Figure 5.
Sdy and Tmem163-KO platelets display significant impairment of DG release and AG secretion. (A-B) Washed platelets from sdy+/−, sdy, B6 (WT, C57BL/6J) or Tmem163-KO mice were stimulated with various concentrations of collagen (1.2, 5, or 20 μg/mL) (A) or thrombin (0.25 or 1 U/mL) (B) and secretion of ATP assessed by ATP determination kit. Data represent the relative ATP release as the percentage (mean ± SEM) of the mean normalized value in maximal dose stimulation for their control mice. (C-D) Washed platelets from sdy+/−, sdy, B6, or Tmem163-KO mice were labeled with fluorescein isothiocyanate-CD62p antibody and stimulated with various concentrations of collagen (0.5 or 1.0 μg/mL, sdy vs Tmem163-KO, n = 6, 6, respectively) (C) or thrombin (0.25 or 0.5 U/mL, sdy vs Tmem163-KO, n = 3, 6, respectively) (D) for 5 minutes in the absence or presence of 10 μM ADP and then measured by flow cytometry for CD62p surface expression. Data represent the percentage of platelets with labeling above the background level observed on unstimulated platelets. (E-F) Washed platelets from sdy+/−, sdy, B6, or Tmem163-KO mice were stimulated with various concentrations of collagen (0.8, 1.2, or 5.0 μg/mL) (E) or thrombin (0, 0.25, or 1 U/mL) (F) for 5 minutes in the absence or presence of 10 μM ADP, and then supernatants were collected and measured by enzyme-linked immunosorbent assay for PF-4 (sdy vs Tmem163-KO, n = 4, 4, respectively.). Data shown represent the relative PF4 release as the percentage (mean ± SEM) of the mean normalized value in maximal dose stimulation for their control mice. *P < .05; **P < .01; ***P < .001.

Sdy and Tmem163-KO platelets display significant impairment of DG release and AG secretion. (A-B) Washed platelets from sdy+/−, sdy, B6 (WT, C57BL/6J) or Tmem163-KO mice were stimulated with various concentrations of collagen (1.2, 5, or 20 μg/mL) (A) or thrombin (0.25 or 1 U/mL) (B) and secretion of ATP assessed by ATP determination kit. Data represent the relative ATP release as the percentage (mean ± SEM) of the mean normalized value in maximal dose stimulation for their control mice. (C-D) Washed platelets from sdy+/−, sdy, B6, or Tmem163-KO mice were labeled with fluorescein isothiocyanate-CD62p antibody and stimulated with various concentrations of collagen (0.5 or 1.0 μg/mL, sdy vs Tmem163-KO, n = 6, 6, respectively) (C) or thrombin (0.25 or 0.5 U/mL, sdy vs Tmem163-KO, n = 3, 6, respectively) (D) for 5 minutes in the absence or presence of 10 μM ADP and then measured by flow cytometry for CD62p surface expression. Data represent the percentage of platelets with labeling above the background level observed on unstimulated platelets. (E-F) Washed platelets from sdy+/−, sdy, B6, or Tmem163-KO mice were stimulated with various concentrations of collagen (0.8, 1.2, or 5.0 μg/mL) (E) or thrombin (0, 0.25, or 1 U/mL) (F) for 5 minutes in the absence or presence of 10 μM ADP, and then supernatants were collected and measured by enzyme-linked immunosorbent assay for PF-4 (sdy vs Tmem163-KO, n = 4, 4, respectively.). Data shown represent the relative PF4 release as the percentage (mean ± SEM) of the mean normalized value in maximal dose stimulation for their control mice. *P < .05; **P < .01; ***P < .001.

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