Evaluation of Antiplatelet Activity of Phenolic Compounds by Flow Cytometry

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  •   Konstantinos D. Kyriakidis

  •   Eyrysthenis G. Vartholomatos

  •   Georgios S. Markopoulos

Abstract

Platelets play a pivotal role in coagulation, or clot formation, resulting in haemostasis, after endothelium injury. Disturbance of platelet activation may lead to pathologic thrombosis. Platelet activation and aggregation are common factors in atherothrombotic events, critical in the atherothrombotic process, and cardiovascular diseases. Several drugs are being used for antiplatelet therapy to prevent and/or treat atherosclerosis and cardiovascular diseases. Synthetic antiplatelet drugs hold possible undesired health consequences (cardiovascular diseases, carcinogenicity, etc.), advocating their replacement with natural, effective, and non-toxic compounds. Many phenolic compounds are created as secondary metabolites of plants, are found in many fruits and vegetables, and constitute a wide family of high-added-value molecules. Their biological activities include antioxidant, anti-platelet, and anti-inflammatory action. Based on the above, we examined five phenolic compounds (ellagic acid, ferulic acid, gallic acid, quercetin, and kaempferol) for their effect on platelet reactivity in whole blood samples using flow cytometry.


Quantification of activated platelet marker CD62-P by flow cytometry showed that all five compounds inhibited platelet activation in vitro, induced by adenosine diphosphate (ADP) and collagen. Interestingly, based on the IC50 values obtained for expression of CD62-P, among ellagic, ferulic, and gallic acid, gallic acid showed significantly higher inhibition than the other two. Kaempferol found to be a more potent inhibitor than quercetin, following previously reported results from aggregometry. Results obtained from our flow cytometry screening indicate antiplatelet activity from novel phenolic compounds and their potential use as drugs for thrombosis and cardiovascular diseases.


Keywords: phenolic acids, flavonols, platelet biomarkers, P-selectin, flow cytometry

References

L.K. Jennings, “Mechanisms of platelet activation: Need for new strategies to protect against platelet-mediated atherothrombosis,” Thromb Haemostas, vol.103, pp. 248-257, 2009.

B. Jr Nagy, I.B. Debreceni, J. Kappelmayer, “Flow cytometric investigation of classical and alternative platelet activation markers,” J of Intern Fed Clin. Chem & Lab. medic, vol. 23, pp.11124-1134, 2012.

J.K. Cha, M.H. Jeong, J.Y. Jang, H.R. Bae, Y.J. Lim, J.S. Kim, S.H. Kim, and J.W Kim, “Serial measurement of surface expressions of CD63, P-selectin, and CD40 ligand on platelets in atherosclerotic ischemic stroke. A possible role of CD40 ligand on platelets in atherosclerotic ischemic stroke,” Cerebrovasc. Dis, vol.16, pp. 376-382, 2003.

N.W. Tsai, W.N, Cheng, C.F. Shaw, C.R. Jan, H.W. Chang, C.R. Huang, S.A.D. Chen, Y.C. Chuang, T.H. Lee, and C.H. Lu “Levels and value of platelet activation markers in different subtypes of acute non-cardio-embolic ischemic stroke,” Thromb Res, vol. 124, pp. 213-218, 2009.

C.L. Campbelli, S. Smyth, G. Montalescot, and S.R. Steinhubl, “Aspirin dose for the prevention of cardiovascular disease: a systematic review,” The J of the Am Med Assoc, vol. 298(18), pp. 2018-2024, 2007.

V.J. Marder, M.H. Rosove, and M. Minning, “Foundation and sites of action of antithrombotic agents, Best Practice and Research,” Clin Haematol, vol. 17, pp. 3-22, 2004.

A. Battaglia, E. Stragliotto, F. Heiman, “Indobufen in the prevention of cerebral ischemic attack (TIA): a prospective randomized, controlled study,” Thromb and Haemost, vol. 69, pp.1349-1393,1993.

P. Libby, “Mechanisms of acute coronary syndromes and their implications for therapy,” The New Eng. J of Medicine, vol. 368 (21), pp. 2004-2013, 2013.

B.E. Nignpense, K.A. Chinkwo, L. Christopher, C.L. Blanchard, and A.B. Santhakumarm, “Polyphenols: Modulators of Platelet Function and Platelet Microparticle Generation?” Int. J. Mol. Sci, vol. 21, pp. 146-153, 2020. https://doi.org/10.3390/ijms21010146

A. Moschona A, K.D. Kyriakidis, A.D. Kleontas, and M. Liakopoulou-Kyriakides, “Comparative study of natural phenolic acids and flavonols as antiplatelet and anti-inflammatory agents,” The Grant Medical J, vol. 2, pp. 57-66, 2017.

