Disease Severity in Moderate-to-Severe COVID-19 Is Associated With Platelet Hyperreactivity and Innate Immune Activation

Abstract

Background: Hemostasis and inflammation are both dysregulated in patients with moderate-to-severe coronavirus disease 2019 (COVID-19). Yet, both processes can also be disturbed in patients with other respiratory diseases, and the interactions between coagulation, inflammation, and disease severity specific to COVID-19 are still vague.

Methods: Hospitalized patients with acute respiratory symptoms and with severe acute respiratory syndrome coronavirus 2 (SARS-CoV2)-positive (COV^pos) and SARS-CoV2-negative (COV^neg) status were included. We assessed adenosine diphosphate (ADP)-, thrombin receptor activator peptide 6 (TRAP)-, and arachidonic acid (AA)-induced platelet reactivity by impedance aggregometry, as well as leukocyte subtype spectrum and platelet-leukocyte aggregates by flow cytometry and inflammatory cytokines by cytometric bead array.

Results: ADP-, TRAP-, and AA-induced platelet reactivity was significantly higher in COV^pos than in COV^neg patients. Disease severity, assessed by sequential organ failure assessment (SOFA) score, was higher in COV^pos than in COV^neg patients and again higher in deceased COV^pos patients than in surviving COV^pos. The SOFA score correlated significantly with the mean platelet volume and TRAP-induced platelet aggregability. A larger percentage of classical and intermediate monocytes, and of CD4^pos T cells (T_H) aggregated with platelets in COV^pos than in COV^neg patients. Interleukin (IL)-1 receptor antagonist (RA) and IL-6 levels were higher in COV^pos than in COV^neg patients and again higher in deceased COV^pos patients than in surviving COV^pos. IL-1RA and IL-6 levels correlated with the SOFA score in COV^pos but not in COV^neg patients. In both respiratory disease groups, absolute levels of B-cell-platelet aggregates and NK-cell-platelet aggregates were correlated with ex vivo platelet aggegation upon stimulation with AA and ADP, respectively, indicating a universal, but not a COVID-19-specific mechanism.

Conclusion: In moderate-to-severe COVID-19, but not in other respiratory diseases, disease severity was associated with platelet hyperreactivity and a typical inflammatory signature. In addition to a severe inflammatory response, platelet hyperreactivity associated to a worse clinical outcome in patients with COVID-19, pointing to the importance of antithrombotic therapy for reducing disease severity.

Keywords: COVID-19; disease severity; immunothrombosis; inflammation; platelet hyperactivity; platelet-leucocyte aggregates; survival.

Graphical Abstract

Depicting study design and main observations.

Figure 1

Platelet aggregation induced by TRAP (A), ADP (B), and AA (C), as assessed by MEA, is higher in COV^pos (n = 50) than in COV^neg (n = 37) patients.

Figure 2

Thrombin-antithrombin complex is higher in COV^pos (n = 50) than in COV^neg (n = 37).

Figure 3

Disease severity indicated by the SOFA score was higher in COV^pos (n = 50) than in COV^neg (n = 37) patients (A) and higher among COV^non-surv (n = 10) than COV^surv (n = 40) COV^pos patients (B). MPV was higher in COV^non-surv than in COV^surv COV^pos patients (C). Within COV^pos, SOFA score correlated with MPV (D) and TRAP-initiated platelet activation (E).

Figure 4

CXCL10, IL-6, MCP-1, and IL-1RA are higher in COV^pos than in COV^neg patients (A) and also higher in deceased than in surviving COV^pos patients (B). A higher percentage of classical and intermediate monocytes and CD4^pos T_H lymphocytes form aggregates with platelets in COV^pos than in COV^neg patients (A). Deceased COV^pos patients showed lower relative abundance of lymphocytes and higher total leukocyte counts than surviving COV^pos patients (B). p-value of 0.05 is indicated by the horizontal dashed-dotted line. p-values higher than 0.05 and less-than-twofold changes between groups are underlaid in grey. plt agg, platelet aggregates; gra, granularity; cla Mo, classical monocytes; int Mo, intermediate monocytes; ncl Mo, nonlassical monocytes.

Figure 5

Spearman’s correlations in COV^pos (A) and COV^neg (B) were expressed as dark grey lines if positively correlated and pink lines if correlated inversely. Correlations with p < 0.05 are depicted. cla Mo, classical monocytes; int Mo, intermediate monocytes; ncl Mo, nonclassical monocytes; T_h, CD4^pos helper T cells; T_cyt, CD8^pos cytoxic T cells; NK-T, natural killer T cells; NK cells, natural killer cells; peak CRP, highest CPR level during hospital stay.