Toksični učinak bakra u kulturi stanica neuroblastoma SH-SY5Y

master's thesis

• Title (english): Toxic effect of copper in neuroblastoma SH-SY5Y cells
• Author: Marijana Batarelo
• Mentor: Maja Jazvinšćak Jembrek (mentor)
• Mentor: Julija Erhardt (sumentor)
• Committee member: Julija Erhardt (predsjednik povjerenstva)
• Committee member: Zoran Marčić (član povjerenstva)
• Committee member: Nada Oršolić (član povjerenstva)
• Committee member: Sandra Hudina (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2021-09-28, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Oksidacijski stres, uzrokovan neravnotežom stvaranja i uklanjanja reaktivnih kisikovih i dušikovih vrsta, ima važnu ulogu u starenju mozga i razvoju neurodegenerativnih bolesti. U ovom radu, istraživala sam toksični učinak bakra na stanice neuroblastoma SH-SY5Y. Testom MTT, tripanskim modrilom i mjerenjem razine ATP-a ustanovila sam značajan pad preživljenja stanica SH-SY5Y nakon tretmana bakrom. Bakar je povećao proizvodnju reaktivnih vrsta, koje pri visokim vrijednostima izazivaju oksidacijsko oštećenje, te smanjio razinu glutationa. Također, izazvao je kondenzaciju kromatina bez aktivacije kaspaza 3 i 7, kao i oštećenja stanične membrane, iz čega zaključujem da je bakar izazvao programiranu staničnu smrt putem neovisnim o aktivaciji kaspaza i nekrozu. Na proteinskoj razini došlo je do povećane ekspresije proteina p53 i PARP-1, dok se ekspresija proteina NME1 smanjila. Primjena inhibitora proteina PARP-1 (PJ34) nije utjecala na toksičnost, dok je primjena inhibitora proteina p53 (pifitrin-α) uzrokovala povećanje toksičnosti bakra. Flavonoid miricetin pojačao je citotoksičnost bakra prooksidacijskim djelovanjem. Dobiveni rezultati razjašnjavaju mehanizam prooksidacijskog djelovanja bakra na staničnoj i molekularnoj razini što bi moglo pridonijeti razvoju novih farmakoloških pristupa u prevenciji i terapiji neurodegenerativnih bolesti.

Abstract (english)

Oxidative stress, caused by an imbalance in the formation and elimination of reactive oxygen and nitrogen species, plays an important role in brain aging and the development of neurodegenerative diseases. In this study, I investigated the toxic effect of copper on SH-SY5Y neuroblastoma cells. By MTT test, trypan blue exclusion assay, and ATP level measurement, I found a significant decline in SH-SY5Y cell survival after copper treatment. Copper has increased the production of reactive species, which at high values cause oxidative damage, and reduced glutathione levels. It also caused chromatin condensation without caspase 3 and 7 activation, as well as cell membrane damage, from which I conclude that copper caused programmed cell death via caspase-independent activation and necrosis. At the protein level, there was increased expression of p53 and PARP-1 proteins, while NME1 protein expression decreased. Administration of the PARP-1 protein inhibitor (PJ34) did not affect toxicity, whereas administration of the p53 protein inhibitor (pifitrin-α) caused an increase in copper toxicity. The flavonoid myricetin enhanced the cytotoxicity of copper by its prooxidative action. The obtained results clarify the mechanism of prooxidative action of copper at the cellular and molecular level, which could contribute to the development of new pharmacological approaches in the prevention and treatment of neurodegenerative diseases.

• Keywords: Oksidacijski stres; redoks metali; ROS; neuroblastom; prooksidansi; antioksidansi; neurodegeneracija
• Keywords (english): Oxidative stress; redox metals; ROS; neuroblastoma; prooxidants; antioxidants; neurodegeneration
• Language: croatian
• URN:NBN: urn:nbn:hr:217:194468
• Study programme: Title: Experimental Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra eksperimentalne biologije (magistar/magistra eksperimentalne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2021-10-20 09:55:42