Ahmet HACIMÜFTÜOĞLU, Betül ÇİÇEK, Ali TAGHİZADEHGHALEHJOUGHİ, Gülşah GÜNDOĞDU, Onur ŞENOL, Kemal Alp NALCI, Alper Kürşat DEMİRKAYA

Glutamat Eksitotoksisitesi Oluşturulan Primer Kortikal Nöron Kültürlerinde Parietinin Nöroprotektif Etkisinin İncelenmesi

Glutamat merkezi sinir sisteminin en önemli nörotransmitter maddesidir. Aşırı glutamat salınımı, glutamat reseptörlerininuzun süreli aktivasyonuna neden olarak eksitotoksisteye yol açar. Parietin, Rheum ribes L.’den izole edilen çeşitli farmakolojiközelliklere sahip bir antrakinondur. Bu çalışmada glutamat eksitotoksisitesine maruz bırakılan primer kortikal nöronkültürlerinde parietinin nöroprotektif etkisinin araştırılması amaçlanmıştır. Primer kortikal nöronlar, yeni doğan SpragueDawley cinsi sıçanlardan elde edildi. Glutamat eksitotoksisitesini stimüle etmek için, kültür medyumuna 10-5 Mkonsantrasyonda glutamat uygulandı. Daha sonra hücrelere 2.5-500 μM konsantrasyonda parietin uygulanarak 24 saatinkübasyona bırakıldı. Hücre canlılık oranı MTT yöntemi ile belirlendi. Aynı zamanda hücrelerde oluşan reaktif oksijen türleritotal antioksidan seviyesi (TAS)- total oksidan seviyesi (TOS) yöntemi ile değerlendirildi. MTT analiz sonuçlarına göre 10 µMparietinin glutamat eksitotoksisitesine karşı nöronlarda anlamlı düzeyde koruyucu etkiye sahip olduğu tespit edildi. TAS-TOSanaliz sonuçlarına göre 10 µM parietinin hücrelerde antioksidan seviyesini anlamlı ölçüde artırırken, parietinin yüksekkonsantrasyonlarının hücrelerdeki oksidan seviyesini anlamlı ölçüde arttırdığı gözlendi. Bu çalışma sonucuna göre parietininprimer kortikal nöron hücrelerinde glutamat eksitotoksisitesine karşı koruyucu etkiye sahip olduğu ve glutamateksitotoksisitesine karşı teropatik bir ajan olarak kullanılabileceği düşünülmektedir.

Investigation of Protective Effect of Parietin Against Glutamate Excitotoxicity in Primary Cortical Neuron Culture

Glutamate is the most important neurotransmitter in the central nervous system. Excitotoxicity is induced by excessive release of glutamate followed by overstimulation of glutamate receptor. Parietin is an anthraquinone from Rheum ribes L. and has been reported to have a variety of pharmacological properties. The present study investigated the neuroprotective effects of parietin in primary cortical neuron cultures against glutamate excitotoxicity. Primary rat cortical neuronal cultures were obtained from new born Sprague Dawley rats. Cultures were subject to 10-5 M glutamate to stimulate glutamate excitotoxicity. After that, cells were treated with 2.5-500 μM concentrations of parietin during 24 h in dose dependent manner. Cell viability was determined using MTT assay. Reactive oxygen species generation was assessed using the total antioxidant status (TAS)-total antioxidant status (TOS) assays. The results of MTT analysis showed that 10 µM parietin effectively protected neuron from glutamate toxicity. According to the results of TAS-TOS analysis, it showed that 10 µM parietin significantly increased the antioxidant level in the cells, whereas high concentrations of parietin significantly increased the oxidant level in the cells. The results of this study suggest that parietin had neuroprotective effect against glutamate excitotoxicity in primary rat cortical neuron cultures and it may be conceive that parietin can be used as a therapeutic agent for glutamate excitotoxicity.

