Increased Circulating Proplatelets and Elevated Plasma TPO Levels: A Novel Pathological Mechanism of Cerebral Infarction

Background: Platelets are produced by megakaryocytes and are primarily regulated by thrombopoietin (TPO). In the conditions of inflammation or stress, increased 5-HT and other inflammatory cytokines may affect liver and bone marrow to increase TPO and platelet production (Yang et al., Stem Cells, 2007; 2014). Mature megakaryocytes are fragmented in pulmonary circulation to generate more platelet intermediate proplatelets, then entered the circulation. We hypothesized that in patients with cerebral infarction, the secretion of inflammatory cytokines increases under stress or inflammatory conditions, increasing circulating TPO levels, followed by an increase in platelet production, and an increase in the number of platelet intermediate proplatelets. At the same time, platelets also have varying degrees of activation. Therefore, the increase in the number of proplatelets in the circulation and the increase in TPO levels may be the new pathological mechanism of cerebral infarction.

Methods: The plasma levels of TPO were detected by ELISA. The in vivo proplatelets from patients were stained with CD41-FITC, which was further confirmed under a fluorescence microscope and flow cytometry. The in vitro proplatelets were observed in a culture system by CD41-FITC staining under a fluorescence microscope and flow cytometry detection.

Results: Our clinical data demonstrated that patients with acute inflammation states, plasma TPO levels were significantly higher compared with healthy subjects (181.1 ± 35.38 vs 96.1 ± 9.7 pg/ml, p<0.001, n=65), which may act as an acute response protein to protect the body. We also detected 20 patients with cerebral infarction and 20 normal controls and found that the plasma TPO levels of cerebral infarction patients (186.2±12.5 pg/ml) was significantly higher than that of the control group (122.3±10.2 pg/ml). The number of platelets in patients with cerebral infarction (222.11±8.55 ×109/L, n=20) was slightly higher than that in the control group (206.55±8.83 ×109/L, n=20). More importantly, we found more large platelets or proplatelets in the circulating blood of patients with cerebral infarction, which was significantly different from the control group (n=8). Furthermore, in vitro study confirmed that in 200 megakaryocytes (MKs), (18±2.5)% of MKs producing proplatelets in the TPO group (100 ng/ml), and in 5-HT group (100 nM), (15±3.2)% of MKs producing proplatelets, while the control group was only (7±3.2)% (n=5). At the same time, the effects of TPO and 5-HT were interfered by the corresponding receptor blockers, confirming that their effects were mediated by its receptors. The study further confirmed that both TPO and 5-HT affect the cytoskeleton by activating p-ERK1/2, reorganizing F-actin to generate proplatelets, and its role was blocked by PD98059.

Conclusions: The study demonstrated that increased numbers of proplatelets in circulation and elevated plasma TPO levels may be novel pathological mechanisms of cerebral infarction