Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

DDAH-1, via regulation of ADMA levels, protects against ischemia-induced blood-brain barrier leakage

Abstract

Dimethylarginine dimethylamino hydrolase-1 (DDAH-1) is an important regulator of nitric oxide (NO) metabolism that has been implicated in the pathogenesis of cardiovascular diseases. Nevertheless, its role in cerebral ischemia still needs to be elucidated. Herein, we examined the expression of DDAH-1 in the brain of rat by double-label immunofluorescence staining. DDAH-1 knock-out (DDAH-1−/−) and wild-type rats underwent middle cerebral artery occlusion/reperfusion (MCAO/R). After 24 h, neurological scores, TTC staining and TUNEL assay were used to evaluate neurological damages. 3 and 7-days infarct outcomes were also shown. Blood-brain-barrier (BBB) permeability was examined via Evans blue extravasation and tight junction (TJ) proteins expression and mRNA levels by western blot and RT-qPCR. The levels of plasma asymmetric dimethylarginine (ADMA), NO and ADMA in brain tissue were also assessed. In addition, supplementation of L-arginine to DDAH-1−/− rats was used to explore its role in regulating NO. DDAH-1 was abundantly distributed in cerebral cortex and basal nuclei, and mainly expressed in neurons and endothelial cells. DDAH-1−/− rats showed aggravated neurological damage and BBB disruption, including decrease of TJ proteins expression but indistinguishable mRNA levels after MCAO/R. DDAH-1 depletion and neurological damages were accompanied with increased ADMA levels and decreased NO concentrations. The supplementation with L-arginine partly restored the neurological damages and BBB disruption. To sum up, DDAH-1 revealed to have a protective role in ischemia stroke (IS) and IS-induced leakage of BBB via decreasing ADMA level and possibly via preventing TJ proteins degradation.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Main distribution of DDAH-1 in basal nuclei, not brainstem.
Fig. 2: Main distribution of DDAH-1 in neurons and endothelial cells.
Fig. 3: DDAH-1−/− rats showed aggravated ischemic damage after MCAO/R.
Fig. 4: DDAH-1−/− rats showed severer apoptosis in brain after MCAO/R.
Fig. 5: DDAH-1−/− rats showed increased EBD extravasation after MCAO/R with lower NO and higher ADMA levels.
Fig. 6: Supplement of L-arginine to DDAH-1−/− rats alleviated partial neurological damages.
Fig. 7: Supplement of L-arginine to DDAH-1−/− rats relieved long-term neurological damages.
Fig. 8: DDAH-1−/− rats showed decreased expressions of tight junction proteins.

References

  1. 1.

    Katan M, Luft A. Global burden of stroke. Semin. Neurol. 2018;38:208–11.

    PubMed  Article  PubMed Central  Google Scholar 

  2. 2.

    Wang W, Jiang B, Sun H, Ru X, Sun D, Wang L, et al. Prevalence, incidence, and mortality of stroke in China. Circulation. 2017;135:759.

    PubMed  Article  PubMed Central  Google Scholar 

  3. 3.

    Chung JW, Oh MJ, Cho YH, Moon GJ, Kim GM, Chung CS, et al. Distinct roles of endothelial dysfunction and inflammation in intracranial atherosclerotic stroke. Eur. Neurol. 2017;77:211–9.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  4. 4.

    Na L, Hans W, Milani D, Shufen C, Karin W. Nitric oxide (NO) and asymmetric dimethylarginine (ADMA): their pathophysiological role and involvement in intracerebral hemorrhage. Neurol. Res. 2011;33:541–8.

    Article  CAS  Google Scholar 

  5. 5.

    Patrick V, James L, Blocking NO. synthesis: how, where and why? Nat. Rev. Drug Discov. 2002;1:939–50.

    Article  CAS  Google Scholar 

  6. 6.

