Antiplatelet aggregation and fibrinolytic activity of Artemisia Vulgaris L. extract

Abstract

Platelet aggregation is involved in atherosclerosis plaque, one of the major causes of cardiovascular disease. The objectives of the present study are to determine antiplatelet and fibrinolytic activities of herbal extracts from Phikud Benjakot. The ethanol and water extracts of each herb in Phikud Benjakot, Kot Soa (Angelica dahurica, AD), Kot Kamao (Atractylodes lancea, AL), Kot Huabua (Ligusticum sinense, LS), Kot Chiang, (Angelica sinensis, AS) and Kot Chulalumpa (Artemisia vulgaris, AV), at the concentration of 1 mg/mL did not affect both viability and DNA damage of human umbilical vein endothelial ECV304 cells. The effect of the herbal extracts at the concentration of 1 mg/mL on platelet aggregation induced by ADP was performed using light transmission aggregometry method. The result demonstrated that antiplatelet activity of ethanol and water extracts were in descending order of AV > AD > AS. Since AV expressed the highest antiplatelet activity, the ethanol and water extracts of AV were chosen for a concentration-dependent study, resulting in exhibiting IC50 values of 0.43 and 0.51 mg/mL, respectively. However, both extracts did not inhibit platelet aggregation induced by arachidonic acid and collagen. The result suggested that the ethanol and water extracts of AV are likely to inhibit platelet aggregation via ADP signaling pathway. To confirm whether the antiplatelet activity of both extracts of AV was not due to platelet damage, platelet viability was then performed using MTT reduction assay. Treatment of platelets with both extracts of AV at the antiplatelet concentration for 1 h did not affect platelet viability. Additionally, AV did not affect platelet function using acid phosphatase assay. The result indicated that antiplatelet activity of AV was not due to their toxicity or interfering platelet function. The effect of both extracts of AV on platelet adhesion to collagen was determined based on acid phosphatase activity. Both extracts of AV did not affect platelet adhesion when allowed to adhere to collagen for 1 h. Since endothelial-derived nitric oxide can inhibit platelet aggregation, the effect of the ethanol extract of AV on phosphorylation of Akt and eNOS in ECV304 cells was studied using Western blot analysis. The result revealed that the ethanol extract of AV at the concentration of 0.5 mg/mL for 1 h treatment could enhance the phosphorylation of eNOS, but not Akt, in ECV304 cells under serum-free condition. Our findings suggested that AV, one of crude drugs in Phikud Benjakot, could be considered as a candidate herb in the treatment of platelet-associated vascular disease.