Opposite Modulatory Effects of Crataegus aronia Aqueous Extract on Platelet Aggregation in Rats

Abstract

Objectives: To reveal the mechanisms behind the dual effects of Crataegus aronia (C. aronia) aqueous extract on platelet aggregation by focusing on function, regulation, expression, and signaling of platelets P₂Y₁₂ receptors.

Methods: Adult male Wistar rats (120 ± 10 g) were classified as control received the vehicle, C. aronia (200 mg/kg), and C. aronia (2,000 mg/kg)-treated rats. After treatments for consecutive 7 days, hematological and molecular experiments were conducted to detect alterations in platelet aggregation, thromboxane B2 (THXB2) and intracellular reactive oxygen species (ROS) content; protein levels of P₂Y₁₂, p-Akt, cyclic adenosine monophosphate (cAMP), phosphorylated vasodilator-stimulated-phosphoprotein (p-VASP), nuclear factor κB (NF-κB), P-selectin, and etc. in platelets were determined by Western blot; mRNA expressions of P₂Y₁₂ and some inflammatory markers were determined by real-time polymerase chain reaction.

Results: At a concentration of 200 mg/kg, C. aronia inhibited platelet aggregation through multiple interconnected mechanisms including downregulation P₂Y₁₂ synthesis and expression, stimulating intracellular cAMP levels and protein levels of p-VASP, inhibiting platelets THXB2 release and protein levels of P-selectin. Also, it inhibited platelets level of ROS and of NF-κB, a major signaling pathway that stimulates the expression of P₂Y₁₂ and THXA2 synthesis. Opposite findings were seen in platelets of rats received C. aronia at a concentration of 2,000 mg/kg. Interestingly, co-administration of N-acetylcysteine prevented all hematological and molecular alterations exerted by the high dose of the extract and inhibited platelet aggregation.

Conclusion: Oral administration of C. aronia at low dose inhibits platelet aggregation by reducing THXB2 release, expression of P-selectin and activating cAMP and Akt signaling through two major mechanisms including downregulation of P₂Y₁₂ and inhibition of ROS-induced activation of NF-κB, an effect that is observed to be in the opposite direction with its high dose.

Keywords: Crataegus aronia; Intracellular cyclic adenosine monophosphate; P2Y12; aggregation; nuclear factor αB; platelets; reactive oxygen species.