Nitric Oxide in β-Thalassemia

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Thanaporn Sriwantana
Sirada Srihirun
Nathawut Sibmooh

Abstract

Nitric oxide (NO) has multiple physiologic functions and its decrease or increase is associated with pathophysiology of diseases. NO is produced by nitric oxide synthase (NOS)-dependent and NOS-independent pathways. The NO concentrations and rate of production from different pathways are determinants of its biological functions. NO at low levels produced by constitutive NOS maintains adequate blood flow and inhibits platelets in normal situation, while NO at high levels produced by inducible NOS plays role in pathophysiologic process. In NOS-independent pathway, NO is produced from nitrite by the nitrite reductase activity of deoxyhemoglobin. In thalassemia, iron overload and oxidative stress lead to endothelial dysfunction and decreased NO, which are associated with platelet hyperactivity and pulmonary hypertension. Here, the preclinical and clinical studies of NO-related and nitrite therapy in β-thalassemia are reviewed.

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References

Cosby K, Partovi KS, Crawford JH, Patel RP, Reiter CD, Martyr S, et al. Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation. Nat Med. 2003 Dec;9(12):1498-505.

Kim-Shapiro DB, Gladwin MT. Mechanisms of nitrite bioactivation. Nitric Oxide. 2014 Apr 30;38:58-68.

Lundberg JO, Weitzberg E, Gladwin MT. The nitrate-nitrite-nitric oxide pathway in physiology and therapeutics. Nat Rev Drug Discov. 2008 Feb;7(2):156-67.

Srihirun S, Sriwantana T, Unchern S, Kittikool D, Noulsri E, Pattanapanyasat K, et al. Platelet inhibition by nitrite is dependent on erythrocytes and deoxygenation. PLoS One. 2012;7(1):e30380.

Beltran B, Mathur A, Duchen MR, Erusalimsky JD, Moncada S. The effect of nitric oxide on cell respiration: A key to understanding its role in cell survival or death. Proc Natl Acad Sci U S A. 2000 Dec 19;97(26):14602-7.

Zhang Z, Naughton D, Winyard PG, Benjamin N, Blake DR, Symons MC. Generation of nitric oxide by a nitrite reductase activity of xanthine oxidase: a potential pathway for nitric oxide formation in the absence of nitric oxide synthase activity. Biochem Biophys Res Commun. 1998 Aug 28;249(3):767-72.

Pannala AS, Mani AR, Spencer JP, Skinner V, Bruckdorfer KR, Moore KP, et al. The effect of dietary nitrate on salivary, plasma, and urinary nitrate metabolism in humans. Free Radic Biol Med. 2003 Mar 1;34(5):576-84.

McKnight GM, Duncan CW, Leifert C, Golden MH. Dietary nitrate in man: friend or foe? Br J Nutr. 1999 May;81(5):349-58.

Bogdan C. Nitric oxide and the immune response. Nat Immunol. 2001 Oct; 2(10): 907-16.

Kim PK, Zamora R, Petrosko P, Billiar TR. The regulatory role of nitric oxide in apoptosis. Int Immunopharmacol. 2001 Aug;1(8):1421-41.

Togashi H, Sakuma I, Yoshioka M, Kobayashi T, Yasuda H, Kitabatake A, et al. A central nervous system action of nitric oxide in blood pressure regulation. J Pharmacol Exp Ther. 1992 Jul;262(1):343-7.

Kim N, Azadzoi KM, Goldstein I, Saenz de Tejada I. A nitric oxide-like factor mediates nonadrenergic-noncholinergic neurogenic relaxation of penile corpus cavernosum smooth muscle. J Clin Invest. 1991 Jul;88(1):112-8.

Gladwin MT, Kim-Shapiro DB. The functional nitrite reductase activity of the hemeglobins. Blood. 2008 Oct 1;112(7):2636-47.

Huang Z, Shiva S, Kim-Shapiro DB, Patel RP, Ringwood LA, Irby CE, et al. Enzymatic function of hemoglobin as a nitrite reductase that produces NO under allosteric control. J Clin Invest. 2005 Aug;115(8):2099-107.

Doyle MP, Pickering RA, DeWeert TM, Hoekstra JW, Pater D. Kinetics and mechanism of the oxidation of human deoxyhemoglobin by nitrites. J Biol Chem. 1981 Dec 10;256(23):12393-8.

