THE HEART OF THE MATTER

Decoding the role of EVs in Cardiovascular Health: a discussion with Dr. Pierre-Michaël Coly and Stephane Mazlan from the Paris Cardiovascular Research Centre

Cardiovascular disease (CVD) is a broad classification for the group of diseases that involve the heart and blood vessels, and is the cause for one in every four deaths.¹ It includes the conditions associated with chest pain, heart failure, hypertension, stroke, heart attack, and coronary artery disease (CAD), with the latter being most common; in fact, CAD is responsible for more than 15.6% of all deaths globally.²

Given the global impact of CAD, much of the research in past decade has focused on understanding the underlying mechanisms of disease development and progression. Dr. Pierre-Michaël Coly and Stephane Mazlan, researchers at the Paris Cardiovascular Research Centre, believe that extracellular vesicles (EVs) may be central to this understanding. They are working together in the lab of Dr. Chantal Boulanger to understand both sides of CAD. Coly’s work focuses on atherosclerosis, which is the process that causes CAD. Mazlan’s work is focused on the effects of CAD, primarily myocardial infarction—commonly referred to as a heart attack.

Dr. Pierre-Michaël Coly

Dr. Pierre-Michaël Coly

The development of atherosclerosis is a gradual process that occurs over many years, but it begins with inflammation in the arterial wall. Over time, this inflammation results in the gradual deposition of white blood cells, lipids, calcium, and fibrous connective tissue along the walls of the arteries. These deposits, or plaques, constrict the flow of blood through the arteries of the heart. Severe plaques can rupture, initiating a coagulation cascade that results in the formation of blood clots that block blood flow completely.³ When the blockage occurs in an artery that feeds the muscles of the heart, the result is a myocardial infarction.
Plaque formation isn’t random–Coly, a post-doctoral researcher in the lab, has found a pattern. “We’ve noticed that the plaques don’t form everywhere in your blood vessels–there are spots that preferentially form them.”

He and others in the lab have shown that the blood flow through an artery exerts varying levels of abrasive pressure, or shear stress, on the cells that make up the artery walls. Atherosclerosis tends to develop in arterial forks and in the inner part of artery curvatures where the blood flow is lower or has been disturbed, while areas that are exposed to high blood flow experience a higher level of shear stress and are free of plaques.4 Coly is studying the role of the autophagy machinery [the proteins and enzymes that regulate the degradation and recycling of cellular components] in the release and uptake of vesicles by the cells of arterial walls under various levels of shear stress. The cells in low-flow areas exhibit pro-inflammatory characteristics that may promote the recruitment of white blood cells and ultimately
“One of our main interests has been finding out that EVs are, in many cases, precursors to the diseases that we study,” says Coly. “We are looking at the types of vesicles released from these cells to see if they could be biomarkers of the early stress that lead to disease”.

Using an in-vitro model of epithelial shear stress and Izon’s qEV columns, Coly is isolating EVs from the cell culture media and looking for these potential biomarkers. This process has recently been aided by the release of Izon’s Automatic Fraction Collector (AFC), an instrument which will allow him to automate the isolation process and enhance the precision and repeatability of EV isolation between experiments.

Being able to detect plaques in the early stages could help decrease the incidence of coronary heart disease and ultimately the number of myocardial infarction events. These events downstream of atherosclerosis are the focus of Stephane Mazlan, a Ph.D. candidate in the lab.

A myocardial infarction triggers the release of EVs from cardiomyocytes and endothelial cells.5 Many of these EVs are carrying microRNAs, which may be the messages involved in repairing the heart tissue after an infarction event. By looking at which microRNAs are up- and down-regulated, Mazlan hopes to uncover effects that may further the development of future therapeutics, “Our work has to benefit the patient at the end of the day. If we find beneficial exomiRs [exosomal microRNAs], we can amplify them and load them with therapeutics. If it’s not beneficial, if it’s detrimental, then we can give an antagomir [molecules used to block microRNAs].”

