 1.5 billion high-quality PSCs per liter—enough for both medical therapy and high-throughput drug-discovery screening.
PSCs grown this way can reliably differentiate into all three of the cell layers, or germlines, formed in early embryonic development, Kuninger says. They also differentiate efficiently into beating heart cells called cardiomyocytes, midbrain dopaminergic neurons for studying Parkinson's disease, and many other cell types. What’s more, they can be easily isolated, stored frozen for future use, and used in closed and automated systems, which facilitates the large-scale manufacture of cell-based therapies.
Scaling Up
It’s still more straightforward to use standard 2-D cell culture to characterize many key cellular attributes, including pluripotency and differentiation efficiency. But “a 3-D system supports the transition from the cells' initial development towards generating robust populations at scale,” Kuninger says.
That means more stem cell treatments to join those already on the market for blood disorders, and those currently in clinical trials, such as the autologous PSCs for macular degeneration, PSCs for treating Parkinson's disease and cardiac repair, and for other indications, which are being tested in dozens of clinical trials.
While the number of cells required for these treatments varies, all of them, as well as early-stage drug discovery efforts, require high-quality cells, and more of them, Kuninger says. "That's what is going to enable scientists who do this kind of translational work."