The in vitro cytotoxic, genotoxic, oxidative damage potential of enoxaparin sodium in human peripheral blood mononuclear cells

Objectives: Enoxaparin sodium, low-molecular weight heparin (LMWH) indicated for the prophylaxis deep vein thrombosis. As far as we know, its cytotoxic, genotoxic and oxidative effects have never been studied on any cell lines. The purpose of the present study is to evaluate the in vitro cytotoxic, genotoxic damage potential and antioxidant/oxidant activity of enoxaparin sodium on primary human whole blood cultures. Methods: After exposure to different doses (from 0.5 to 100 mg/L) of enoxaparin sodium, cell viability was assessed by the cytotoxicity tests including MTT (3, (4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide) and lactate dehydrogenase (LDH) release assays. The antioxidant activity was measured by the total antioxidant capacity (TAC) and total oxidative stress (TOS) parameters. To determine the genotoxic damage potential, the rate of chromosomal aberrations (CAs) and 8-oxo-2'-deoxyguanosine (8-oxo-dG) levels were also assessed. Results: Cytotoxicity assays showed that treatment with enoxaparin sodium caused significant decreases in the cellular viability in a clear dose-dependent manner. Also, it was found that enoxaparin sodium did not alter the TAC and TOS levels. The genotoxicity assay showed that the formation of CAs was not observed in the lymphocytes. Likewise, the levels of 8-oxo-dG did not change in treated cultures as compared to control values. Conclusions: Enoxaparin sodium appeared to exhibit cytotoxic but not oxidative and genotoxic damage potentials in cultured human blood cells.

Keywords:

enoxaparin sodium, low-molecular-weight heparin, cytotoxicity, oxidative stress, DNA damage human peripheral blood mononuclear cells,