PDF

___

1. Albright TD., Jessell TM., Kandel ER., Posner MI., 2000. Neural science: a century of progress and the mysteries that remain. Cell, 100, 1-55.
2. Elmann A., Telerman A., Ofir R., Kashman Y., 2017. Glutamate Toxicity to Differentiated Neuroblastoma N2a Cells Is Prevented by the Sesquiterpene Lactone Achillolide A and the Flavonoid 3, 5, 4′-Trihydroxy-6, 7, 3′- Trimethoxyflavone from Achillea fragrantissima. J Mol Neurosci, 62, 99-105.
3. Wen SY., Li AM., Mi KQ., Wang RZ., Li H., Liu HX., Xing Y., 2017. In vitro neuroprotective effects of ciliary neurotrophic factor on dorsal root ganglion neurons with glutamate-induced neurotoxicity. Neural Regen Res, 12, 1716-1721.
4. Hacimuftuoglu A., Tatar A., Cetin D., Taspinar N., Saruhan F., Okkay U., Turkez H., Unal D., Stephens RL., Suleyman H., 2016. Astrocyte/neuron ratio and its importance on glutamate toxicity: an in vitro voltammetric study. Cytotechnology, 68, 1425-1433.
5. Kanamori K., 2016. In vivo N-15 MRS study of glutamate metabolism in the rat brain. Analytical biochemistry, 529, 179-192.
6. Zhao J., Verwer R., van Wamelen D., Qi XR., Gao SF., Lucassen P., Swaab D., 2016. Prefrontal changes in the glutamate-glutamine cycle and neuronal/glial glutamate transporters in depression with and without suicide. Journal of psychiatric research, 82, 8-15.
7. Kostic M., Zivkovic N., Cvetanovic A., Stojanovic I., Colic M., 2017. IL-17 signalling in astrocytes promotes glutamate excitotoxicity: Indications for the link between inflammatory and neurodegenerative events in multiple sclerosis. Multiple sclerosis and related disorders, 11, 12- 17.
8. Song JH., Kang K., Choi Y., 2017. Protective effect of casuarinin against glutamate-induced apoptosis in HT22 cells through inhibition of oxidative stress-mediated MAPK phosphorylation. Bioorganic & Medicinal Chemistry Letters, 27, 5109-5113.
9. Zhao H., Ji ZH., Liu C., Yu XY., 2015. Neuroprotective mechanisms of 9-hydroxy epinootkatol against glutamate-induced neuronal apoptosis in primary neuron culture. J Mol Neurosci, 56, 808-814.
10. Jia N., Sun Q., Su Q., Chen G., 2016. SIRT1- mediated deacetylation of PGC1α attributes to the protection of curcumin against glutamate excitotoxicity in cortical neurons. Biochemical and biophysical research communications, 478, 1376-1381.
11. Floyd RA., Hensley K., 2002. Oxidative stress in brain aging: implications for therapeutics of neurodegenerative diseases. Neurobiology of aging, 23, 795-807.
12. Kim HT., Prochiantz A., Kim JW., 2016. Donating Otx2 to support neighboring neuron survival. BMB reports, 49, 69-70.
13. Mehta A., Prabhakar M., Kumar P., Deshmukh R., Sharma P., 2013. Excitotoxicity: bridge to various triggers in neurodegenerative disorders. Eur J Pharmacol, 698, 6-18.
14. Verrier J., Kochanek P., Jackson T., 2013. Anthraquinone-2-sulfonic acid (AQ2S) is a novel neurotherapeutic agent. Cell death & Disease, 4, 451-459.
15. Wei G., Wu Y., Gao Q., Zhou C., Wang K., Shen C., Wang G., Wang K., Sun X., Li X., 2017 Effect of Emodin on Preventing Postoperative IntraAbdominal Adhesion Formation. Oxid Med Cell Longev, 2017.
16. Turkmen O., Crka M., Suat E., 2005. Initial evaluation of a new edible wild rhubarb species (Rheum ribes L.) with a modified weighted scaling index method. Pakistan Journal of Biological Sciences, 8, 763-765.
17. Hong JY., Chung HJ., Bae SY., Trung TN., Bae K., Lee SK., 2014. Induction of cell cycle arrest and apoptosis by physcion, an anthraquinone isolated from rhubarb (rhizomes of Rheum tanguticum), in MDA-MB-231 human breast cancer cells. Journal of cancer prevention, 19, 273.
18. Pang MJ., Yang Z., Zhang XL., Liu ZF., Fan J., Zhang HY., 2016. Physcion, a naturally occurring anthraquinone derivative, induces apoptosis and autophagy in human nasopharyngeal carcinoma. Acta Pharmacologica Sinica, 37, 1623-1640.