Shiva S, Huang Z, Grubina R, Sun J, Ringwood LA, MacArthur PH, et al. Deoxymyoglobin is a nitrite reductase that generates nitric oxide and regulates mitochondrial respiration. Circ Res. 2007 Mar 16;100(5):654-61.

Tiso M, Tejero J, Basu S, Azarov I, Wang X, Simplaceanu V, et al. Human neuroglobin functions as a redox-regulated nitrite reductase. J Biol Chem. 2011 May 20;286(20):18277-89.

Basu S, Azarova NA, Font MD, King SB, Hogg N, Gladwin MT, et al. Nitrite reductase activity of cytochrome c. J Biol Chem. 2008 Nov 21;283(47):32590-7.

Vanin AF, Bevers LM, Slama-Schwok A, van Faassen EE. Nitric oxide synthase reduces nitrite to NO under anoxia. Cell Mol Life Sci. 2007 Jan;64(1):96-103.

Gautier C, van Faassen E, Mikula I, Martasek P, Slama-Schwok A. Endothelial nitric oxide synthase reduces nitrite anions to NO under anoxia. Biochem Biophys Res Commun. 2006 Mar 17;341(3):816-21.

Mikula I, Durocher S, Martasek P, Mutus B, Slama-Schwok A. Isoform-specific differences in the nitrite reductase activity of nitric oxide synthases under hypoxia. Biochem J. 2009 Mar 15;418(3):673-82.

Millar TM, Stevens CR, Benjamin N, Eisenthal R, Harrison R, Blake DR. Xanthine oxidoreductase catalyses the reduction of nitrates and nitrite to nitric oxide under hypoxic conditions. FEBS Lett. 1998 May 8;427(2):225-8.

Li H, Kundu TK, Zweier JL. Characterization of the magnitude and mechanism of aldehyde oxidase-mediated nitric oxide production from nitrite. J Biol Chem. 2009 Dec 4;284(49):33850-8.

Sparacino-Watkins CE, Tejero J, Sun B, Gauthier MC, Thomas J, Ragireddy V, et al. Nitrite reductase and nitric-oxide synthase activity of the mitochondrial molybdopterin enzymes mARC1 and mARC2. J Biol Chem. 2014 Apr 11;289(15): 10345-58.

Xu Q, Wink DA, Colton CA. Nitric oxide production and regulation of neuronal NOS in tyrosine hydroxylase containing neurons. Exp Neurol. 2004 Aug;188(2): 341-50.

Kelley JB, Anderson KL, Altmann SL, Itzhak Y. Long-term memory of visually cued fear conditioning: roles of the neuronal nitric oxide synthase gene and cyclic AMP response element-binding protein. Neuroscience. 2011 Feb 3;174: 91-103.

Sanders KM, Ward SM. Nitric oxide and its role as a non-adrenergic, non-cholinergic inhibitory neurotransmitter in the gastrointestinal tract. Br J Pharmacol. 2019 Jan;176(2):212-27.

Rajfer J, Aronson WJ, Bush PA, Dorey FJ, Ignarro LJ. Nitric oxide as a mediator of relaxation of the corpus cavernosum in response to nonadrenergic, noncholinergic neurotransmission. N Engl J Med. 1992 Jan 9;326(2):90-4.

Li LM, Kilbourn RG, Adams J, Fidler IJ. Role of nitric oxide in lysis of tumor cells by cytokine-activated endothelial cells. Cancer Res. 1991 May 15;51(10): 2531-5.

Connelly ST, Macabeo-Ong M, Dekker N, Jordan RC, Schmidt BL. Increased nitric oxide levels and iNOS over-expression in oral squamous cell carcinoma. Oral Oncol. 2005 Mar;41(3):261-7.

D'Ambrosio SM, Gibson-D'Ambrosio RE, Brady T, Oberyszyn AS, Robertson FM. Mechanisms of nitric oxide-induced cytotoxicity in normal human hepatocytes. Environ Mol Mutagen. 2001;37(1):46-54.

Green SJ, Mellouk S, Hoffman SL, Meltzer MS, Nacy CA. Cellular mechanisms of nonspecific immunity to intracellular infection: cytokine-induced synthesis of toxic nitrogen oxides from L-arginine by macrophages and hepatocytes. Immunol Lett. 1990 Aug;25(1-3):15-9.

