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Neuroprotective effect of heparin Trisulfated disaccharide on ischemic stroke

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Abstract

Cells undergoing hypoxia experience intense cytoplasmic calcium (Ca2+) overload. High concentrations of intracellular calcium ([Ca2+]i) can trigger cell death in the neural tissue, a hallmark of stroke. Neural Ca2+ homeostasis involves regulation by the Na+/Ca2+ exchanger (NCX). Previous data published by our group showed that a product of the enzymatic depolymerization of heparin by heparinase, the unsaturated trisulfated disaccharide (TD; ΔU, 2S-GlcNS, 6S), can accelerate Na+/Ca2+ exchange via NCX, in hepatocytes and aorta vascular smooth muscle cells. Thus, the objective of this work was to verify whether TD could act as a neuroprotective agent able to prevent neuronal cell death by reducing [Ca2+]i. Pretreatment of N2a cells with TD reduced [Ca2+]i rise induced by thapsigargin and increased cell viability under [Ca2+]I overload conditions and in hypoxia. Using a murine model of stroke, we observed that pretreatment with TD decreased cerebral infarct volume and cell death. However, when mice received KB-R7943, an NCX blocker, the neuroprotective effect of TD was abolished, strongly suggesting that this neuroprotection requires a functional NCX to happen. Thus, we propose TD-NCX as a new therapeutic axis for the prevention of neuronal death induced by [Ca2+]i overload.

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References

  1. Benjamin, E.J., Blaha, M.J., Chiuve, S.E., Cushman, M., Das, S.R., Deo, R., de Ferranti, S.D., Floyd, J., Fornage, M., Gillespie, C., Isasi, C.R., Jiménez, M.C., Jordan, L.C., Judd, S.E., Lackland, D., Lichtman, J.H., Lisabeth, L., Liu, S., Longenecker, C.T., Mackey, R.H., Matsushita, K., Mozaffarian, D., Mussolino, M.E., Nasir, K., Neumar, R.W., Palaniappan, L., Pandey, D.K., Thiagarajan, R.R., Reeves, M.J., Ritchey, M., Rodriguez, C.J., Roth, G.A., Rosamond, W.D., Sasson, C., Towfighi, A., Tsao, C.W., Turner, M.B., Virani, S.S., Voeks, J.H., Willey, J.Z., Wilkins, J.T., Wu, J.H., Alger, H.M., Wong, S.S., Muntner, P., American Heart Association Statistics Committee and Stroke Statistics Subcommittee: Heart disease and stroke Statistics-2017 update: a report from the American Heart Association. Circulation. 135(10), e146–e603 (2017). https://doi.org/10.1161/CIR.0000000000000485

    Article  PubMed  PubMed Central  Google Scholar 

  2. Neumar, R.W.: Molecular mechanisms of ischemic neuronal injury. Ann. Emerg. Med. 36(5), 483–506 (2000). https://doi.org/10.1067/mem.2000.110995

    Article  CAS  PubMed  Google Scholar 

  3. Kalogeris, T., Baines, C.P., Krenz, M., Korthuis, R.J.: Ischemia/Reperfusion. Compr Physiol. 7(1), 113–170 (2016). https://doi.org/10.1002/cphy.c160006

    Article  PubMed  PubMed Central  Google Scholar 

  4. Carini, R., Bellomo, G., Dianzani, M.U., Albano, E.: Evidence for a sodium-dependent calcium influx in isolated rat hepatocytes undergoing ATP depletion. Biochem. Biophys. Res. Commun. 202(1), 360–366 (1994). https://doi.org/10.1006/bbrc.1994.1936

    Article  CAS  PubMed  Google Scholar 

  5. Song, H.A., Kim, Y.S., Cho, H.J., Kim, S.I., Kang, M.J., Kim, J.H., Min, H.J., Kang, J.W., Yoon, J.H., Kim, C.H.: Hypoxia modulates epithelial permeability via regulation of vascular endothelial growth factor in airway epithelia. Am. J. Respir. Cell Mol. Biol. 57(5), 527–535 (2017). https://doi.org/10.1165/rcmb.2016-0080OC