19. Li W., Li F., Zhu Y., Song D., 2017. Physcion 8-O-βglucopyranosideregulates cell cycle, apoptosis, and invasion in glioblastoma cells through modulating Skp2. Biomedicine & Pharmacotherapy, 95, 1129-1138.
20. Wang Q., Jiang Y., Guo R., Lv R., Liu T., Wei H., Ming H., Tian X., 2017. Physcion 8-O-βglucopyranoside suppresses tumor growth of Hepatocellular carcinoma by downregulating PIM1. Biomedicine & Pharmacotherapy, 92, 451- 458.
21. Michaels R., Rothman SM., 1990. Glutamate neurotoxicity in vitro: antagonist pharmacology and intracellular calcium concentrations. Journal of Neuroscience, 10, 283-292.
22. Sasaki-Hamada S., Suzuki A., Sanai E., Matsumoto K., Oka JI., 2017. Neuroprotection by chotosan, a Kampo formula, against glutamate excitotoxicity involves the inhibition of GluN2B-, but not GluN2A-containing NMDA receptor-mediated responses in primary cultured cortical neurons. J Pharmacol Sci, 135, 134-137.
23. Coyle JT., Puttfarcken P., 1993. Oxidative stress, glutamate, and neurodegenerative disorders. Science-New York Then Washington, 262, 689- 699.
24. Depp C., Bas-Orth C., Schroeder L., Hellwig A., Bading H., 2017. Synaptic activity protects neurons against calcium-mediated oxidation and contraction of mitochondria during excitotoxicity. Antioxidants & Redox Signaling, 28, 530-537.
25. Lipton SA., Rosenberg PA., 1994. Excitatory amino acids as a final common pathway for neurologic disorders. New England Journal of Medicine, 330, 613-622.
26. Kortagere S., Mortensen OV., Xia J., Lester W., Fang Y., Srikanth YV., Salvino JM., Fontana ACK., 2018. Identification of novel allosteric modulators of glutamate transporter EAAT2. ACS Chemical Neuroscience, 9 (3), 522-534.
27. Gao F., Liu W., Guo Q., Bai Y., Yang H., Chen H., 2017. Physcion blocks cell cycle and induces apoptosis in human B cell precursor acute lymphoblastic leukemia cells by downregulating HOXA5. Biomedicine & Pharmacotherapy, 94, 850-857.
28. Sylvester PW., 2011. Optimization of the tetrazolium dye (MTT) colorimetric assay for cellular growth and viability. Drug Design and Discovery: Methods and Protocols, 32, 157-168.
29. Wang Q., Wang Y., Xing Y., Yan Y., Guo P., Zhuang J., Qin F., Zhang J., 2016. Physcion 8-O-βglucopyranoside induces apoptosis, suppresses invasion and inhibits epithelial to mesenchymal transition of hepatocellular carcinoma HepG2 cells. Biomedicine & Pharmacotherapy, 83, 372- 380.
30. Büyükokuroğlu ME., Gepdiremen A., Hacimüftüoğlu A., Oktay M., 2003. The effects of aqueous extract of Lavandula angustifolia flowers in glutamate-induced neurotoxicity of cerebellar granular cell culture of rat pups. Journal of Ethnopharmacology, 84, 91-94.
31. Ghezzi F., Monni L., Corsini S., Rauti R., Nistri A., 2017. Propofol Protects Rat Hypoglossal Motoneurons in an In Vitro Model of Excitotoxicity by Boosting GABAergic Inhibition and Reducing Oxidative Stress. Neuroscience, 367, 15-33.
32. Hara Y., McKeehan N., Dacks P.A., Fillit H.M., 2017. Evaluation of the Neuroprotective Potential of N-Acetylcysteine for Prevention and Treatment of Cognitive Aging and Dementia. J Prev Alzheimers Dis, 4, 201-206.
33. Brkanac SR., Geric M., Gajski G., Vujcic V., GarajVrhovac V., Kremer D., Domijan AM., 2015. Toxicity and antioxidant capacity of Frangula alnus Mill. bark and its active component emodin. Regulatory Toxicology and Pharmacology, 73, 923-929.
34. Sun YN., Li W., Lee SH., Jang HD., Ma JY., Kim YH., 2017. Antioxidant and anti-osteoporotic effects of anthraquinones and related constituents from the aqueous dissolved Aloe exudates. Natural Product Research, 23, 1-4.
35. Gu JW., Hasuo H., Takeya M., Akasu T., 2005. Effects of emodin on synaptic transmission in rat hippocampal CA1 pyramidal neurons in vitro. Neuropharmacology, 49, 103-111.