A. Moschona, K. Rouptsiou, S. Theodoridou, and M. Liakopoulou-Kyriakides, “Studies on polyphenols from sea buckthorn berries and pomegranate peels extracts. Recovery, bioactivities and encapsulation into polymers,” Int. J Pharm & Biol Sci, vol. 10, pp.158-171, 2020.

D. Menssen, K. Melber, N. Brandt, and E. Thiel, “The Use of Hirudin as Universal Anticoagulant in Haematology. Clinical Chemistry and Blood Grouping,” Clin Chem & Lab Medicine, vol. 39(12), pp. 1267-1277, 2002.

N.H. Wallén, M. Ladjevardi, J. Albert, and A. Bröijersén, “Influence of Different Anticoagulants on Platelet Aggregation in Whole Blood; A Comparison Between Citrate, Low Molecular Mass Heparin and Hirudin,” Thromb Res, vol. 87, pp.151-157, 1997.

M.L. Kalb, L. Potura, G. Scharbert, and S.A. Kozek-Langenecker, “The effect of ex vivo anticoagulants on whole blood platelet aggregation,” Platelets, vol. 20, pp. 7-11, 2009. doi: 10.1080/09537100802364076 PMID:19172515.

J. Kappelmayer, B. Jr Nagy, K. Miszti-Blasius, Z. Havessy, and H. Sertidi, “The emerging value of P-selectin as a disease marker,” Clin Chem Lab Med, vol. 42, pp. 475-486, 2004.

W. Russel, G. Duthle, “Plant secondary metabolites and gut health: the case for phenolic acids,” The proceedings of the Nutrition Society, vol. 10(2), pp. 389-396, 2011.

E. Barone, V. Calabrese, and C. Mancuso, “Ferulic acid its therapeutic potential as a hormetin for age-related diseases,” Biotechnol, vol. 10, pp. 97-108, 2009.

H.X. Zhang, H. Lin, C. Qu, YP. Tang, N.G. Li, J. Kai, G. Shang, and B. Li, “Design, synthesis, and in vitro antiplatelet aggregation Activities of Ferulic acid derivatives,” J of Chem, hindawi publishing corporation, article id 376527, 2015.

M. Modi, T. Goel, T. Das, S. Malik, S. Suri, A.K. Rawat, et al, “Ellagic acid & gallic acid from Lagerstroemia speciosa L inhibit HIV-1 infection through inhibition of HIV-1 protease & reverse transcriptase activity,” Indian J Med Res, vol. 137, pp. 540-548, 2013.

D. Lee, S. Eom, Y. Kim, H. Kim, M. Yim, S. Lee, et al, “Antibacterial and synergic effects of gallic acid-grafted-chitosan with beta-lactams against methicillin-resistant Staphylococcus aureus (MRSA),” Can J Microbiol, vol. 60, pp. 629-638, 2014.

C. Liao, K. Lai, A. Huang, J. Yang, J. Lin, S. Wu, et al, “Gallic acid inhibits migration and invasion in human osteosarcoma U-2 OS cells through suppressing the matrix metalloproteinase-2/-9, protein kinase B (PKB) and PKC signaling pathways,” Food Chem Toxicol, vol. 50, pp. 1734-1740, 2012.

A. Borges, C. Ferreira, M. Saavedra, and M. Simoes, “Antibacterial activity and mode of action of ferulic and gallic acids against pathogenic bacteria,” Microb Drug Resist, vol. 19, pp. 256–265, 2013.

A. Sahebkar, M. Reza Zirak, and A. Sahebkar, “Ellagic Acid: A Logical Lead for Drug Development?” Cur Pharmac Design, vol. 24(2), pp. 106-122, 2018.

C. Guo, S. Liu, Y. Guo, Y. Yin, J. Lin, X. Chen, and MA. Sun, “Comparative function-structural analysis of antiplatet and antiradical activities of flavonoid phytochemicals,” J Anim Plant Sci, vol. 4, pp. 926-935, 2014.

M. Bijak, R. Ziewiecki, J. Saluk, M. Ponczek, I. Pawlaczyk, H. Krotkiewski, P. Wachowicz, and P. Nowak, “Thrombin inhibitory activity of some polyphenolic compounds,” Med. Chem. Res, vol. 23, pp. 2324-2337, 2014.

G.P. Hubbard, S. Wolffram, J.A. Lovegrove, and J.M. Gibbins, “Ingestion of quercetin inhibits platelet aggregation and essential components of the collagen-stimulated platelet activation pathway in humans,” J Thromb Haemost, vol. 2(12), pp. 2138-2145, 2004.