    Article  CAS  PubMed  Google Scholar 

  6. Giladi, M., Lee, S.Y., Ariely, Y., Teldan, Y., Granit, R., Strulovich, R., Haitin, Y., Chung, K.Y., Khananshvili, D.: Structure-based dynamic arrays in regulatory domains of sodium-calcium exchanger (NCX) isoforms. Sci. Rep. 7(1), 993 (2017). https://doi.org/10.1038/s41598-017-01102-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Ren, X., Philipson, K.D.: The topology of the cardiac Na+/Ca2+ exchanger, NCX1. J. Mol. Cell. Cardiol. 57, 68–71 (2013). https://doi.org/10.1016/j.yjmcc.2013.01.010

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Shenoda, B.: The role of Na+/Ca2+ exchanger subtypes in neuronal ischemic injury. Transl. Stroke Res. 6(3), 181–190 (2015). https://doi.org/10.1007/s12975-015-0395-9

    Article  CAS  PubMed  Google Scholar 

  9. Silva, M.E., Dietrich, C.P.: Structure of heparin. Characterization of the products formed from heparin by the action of a heparinase and a heparitinase from Flavobacterium heparinum. J. Biol. Chem. 250(17), 6841–6846 (1975)

    Article  CAS  Google Scholar 

  10. Vasques, E.R., Cunha, J.E., Coelho, A.M., Sampietre, S.N., Patzina, R.A., Abdo, E.E., et al.: Trisulfate disaccharide decreases calcium overload and protects liver injury secondary to liver ischemia/reperfusion. PLoS One. 11(2), e0149630 (2016). https://doi.org/10.1371/journal.pone.0149630

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Shinjo, S.K., Tersariol, I.L., Oliveira, V., Nakaie, C.R., Oshiro, M.E., Ferreira, A.T., et al.: Heparin and heparan sulfate disaccharides bind to the exchanger inhibitor peptide region of Na+/Ca2+ exchanger and reduce the cytosolic calcium of smooth muscle cell lines. Requirement of C4-C5 unsaturation and 1--> 4 glycosidic linkage for activity. J. Biol. Chem. 277(50), 48227–48233 (2002). https://doi.org/10.1074/jbc.M205867200

    Article  CAS  PubMed  Google Scholar 

  12. Nader, H.B., Tersariol, I.L., Dietrich, C.P.: Antihemostatic activity of heparin disaccharides and oligosaccharides obtained by chemical and enzymatic fragmentation: reversal of the hemorrhagic activity by ATP and myosin. Thromb. Res. 54(3), 207–214 (1989). https://doi.org/10.1016/0049-3848(89)90228-4

    Article  CAS  PubMed  Google Scholar 

  13. de Godoy, C.M.G., Vasques, Ê., Caricati-Neto, A., Tavares, J.G.P., Alves, B.J., Duarte, J., et al.: Heparin oligosaccharides have antiarrhythmic effect by accelerating the sodium-calcium exchanger. Front Cardiovasc Med. 5, 67 (2018). https://doi.org/10.3389/fcvm.2018.00067

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Nader, H.B., Tersariol, I.L., Dietrich, C.P.: Structural requirements of heparin disaccharides responsible for hemorrhage: reversion of the antihemostatic effect by ATP. FASEB J. 3(12), 2420–2424 (1989). https://doi.org/10.1096/fasebj.3.12.2529161

    Article  CAS  PubMed  Google Scholar 

  15. Nader, H.B., Porcionatto, M.A., Tersariol, I.L., Pinhal, M.A., Oliveira, F.W., Moraes, C.T., Dietrich, C.P.: Purification and substrate specificity of heparitinase I and heparitinase II from Flavobacterium heparinum. Analyses of the heparin and heparan sulfate degradation products by 13C NMR spectroscopy. J. Biol. Chem. 265(28), 16807–16813 (1990)

