Figure 3
Figure 3. OxLDL signals through CD36 to induce platelet shape change. (A) Platelets (2.5 × 108/mL) were treated with FA6.152 (1 µg/mL) or control IgG (1 µg/mL) for 15 minutes in the presence of apyrase (2 U/mL), indomethacin (10 µM), and EGTA (1 mM) followed by stimulation with oxLDL (50 µg/mL) or thrombin (0.05 U/mL). Traces were recorded for 2 minutes. Shown are representative traces of 7 independent experiments. (B) Samples were processed as in (A), except that platelets were treated with SSO (50 μM) or DMSO. Shown are representative traces of 3 separate experiments. (C) Platelets (5 × 107/mL) were adhered to nLDL or oxLDL (50 μg/mL) slides in the presence or absence of SSO (50 μM), FA6.152 (1 µg/mL), or control IgG (1 µg/mL), for 30 minutes and were viewed by fluorescence microscopy. Representative images of 3 independent experiments were taken under ×60 magnification. Bar = 20 µm. (D) Platelets (3 × 108/mL) were treated with FA6.152 (1 µg/mL) or control IgG (1 µg/mL), SSO (50 μM), or fucoidan (5 μg/mL) for 15 minutes in the presence of apyrase (2 U/mL), indomethacin (10 µM), and EGTA (1 mM) followed by stimulation with oxLDL (50 µg/mL) for 15 seconds. Platelets were lysed and separated by SDS-PAGE and immunoblotted for phospho-MLCSer19, followed by reprobing for β-tubulin. (Di) Representative blots. (Dii) Densitometric analysis of 3 independent experiments. Data are presented as mean ± SEM. *P < .05.

OxLDL signals through CD36 to induce platelet shape change. (A) Platelets (2.5 × 108/mL) were treated with FA6.152 (1 µg/mL) or control IgG (1 µg/mL) for 15 minutes in the presence of apyrase (2 U/mL), indomethacin (10 µM), and EGTA (1 mM) followed by stimulation with oxLDL (50 µg/mL) or thrombin (0.05 U/mL). Traces were recorded for 2 minutes. Shown are representative traces of 7 independent experiments. (B) Samples were processed as in (A), except that platelets were treated with SSO (50 μM) or DMSO. Shown are representative traces of 3 separate experiments. (C) Platelets (5 × 107/mL) were adhered to nLDL or oxLDL (50 μg/mL) slides in the presence or absence of SSO (50 μM), FA6.152 (1 µg/mL), or control IgG (1 µg/mL), for 30 minutes and were viewed by fluorescence microscopy. Representative images of 3 independent experiments were taken under ×60 magnification. Bar = 20 µm. (D) Platelets (3 × 108/mL) were treated with FA6.152 (1 µg/mL) or control IgG (1 µg/mL), SSO (50 μM), or fucoidan (5 μg/mL) for 15 minutes in the presence of apyrase (2 U/mL), indomethacin (10 µM), and EGTA (1 mM) followed by stimulation with oxLDL (50 µg/mL) for 15 seconds. Platelets were lysed and separated by SDS-PAGE and immunoblotted for phospho-MLCSer19, followed by reprobing for β-tubulin. (Di) Representative blots. (Dii) Densitometric analysis of 3 independent experiments. Data are presented as mean ± SEM. *P < .05.

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