The role of xanthine oxidoreductase in nitrate-nitric oxide transformation in vascular endothelial cells of hypoxia injury

doi:10.19438/j.cjoms.2021.04.001

Abstract

Abstract: PURPOSE: To investigate the mechanism of nitrate-nitric oxide (NO) transformation in vascular endothelial cells under hypoxia injury and the role of xanthine oxidoreductase(XOR). METHODS: Model of hypoxia reoxygenation injury of human vascular endothelial cells was established. The level of nitric oxide and the content of sodium nitrate were detected in cells cultured in medium with different concentrations of sodium nitrate and sodium nitrite. The expression of endothelial nitric oxide synthase(eNOS) and XOR were detected by real-time quantitative PCR and Western blot. After eNOS inhibitor L-NMMA and XOR inhibitor allopurinol were used respectively, the levels of NO in cells cultured in medium with sodium nitrate were detected again. The data were analyzed with GraphPad Prism 8.0 software package. RESULTS: Higher concentration of extracellular nitrate increased intracellular nitrate content in both normoxia and hypoxia conditions. Nitrite and nitrate extracellular only increased levels of intracellular NO in hypoxia condition, and with allopurinol, XOR inhibitor, extracellular nitrate did not increase intracellular NO level. CONCLUSIONS: Nitrate-NO transformation in human vascular endothelial cells occurs in condition of hypoxia, which is mainly dependent on XOR.

Key words: Vascular endothelial cells, Hypoxia injury, Nitrate-nitrite-nitric oxide pathway, Xanthine oxidoreductase

CLC Number:

YANG Yang, NIU Qi-fang, LI Delong, ZHANG Shu, QIN Li-zheng, HAN Zheng-xue. The role of xanthine oxidoreductase in nitrate-nitric oxide transformation in vascular endothelial cells of hypoxia injury[J]. China Journal of Oral and Maxillofacial Surgery, 2021, 19(4): 289-294.

References

[1] 韩正学, 李金忠, 李华, 等. 游离组织瓣移植重建颅内外沟通性缺损的临床研究[J]. 中华神经外科杂志, 2012, 28(8): 772-774.
[2] 韩正学, 李华, 李金忠, 等. 口腔颌面缺损游离组织移植修复138例临床分析[J]. 北京口腔医学, 2010, 18(4): 225-227.
[3] Futran ND, Alsarraf R.Microvascular free-flap reconstruction in the head and neck[J]. JAMA, 2000, 284(14): 1761-1763.
[4] Chinn SB, Myers JN.Oral cavity carcinoma: current management, controversies, and future directions[J]. J Clin Oncol, 2015, 33(29): 3269-3276.
[5] 欧阳嘉杰, 牛其芳, 韩正学. 非酶源一氧化氮及其在游离皮瓣低血再灌注损伤中的作用[J]. 国际口腔医学杂志, 2015, 42(1): 106-110.
[6] Lundberg JO, Weitzberg E, Gladwin MT.The nitrate-nitrite-nitric oxide pathway in physiology and therapeutics[J]. Nat Rev Drug Discov, 2008, 7(2): 156-167.
[7] Kapil V, Weitzberg E, Lundberg JO, et al.Clinical evidence demonstrating the utility of inorganic nitrate in cardiovascular health[J]. Nitric Oxide, 2014, 38: 45-57.
[8] Ingram TE, Fraser AG, Bleasdale RA, et al.Low-dose sodium nitrite attenuates myocardial ischemia and vascular ischemia-reperfusion injury in human models[J]. J Am Coll Cardiol, 2013, 61(25): 2534-2541.
[9] Li W, Meng Z, Liu Y, et al.The hepatoprotective effect of sodium nitrite on cold ischemia-reperfusion injury[J]. J Transplant, 2012: 635179.
[10] Carlström M, Lundberg JO, Weitzberg E.Mechanisms underlying blood pressure reduction by dietary inorganic nitrate[J]. Acta Physiol (Oxf), 2018, 224(1): e13080.
[11] Waltz P, Escobar D, Botero AM, et al.Nitrate/nitrite as critical mediators to limit oxidative injury and inflammation[J]. Antioxid Redox Signal, 2015, 23(4): 328-339.
[12] Jin L, Qin L, Xia D, et al.Active secretion and protective effect of salivary nitrate against stress in human volunteers and rats[J]. Free Radic Biol Med, 2013, 57(1): 61-67.
[13] Huang L, Borniquel S, Lundberg JO.Enhanced xanthine oxidoreductase expression and tissue nitrate reduction in germ free mice[J]. Nitric Oxide, 2010, 22(2): 191-195.
[14] Carlström M, Liu M, Yang T, et al.Cross-talk between nitrate-nitrite-NO and NO synthase pathways in control of vascular NO homeostasis[J]. Antioxid Redox Signal, 2015, 23(4): 295-306.
[15] Peleli M, Zollbrecht C, Montenegro MF, et al.Enhanced XOR activity in eNOS-deficient mice: effects on the nitrate-nitrite-NO pathway and ROS homeostasis[J]. Free Radic Biol Med, 2016, 99: 472-484.
[16] Yang Y, Li S, Qu Y, et al.Nitrate partially inhibits lipopolysaccharide-induced inflammation by maintaining mitochondrial function[J]. J Int Med Res, 2020, 48(2):030006052090260.
[17] 牛其芳, 李德龙, 杨杨, 等. 人血管内皮细胞缺氧复氧损伤细胞模型的建立[J]. 中国口腔颌面外科杂志, 2019, 17(4): 295-299.
[18] Cauwels A, Buys ES, Thoonen R, et al.Nitrite protects against morbidity and mortality associated with TNF- or LPS-induced shock in a soluble guanylate cyclase-dependent manner[J]. J Exp Med, 2009, 206(13): 2915-2924.
[19] Vitturi DA, Patel RP.Current perspectives and challenges in understanding the role of nitrite as an integral player in nitric oxide biology and therapy[J]. Free Radic Biol Med, 2011, 51(4): 805-812.
[20] Shiva S, Sack MN, Greer JJ, et al.Nitrite augments tolerance to ischemia/reperfusion injury via the modulation of mitochondrial electron transfer[J]. J Exp Med, 2007, 204(9): 2089-2102.
[21] Kleinbongard P, Dejam A, Lauer T, et al.Plasma nitrite reflects constitutive nitric oxide synthase activity in mammals[J]. Free Radic Biol Med, 2003, 35(7): 790-796.
[22] Larsen FJ, Schiffer TA, Borniquel S, et al.Dietary inorganic nitrate improves mitochondrial efficiency in humans[J]. Cell Metab, 2011, 13(2): 149-159.
[23] Monaco CMF, Miotto PM, Huber JS, et al.Sodium nitrate supplementation alters mitochondrial H₂O₂ emission but does not improve mitochondrial oxidative metabolism in the heart of healthy rats[J]. Am J Physiol Regul Integr Comp Physiol, 2018, 315(2): R191-R204.
[24] Asai R, Nishino T, Matsumura T, et al.Two mutations convert mammalian xanthine oxidoreductase to highly superoxide-productive xanthine oxidase[J]. J Biochem, 2007, 141(4): 525-534.
[25] Nishino T, Okamoto K, Eger BT, et al.Mammalian xanthine oxidoreductase-mechanism of transition from xanthine dehydrogenase to xanthine oxidase[J]. FEBS J, 2008, 275(13): 3278-3289.
[26] Kelley EE.A new paradigm for XOR-catalyzed reactive species generation in the endothelium[J]. Pharmacol Rep, 2015, 67(4): 669-674.