Stephane Mazlan

Understanding the function and biological effects of these EV cargos will be key to creating the first generations of EV diagnostics and therapeutics, but there are still many obstacles to overcome before EVs will be ready for the clinic. One of the recurring themes in the literature is a need for standardization of methods so that EV research can be more readily comparable. “The biggest challenge right now is just having a common ground; a universal way for how the vesicles are isolated, the way they are analysed,” Coly says “The field is still very new, so it is still going through the hurdles that are inherent to this type of work.”

Quantification and characterization of EV subpopulations will be critical for identifying novel biomarkers of disease progression, and there is mounting evidence that the quantity of EVs correlates with the severity of cardiac injury. Several studies have shown that the levels of circulating EVs can be used to identify patients may be at risk of developing cardiovascular complications.6 One study even showed that the number of endothelium-derived EVs was sufficient to discriminate between patients that had stable angina (chest pain), versus a first or recurring myocardial infarction.7

All of these data together make the standardization of EV quantification methods a primary concern for the field. Both Mazlan and Coly are using the qNano system and TRPS to quantify EVs as a part of their work, and hope that these contributions will ultimately improve patient outcomes in the future. “Being able to detect plaques in the early stage by something as simple as a blood draw–it is still a way down the road, but it might be the first step,” says Coly.

While the field may still be in its adolescence, one thing is clear: understanding the role of EVs and the cargos they carry will be pivotal to uncovering the molecular mechanisms of cardiovascular disease. “Decoding these messages and seeing whether they are pro-help or anti-help to heal the heart, that’s the ultimate goal,” says Mazlan. “And at the end of the day, these EVs are not only a matter of the heart, but the heart of the matter.”

REFERENCES

  1. Zhang, Y., Hu, Y., Zheng, L., & Wang, Q. (2017). Characteristics and Roles of Exosomes in Cardiovascular Disease. DNA and Cell Biology,36(3), 202-211. doi:10.1089/dna.2016.3496
  2. Wang, H., Naghavi, M., Allen, C., Barber, R. M., Bhutta, Z. A., Carter, A., . . . Murray, C. J. (2016). Global, regional, and national life expectancy, all-cause mortality, and cause-specific mortality for 249 causes of death, 1980–2015: A systematic analysis for the Global Burden of Disease Study 2015. The Lancet,388(10053), 1459-1544. doi:10.1016/s0140-6736(16)31012-1
  3. Boulanger, C. M., Loyer, X., Rautou, P., & Amabile, N. (2017). Extracellular vesicles in coronary artery disease. Nature Reviews Cardiology,14(5), 259-272. doi:10.1038/nrcardio.2017.7
  4. Vion, A., Kheloufi, M., Hammoutene, A., Poisson, J., Lasselin, J., Devue, C., . . . Rautou, P. (2017). Autophagy is required for endothelial cell alignment and atheroprotection under physiological blood flow. Proceedings of the National Academy of Sciences, 114(41). doi:10.1073/pnas.1702223114
  5. Loyer, X., Zlatanova, I., Devue, C., Yin, M., Howangyin, K., Klaihmon, P., . . . Boulanger, C. M. (2018). Intra-Cardiac Release of Extracellular Vesicles Shapes Inflammation Following Myocardial Infarction. Circulation Research,123(1), 100-106. doi:10.1161/circresaha.117.311326
  6. Dignat-George, F., & Boulanger, C. M. (2011). The Many Faces of Endothelial Microparticles. Arteriosclerosis, Thrombosis, and Vascular Biology,31(1), 27-33. doi:10.1161/atvbaha.110.218123
  7. Sluijter, J. P., Davidson, S. M., Boulanger, C. M., Buzás, E. I., Kleijn, D. P., Engel, F. B., . . . Ferdinandy, P. (2017). Extracellular vesicles in diagnostics and therapy of the ischaemic heart: Position Paper from the Working Group on Cellular Biology of the Heart of the European Society of Cardiology. Cardiovascular Research,114(1), 19-34. doi:10.1093/cvr/cvx211