Cytocentric Visionaries: Dr. Adrien Moya, University of Miami
Part 1: Oxygen and Glucose for Engineering Tissues
Dr. Adrien Moya has just started a post-doc at the University of Miami in the lab of Paul Schiller. Here we discuss Dr. Moya’s previous work in the CNRS lab at Diderot University in Paris. His recent publication in Stem Cells reports use of the Cytocentric platform for to control conditions for MSC used in bone regenerative medicine applications. His paper is entitled, “Human Mesenchymal Stem Cell Failure to adapt to Glucose Shortage and Rapidly Use Intracellular Energy Reserves through Glycolysis Explains Poor Cell Survival after Implantation.” 
Congratulations on your publication! We see you as a Cytocentric Visionary because of your use of physiologically relevant conditions to study cell fate in hydrogel implants for bone regeneration.
Thank you very much.
What made you in particular want to look at Multipotent Stromal Cells (MSC) and glucose metabolism for tissue engineering?
MSC and glucose metabolism in tissue engineering in our lab started about ten years ago with a PhD student Mickael Deschepper. He reported that the survival and the function of the MSC depended on glucose to overcome exposure and long term anoxia (Deschepper et al., 2011 and 2013). Joseph Paquet, a post doc at that time, then published a paper in 2015 on oxygen tensions and paracrine function regulating MSC in hypoxia. I came along for my PhD from 2012 to 2016.
Originally I was working on MSC pre-conditioning and how to have them survive without glucose. But even without exogenous glucose I found out that the MSC still rely strongly on their glycolytic reserve, so at that time, I already had some insights to put in this last paper. All these papers on MSC, glucose metabolism and hypoxia/ischemia were a team effort. I would also like to thank the CNRS (centre national de la recherche scientifique) and the DGA (direction generale de l'armement) for funding this research.
What do you want people to know about this work?
Three key points. The first one would be that if you think that, based on their own natural energetic reserve, MSC are going to survive when you implant them in vivo, that’s actually false. Most of them (80-90%) are not going to survive.
A second one is that 1% oxygen tension does not equal 0.1% oxygen tension. Near-anoxia (0.1%) is really different from hypoxia (1%). We have found that in vivo, at the core of the tissue engineering constructs, the oxygen tension for the implanted cells is at near-anoxia rather than hypoxia. The cellular markers and how the MSC behave in that environment is different from 1% oxygen tension.
The third one is that MSC do not adapt to exogenous glucose levels. If they are lacking glucose they will not reduce their glucose consumption. However, there seems to be an optimum around 1g/L of external glucose, which could be because it is physiological, but even between 0.5 g/L and 5 g/L, they will still use the same amount of glucose.
So, in one key phrase, if we are trying to improve tissue engineering constructs and improve MSC survival in them, we should focus on how to deliver glucose to them in vivo.
You tried some other intermediate metabolites right? Pyruvate and some amino acids.
Yes, if MSC have glucose they survive relying on glycolysis and producing pyruvate. If they had no external glucose, they could use the external pyruvate to survive a bit longer, but really it is dying less quickly. Pyruvate has a slight benefit, but is not as good as glucose. In hypoxia, I’m suspecting that MSC can in fact use pyruvate to somehow create a feedback loop to glycolysis using some steps of the neoglucogenesis pathway. This is especially true when the MSC are in a quiescent state. I really think that it’s a key feature in how MSC behave in a hypoxic or anoxic environment.
So there are still new things still to discover at very low oxygen levels? We’ve been using room air oxygen for all of our work for so long.
Yes, I really agree. We have just barely scratched the surface. Now we have systems that allow us to precisely control the cells’ environment. Just the fact that we can now culture cells and perform experiments at low oxygen tensions without oxygen tension variation is huge. In fact, when you abruptly take out the cells from the hypoxic chamber, you create a huge oxidative stress on the cells that can possibly modify all their behavior.
Now that we can maintain hypoxic conditions I think we’re going to first discover new things, and second, we are going to be able to more accurately understand what is going on in vivo. Mainly because the in vivoenvironment for these cells is supposed to be hypoxic, it should be called physiological oxygen tension. When we are using air as a reference for oxygen tensions, the cells are actually experiencing hyperoxia.
