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Introduction to Stem Cells, Culture, and Surface Modification
Surface modified biomaterials may be an excellent addition to suspension culture of pluripotent stem cells…But before we get there, let’s start by understanding what pluripotent stem cells are: human cells that can “differentiate” into other cells in the body (hence they are pluripotent). Also called HPSCs, these cells can be embryonic stem cells (collected when the human was a baby) or induced pluripotent stem cells (stem cells generated from adult cells). HPSCs are incredibly valuable because they can be used in cell therapies (replace cells in the body with new cells) and for drug discovery modeling (models of tissues so we can develop new drugs). Right now there are about 234 different HPSC stem cell lines at the NIH which can be used in research and the development of future therapies.
However, the use of these cells is currently limited because there are several challenges that face researchers who want to study them. First and foremost, growing or “culturing” stem cells is difficult. Second, there is a lack of standard methods for using stem cells in the above applications (cell therapy and drug discovery). And third, stem cells that are grown have impurities or differentiated cells (no longer pluripotent) amidst them. These differentiated cells may be carcinogenic if implanted as part of a cell therapy regime.
Challenges with Suspension Culture of Pluripotent Stem Cells
Culturing stem cells is a challenge because they tend to differentiate in normal culture media. Maintaining pluripotency in these cell lines requires the interplay between FGF-2, Noggin, Activin/Nodal, and TGF-beta signaling pathways and is accomplished by including these in cell culture media. In a small scale, all these elements are easily available to cells. However, in a large scale, maintaining pluripotency is challenging because of mixing and signal availability. Here, the tight maintenance of nutrient mixing and oxygenation is balanced by minimizing cell shear stresses which can induce cell death. Bioreactors and stirrer flasks are the classic technique for suspension culture of stem cells.
Cells in suspension culture tend to aggregate and form natural multicellular colonies. Formation of these colonies helps maintain higher proliferation rates but also confounds growth analysis because growth rate varies based on aggregation state. This is also a challenge because cells naturally aggregate, grow, apoptose and spontaneously differentiate. As stated before, differentiated cells are carcinogenic.
Use of Surface Modified Biomaterials with HPSC Suspension Culture
Biomaterials that have modified surfaces for binding differentiated HPSCs may help suspension cell culture. Researchers have now developed ideal media for maintaining “stem-ness” in multiple HPSC lines. Perhaps the addition of a coating to bioreactors and stirring flasks may also solve other challenges that are faced.
- Aldehyde containing membranes with antibodies immobilized on the surface could be used to bind differentiated cells as they are produced. Naturally, it is expected that one researches the ideal biomarkers for capturing differentiated cells and the best stirring speed/load ratio where the shear rate is not too high for capture.
- Amine containing membranes could have immobilized ligands and proteins on the surface that provide cues to maintain stem-ness. Mixing is a challenge since cells can be sheared at too high an impeller speed. Perhaps having FGF-2 or TGF-beta (or both) on the walls of the vessel can improve their availability to suspended cells.
- Similar to capturing circulating tumor cells, filters could be utilized in a flow system (combined with mechanical sieves) to harvest aggregates that become too large and are prone to apoptosis or spontaneous differentiation.