Fehsel K, Jalowy A, Qi S, Burkart V, Hartmann B, Kolb H. Islet cell DNA is a target of inflammatory attack by nitric oxide. Diabetes. 1993 Mar;42(3):496-500.

Jaiswal M, LaRusso NF, Burgart LJ, Gores GJ. Inflammatory cytokines induce DNA damage and inhibit DNA repair in cholangiocarcinoma cells by a nitric oxide-dependent mechanism. Cancer Res. 2000 Jan 1;60(1):184-90.

Forstermann U, Mulsch A, Bohme E, Busse R. Stimulation of soluble guanylate cyclase by an acetylcholine-induced endothelium-derived factor from rabbit and canine arteries. Circ Res. 1986 Apr;58(4):531-8.

Rapoport RM, Draznin MB, Murad F. Endothelium-dependent relaxation in rat aorta may be mediated through cyclic GMP-dependent protein phosphorylation. Nature. 1983 Nov 10-16;306(5939):174-6.

Liao JK. Linking endothelial dysfunction with endothelial cell activation. J Clin Invest. 2013 Feb;123(2):540-1.

Sato J, Nair K, Hiddinga J, Eberhardt NL, Fitzpatrick LA, Katusic ZS, O'Brien T. eNOS gene transfer to vascular smooth muscle cells inhibits cell proliferation via upregulation of p27 and p21 and not apoptosis. Cardiovasc Res. 2000 Sep; 47(4):697-706.

Tanner FC, Meier P, Greutert H, Champion C, Nabel EG, Luscher TF. Nitric oxide modulates expression of cell cycle regulatory proteins: a cytostatic strategy for inhibition of human vascular smooth muscle cell proliferation. Circulation. 2000 Apr 25;101(16):1982-9.

Fukumura D, Gohongi T, Kadambi A, Izumi Y, Ang J, Yun CO, et al. Predominant role of endothelial nitric oxide synthase in vascular endothelial growth factor-induced angiogenesis and vascular permeability. Proc Natl Acad Sci U S A. 2001 Feb 27;98(5):2604-9.

Rodriguez C, Vitturi DA, He J, Vandromme M, Brandon A, Hutchings A, et al. Sodium nitrite therapy attenuates the hypertensive effects of HBOC-201 via nitrite reduction. Biochem J. 2009 Aug 27;422(3):423-32.

Rassaf T, Flogel U, Drexhage C, Hendgen-Cotta U, Kelm M, Schrader J. Nitrite reductase function of deoxymyoglobin: oxygen sensor and regulator of cardiac energetics and function. Circ Res. 2007 Jun 22;100(12):1749-54.

Hendgen-Cotta UB, Merx MW, Shiva S, Schmitz J, Becher S, Klare JP, et al. Nitrite reductase activity of myoglobin regulates respiration and cellular viability in myocardial ischemia-reperfusion injury. Proc Natl Acad Sci U S A. 2008 July 22;105(29):10256-61.

44. Bjorne HH, Petersson J, Phillipson M, Weitzberg E, Holm L, Lundberg JO. Nitrite in saliva increases gastric mucosal blood flow and mucus thickness.

J Clin Invest. 2004 Jan;113(1):106-14.

Kim DJ, Roe CA, Somani YB, Moore DJ, Barrett MA, Flanagan M, et al. Effects of acute dietary nitrate supplementation on aortic blood pressures and pulse wave characteristics in post-menopausal women. Nitric Oxide. 2019 Apr 1; 85:10-16.

Petersson J, Carlstrom M, Schreiber O, Phillipson M, Christoffersson G, Jagare A, et al. Gastroprotective and blood pressure lowering effects of dietary nitrate are abolished by an antiseptic mouthwash. Free Radic Biol Med. 2009 Apr 15; 46(8):1068-75.

Webb AJ, Patel N, Loukogeorgakis S, Okorie M, Aboud Z, Misra S, et al. Acute blood pressure lowering, vasoprotective, and antiplatelet properties of dietary nitrate via bioconversion to nitrite. Hypertension. 2008 Mar;51(3):784-90.

Jia L, Bonaventura C, Bonaventura J, Stamler JS. S-nitrosohaemoglobin: a dynamic activity of blood involved in vascular control. Nature. 1996 Mar 21; 380(6571):221-6.

