Platelet soluble CD40L and matrix metalloproteinase 9 activity are proinflammatory mediators in Behçet disease patients

Abstract

Platelets are the major source of plasma-soluble CD40L (sCD40L), an important inflammatory mediator. This study explored the impact of platelet-derived sCD40L on Behçet disease (BD), an autoinflammatory vasculitis. We also searched for influences by platelet matrix metalloproteinases (MMP) -2 and MMP-9, implicated in several inflammatory diseases, on CD40L shedding from platelet membrane. Platelet activation were studied by flow cytometry and aggregometry, surface expression of CD40L and platelet-leukocyte aggregates by flow cytometry, sCD40L by ELISA, cellular CD40L and CD40 levels by Western blot and MMPs activity by gelatin zymography. The effect of sCD40L on MMP9 expression was studied in cultured MEG-01 cells. Plasma and platelet-released sCD40L levels were higher in BD patients. No differences in platelet activation and in platelet-leukocyte aggregates formation were observed between BD patients and controls. Plasma and platelet MMP-9 levels were increased in BD patients, whereas there was no difference in platelet MMP-2 activity. Since a correlation between plasma sCD40L and platelet MMP-9 activity was observed, we studied the influence of sCD40L on MMP-9 levels in the megakaryoblastic cell line MEG-01. Treatment of MEG-01 cells with recombinant sCD40L increased MMP-9 but did not change MMP-2 levels. In conclusion, sCD40L release from platelets was mediated by MMP-9, and MMP-9 expression was in turn upregulated by sCD40L in the MEG-01 cell line. We conclude that platelets and megakaryocytes might participate in a positive feedback loop occurring between sCD40L and MMP-9 which would contribute to the proinflammatory state observed in BD.

Behçet’s disease: new insight into the relationship between procoagulant state, endothelial activation/damage and disease activity

“…However, we did not find any differences in PTS between the patients and the controls. Although higher platelet activation and response to stimulus have been reported in BD patients [13,32], our research group recently published a study of platelet function in the same group of BD patients included in the present study and found no differences in platelet activation markers between the controls and the BD patients, either at baseline conditions or after stimulation with agonists (TRAP and ADP) [33]. After considering this finding and the lack of differences in PTS and platelet contribution to the ROTEM trace between the groups, we conclude that platelets are not the cause of the deviation in the ROTEM results.…”

The Signaling Role of CD40 Ligand in Platelet Biology and in Platelet Component Transfusion

“…Here, MMP9 was involved in the shedding of CD40L after platelet-neutrophil interaction. Again, different experimental approaches confirmed a role for MMP9 [ 127 , 128 , 129 , 130 ].…”

Soluble CD40L is associated with increased oxidative burst and neutrophil extracellular trap release in Behçet’s disease

“…CD40L protein expression was determined by flow cytometry as previously described [ 14 , 26 ]. Platelets were obtained and assessed by flow cytometry for CD40L protein expression before and after stimulation with 0.2 U/ml thrombin for 5 minutes at 37 °C in a 5% CO 2 atmosphere [ 14 ].…”

“…Fernández-Bello et al [ 14 ] showed higher levels of plasma and platelet-derived sCD40L in a group of patients with BD, although no differences in platelet activation or platelet-leukocyte aggregate formation were observed. In normal mouse neutrophils, sCD40L strongly stimulates the oxidative burst via a CD40-dependent phosphoinositide 3-kinase/NF-κB pathway [ 15 ].…”