    Article  CAS  Google Scholar 

  16. Mao, A.J., Bechberger, J., Lidington, D., Galipeau, J., Laird, D.W., Naus, C.C.: Neuronal differentiation and growth control of neuro-2a cells after retroviral gene delivery of connexin43. J. Biol. Chem. 275(44), 34407–34414 (2000). https://doi.org/10.1074/jbc.M003917200

    Article  CAS  PubMed  Google Scholar 

  17. Wei, J., Xu, H., Shi, L., Tong, J., Zhang, J.: Trimetazidine protects cardiomyocytes against hypoxia-induced injury through ameliorates calcium homeostasis. Chem. Biol. Interact. 236, 47–56 (2015). https://doi.org/10.1016/j.cbi.2015.04.022

    Article  CAS  PubMed  Google Scholar 

  18. Labat-gest, V., Tomasi, S.: Photothrombotic ischemia: a minimally invasive and reproducible photochemical cortical lesion model for mouse stroke studies. J. Vis. Exp. 76, (2013). https://doi.org/10.3791/50370

  19. Paxinos, G., Franklin, K.B.J.: The Mouse Brain in Stereotaxic Coordinates, 2nd edn. Academic Press, San Diego (2001)

  20. Kramer, M., Dang, J., Baertling, F., Denecke, B., Clarner, T., Kirsch, C., Beyer, C., Kipp, M.: TTC staining of damaged brain areas after MCA occlusion in the rat does not constrict quantitative gene and protein analyses. J. Neurosci. Methods. 187(1), 84–89 (2010). https://doi.org/10.1016/j.jneumeth.2009.12.020

    Article  PubMed  Google Scholar 

  21. Hankey, G.J.: Stroke. Stroke. Lancet. 389(10069), 641–654 (2017). https://doi.org/10.1016/S0140-6736(16)30962-X

    Article  PubMed  Google Scholar 

  22. Carmichael, S.T.: The 3 Rs of stroke biology: radial, relayed, and regenerative. Neurotherapeutics. 13(2), 348–359 (2016). https://doi.org/10.1007/s13311-015-0408-0

    Article  CAS  PubMed  Google Scholar 

  23. Zündorf, G., Reiser, G.: Calcium dysregulation and homeostasis of neural calcium in the molecular mechanisms of neurodegenerative diseases provide multiple targets for neuroprotection. Antioxid. Redox Signal. 14(7), 1275–1288 (2011). https://doi.org/10.1089/ars.2010.3359

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Jeon, D., Chu, K., Jung, K.H., Kim, M., Yoon, B.W., Lee, C.J., Oh, U., Shin, H.S.: Na(+)/Ca(2+) exchanger 2 is neuroprotective by exporting Ca(2+) during a transient focal cerebral ischemia in the mouse. Cell Calcium. 43(5), 482–491 (2008). https://doi.org/10.1016/j.ceca.2007.08.003

    Article  CAS  PubMed  Google Scholar 

  25. Molinaro, P., Cuomo, O., Pignataro, G., Boscia, F., Sirabella, R., Pannaccione, A., Secondo, A., Scorziello, A., Adornetto, A., Gala, R., Viggiano, D., Sokolow, S., Herchuelz, A., Schurmans, S., di Renzo, G., Annunziato, L.: Targeted disruption of Na+/Ca2+ exchanger 3 (NCX3) gene leads to a worsening of ischemic brain damage. J. Neurosci. 28(5), 1179–1184 (2008). https://doi.org/10.1523/JNEUROSCI.4671-07.2008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Molinaro, P., Cantile, M., Cuomo, O., Secondo, A., Pannaccione, A., Ambrosino, P., Pignataro, G., Fiorino, F., Severino, B., Gatta, E., Sisalli, M.J., Milanese, M., Scorziello, A., Bonanno, G., Robello, M., Santagada, V., Caliendo, G., di Renzo, G., Annunziato, L.: Neurounina-1, a novel compound that increases Na+/Ca2+ exchanger activity, effectively protects against stroke damage. Mol. Pharmacol. 83(1), 142–156 (2013). https://doi.org/10.1124/mol.112.080986