Yamsri S, Singha K, Prajantasen T, Taweenan W, Fucharoen G, Sanchaisuriya K, Fucharoen S. A large cohort of β(+)-thalassemia in Thailand: molecular, hematological and diagnostic considerations. Blood Cells Mol Dis. 2015 Feb; 54(2):164-9.

Olivieri NF. The β-thalassemias. N Engl J Med. 1999 July 8;341(2):99-109.

Mokhtar GM, Gadallah M, El Sherif NH, Ali HT. Morbidities and mortality in transfusion-dependent β-thalassemia patients (single-center experience). Pediatr Hematol Oncol. 2013 Mar;30(2):93-103.

Rajaeefard A, Hajipour M, Tabatabaee HR, Hassanzadeh J, Rezaeian S, Moradi Z, et al. Analysis of survival data in thalassemia patients in Shiraz, Iran. Epidemiol Health. 2015 July 7;37:e2015031.

Zamani R, Khazaei S, Rezaeian S. Survival analysis and its associated factors of β thalassemia major in hamadan province. Iran J Med Sci. 2015 May;40(3):233-9.

Schrier SL. Thalassemia: pathophysiology of red cell changes. Annu Rev Med. 1994;45:211-8.

Thein SL. Pathophysiology of β thalassemia--a guide to molecular therapies. Hematology Am Soc Hematol Educ Program. 2005:31-7.

Kremastinos DT, Farmakis D, Aessopos A, Hahalis G, Hamodraka E, Tsiapras D, et al. β-Thalassemia cardiomyopathy: history, present considerations, and future perspectives. Circ Heart Fail. 2010 May;3(3):451-8.

Kolios M, Korantzopoulos P, Vlahos AP, Kapsali E, Briasoulis E, Goudevenos JA. Electrocardiographic abnormalities and arrhythmic risk markers in adult patients with β thalassemia major. Int J Cardiol. 2016 Oct 15;221:932-6.

Amoozgar H, Zeighami S, Haghpanah S, Karimi M. A comparison of heart function and arrhythmia in clinically asymptomatic patients with β-thalassemia intermedia and β-thalassemia major. Hematology. 2017 Jan;22(1):25-9.

Rago A, Russo V, Papa AA, Ciardiello C, Pannone B, Mayer MC, et al. The role of the atrial electromechanical delay in predicting atrial fibrillation in β-thalassemia major patients. J Interv Card Electrophysiol. 2017 Mar;48(2):147-57.

Russo V, Rago A, Pannone B, Papa AA, Mayer MC, Spasiano A, et al. Atrial fibrillation and β thalassemia major: The predictive role of the 12-lead electro-cardiogram analysis. Indian Pacing Electrophysiol J. 2014 May 25;14(3):121-32.

Kremastinos DT, Tiniakos G, Theodorakis GN, Katritsis DG, Toutouzas PK. Myocarditis in β-thalassemia major. A cause of heart failure. Circulation. 1995 Jan 1;91(1):66-71.

Eldor A, Rachmilewitz EA. The hypercoagulable state in thalassemia. Blood. 2002 Jan 1;99(1):36-43.

Taher AT, Otrock ZK, Uthman I, Cappellini MD. Thalassemia and hyper-coagulability. Blood Rev. 2008 Sep;22(5):283-92.

Atichartakarn V, Angchaisuksiri P, Aryurachai K, Chuncharunee S, Thakkinstian A. In vivo platelet activation and hyperaggregation in hemoglobin E/β-thalassemia: a consequence of splenectomy. Int J Hematol. 2003 Apr;77(3):299-303.

Opartkiattikul N, Funahara Y, Fucharoen S, Talalak P. Increase in spontaneous platelet aggregation in β-thalassemia/hemoglobin E disease: a consequence of splenectomy. Southeast Asian J Trop Med Public Health. 1992;23 Suppl 2:36-41.

Ruf A, Pick M, Deutsch V, Patscheke H, Goldfarb A, Rachmilewitz EA, et al. In-vivo platelet activation correlates with red cell anionic phospholipid exposure in patients with β-thalassaemia major. Br J Haematol. 1997 July;98(1):51-6.

Fayed MA, Abdel-Hady HE, Hafez MM, Salama OS, Al-Tonbary YA. Study of platelet activation, hypercoagulable state, and the association with pulmonary hypertension in children with β-thalassemia. Hematol Oncol Stem Cell Ther. 2018 Jun;11(2):65-74.

