Kynurenic acid and its derivatives are able to modulate the adhesion and locomotion of brain endothelial cells
The neuroprotective actions of kynurenic acid (KYNA) and its derivatives in several neurodegenerative disorders [characterized by damage to the cerebral endothelium and to the blood–brain barrier (BBB)] are well established. Cell–extracellular matrix (ECM) adhesion is supposedly involved in recovery of impaired cerebral endothelium integrity (endothelial repair). The present work aimed to investigate the effects of KYNA and its synthetic derivatives on cellular behaviour (e.g. adhesion and locomotion) and on morphology of the GP8 rat brain endothelial cell line, modeling the BBB endothelium. The effects of KYNA and its derivatives on cell adhesion were measured using an impedance-based technique, the xCELLigence SP system. Holographic microscopy (Holomonitor™ M4) was used to analyse both chemokinetic responses and morphometry. The GP8 cells proved to be a suitable model cell line for investigating cell adhesion and the locomotion modulator effects of kynurenines. KYNA enhanced cell adhesion and spreading, and also decreased the migration/motility of GP8 cells at physiological concentrations (10−9 and 10−7 mol/L). The derivatives containing an amide side-chain at the C2 position (KYNA-A1 and A2) had lower adhesion inducer effects compared to KYNA. All synthetic analogues (except KYNA-A5) had a time-dependent inhibitory effect on GP8 cell adhesion at a supraphysiological concentration (10−3 mol/L). The immobilization promoting effect of KYNA and the adhesion inducer activity of its derivatives indicate that these compounds could contribute to maintaining or restoring the protective function of brain endothelium; they also suggest that cell–ECM adhesion and related cell responses (e.g. migration/motility) could be potential new targets of KYNA.
KeywordsEndothelial repair Kynurenic acid Holographic microscopy Impedimetry Synthetic kynurenines Brain endothelial cell Kynurenic acid (PubChem CID: 3845) l-Kynurenine (Pubchem CID: 161166)
Authors express their gratitude to Auro-Science Consulting Kft for their expert technical assistance, and to Mr. P. Samotik and Mr. Z. Ambrus for proofreading and technical language support.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
This work was supported by the following grants: GINOP-2.3.2-15-2016-00034, MTA-SZTE Neuroscience Research Group. The funding sources were in no way involved in the conduct of the research, nor in the analysis or interpretation of data, nor in preparation of the article.
- Andras IE, Deli MA, Veszelka S, Hayashi K, Hennig B, Toborek M (2007) The NMDA and AMPA/KA receptors are involved in glutamate-induced alterations of occludin expression and phosphorylation in brain endothelial cells. J Cereb Blood Flow Metab 27:1431–1443. https://doi.org/10.1038/sj.jcbfm.9600445 CrossRefPubMedGoogle Scholar
- Antal O, Hackler JL, Shen J, Mán I, Hideghéty K, Kitajka K, Puskás LG (2014) Combination of unsaturated fatty acids and ionizing radiation on human glioma cells: cellular, biochemical and gene expression analysis. Lipids Health Dis 13:142–157. https://doi.org/10.1186/1476-511x-13-142 CrossRefPubMedPubMedCentralGoogle Scholar
- Beal MF, Matson WR, Swartz KJ, Gamache PH, Bird ED (1990) Kynurenine pathway measurements in Huntington’s disease striatum: evidence for reduced formation of kynurenic acid. J Neurochem 55:1327–1339. https://doi.org/10.1111/j.1471-4159.1990.tb03143.x CrossRefPubMedGoogle Scholar
- Cosi C et al (2011) G-protein coupled receptor 35 (GPR35) activation and inflammatory pain: studies on the antinociceptive effects of kynurenic acid and zaprinast. Neuropharmacology 60:1227–1231. https://doi.org/10.1016/j.neuropharm.2010.11.014S0028-3908(10)00313-8 CrossRefPubMedGoogle Scholar
- Dehouck MP, Meresse S, Delorme P, Fruchart JC, Cecchelli R (1990) An easier, reproducible, and mass-production method to study the blood–brain barrier in vitro. J Neurochem 54:1798–1801. https://doi.org/10.1111/j.1471-4159.1990.tb01236.x CrossRefPubMedGoogle Scholar
- del Zoppo GJ, Milner R (2006) Integrin–matrix interactions in the cerebral microvasculature. Arterioscler Thromb Vasc Biol 26:1966–1975. https://doi.org/10.1161/01.ATV.0000232525.65682.a2 CrossRefPubMedGoogle Scholar
- Demeter I, Nagy K, Gellert L, Vecsei L, Fulop F, Toldi J (2012) A novel kynurenic acid analog (SZR104) inhibits pentylenetetrazole-induced epileptiform seizures. An electrophysiological study: special issue related to kynurenine. J Neural Transm 119:151–154. https://doi.org/10.1007/s00702-011-0755-x CrossRefPubMedGoogle Scholar
