ITAM upregulation in platelets after major surgery

Introduction: Perioperative platelet consumption due to blood loss and coagulation stimulates de novo synthesis of platelets. The juvenile platelets are RNA- (reticulated) and HLA-I rich and more reactive. This might contribute to the increased thrombotic risk after major surgery. Besides the presumed increased platelet reactivity, little is known about the altered platelet phenotype and function post-surgery. We analyzed hemostatic and non-hemostatic platelet functions, before and after major orthopedic surgery, with a special focus on juvenile platelets.

Method: Washed platelets from 27 patients before (day 0) and after total hip or knee replacement or periacetabular osteotomy (day 1, 3, 5) were analyzed by flow cytometry. We assessed surface expressions of platelet receptors GPVI, PAR-1, P2Y12 and FcyIIa and activation of platelets: CD62p expression, PAC-1 binding and Phosphatidyl serine (PS) exposure after stimulation with platelet agonists (TRAP (+/- Convulxin), CRP, ADP. The juvenile platelet fraction was identified by HLA-I and SYTO13 staining (RNA). Calcium release from intracellular stores was assessed by Fluo-4 labeling of platelets+/- calcium pump-inhibitor Thapsigargin or TBHQ. Platelet ITAM activation was assessed by FcyRIIa activation in a modified HIPA-test, using the monoclonal antibody KKO.

Results: HLA-I expression, SYTO13 staining, platelet activation markers and platelet response to agonists, including the proportion of procoagulant platelets (PS+/CD62p+), remained stable at any timepoint after surgery. Juvenile (HLA top 20%) platelets expressed 4 times more CD62p compared to HLA poor (lower 80%), p<0.001. Activation of both subfractions did not change after surgery. While surface expression of GPVI and P2Y12 remained stable, expression of PAR1 (p=0.0016) and FcyRIIa (p=0.0107) increased by a factor of 1.4 at day 5 after surgery. This corresponded to a shortened lag time of FcyRIIa dependent platelet aggregation (28 min at day 0 to 22 min at day 3) after surgery. In addition, platelets stored more calcium in their intracellular compartments with a maximum on day 3 post surgery (Thapsigargin: 1.4-fold increase, TBHQ: 1.3-fold increase).

Conclusion: We found no major increase of circulating juvenile platelets post-surgery. However, platelet calcium storage capacity and non-hemostatic functions of platelets (ITAM signaling via FcγRIIa) are upregulated after surgery

Publikationsverlauf

Artikel online veröffentlicht:
13. Februar 2025