    Nadja S, Anja S, Jens ML, Thomas K, Gulistan T, Thomas L, et al. Endothelial dysfunction of the peripheral vascular bed in the acute phase after ischemic stroke. Cerebrovasc. Dis. 2012;33:37–46.

    Article  CAS  Google Scholar 

  7. 7.

    Leiper J, Nandi M, Torondel B, Murray-Rust J, Malaki M, O’Hara B, et al. Disruption of methylarginine metabolism impairs vascular homeostasis. Nat. Med. 2007;13:198–203.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  8. 8.

    Pope AJ, Karrupiah KP. Role of dimethylarginine dimethylaminohydrolases in the regulation of endothelial nitric oxide production. J. Biol. Chem. 2009;284:35338–47.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  9. 9.

    Pope AJ, Karuppiah K, Cardounel AJ. Role of the PRMT–DDAH–ADMA axis in the regulation of endothelial nitric oxide production. Pharmacol. Res. Off. J. Ital. Pharmacol. Soc. 2009;60:461–5.

    CAS  Google Scholar 

  10. 10.

    Zhang Y, Fan D, Zhang N. The relationship between serum asymmetric dimethylarginine and ABCD2 score in transient ischemic attack patients. Zhonghua NeiKe ZaZhi. 2014;53:876–9.

    CAS  PubMed  Google Scholar 

  11. 11.

    Chen S, Li N, Deb-Chatterji M, Dong Q, Kielstein JT, Weissenborn K, et al. Asymmetric dimethyarginine as marker and mediator in ischemic stroke. Int. J. Mol. Sci. 2012;13:15983.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  12. 12.

    Greco R, Ferrigno A, Demartini C, Zanaboni A, Mangione AS, Blandini F, et al. Evaluation of ADMA-DDAH-NOS axis in specific brain areas following nitroglycerin administration: study in an animal model of migraine. J. Headache Pain. 2015;16:74.

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  13. 13.

    Molnar T, Pusch G, Nagy L, Keki S, Berki T, Illes Z. Correlation of the L-arginine pathway with thrombo-inflammation may contribute to the outcome of acute ischemic stroke. J. Stroke Cerebrovasc. Dis. 2016;25:2055–60.

    PubMed  Article  Google Scholar 

  14. 14.

    Nakayama Y, Ueda S, Yamagishi SI, Obara N, Taguchi K, Ando R, et al. Asymmetric dimethylarginine accumulates in the kidney during ischemia/reperfusion injury. Kidney Int. 2014;85:570–8.

    CAS  PubMed  Article  Google Scholar 

  15. 15.

    Segarra G, Cortina B, Mauricio MD, Novella S, Lluch P, Navarrete-Navarro J, et al. Effects of asymmetric dimethylarginine on renal arteries in portal hypertension and cirrhosis. World J. Gastroenterol. 2016;22:10545–56.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  16. 16.

    Chih-Min T, Hsuan-Chang K, Chien-Ning H, Li-Tung H, You-Lin T. Metformin reduces asymmetric dimethylarginine and prevents hypertension in spontaneously hypertensive rats. Transl. Res. 2014;164:452–9.

    Article  CAS  Google Scholar 

  17. 17.

    Yokoro M, Nakayama Y, Yamagishi SI, Ando R, Sugiyama M, Ito S, et al. Asymmetric dimethylarginine contributes to the impaired response to erythropoietin in CKD-anemia. J. Am. Soc. Nephrol. 2017;28:2670.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  18. 18.

    Monti L, Morbidelli L, Bazzani L, Rossi A. Influence of circulating endothelin-1 and asymmetric dimethylarginine on whole brain circulation time in multiple sclerosis. Biomarker Insights. 2017;12:1177271917712514.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  19. 19.

    Chertow JH, Alkaitis MS, Nardone G, Ikeda AK, Cunnington AJ, Okebe J, et al. Plasmodium infection is associated with impaired hepatic dimethylarginine dimethylaminohydrolase activity and disruption of nitric oxide synthase inhibitor/substrate homeostasis. PLoS Pathog. 2015;11:e1005119.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  20. 20.