- Greenwood J, Pryce G, Devine L, Male DK, dos Santos WL, Calder VL, Adamson P (1996) SV40 large T immortalised cell lines of the rat blood–brain and blood–retinal barriers retain their phenotypic and immunological characteristics. J Neuroimmunol 71:51–63. https://doi.org/10.1016/S0165-5728(96)00130-0 CrossRefPubMedGoogle Scholar
- Hilmas C, Pereira EF, Alkondon M, Rassoulpour A, Schwarcz R, Albuquerque EX (2001) The brain metabolite kynurenic acid inhibits alpha7 nicotinic receptor activity and increases non-alpha7 nicotinic receptor expression: physiopathological implications. J Neurosci 21:7463–7473CrossRefPubMedGoogle Scholar
- Krizbai IA, Deli MA, Pestenacz A, Siklos L, Szabo CA, Andras I, Joo F (1998) Expression of glutamate receptors on cultured cerebral endothelial cells. J Neurosci Res 54:814–819. https://doi.org/10.1002/(sici)1097-4547(19981215)54:6<814::aid-jnr9>3.0.co;2-3
- Lajkó E, Bányai P, Zámbó Z, Kursinszki L, Szőke É, Kőhidai L (2015) Targeted tumor therapy by Rubia tinctorum L.: analytical characterization of hydroxyanthraquinones and investigation of their selective cytotoxic, adhesion and migration modulator effects on melanoma cell line (A2058 and HT168-M1). Cancer Cell. https://doi.org/10.1186/s12935-015-0271-4 Google Scholar
- Persson J, Mölder A, Pettersson SG, Alm K (2010) Cell motility studies using digital holographic microscopy. In: Vilas M, Díaz J (eds) Microscopy: science, technology, applications and education, vol 4. Formatex Research Center, Badajoz, Spain, pp 1063–1072Google Scholar
- Nagy K et al (2011) Synthesis and biological effects of some kynurenic acid analogs. Bioorg Med Chem 19:7590–7596. https://doi.org/10.1016/j.bmc.2011.10.029S0968-0896(11)00837-6 CrossRefPubMedGoogle Scholar
- Owe-Young R et al (2008) Kynurenine pathway metabolism in human blood–brain-barrier cells: implications for immune tolerance and neurotoxicity. J Neurochem 105:1346–1357. https://doi.org/10.1111/j.1471-4159.2008.05241.xJNC5241 CrossRefPubMedGoogle Scholar
- Pawlak K, Kowalewska A, Mysliwiec M, Pawlak D (2009) Kynurenine and its metabolites–kynurenic acid and anthranilic acid are associated with soluble endothelial adhesion molecules and oxidative status in patients with chronic kidney disease. Am J Med Sci 338:293–300. https://doi.org/10.1097/MAJ.0b013e3181aa30e6 CrossRefPubMedGoogle Scholar
- Peter B et al (2015) Incubator proof miniaturized Holomonitor to in situ monitor cancer cells exposed to green tea polyphenol and preosteoblast cells adhering on nanostructured titanate surfaces: validity of the measured parameters and their corrections. J Biomed Opt 20:067002. https://doi.org/10.1117/1.jbo.20.6.067002 CrossRefPubMedGoogle Scholar
- Phase Holographic Imaging PHI AB (2015) HoloMonitor application note on Label-free Cell Motility. Phase Holographic Imaging PHI AB. http://www.phiab.se/reports/2014/CellMotilityAppNotePHI-140919.pdf. Accessed 29 Sept 2017
- Silva-Adaya D et al (2011) Protective effect of l-kynurenine and probenecid on 6-hydroxydopamine-induced striatal toxicity in rats: implications of modulating kynurenate as a protective strategy. Neurotoxicol Teratol 33:303–312. https://doi.org/10.1016/j.ntt.2010.10.002S0892-0362(10)00171-6 CrossRefPubMedGoogle Scholar
- Tang W, Song H, Cai W, Shen X (2015) Real time monitoring of inhibition of adipogenesis and angiogenesis by (−)-epigallocatechin-3-gallate in 3T3-L1 adipocytes and human umbilical vein endothelial cells. Nutrients 7:8871–8886. https://doi.org/10.3390/nu7105437 CrossRefPubMedPubMedCentralGoogle Scholar
- Tiszlavicz Z, Nemeth B, Fulop F, Vecsei L, Tapai K, Ocsovszky I, Mandi Y (2011) Different inhibitory effects of kynurenic acid and a novel kynurenic acid analogue on tumour necrosis factor-alpha (TNF-alpha) production by mononuclear cells, HMGB1 production by monocytes and HNP1-3 secretion by neutrophils. Naunyn Schmiedebergs Arch Pharmacol 383:447–455. https://doi.org/10.1007/s00210-011-0605-2 CrossRefPubMedGoogle Scholar
- Varga G et al (2010) N-Methyl-d-aspartate receptor antagonism decreases motility and inflammatory activation in the early phase of acute experimental colitis in the rat. Neurogastroenterol Motil 22(217–225):e268. https://doi.org/10.1111/j.1365-2982.2009.01390.xNMO1390 Google Scholar
- Wilhelm I, Fazakas C, Krizbai IA (2011) In vitro models of the blood–brain barrier. Acta Neurobiol Exp (Wars) 71:113–128Google Scholar
- Zwilling D et al (2011) Kynurenine 3-monooxygenase inhibition in blood ameliorates neurodegeneration. Cell 145:863–874. https://doi.org/10.1016/j.cell.2011.05.020S0092-8674(11)00581-2 CrossRefPubMedPubMedCentralGoogle Scholar