It’s those oxygen changes which throw the cells off?
MSC are really tough cells. They can adapt to a lot of conditions, but what they don’t like is sudden, harsh changes. By provoking those changes, it often results in a good amount of the cell population dying so you could end up isolating a smaller subpopulation of the MSC. Then, there is no telling if the cells isolated qualify as MSC anymore, so we should be careful about that.
Have you done any work with cells that were isolated from the organ under physiologic oxygen the whole time?
I wish I had. I did not, but I know a group of researchers that has done it. You might be familiar with Schiller’s group work in Miami. They called their cells, Marrow-isolated adult multilineage inducible cells, MIAMI cells for short. They isolated bone marrow MSC at 3% oxygen with low FBS.
MSC should be quiescent most of the time in vivo, waiting to be awakened. Upon being triggered by a signal, for example, if there is a need for them to perform their job in homeostasis, then they would exit the quiescent state, activate, and migrate to the site of injury.
So by isolating these cells in an environment that better represents their physiological environment, they demonstrated that these cells have more stemness than the regular room air isolated MSC. I think that’s very fascinating. After one or two passages in normoxia (21% oxygen), we don’t know how much their metabolism has changed already.
Nowadays, we have the technology to properly isolate these cells, so I wouldn’t be surprised to see more groups isolate their MSC under what we call now hypoxia.
I think that we are going to see a new understanding of biology if more people do this.
Also, something peculiar that I discovered in the quiescence paper  is that when removing the FBS from the cell culture media for 48 hours, the MSC shift from an OxPhos metabolism to an almost exclusive glycolytic metabolism even in the presence of oxygen (21%). So that seems to indicate that when quiescent MSC would rather use glycolysis than mitochondrial respiration to fuel their metabolism. Maybe when they are in their natural niche they don’t use as much oxygen as we think, they just rely on glycolysis to control their metabolism. But, because we culture them in 21% oxygen we did not see this phenomenon before.
So there could be room air oxygen artifact that we are introducing into this system?
My guess is that indeed it is an artifact that we are introducing. There is extensive literature that shows that stem cells tend to be more glycolytic than differentiated cells. The TCA provides a lot more energy to the cells, but it comes with drawbacks. For example, the cells have to deal with reactive oxygen species.
When MSC go towards the osteoblastic lineage, they became more and more OxPhos-based. The reason is that they need greater amount of energy in order to accomplish their function. In contrast, the stem cells, whose main job is to self-renew do not need as much energy as the differentiated cells.
So I am a big fan of low oxygen isolation of MSC.
In Part Two, we talk with Dr. Moya about the fate of cells in regenerative medicine implants.
If you would like to be featured in our Cytocentric Visionary Series, contact us. We would love to hear about your work.
- Moya A, Paquet J, Deschepper M, Larochette N, Oudina K, Denoeud C, Bensidhoum M, Logeart‐Avramoglou D, Petite H: Human Mesenchymal Stem Cell Failure to Adapt to Glucose Shortage and Rapidly use Intracellular Energy Reserves through Glycolysis Explains Poor Cell Survival After Implantation. STEM CELLS 2017.
- Moya A, Larochette N, Paquet J, Deschepper M, Bensidhoum M, Izzo V, Kroemer G, Petite H, Logeart-Avramoglou D: Quiescence Preconditioned Human Multipotent Stromal Cells Adopt a Metabolic Profile Favorable for Enhanced Survival under Ischemia. Stem Cells 2017, 35(1):181-196.
About the Author
Alicia D Henn, PhD, MBA
Alicia Henn has been the Chief Scientific Officer of BioSpherix, Ltd for two years. Previously, she was a researcher at the Center for Biodefense Immune Modeling in Rochester, NY. Alicia obtained her PhD in molecular pharmacology and cancer therapeutics from Roswell Park Cancer Institute in Buffalo, NY and her MBA from the Simon School at University of Rochester in Rochester, NY.