Natesirinilkul R, Charoenkwan P, Nawarawong W, Boonsri S, Tantivate P, Wongjaikum S, et al. Hypercoagulable state as demonstrated by thrombo-elastometry in hemoglobin E/ β-thalassemia patients: Association with clinical severity and splenectomy status. Thromb Res. 2016 Apr;140:125-31.

Klaihmon P, Phongpao K, Kheansaard W, Noulsri E, Khuhapinant A, Fucharoen S, et al. Microparticles from splenectomized β-thalassemia/HbE patients play roles on procoagulant activities with thrombotic potential. Ann Hematol. 2017 Feb;96(2):189-98.

Aessopos A, Stamatelos G, Skoumas V, Vassilopoulos G, Mantzourani M, Loukopoulos D. Pulmonary hypertension and right heart failure in patients with β-thalassemia intermedia. Chest. 1995 Jan;107(1):50-3.

Fraidenburg DR, Machado RF. Pulmonary hypertension associated with thalassemia syndromes. Ann N Y Acad Sci. 2016 Mar;1368(1):127-39.

Aessopos A, Farmakis D, Karagiorga M, Voskaridou E, Loutradi A, Hatziliami A, et al. Cardiac involvement in thalassemia intermedia: a multicenter study. Blood. 2001 Jun 1;97(11):3411-6.

Derchi G, Galanello R, Bina P, Cappellini MD, Piga A, Lai ME, et al. Prevalence and risk factors for pulmonary arterial hypertension in a large group of β-thalassemia patients using right heart catheterization: a Webthal study. Circulation. 2014 Jan 21;129(3):338-45.

Ryan JJ, Thenappan T, Luo N, Ha T, Patel AR, Rich S, Archer SL. The WHO classification of pulmonary hypertension: A case-based imaging compendium. Pulm Circ. 2012 Jan-Mar;2(1):107-21.

Anthi A, Orfanos SE, Armaganidis A. Pulmonary hypertension in β-thalassaemia. Lancet Respir Med. 2013 Aug;1(6):488-96.

Morris CR, Kim HY, Trachtenberg F, Wood J, Quinn CT, Sweeters N, et al. Risk factors and mortality associated with an elevated tricuspid regurgitant jet velocity measured by Doppler-echocardiography in thalassemia: a Thalassemia Clinical Research Network report. Blood. 2011 Oct 6;118(14):3794-802.

Taher AT, Musallam KM, Karimi M, El-Beshlawy A, Belhoul K, Daar S, et al. Splenectomy and thrombosis: the case of thalassemia intermedia. J Thromb Haemost. 2010 Oct;8(10):2152-8.

Mannucci PM. Red cells playing as activated platelets in thalassemia intermedia. J Thromb Haemost. 2010 Oct;8(10):2149-51.

Atichartakarn V, Angchaisuksiri P, Aryurachai K, Onpun S, Chuncharunee S, Thakkinstian A, et al. Relationship between hypercoagulable state and erythrocyte phosphatidylserine exposure in splenectomized haemoglobin E/β-thalassaemic patients. Br J Haematol. 2002 Sep;118(3):893-8.

Palkar AV, Agrawal A, Verma S, Iftikhar A, Miller EJ, Talwar A. Post splenectomy related pulmonary hypertension. World J Respirol. 2015 Jul 28;5(2):69-77.

Singer ST, Kuypers F, Fineman J, Gildengorin G, Larkin S, Sweeters N, et al. Elevated tricuspid regurgitant jet velocity in subgroups of thalassemia patients: insight into pathophysiology and the effect of splenectomy. Ann Hematol. 2014 Jul;93(7):1139-48.

Srihirun S, Tanjararak N, Chuncharunee S, Sritara P, Kaewvichit R, Fucharoen S, Pattanapanyasat K, Sibmooh N. Platelet hyperactivity in thalassemia patients with elevated tricuspid regurgitant velocity and the association with hemolysis. Thromb Res. 2015 Jan;135(1):121-6.

Rachmilewitz EA, Giardina PJ. How I treat thalassemia. Blood. 2011 Sep 29; 118(13): 3479-88.

Prati D. Benefits and complications of regular blood transfusion in patients with β-thalassaemia major. Vox Sang. 2000;79(3):129-37.