    Donia A, Anne M, Sabine GS, Monique P, Bernard B, Philippe V, et al. Cerebral changes occurring in arginase and dimethylarginine dimethylaminohydrolase (DDAH) in a rat model of sleeping sickness. Plos One. 2011;6:e16891.

    Article  CAS  Google Scholar 

  21. 21.

    Adel B, Oleg P, Ashraf T, Farid H, Ramadan H, Wiebke J, et al. Effects of dimethylarginine dimethylaminohydrolase-1 overexpression on the response of the pulmonary vasculature to hypoxia. Am J Respir Cell Mol Biol. 2013;49:491–500.

    Article  CAS  Google Scholar 

  22. 22.

    Hu D, Bin W, Hu W, Zhilan L, Jiangtao Y, Xiaojing W, et al. A novel loss-of-function DDAH1 promoter polymorphism is associated with increased susceptibility to thrombosis stroke and coronary heart disease. Circ. Res. 2010;106:1145–52.

    Article  CAS  Google Scholar 

  23. 23.

    Dê¼Mello R, Sand CA, Pezet S, Leiper JM, Gaurilcikaite E, Mcmahon SB, et al. Dimethylarginine dimethylaminohydrolase 1 is involved in spinal nociceptive plasticity. Pain. 2015;156:2052–60.

    Article  CAS  Google Scholar 

  24. 24.

    Mookerjee RP, Gautam M, Vairappan B, Mohamed FEZ, Nathan D, Vikram S, et al. Hepatic dimethylarginine-dimethylaminohydrolase1 is reduced in cirrhosis and is a target for therapy in portal hypertension. J. Hepatol. 2015;62:325–31.

    CAS  PubMed  Article  Google Scholar 

  25. 25.

    Speranza L, Franceschelli S, D’Orazio N, Gaeta R, Bucciarelli T, Felaco M, et al. The biological effect of pharmacological treatment on dimethylaminohydrolases (DDAH-1) and cationic amino acid transporter-1 (CAT-1) expression in patients with acute congestive heart failure. Microvasc. Res. 2011;82:391–6.

    CAS  PubMed  Article  Google Scholar 

  26. 26.

    Bell T, Araujo M, Luo Z, Tomlinson J, Leiper J, Welch WJ, et al. Regulation of fluid reabsorption in rat or mouse proximal renal tubules by asymmetric dimethylarginine (ADMA) & dimethylarginine dimethylaminohydrolase (DDAH) 1. Am. J. Physiol. Renal Physiol. 2018;315:F74–8.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  27. 27.

    Shahin NN, Abdelkader NF, Safar MM. A novel role of irbesartan in gastroprotection against indomethacin-induced gastric injury in rats: targeting DDAH/ADMA and EGFR/ERK signaling. Sci Rep. 2018;8:4280.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  28. 28.

    Frank L, Chi-Un C, Mathias G, Eike-Christin VL, Dorothee A, Edzard S, et al. Dimethylarginine dimethylaminohydrolase-1 transgenic mice are not protected from ischemic stroke. Plos One. 2009;4:e7337.

    Article  CAS  Google Scholar 

  29. 29.

    Nannan C, Changhong S, Lu G, Dan Z, Xiufei X, Xiaojun Z, et al. Genome editing with RNA-guided Cas9 nuclease in zebrafish embryos. Cell Res. 2013;23:465–72.

    Article  CAS  Google Scholar 

  30. 30.

    Haley MJ, Lawrence CB. The blood-brain barrier after stroke: Structural studies and the role of transcytotic vesicles. J. Cereb. Blood Flow Metab. 2016;37:0271678X16629976.

    Google Scholar 

  31. 31.

