Autor:innen:
T. Flieder (Bad Oeynhausen, DE)
M. Wolny (Bad Oeynhausen, DE)
K. von Bargen (Bad Oeynhausen, DE)
J. Schmitz (Bielefeld, DE)
A. Vollmer (Bad Oeynhausen, DE)
C. Knabbe (Bad Oeynhausen, DE)
I. Birschmann (Bad Oeynhausen, DE)
Introduction
The main function of platelets is within hemostasis, but they also play a major role in inflammatory processes. For example, together with leukocytes they can form so-called platelet-leukocyte aggregates (PLA), which can be distinguished in subgroups like platelet-neutrophile aggregates (PNA) or platelet-monocyte aggregates (PMA). The occurrence of such aggregates is predominantly described in connection with inflammatory or thrombotic situations. Therefore, they are also considered as potential biomarkers for some diseases such as chronic obstructive pulmonary disease, consumptive coagulopathy, or sepsis.
In patients with implanted extracorporeal membrane oxygenation (ECMO) or left ventricular assist device (LVAD), who also showed increased PLA levels, the occurrence of these PLA has been associated with inflammatory processes. We investigated whether the shear stress exerted on cells by ECMO or LVAD can also lead to PLA formation using a flow chamber model.
Methods
D-Phenylalanyl-L-prolyl-L-arginine chloromethyl ketone (PPACK) anticoagulated blood was collected from healthy donors and for part of the experiments a platelet inhibitor was added to the blood 15 min before shear stress exposure. Whole blood was pumped at a defined rate through a PDMS-glass-based flow chamber (height 100 µm, width 1000 µm, length 5 cm, with or without a 75% stenosis), causing a defined shear stress on the cells. We chose a low shear rate of 500s-1 and a high shear rate of 5000 s-1. In the stenosis area, the shear rates were 2000 s-1 and 20000 s-1. After shear stress exposure, PLA, PNA and PMA were determined by flow cytometry.
The study received a positive vote from the ethics committee (AZ: 2022-1015).
Results
At shear rates of 500 s-1 and 5000 s-1 in the flow chamber without stenosis the measurements showed a significant increase in PNA and PMA relative to the respective control. An increased appearance of PLA was observed only at 5000 s-1. Expression of CD62P on platelets was not increased by shear rates.
In the flow chamber with a 75% stenosis, it was shown that PLA, PNA, and PMA were significantly increased at 500 s-1 and 5000 s-1. Increased CD62P expression was only observed at 5000 s-1.
When comparing the samples with 5000 s-1 shear rate of the model without stenosis and the samples with 500 s-1 shear rate and the model with stenosis (2000 s-1 at the stenosis), it was shown that significantly more PLA, PNA, and PMA were formed in the samples from the stenotic flow chamber.
Conclusion
This study demonstrated that shear stress exposure can lead to increased formation of PLA, PNA, and PMA. Thereby, platelet activation does not seem to be necessary. However, acceleration of cells due to stenosis seems to have a higher influence on PLA formation than the degree of shear rate alone. Whether these results can also be confirmed in patients with ECMO or LVAD needs to be investigated in further studies.