Saliba AN, Harb AR, Taher AT. Iron chelation therapy in transfusion-dependent thalassemia patients: current strategies and future directions. J Blood Med. 2015 Jun 17;6:197-209.

Fucharoen S, Siritanaratkul N, Winichagoon P, Chowthaworn J, Siriboon W, Muangsup W, et al. Hydroxyurea increases hemoglobin F levels and improves the effectiveness of erythropoiesis in β-thalassemia/hemoglobin E disease. Blood. 1996 Feb 1;87(3):887-92.

Thein SL. The emerging role of fetal hemoglobin induction in non-transfusion-dependent thalassemia. Blood Rev. 2012 Apr;26 Suppl 1:S35-9.

Kohne E. Hemoglobinopathies: clinical manifestations, diagnosis, and treatment. Dtsch Arztebl Int. 2011 Aug;108(31-32):532-40.

Angelucci E, Matthes-Martin S, Baronciani D, Bernaudin F, Bonanomi S, Cappellini MD, et al. Hematopoietic stem cell transplantation in thalassemia major and sickle cell disease: indications and management recommendations from an international expert panel. Haematologica. 2014 May;99(5):811-20.

Baronciani D, Angelucci E, Potschger U, Gaziev J, Yesilipek A, Zecca M, et al. Hemopoietic stem cell transplantation in thalassemia: a report from the European Society for Blood and Bone Marrow Transplantation Hemoglobin-opathy Registry, 2000-2010. Bone Marrow Transplant. 2016 Apr;51(4):536-41.

de Dreuzy E, Bhukhai K, Leboulch P, Payen E. Current and future alternative therapies for β-thalassemia major. Biomed J. 2016 Feb;39(1):24-38.

Thompson AA, Walters MC, Kwiatkowski J, Rasko JEJ, Ribeil JA, Hongeng S, et al. Gene therapy in patients with transfusion-dependent β-thalassemia.

N Engl J Med. 2018 Apr 19;378(16):1479-93.

Kleinbongard P, Dejam A, Lauer T, Rassaf T, Schindler A, Picker O, et al. Plasma nitrite reflects constitutive nitric oxide synthase activity in mammals. Free Radic Biol Med. 2003 Oct 1;35(7):790-6.

Cheung YF, Chan GC, Ha SY. Arterial stiffness and endothelial function in patients with β-thalassemia major. Circulation. 2002 Nov 12;106(20):2561-6.

Kukongviriyapan V, Somparn N, Senggunprai L, Prawan A, Kukongviriyapan U, Jetsrisuparb A. Endothelial dysfunction and oxidant status in pediatric patients with hemoglobin E-β-thalassemia. Pediatr Cardiol. 2008 Jan;29(1):130-5.

Dejam A, Hunter CJ, Pelletier MM, Hsu LL, Machado RF, Shiva S, et al. Erythrocytes are the major intravascular storage sites of nitrite in human blood. Blood. 2005 July 15;106(2):734-9.

Kleinbongard P, Rassaf T, Dejam A, Kerber S, Kelm M. Griess method for nitrite measurement of aqueous and protein-containing samples. Methods Enzymol. 2002;359:158-68.

Suvachananonda T, Wankham A, Srihirun S, Tanratana P, Unchern S, Fucharoen S, et al. Decreased nitrite levels in erythrocytes of children with β-thalassemia/ hemoglobin E. Nitric Oxide. 2013 Sep 1;33:1-5.

Sriwantana T, Vivithanaporn P, Paiboonsukwong K, Rattanawonsakul K, Srihirun S, Sibmooh N. Deferiprone increases endothelial nitric oxide synthase phosphoryl- ation and nitric oxide production. Can J Physiol Pharmacol. 2018 Sep;96(9): 879-85.

Chamchoi A, Srihirun S, Paiboonsukwong K, Sriwantana T, Sathavorasmith P, Pattanapanyasat K, et al. Decreased nitrite reductase activity of deoxy-hemoglobin correlates with platelet activation in hemoglobin E/ss-thalassemia subjects. PLoS One. 2018 Sep 20;13(9):e0203955.

Cokic VP, Beleslin-Cokic BB, Tomic M, Stojilkovic SS, Noguchi CT, Schechter AN. Hydroxyurea induces the eNOS-cGMP pathway in endothelial cells. Blood. 2006 Jul 1;108(1):184-91.