PDF

___

1. Parakh RS, Sabath DE. Venous thromboembolism: role of the clinical laboratory in diagnosis and management. J Appl Lab Med 2019;3:870-82.
2. Akhtar F, Wan X, Wu G, Kesse S, Wang S, He S. Low-molecular-weight heparins: reduced size particulate systems for improved therapeutic outcomes. Molecules 2018;23:1757.
3. Nadi S, Vreugdenburg TD, Atukorale Y, Ma N, Maddern G, Rovers M. Safety and effectiveness of aspirin and enoxaparin for venous thromboembolism prophylaxis after total hip and knee arthroplasty: a systematic review. ANZ J Surg 2019;89:1204-10.
4. Jupalli A, Iqbal AM. Enoxaparin. In StatPearls [Internet]. StatPearls Publishing. 2020. https://www.ncbi.nlm.nih.gov/books/NBK539865/.
5. Caprini JA, Arcelus J, Sehgal LR, Cohen EB, Reyna JJ. The use of low molecular weight heparins for the prevention of postoperative venous thromboembolism in general surgery. a survey of practice in the United States. Int Angiol 2002;21:78-85.
6. Tapson VF, Hyers TM, Waldo AL, Ballard DJ, Becker RC, Caprini J, et al. Antithrombotic therapy practices in US hospitals in an era of practice guidelines. Arch Intern Med 2005;165:1458-64.
7. MacDougall DA, Feliu AL, Boccuzzi SJ, Lin J. Economic burden of deep-vein thrombosis, pulmonary embolism, and post-thrombotic syndrome. Am J Health Syst Pharm 2006;63:5-15.
8. Tapson VF, Decousus H, Pin M, Chong BH, Froehlich JB, Monreal M, et al. Venous thromboembolism prophylaxis in acutely ill hospitalized medical patients: findings from the International Medical Prevention Registry on Venous Thromboembolism. Chest 2007;132:936-45.
9. Vats V, Nutescu EA, Theobald JC, Wojtynek JE, Schumock GT. Survey of hospitals for guidelines, policies, and protocols for anticoagulants. Am J Heal Pharm 2007;64:1203-8.
10. Haines ST, DiPiro JT, Talbert RL, Yee GC, Matzke GR, Wells BG, et al. TED Pharmacotherapy. 5th Edition. Vol. 1. Chapter 26. Venous Thromboembolism. McGraw-Hill Global Education Holdings. LLC., 2002: pp. 373-413.
11. Fareed J, Leong W, Hoppensteadt D. Generic low-molecular-weight heparins: some practical considerations. Semin Thromb Hemost 2004;30:703-13.
12. Garcia DA, Baglin TP, Weitz JI, Samama MM. Parenteral anticoagulants: Anthithrombotic Therapy and Prevention of Thrombosis, 9th ed: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines. Chest 2012;141(2 Suppl):e24S-e43S.
13. Fareed J, Fu K, Yang LH, Hoppensteadt DA. Pharmacokinetics of low molecular weight heparins in animal models. Semin Thromb Hemost 1999;25:51-5.
14. Merli GJ, Vanscoy GJ, Rihn TL, Groce JB 3rd, McCormick W. Applying scientific criteria to therapeutic interchange: A balanced analysis of low-molecular-weight heparins. J Thromb Thrombolysis 2001;11:247-59.
15. Mourier PAJ, Agut C, Souaifi-Amara H, Herman F, Viskov C. Analytical and statistical comparability of generic enoxaparins from the US market with the originator product. J Pharm Biomed Anal 2015;115:431-42.
16. Drouet L. Low molecular weight heparin biosimilars: how much similarity for how much clinical benefit? Target Oncol 2012;7:35-42.
17. Turkez H, Tozlu OO, Lima TC, de Brito AEM, de Sousa DP. A comparative evaluation of the cytotoxic and antioxidant activity of mentha crispa essential oil, its major constituent rotundifolone, and analogues on human glioblastoma. Oxid Med Cell Longev 2018:2083923.
18. Handschin AE, Trentz OA, Hoerstrup SP, Kock HJ, Wanner GA, Trentz O. Effect of low molecular weight heparin (dalteparin) and fondaparinux (Arixtra®) on human osteoblasts in vitro. Br J Surg 2005;92:177-83.
19. Erel O. A novel automated direct measurement method for total antioxidant capacity using a new generation, more stable ABTS radical cation. Clin Biochem 2004;37:277-8.
20. Turkez H, Aydin E, Aslan A. Xanthoria elegans (Link) (lichen) extract counteracts DNA damage and oxidative stress of mitomycin C in human lymphocytes. Cytotechnol 2012;64:679-86.
21. Erel O. A new automated colorimetric method for measuring total oxidant status. Clin Biochem 2005;38:1103-11.
22. Aydin E, Türkez H, Taşdemir S. Anticancer and antioxidant properties of terpinolene in rat brain cells. Arh Hig Rada Toksikol 2013;64:415-24.
23. IPCS 1985. International Program on Chemical Safety Environmental Health Criteria 46. Guidelines for the Study of Genetic Effects in Human Populations. Geneva, Switzerland: WHO, 1985: pp. 45-54.
24. Geyikoglu F, Turkez H. Protective effect of sodium selenite on genotoxicity to human whole blood cultures induced by aflatoxin B-1. Brazil Arch Biol Technol 2005;48:905-10.
25. Floyd RA, Watson JJ, Wong PK, Altmiller DH, Rickard RC. Hydroxyl free radical adduct of deoxyguanosine: sensitive detection and mechanisms of formation. Free Radic Res Commun 1986;1:163-72.
26. Hemeda H, Kalz J, Walenda G, Lohmann M, Wagner W. Heparin concentration is critical for cell culture with human platelet lysate. Cytotherapy 2013;15:1174-81.
27. Castellot JJ, Cochran DL, Karnovsky MJ. Effect of heparin on vascular smooth muscle cells. I. Cell metabolism. J Cell Physiol 1985;124:21-8.
28. Castellot JJ Jr, Wong K, Herman B. Binding and internalization of heparin by vascular smooth muscle cells. J Cell Physiol 1985;124:13-20.
29. Clowes AW, Reidy MA, Clowes MM. Kinetics of cellular proliferation after arterial injury. I. Smooth muscle growth in the absence of endothelium. Lab Invest 1983;49:327-33.
30. Clowes AW, Clowes MM, Reidy MA. Kinetics of cellular proliferation after arterial injury. III. Endothelial and smooth muscle growth in chronically denuded vessels. Lab Invest 1986;54:295-303.
31. Karnovsky MJ, Ch B, Sc D. Regulation of vascular smooth muscle cell growth by heparin and heparan sulfates. Semin Thromb 1987;13:489-503.
32. Shastri MD, Stewart N, Eapen M, Peterson GM, Zaidi STR, Gueven N, et al. Opposing effects of low molecular weight heparins on the release of inflammatory cytokines from peripheral blood mononuclear cells of asthmatics. PLoS One 2015;10:e0118798.
33. Shastri MD, Stewart N, Horne J, Zaidi ST, Sohal SS, Peterson GM, et al. Non-anticoagulant fractions of enoxaparin suppress inflammatory cytokine release from peripheral blood mononuclear cells of allergic asthmatic individuals. Plos One 2015;10:e0128803.
34. Poyrazoglu OK, Dogukan A, Yalniz M, Seckin D, Gunal AL. Acute effect of standard heparin versus low molecular weight heparin on oxidative stress and inflammation in hemodialysis patients. Ren Fail 2006;28:723-7.
35. Deepa PR, Varalakshmi P. Protective effect of low molecular weight heparin on oxidative injury and cellular abnormalities in adriamycin-induced cardiac and hepatic toxicity. Chem Biol Interact 2003;146:201-10.
36. Uysal II, Karabulut AK, Ozdemir K, Aksoy M, Altunkeser BB, Acar H. Investigation of direct toxic and teratogenic effects of anticoagulants on rat embryonic development using in vitro culture method and genotoxicity assay. Anat Histol Embryol 2006;35:84-92.