Enhancing Cell Culture Media to Better Mimic Human Nutrients
For decades, scientists have relied on cell culture media to grow cells in the lab. A recent study suggests it's time for a major upgrade.
Most of us know that scientists grow cells in the laboratory using media. But have any of us ever stopped to think about what is inside that media?
In laboratory research, the quality of cell culture media can significantly impact experimental outcomes. And while cell culture media has long been a staple in research and has been relatively unchanged over the past several decades, new findings suggest they might be missing a crucial element: physiological accuracy.
The development of cell culture media dates back to the mid-20th century, when researchers sought methods to grow living animal cells outside of their host organism. Given this goal, early formulations simply aimed to maintain cell viability rather than accurately mimic physiological conditions. For example, in 1913, French surgeon Alexis Carrel discovered that a solution containing embryonic extract and blood plasma allowed chicken embryo heart fibroblasts to proliferate outside the body, even though the exact amounts of the lymph, plasma, and embryonic extract in this medium did not match physiological concentrations (Yao, 2017).
The beginnings of tissue culture. The figure shows two technicians working in the Carrel laboratory at the Rockefeller Institute. They are wearing full-length black, hooded gowns that were adopted by Carrell. (Source: HarperCollins)
Nonetheless, given how effective this medium was at maintaining cell growth ex vivo, it was used for several decades. Even today, the development of cell culture media tends to focus on simply supporting cell proliferation, rather than trying to accurately reflect physiological levels of metabolites and nutrients in the body. While this discrepancy in metabolite levels usually does significantly alter cellular pathways, it has been found to sometimes have a large effect on specific metabolic pathways. For instance, when cancer cells are cultured in standard media, they tend to be sensitive to glutaminase inhibitors. However, when cancer cells are cultured in physiologically-relevant (serum-like) media or are grown as tumors in mice, they actually are no longer killed by glutaminase inhibitors (Muir, 2017).
This is just one example of how different culture media can affect a cell’s response to a therapeutic. While the differential effect of glutaminase inhibitors in standard media versus serum-like media is now understood, however, most cancer therapeutics that were only studied in the context of standard media. The effect that culture media has on in vitro metabolic findings remains widely underappreciated, and many research studies in metabolism still fail to acknowledge the effect that the choice of cell culture medium has on their findings. As a result, many researchers may be putting substantial effort in testing cancer therapeutics in mouse models and clinical trials that are ultimately not effective, when they could have tested the compound in serum-like media in the first place.
To address this issue, a recent study led by Matthew Vander Heiden, MD, PhD, the Lester Wolfe Professor of Molecular Biology at MIT and Director of Koch Institute for Integrative Cancer Research, reported the development a high-throughput screening method that researchers can use to identify metabolism-targeting compounds that are differentially effective at killing cancer in serum-like media versus in standard media. The team created a new cell culture medium called ftABS, which is designed to more closely mimic the nutrient levels found in the human body. They did this by filtering and mixing different types of serum and adding specific nutrients. When they compared this new medium to the traditional RPMI medium, they found that ftABS had more realistic levels of nutrients.
The researchers tested how well various drugs worked against lung cancer cells grown in both media. Most drugs worked similarly in both media, but some drugs that target specific cellular processes worked better in RPMI. For instance, ftABS contains certain nutrients that make the cells less sensitive to inhibitors of key enzymes involved in cell growth, which were more effective in RPMI. They also found that adding extra cystine (a nutrient) to ftABS made it more like RPMI and restored the effectiveness of certain drugs. Additionally, ftABS reduced the activity of a protein called mTOR, which usually helps cells grow. Surprisingly, even though mTOR activity was lower, the cancer cells didn’t grow any slower in ftABS compared to RPMI. This suggests that mTOR may behave differently in this new medium.
The researchers developed a serum-derived culture medium and found that compounds targeting metabolic enzymes were differentially effective in standard compared to serum-derived medium. This approach can be used to screen potential therapeutics and understand how their efficacy is altered by available nutrients.
There are several limitations of this study. One fundamental problem is that while it clearly lays out the differences between the serum-like ftABS medium and standard cell culture media, it does not verify whether the cell behavior in ftABS actually reflects how they would behave in vivo. The absence of in vivo validation raises questions about the translatability of the observed cellular responses to drug treatments. Additionally, the study primarily focuses on the A549 lung cancer cell line, and the generalizability of the findings to other cancers or tissues remains uncertain. Moreover, physiologic media like ftABS have inherent limitations given that nutrient levels in vivo can vary over time and space due to factors such as tissue location, diet, genetics, and circadian rhythm. While ftABS accurately reflects physiological metabolic conditions on average, caution should be taken in interpreting proliferation rates in mixed cell culture systems. Having a system that can adjust nutrient conditions in physiologic media in real-time may be necessary for robust screening results, especially for serum-free conditions such as patient-derived models and organoids.
The method developed by Abbott et al. pose interesting next questions facing the field. While this paper focused on identifying cancer therapeutics that are less effective in physiologic media compared to standard cell culture media, there can also be a focus on identifying compounds that are more effective in physiologic media compared to standard cell culture media. That is, there possibly exist underappreciated compounds that are very effective at killing cancer in vivo and in serum-like media, but we do not know about them because they have not been found to be effective at killing cancer in standard cell culture media, which is used for most modern research studies.
The development of more physiologically accurate cell culture media represents a pivotal advancement in biomedical research. By incorporating a richer array of nutrients and mimicking the dynamic conditions of human tissues, these new media formulations hold the potential to enhance the fidelity of experimental models significantly. As researchers continue to refine these media, the next generation of cell culture systems will likely yield deeper insights into human health and disease, ultimately bridging the gap between in vitro studies and clinical applications.
Original Article: https://doi.org/10.1016/j.chembiol.2023.08.007
References
Huang, J. & Manning, B. D. The TSC1–TSC2 complex: a molecular switchboard controlling cell growth. Biochem. J. 412, 179–190 (2008).
Muir, A. et al. Environmental cystine drives glutamine anaplerosis and sensitizes cancer cells to glutaminase inhibition. eLife 6, e27713 (2017).
Valvezan, A. J. et al. mTORC1 Couples Nucleotide Synthesis to Nucleotide Demand Resulting in a Targetable Metabolic Vulnerability. Cancer Cell 32, 624-638.e5 (2017).
Yao, T. & Asayama, Y. Animal‐cell culture media: History, characteristics, and current issues. Reprod. Med. Biol. 16, 99–117 (2017).
