Figure 4: ABCC4 Affects Dense Granule Release and Platelet cAMP Homeostasis in HIV

(A) Mepacrine staining was assessed via flow cytometry in platelets from controls (C) and HIV before and after activation with 0.05 or 0.25 U/ml thrombin. A representative histogram of 1 healthy control is shown. Quantitative analysis of 8 controls and 9 HIV is shown. *p < 0.05 and ***p < 0.0001. (B) Mepacrine staining via flow cytometry of platelets from controls (n = 9) and HIV (n = 8) at basal state. **p< 0.01. (C) Platelets from controls (n = 5) and HIV subjects (n = 4) were pre-treated with Ceefourin 2 (Ceefourin, 10 μmol/l), activated with 0, 0.05, or 0.25 U/ml thrombin and assessed for mepacrine staining. **p < 0.01 and ***p < 0.0001. (D) cAMP was quantified in the supernatants of activated (0.05 U/ml thrombin, 5 min) platelets from controls (n = 9) and subjects with HIV (n = 13). Levels are reported as pg/ml. **p < 0.01. (E) Platelets from HIV (n = 3) and controls (n = 3) were harvested for Western blot analysis. Blots were probed with anti P-VASP Ser 157 and anti-VASP, and loading was controlled with anti-GAPDH. Quantitative analysis was obtained by densitometry using ImageJ 1.51n software (NIH, Bethesda, Maryland). *p < 0.05. cAMP = cyclic adenosine monophosphate; GAPDH = glyceraldehyde 3-phoshate dehydrogenase; MFI = mean fluorescence intensity; VASP = vasodilator-stimulated phosphoprotein; other abbreviations as in Figure 1.

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