    Article  CAS  PubMed  Google Scholar 

  27. Cerullo, P., Brancaccio, P., Anzilotti, S., Vinciguerra, A., Cuomo, O., Fiorino, F., Severino, B., di Vaio, P., di Renzo, G., Annunziato, L., Pignataro, G.: Acute and long-term NCX activation reduces brain injury and restores behavioral functions in mice subjected to neonatal brain ischemia. Neuropharmacology. 135, 180–191 (2018). https://doi.org/10.1016/j.neuropharm.2018.03.017

    Article  CAS  PubMed  Google Scholar 

  28. He, Z., Feng, S., Tong, Q., Hilgemann, D.W., Philipson, K.D.: Interaction of PIP(2) with the XIP region of the cardiac Na/Ca exchanger. Am J Physiol Cell Physiol. 278, C661–C666 (2000). https://doi.org/10.1152/ajpcell.2000.278.4.c661

    Article  CAS  PubMed  Google Scholar 

  29. Wu, G., Xie, X., Lu, Z.H., Ledeen, R.W.: Sodium-calcium exchanger complexed with GM1 ganglioside in nuclear membrane transfers calcium from nucleoplasm to endoplasmic reticulum. Proc. Natl. Acad. Sci. U. S. A. 106, 10829–10834 (2009). https://doi.org/10.1073/pnas.0903408106

    Article  PubMed  PubMed Central  Google Scholar 

  30. Buenaflor, F.G.: Recurrence rate of ischemic stroke: a single center experience. J. Neurol. Sci. 381, 399 (2017). https://doi.org/10.1016/j.jns.2017.08.3340

    Article  Google Scholar 

  31. Diener, H.C., Weimar, C., Weber, R.: Antiplatelet therapy in secondary stroke prevention--state of the art. J. Cell. Mol. Med. 14(11), 2552–2560 (2010). https://doi.org/10.1111/j.1582-4934.2010.01163.x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Whelihan, M.F., Cooley, B., Xu, Y., Pawlinski, R., Liu, J., Key, N.S.: In vitro and in vivo characterization of a reversible synthetic heparin analog. Thromb. Res. 138, 121–129 (2016). https://doi.org/10.1016/j.thromres.2015.12.007

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by Fundação de Amparo à Pesquisa de São Paulo - FAPESP (2011/00526-7; 2015/03964-6; 2017/18765-4), Conselho Nacional de Desenvolvimento Científico e Tecnológico - CNPq (402391/2013-3; 465656/2014-5) and National Institute of General Medical Sciences (GM23244), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – CAPES (Finance Code 001).

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Authors and Affiliations

  1. Laboratory of Molecular Neurobiology, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Gabrielly M. D. Chiarantin, Lina M. Delgado-Garcia, Laura N. Zamproni & Marimélia Porcionatto

  2. Department of Biochemistry, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Gabrielly M. D. Chiarantin, Lina M. Delgado-Garcia, Laura N. Zamproni, Marcelo A. Lima, Helena B. Nader, Ivarne L. S. Tersariol & Marimélia Porcionatto

  3. Molecular & Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, ST5 5BG, UK

    Marcelo A. Lima

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  1. Gabrielly M. D. Chiarantin
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Correspondence to Ivarne L. S. Tersariol or Marimélia Porcionatto.

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Chiarantin, G.M.D., Delgado-Garcia, L.M., Zamproni, L.N. et al. Neuroprotective effect of heparin Trisulfated disaccharide on ischemic stroke. Glycoconj J 38, 35–43 (2021). https://doi.org/10.1007/s10719-020-09966-4

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