    Albert-Weißenberger C, Várrallyay C, Raslan F, Kleinschnitz C, Sirén AL. An experimental protocol for mimicking pathomechanisms of traumatic brain injury in mice. Exp. Transl. Stroke Med. 2012;4:1–5.

    PubMed  PubMed Central  Article  Google Scholar 

  32. 32.

    Wang D, Li H, Weir E, Xu Y, Xu D, Chen Y. Dimethylarginine dimethylaminohydrolase 1 deficiency aggravates monocrotaline-induced pulmonary oxidative stress, pulmonary arterial hypertension and right heart failure in rats. Int. J. Cardiol. 2019;295:14–20.

    PubMed  Article  PubMed Central  Google Scholar 

  33. 33.

    Palm F, Onozato ML, Luo Z, Wilcox CS. Dimethylarginine dimethylaminohydrolase (DDAH): expression, regulation, and function in the cardiovascular and renal systems. Am. J. Physiol. Heart Circ. Physiol. 2007;293:H3227.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  34. 34.

    Zhao Y, Zhou Y, Ma X, Liu X, Zhao Y, Liu X. DDAH-1 via HIF-1 target genes improves cerebral ischemic tolerance after hypoxic preconditioning and middle cerebral artery occlusion-reperfusion. Nitric Oxide. 2020;95:17–28.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  35. 35.

    Kielstein JT, Donnerstag F, Gasper S, Menne J, Kielstein A, Martens-Lobenhoffer J, et al. ADMA increases arterial stiffness and decreases cerebral blood flow in humans. Stroke. 2006;37:2024–9.

    PubMed  Article  PubMed Central  Google Scholar 

  36. 36.

    Czarnecka A, Milewski K, Zielińska M. Asymmetric dimethylarginine and hepatic encephalopathy: cause, effect or association? Neurochem Res. 2017;42:750–61.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  37. 37.

    Czarnecka A, Aleksandrowicz M, Jasiński K, Jaźwiec R, Kalita K, Hilgier W, et al. Cerebrovascular reactivity and cerebral perfusion of rats with acute liver failure: role of L-glutamine and asymmetric dimethylarginine in L-arginine-induced response. J. Neurochem. 2018;147:692–704.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. 38.

    Behrouzifar S, Vakili A, Bandegi AR, Kokhaei P. Neuroprotective nature of adipokine resistin in the early stages of focal cerebral ischemia in a stroke mouse model. Neurochem. Int. 2018;114:99–107.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  39. 39.

    Pei Z, Fung PC, Cheung RT. Melatonin reduces nitric oxide level during ischemia but not blood-brain barrier breakdown during reperfusion in a rat middle cerebral artery occlusion stroke model. J. Pineal Res. 2003;34:110–8.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  40. 40.

    Hayan D, Rodionov RN, Cynthia L, Cooke JP, Erland A, Teodoro B, et al. Overexpression of dimethylarginine dimethylaminohydrolase inhibits asymmetric dimethylarginine-induced endothelial dysfunction in the cerebral circulation. Stroke. 2008;39:180–4.

    Article  CAS  Google Scholar 

  41. 41.

    Murphy RB, Tommasi S, Lewis BC, Mangoni AA. Inhibitors of the hydrolytic enzyme dimethylarginine dimethylaminohydrolase (DDAH): discovery, synthesis and development. Molecules. 2016;21:615.

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  42. 42.

    Böger RH. L-arginine improves vascular function by overcoming the deleterious effects of ADMA, a novel cardiovascular risk factor. Altern. Med. Rev. Altern. Med. Rev. 2005;10:14.

    PubMed  Google Scholar 

  43. 43.

    Jabecka A, Ast J, Bogdaski P, Drozdowski M, Pawlak-Lemaska K, Cielewicz AR, et al. Oral L-arginine supplementation in patients with mild arterial hypertension and its effect on plasma level of asymmetric dimethylarginine, L-citruline, L-arginine and antioxidant status. Eur. Rev. Med. Pharmacol. Sci. 2012;16:1665–74.