K-H. Lee, E. Park, H-J. Lee, et al, “Effects of daily quercetin-rich supplementation on cardiometabolic risks in male smokers,” Nutr Res Pract, vol. 5(01), pp. 28–33, 2011.

A.R. Stainer, P. Sasikumar, A.P. Bye, A.J. Unswort, L.M. Holbrook, M. Tindall, J.A. Lovegrove, and J.M. Gibbins, “The Metabolites of the Dietary Flavonoid Quercetin Possess Potent Antithrombotic Activity, and Interact with Aspirin to Enhance Antiplatelet Effects,” TH Open, vol. 3(3), pp. e244–e258, 2019.

S.Cid-Ortega, and J.A. Monroy-Rivera, “Extraction of Kaempferol and Its Glycosides Using Supercritical Fluids from Plant Sources: A Review,” Food Technol Biotechnol, vol. 56(4), pp. 480-493, 2018.

A.Y. Chen, and Y.C. Chen, “A review of the dietary flavonoid, kaempferol on human health and cancer chemoprevention,” Food Chem, vol. 138(4), pp. 2099-2107, 2013. https://doi.org/10.1016/j.foodchem.2012.11.13.

Y. Cui, H. Morgenstern, S. Greenland, D.P. Tashkin, J.T. Mao, L. Cai, et al, “Dietary flavonoid intake and lung cancer -A population-based case-control study,” Cancer, vol. 112 (10), 2241-2248, 2008. https://doi.org/10.1002/cncr.2339831.

J.M. Calderon-Montano, E. Burgos-Morón, C. Pérez-Guerrero, and M. López-Lázaro, “A review on the dietary flavonoid kaempferol,” Mini Rev Med Chem, vol. 11(4), pp. 298-344, 2011. https://doi.org/10.2174/138955711795305335.

N.W. Tsai, W.N. Chang, C.F. Shaw, C.R. Jan, C.R. Chang, S.D. Chem, Y.C. Chuang, T.H. Lee, H.C. Wang, and C.H. Lu, “Levels and value of platelet activation markers in different sub types of acutenon-cardio-embopolic ischaemic stroke,” Thromb Res, vol. 124, pp. 213-218, 2009.

D.J. McCabe, P. Harrison, I.J. Macklie, PS. Sidhu, G. Purdy, A.S. Lawrie, H. Watt, M.M. Brown, and S.J. Machin, “Platelet degranulation and monocyte-platelet complex formation are increased in the acute and convalescent phases after ischaemic stroke or transient ischaemic attack,” Br J Haematol, vol. 125, pp. 777-787, 2004.

R.A. Preston, J.O. Coffey, B.J. Materson, M. Ledfordand, and A.B. Alonso, “Elevated platelet P-selectin expression and platelet activation in high risk patients with uncontrolled severe hypertension,” Atherosclerosis, vol. 192, pp. 148-154, 2007.

M. Lukasik, G. Dworacki, J. Kufel-Grabowska, C. Watala, W. Kozubski, “Chronic hyper-reactivity of platelets resulting in enhanced monocyte recruitment in patients after ischaemic stroke,” Platelets, vol. 20, pp. 235-241, 2009.

S. Fateh-Moghadam, P. Htum, B. Tomandi, D. Sander, K. Stellios, et al, “Hyper responsiveness of platelets in ischaemic stroke,” Thromb Haemost, vol. 97, pp. 974-978, 2007.

E. Csongrady, B. Jr Nagy, T. Fulop, Z. Varga, Z. Karanyl, M.T. Magyar, L. Olah, et al, “Increased levels of platelet activation markers are positively associated with carotid wall thickness and other atherosclerotic risk factors in obese patients,” Thromb Haemost, vol. 106, pp. 683-692, 2011.

F. Zeigel, S. Stephan, G. Hoheisel, D. Pfeiffer, C. Riehlmann, and M. Koksch, “P-selectin expression, platelet aggregation, and platelet-derived microparticle formation are increased in peripheral arterial disease,” Blood Coagul Fibrinol, vol. 11, pp.723-728, 2000.

T. Gremmel, R. Koppensteiner, and S. Panze, “Comparison of Aggregometry with Flow Cytometry for the Assessment of Agonists-Induced Platelet Reactivity in Patients on Dual Antiplatelet Therapy,” PLoS ONE 10:e0129666, 2015.

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How to Cite
Kyriakidis, K. D., Vartholomatos, E. G., & Markopoulos, G. S. (2021). Evaluation of Antiplatelet Activity of Phenolic Compounds by Flow Cytometry. European Journal of Medical and Health Sciences, 3(1), 165-170. https://doi.org/10.24018/ejmed.2021.3.1.703