Ikeda Y, Tajima S, Yoshida S, Yamano N, Kihira Y, Ishizawa K, et al. Deferox-amine promotes angiogenesis via the activation of vascular endothelial cell function. Atherosclerosis. 2011 Apr;215(2):339-47.

Bayraktar N, Erkurt MA, Aydogdu I, Basaran Y. The levels of nitric oxide in β-thalassemia minor. Turk J Haematol. 2008 Dec 5;25(4):187-9.

Satitthummanid S, Uaprasert N, Songmuang SB, Rojnuckarin P, Tosukhowong P, Sutcharitchan P, Srimahachota S. Depleted nitric oxide and prostaglandin E2 levels are correlated with endothelial dysfunction in beta-thalassemia/HbE patients. Int J Hematol. 2017 Sep;106(3):366-374.

El-Hady SB, Farahat MH, Atfy M, Elhady MA. Nitric oxide metabolites and arginase I levels in β-thalassemic patients: an Egyptian study. Ann Hematol. 2012 Aug;91(8):1193-200.

Uaprasert N, Satitthummanid S, Akkawat B, Sutcharitchan P, Rojnuckarin P. Vascular and hemostatic alterations associated with pulmonary hypertension in β-thalassemia hemoglobin E patients receiving regular transfusion and iron chelation. Thromb Res. 2019 Feb;174:104-12.

Atichartakarn V, Chuncharunee S, Archararit N, Udomsubpayakul U, Aryurachai K. Intravascular hemolysis, vascular endothelial cell activation and thrombophilia in splenectomized patients with hemoglobin E/β-thalassemia disease. Acta Haematol. 2014;132(1):100-7.

Liu X, Miller MJ, Joshi MS, Sadowska-Krowicka H, Clark DA, Lancaster JR, Jr. Diffusion-limited reaction of free nitric oxide with erythrocytes. J Biol Chem. 1998 Jul 24;273(30):18709-13.

Morris CR, Kim HY, Klings ES, Wood J, Porter JB, Trachtenberg F, et al. Dysregulated arginine metabolism and cardiopulmonary dysfunction in patients with thalassaemia. Br J Haematol. 2015 Jun;169(6):887-98.

Roche CJ, Malashkevich V, Balazs TC, Dantsker D, Chen Q, Moreira J, et al. Structural and functional studies indicating altered redox properties of hemoglobin E: implications for production of bioactive nitric oxide. J Biol Chem. 2011 Jul 1;286(26):23452-66.

Littera R, La Nasa G, Derchi G, Cappellini MD, Chang CY, Contu L. Long-term treatment with sildenafil in a thalassemic patient with pulmonary hypertension. Blood. 2002 Aug 15;100(4):1516-7.

Correale M, De Rosa F, Ieva R, Di Biase M, Brunetti ND. Long-term treatment with high-dose of sildenafil in a thalassemic patient with pulmonary hypertension. Monaldi Arch Chest Dis. 2012 Jun;78(2):105-6.

Derchi G, Forni GL, Formisano F, Cappellini MD, Galanello R, D'Ascola G, et al. Efficacy and safety of sildenafil in the treatment of severe pulmonary hyper-tension in patients with hemoglobinopathies. Haematologica. 2005 Apr;90(4): 452-8.

Morris CR, Kim HY, Wood J, Porter JB, Klings ES, Trachtenberg FL, et al. Sildenafil therapy in thalassemia patients with Doppler-defined risk of pulmonary hypertension. Haematologica. 2013 Sep;98(9):1359-67.

Hayward CS, Kelly RP, Macdonald PS. Inhaled nitric oxide in cardiology practice. Cardiovasc Res. 1999 Aug 15;43(3):628-38.

Yingchoncharoen T, Rakyhao T, Chuncharunee S, Sritara P, Pienvichit P, Paiboonsukwong K, et al. Inhaled nebulized sodium nitrite decreases pulmonary artery pressure in β-thalassemia patients with pulmonary hypertension. Nitric Oxide. 2018 Jun 1;76:174-8.

Parakaw T, Suknuntha K, Vivithanaporn P, Schlagenhauf A, Topanurak S, Fucharoen S, et al. Platelet inhibition and increased phosphorylated vasodilator- stimulated phosphoprotein following sodium nitrite inhalation. Nitric Oxide. 2017 Jun 1;66:10-16.