    CAS  PubMed  Google Scholar 

  44. 44.

    Jiang, X, Andjelkovic, AV, Zhu, L, Yang, T, Bennett, MVL, Chen, J et al. Blood-brain barrier dysfunction and recovery after ischemic stroke. Prog. Neurobiol. 2018;163–4:144–71.

  45. 45.

    Prakash R, Carmichael ST. Blood-brain barrier breakdown and neovascularization processes after stroke and traumatic brain injury. Curr. Opin. Neurol. 2015;28:556–64.

    PubMed  PubMed Central  Article  Google Scholar 

  46. 46.

    Campisi M, Shin Y, Osaki T, Hajal C, Chiono V, Kamm RD. 3D self-organized microvascular model of the human blood-brain barrier with endothelial cells, pericytes and astrocytes. Biomaterials. 2018;180:117.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  47. 47.

    Gao W, Li F, Liu L, Xu X, Zhang B, Wu Y, et al. Endothelial colony-forming cell-derived exosomes restore blood-brain barrier continuity in mice subjected to traumatic brain injury. Exp. Neurol. 2018;307:99–108.

    CAS  PubMed  Article  Google Scholar 

  48. 48.

    Panahpour, H, Farhoudi, M, Omidi, Y & Mahmoudi, J. An in vivo assessment of blood-brain barrier disruption in a rat model of ischemic stroke. J Vis Exp. 2018;133:57156.

  49. 49.

    Chen FQ, Li Q, Pan CS, Liu YY, Yan L, Sun K, et al. Kudiezi Injection(®) alleviates blood-brain barrier disruption after ischemia-reperfusion in rats. Microcirculation. 2016;23:426–37.

    CAS  PubMed  Article  Google Scholar 

  50. 50.

    Abdullahi W, Tripathi D, Ronaldson P. Blood-brain barrier dysfunction in ischemic stroke: targeting tight junctions and transporters for vascular protection. Am. J. Physiol. Cell Physiol. 2018;315:C343–C356.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  51. 51.

    Ping Z, Xinli H, Xin X, Yingjie C, Bache RJ. Dimethylarginine dimethylaminohydrolase 1 modulates endothelial cell growth through nitric oxide and Akt. Arterioscler. Thromb. Vasc. Biol. 2011;31:890.

    Article  CAS  Google Scholar 

  52. 52.

    Choi S, Singh I, Singh A, Khan M, Won J. Asymmetric dimethylarginine exacerbates cognitive dysfunction associated with cerebrovascular pathology. FASEB J. 2020;34:6808–23.

    CAS  PubMed  Article  Google Scholar 

  53. 53.

    Chen Y, Xu X, Sheng M, Zheng Z, Gu Q. Effects of asymmetric dimethylarginine on bovine retinal capillary endothelial cell proliferation, reactive oxygen species production, permeability, intercellular adhesion molecule-1, and occludin expression. Mol. Vis. 2011;17:332–40.

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

DDAH-1 knockout SD rats were provided by Professor Da-Chun Xu (Department of Cardiology, Shanghai Tenth People’s Hospital, Tongji University School of Medicine).

Funding

The present study was supported by the Science and Technology Commission of Shanghai Municipality (grant numbers 18140901900 and 20ZR1443500), National Natural Science Foundation of China (Grant no. 8207052336), and Shanghai Municipal Key Clinical Specialty (Grant no. shslczdzk06102).

Author information

Affiliations

Authors

Corresponding authors

Correspondence to Xueyuan Liu or Yanxin Zhao.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Zhao, Y., Ma, X., Zhou, Y. et al. DDAH-1, via regulation of ADMA levels, protects against ischemia-induced blood-brain barrier leakage. Lab Invest (2021). https://doi.org/10.1038/s41374-021-00541-5

Download citation

Search

Quick links