Fig 2: Decreased functions of platelets in Tet2-deficient mice. (A) The blood routine results of WT and Tet2 KO mice showed no significant differences in their WBC, RBC, and PLT counts (left panel), and in P-LCR (middle panel) and MPV (right panel), (Tet2 KO, n = 7; WT, n = 7). Only PDW (middle panel) was increased moderately. (B) Flow cytometry was used to evaluate PB platelet activation with agonists in Tet2 KO (n = 4) and WT (n = 5) mice. CD62P (left) and aIIbß3 (right) were the surface markers used for detection of platelet activation. (C) Aggregation assays of washed platelets (2 × 108/ml) were performed with Chrono-log Model 700 Whole Blood/Optical Lumi-Aggregometers. When stimulated by ADP (right) and medium concentrations of thrombin (left), the aggregation of washed platelets of Tet2 KO mice was lower than WT mice. Black curve and red arrow represent Tet2 KO platelets while blue one represents WT platelets. MaxA, maximum aggregation (%). (D and E) Phalloidin-FITC staining of washed platelets (3 × 107/ml) with or without agonists from PB of Tet2 KO and WT mice. The representative images were displayed (D). Scale bar: 6 µm. Quantification of the area of each platelet (E, n = 3). Data was processing with Volocity software. MPV, mean platelet volume; PB, peripheral blood; PDW, platelet distribution width; P-LCR, platelet-larger cell ratio; PLT, platelet; RBC, red blood cell; WBC, white blood cells. The data were shown as means ± SEM. Statistical analysis performed by unpaired, multiple Student's t test. P value:*P < .05, ***P < .001; ns, not significant.

Fig 3: Biocompatibility evaluation of the anticoagulant sponges in vitro.a Blood cell count for pristine blood and blood after incubation with MS@D-HMP (n = 6 biologically independent samples, mean ± SD. Unpaired, two-tailed student’s t-test). b Differential white blood cell count (DIFF) scatter (forward (FS), side (SS)) charts of pristine blood and blood after incubation with MS@D-HMP. Monocyte (pink), lymphocyte (green), neutrophil (cyan), eosinophil (red). c Hemolysis ratio of the blood after incubation with MS@D-HMP (n = 3 biologically independent samples, mean ± SD. Unpaired, two-tailed student’s t-test). d Quantitative evaluation of adhered platelets on the sponges by LDH assay (n = 8 biologically independent samples, mean ± SD. One-way ANOVA with Bonferroni post-hoc tests). e Generation of PF4 in the blood after incubation with sponges (n = 3 biologically independent samples, mean ± SD. Kruskal–Wallis test with Dunn’s multiple comparisons). f Platelet activation in PRP based on the expression of platelet activation marker CD62p. Thrombin receptor activating peptide (TRAP, 0.1 mM) and control PRP were used as positive and negative control, respectively (n = 5 biologically independent samples, mean ± SD. One-way ANOVA with Bonferroni post-hoc tests). g Monocyte activation in whole blood based on the expression of monocyte activation marker CD11b. Lipopolysaccharide (10 ng/mL) and pristine blood were used as positive and negative control, respectively (n = 4 biologically independent samples, mean ± SD. One-way ANOVA with Bonferroni post-hoc tests). h Concentration of D-dimer in the citrate-anticoagulated plasma after incubation with sponge (n = 4 biologically independent samples for PPP and MS@D-HMP, n = 3 biologically independent samples for Positive, mean ± SD. One-way ANOVA with Bonferroni post-hoc tests). Generation of C3a (i) and C5a (j) in the blood after incubation with sponges. Cobra-venom factor (CVF, 25 µg/mL) and PBS were used as positive and negative controls, respectively (n = 6 biologically independent samples for Blood, MS, MS@D and MS@D-HMP, n = 4 biologically independent samples for Positive, mean ± SD. One-way ANOVA with Bonferroni post-hoc tests for (i), Kruskal–Wallis test with Dunn’s multiple comparisons for (j)). Source data are provided as a Source Data file.

Fig 4: Effect of MF on the suppression of platelet activation in mice with CLP-induced sepsis. (a) Platelet activation was determined based on CD41 and CD62P (P-selectin) expression. (b) Bar graphs for the percentage of CD62P-positive platelets in each group. Percentages of CD62P-positive platelets are presented as mean ± SD (n = 5 mice). Representative FACS data from one of three separate experiments are shown. ** p < 0.01 compared with the CLP group. MF, Myristica fragrans; OS, oseltamivir phosphate.

Fig 5: Effects of MF on the suppression of sialidase-mediated platelet hyper-reactivity and activation in washed platelets. (a) CD62P expression in washed platelets was measured after stimulation with 5 mU sialidase and 125 µM ADP in the presence of MF or OS at different concentrations. Representative FACS data from one of three separate experiments are shown. (b) Inhibition of CD62P expression using OS in Sia + ADP-stimulated washed platelets. (c) Inhibition of CD62P expression using MF in Sia + ADP-stimulated washed platelets. The bar graphs presented indicate mean ± standard deviation (SD) of three samples. All experiments were repeated thrice, and similar outcomes were observed. * p < 0.05 and ** p < 0.01 compared with the Sia + ADP group. ADP, adenosine diphosphate; MF, Myristica fragrans; OS, oseltamivir phosphate; Sia, sialidase.