Intercellular communication and immunomodulatory activities of platelet microvesicles in vascular inflammation (LSBR 1638)
Project leader: Dr. Rory R. Koenen (CARIM – Maastricht University)
PhD Student: Alexandra Heinzmann (Jan. 2017 – Jan. 2021)
Blood platelets are small cell fragments that do not have a nucleus and are essential for the prevention of blood loss. Upon injury, platelets rapidly form blood clots. This leads to the sealing of the wound and to the arrest of the bleeding. Most textbooks describe platelets as “hemostatic” (blood clotting) cells. Much less well known is that platelets can also act as immune cells.
During the past two decades, this “alternative” function of platelets has received increasing attention. Platelets can bind to many molecules and cells in the vascular environment and carry a multitude of factors that can modify the immune system. In addition, there is compelling evidence that platelets can even synthesize proteins, even without having a nucleus. One of these proteins is interleukin-1, which is a central cytokine in inflammation.
Interestingly, recent studies have also provided evidence that platelet might act as carriers of molecular information. They can take up nucleic acids, for example RNA from tumors. Moreover, they can transport this information to other cells. Involved in this process are so called extracellular vesicles (EV). These are small membrane-enclosed “bubbles” that contain many proteins of the parent cell from which they originate. Platelets were the first cells known to release EVs, but it is currently established that all cells can release EVs.
The release of EVs from platelets is relevant for transfusion medicine, since platelets stored in concentrates (“platelet packs”) continuously release EVs. The activation of platelets also leads to a massive release of EVs. The exact function of EVs is still unknown, but evidence exists that they can modify the phenotype of cells with which they interact. Like platelets, the platelet-derived EV can bind to white blood cells e.g. monocytes, neutrophils and T cells. They also interact with cells of the vessel wall, like endothelial cells and smooth muscle cells. The EVs are thought to be involved in the transfer of information
from the platelet to a recipient cell.
In this project, we aimed to characterize this transfer of information by EVs to cells of the immune and of the vascular systems. In order to unravel the mechanisms underlying the release of EVs by platelets, platelets were activated with different stimuli and also treated with different clinically used antiplatelet medication. The release of EVs from platelets was found to require 3 steps: 1. integrin activation, 2. cytoskeleton rearrangement, 3. exposure of negatively charged phospholipids. Although common antiplatelet drugs such as aspirin and ADP-receptor antagonists did not strongly influence the release of EVs after platelet activation, they did lower the amounts of chemokines released. Antiplatelet drugs targeting surface integrin GPIIbIIIa (aIIbb3) and also inhibitors of intracellular phosphodiesterases were able to strongly reduce the release of EVs from platelets. The inhibition of EV release from platelets would be of interest for longer term storage as EVs might have detrimental effects on vascular function.
For example, we found that platelet-derived EVs strongly bound to smooth muscle cells. This binding depended on surface molecules on the platelets, but also on the surface of smooth muscle cells. Healthy smooth muscle cells are contractile and regulate vascular tone, but when in contact with EVs from platelets, the smooth muscle cells started to divide more rapidly and changed their appearance and properties towards a synthetic and also an inflammatory phenotype.
Also endothelial cells were found to be influenced by platelet-derived EVs. They take up these EVs, leading to increased intracellular signaling and also to increased endothelial inflammation. Both endothelial and smooth muscle cells were found to attract more immune cells after contact with platelet EVs than under resting conditions. In addition, the chemokines CCL5 and CXCL4, which are carried on platelet EVs are rapidly internalized into endothelial and smooth muscle cells. This was found to induce both a pro-inflammatory (endothelial cells) and a pro-calcifying (smooth muscle cells) phenotype
in these cells.
Taken together, platelet-derived EVs can change the behavior of vascular cells, which is relevant for transfusion. When patients receive platelet concentrates, EVs are co-transfused along with the platelets. This might lead to changes of the phenotype of vascular cells, with potentially detrimental longterm effects (e.g. vascular remodeling). Antiplatelet drugs are known to inhibit blood coagulation, but their pronounced anti-inflammatory effects should also be taken into account and these might be implemented to prevent chronic vascular diseases e.g. atherosclerosis.