CXCR7 agonist counteracts thrombotic and thromboinflammatory response elicited by HIT sera/IgG. Whole blood assays with heparinized (0.2 IU/mL) blood from healthy donors (n = 5) was carried out in presence of control sera (n = 3) and HIT⁺ patient sera (n = 3) incubated at 1:10 vol/vol dilution for 1 hour at room temperature. CXCR7 agonist (100 µg/mL) or vehicle control (1% DMSO) was given as a pretreatment for 30 minutes at room temperature. HIT⁺ patient sera induced (A) platelet CD62P surface expression and (B) PAC-1 binding denoting α_IIbβ₃-integrin activation ex vivo. (C) HIPA assay performed with isolated washed platelets from 3 healthy donors in presence/absence of CXCR7 agonist (25 and 100 µg/mL) or vehicle control shows an inhibitory effect of CXCR7 agonist. (D) Ex vivo thrombus formation was performed in heparinized (0.2 IU/mL) blood from healthy donors (n = 5) over collagen-coated surface in presence of control sera (n = 3) and HIT⁺ patient sera (n = 3) in presence/absence of CXCR7 agonist (100 µg/mL) or vehicle control. In panels A through D, *P < .05, ***P < .001, ****P < .0001 using ANOVA followed by Sidak’s multiple comparison test. (E) Targeted lipidomics analysis for oxylipins showed that ex vivo treatment with control sera (n = 3), HIT⁺ patient sera (n = 3), HIT⁺ patient sera (n = 3) for 1 hour at room temperature induced generation of COX-1–derived TxA2 and platelet 12-LOX–derived 12-HETE, which was significantly counteracted in the presence of CXCR7 agonist (100 µg/mL) given as a pretreatment of 30 minutes at room temperature. *P < .05, #P < .05, ##P < .01, ***P < .001 with Wilcoxon signed rank test. (F) Neutrophil CD11b (anti-human active CD11b-FITC) activation ex vivo in heparinized (0.2 IU/mL) blood from healthy donors (n = 5) in response to control sera (n = 3) or sera from HIT⁺ patients (n = 3) (at 1:10 vol/vol dilution) in presence/absence of CXCR7 agonist (100 µg/mL) or vehicle control. Diagram showing formation of CD42b⁺ (anti-human CD42b-FITC)-CD11c⁺ (anti-human CD11c-APC) platelet-neutrophil aggregates ex vivo in heparinized (0.2 IU/mL) blood from healthy donors (n = 5 donors) treated with (Gi) sera or (Gii) corresponding isolated IgG fractions from control healthy subjects and HIT⁺ patients (n = 3 each) at 1:10 vol/vol dilution for 1 hour at room temperature in presence/absence of CXCR7 agonist (100 µg/mL). In panels F through G, *P < .05, ***P < .001, ****P < .0001 using ANOVA followed by Sidak’s multiple comparison test. Washed human platelets (200 × 10⁶/sample) from n = 5 donors were pretreated with CXCR7 agonist (100 µg/mL) or vehicle control for 30 minutes at room temperature and then with IgG fractions from control or HIT⁺ patient sera (n = 3 each) at 1:10 vol/vol for 1 hour at room temperature. Activated platelet supernatants were collected and analyzed by cytometric bead array (Legendplex 13-plex human inflammation panel, 10-plex human thrombosis panel). (H) Violin plots (solid line denotes median) showing levels of proinflammatory mediators and chemoattractants in platelet releasate. Data represent mean ± SEM; #P < .05, ##P < .01 vs control IgG; *P < .05, **P < .01, ***P < .001, ****P < .0001 vs HIT⁺ IgG using Mann-Whitney U test for each inflammatory mediator.