Utjecaj metabolita mikroalgi Pseudo-nitzschia na parametre oksidacijskog stresa stanica krvi

master's thesis

• Title (english): Impact of microalgae Pseudo-nitzschia metabolites on oxidative stress parameters in blood cells
• Author: Melissa Kuralić
• Mentor: Ana-Marija Domijan (mentor)
• Mentor: Mirta Smodlaka Tanković (mentor)
• Committee member: Ana-Marija Domijan (predsjednik povjerenstva)
• Committee member: Mirta Smodlaka Tanković (član povjerenstva)
• Committee member: Erim Bešić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of pharmaceutical botany); Zagreb
• Defense date and country: 2022-06-13, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Pharmacy

Abstract

Mikroalge roda Pseudo-nitzschia prisutne su u gotovo svim morskim i estuarijskim ekosustavima te čine bazu hranidbenoga lanca. Pojedine vrste roda Pseudo-nitzschia izlučuju neurotoksin, domoičnu kiselinu (DK). Poremećaj koji se razvija u ljudi koji su hranom unijeli u organizam DK naziva se amnezijsko trovanje školjkašima. Oksidacijski stres predstavlja neravnotežu u oksido-redukcijskim reakcijama u organizmu te se povezuje s razvojem niza bolesti u ljudi. Cilj ovoga istraživanja bio je ispitati oksidacijski stres kao mehanizam toksičnosti metabolita pojedinih vrsta roda Pseudo-nitzschia u ljudskim stanicama krvi. Prvo su uzgojene tri vrste roda Pseudo-nitzschie, P. delicatissima (PN1), P. pseudodelicatissima (PN2) i P. calliantha (PN3) te je sakupljen hranjivi medij u kojem su rasle mikroalge. Potom su uzorci krvi zdravih dobrovoljnih davatelja tretirani medijem u kojem su rasle Pseudo-nitzschia PN1, PN2 i PN3 te morem i morem i medijem (kontrole) u razdoblju od 4 i 24 sata. Nakon tretmana u uzorcima krvi određene su koncentracije: glutationa (GSH), mjera anti-oksidacijske obrane, malondialdehida (MDA), pokazatelj lipidne peroksidacije te proteinskih karbonili (PC), pokazatelji oksidacijskog oštećenja proteina, spektrofotometrijski. Dobiveni rezultati statistički su obrađeni Studentovim t-testom (Excel, MS Office), a razina značajnosti postavljena je na p ≤ 0,05.
Tretman krvi (4 i 24 sata) s medijem u kojem su rasle Pseudo-nitzschie PN1, PN2 i PN3 doveo je do porasta koncentracije GSH, ali je samo nakon 24-satnog tretmana s P. pseudodelicatissima (PN2) zabilježena značajno viša koncentracija GSH u odnosu na kontrolu (p ≤ 0,05). Tretman hranjivim medijem Pseudo-nitzschie snizio je koncentraciju MDA, a značajan pad koncentracije MDA u krvi u odnosu na kontrolnu skupinu zabilježen je nakon 24-satnog tretmana medijem u kojem je rasla P. delicatissima (PN1) (p ≤ 0,05). Tretman krvi medijem u kojem su rasle PN1, PN2 i PN3 doveo je do pada koncentracije PC, no taj pad nije bio značajan u odnosu na kontrolu. Zabilježeni porast koncentracije GSH nakon tretmana medijem u kojem su rasle Pseudo-nitzschie pokazuje da je došlo do aktivacije obrane od oksidacijskog stresa. Sniženje koncentracija MDA i PC može se povezati s povećanom koncentracijom GSH koja je spriječila oksidacijska oštećenja lipida i proteina. Konačno, može se zaključiti da ispitivane vrste Pseudo-nitzschie u hranjivi medij izlučuju metabolit(e) koji mogu izazvati oksidacijski stres u stanicama krvi.

Abstract (english)

Microalgae of the genus Pseudo-nitzschia are present in almost all marine and estuarine ecosystems and are the basis of the food chain. Some species of the genus Pseudo-nitzschia produce neurotoxin, domoic acid (DA). Individuals who have consumed food containing DA develop disorder called amnesic shellfish poisoning. Oxidative stress is result of an imbalance in oxido-reduction reactions in the body and in humans is associated with the development of numerous diseases. The aim of this study was to explore on human blood cells oxidative stress as a mechanism of toxicity of metabolites of some Pseudo-nitzschia species. In the first step of the study three species of the genus Pseudo-nitzschie were grown: P. delicatissima (PN1), P. pseudodelicatissima (PN2) and P. calliantha (PN3), and the nutrient medium in which the microalgae grew was collected. In the next step blood samples from healthy voluntary donors were treated with collected medium as well as with sea and sea and media (that served as controls) for 4 and 24 hours. After treatment in blood samples glutathione (GSH) as marker of anti-oxidative defence, malondialdehyde (MDA) as marker of lipid peroxidation and protein carbonyls (PC) as marker of oxidative damage to proteins were determined spectrophotometrically. For statistical analysis Student t-test (Excel, MS Office) was used, and the significance level was set at p ≤ 0.05.
In blood samples exposed for 4- and 24-hours to medium in which Pseudo-nitzschie PN1, PN2 and PN3 grew an increase in GSH level was observed. However, only in blood samples exposed for 24-hours to medium in which P. pseudodelicatissima (PN2) grew significantly higher GSH level in comparison to control is recorded (p ≤ 0.05). Exposure to medium in which Pseudo-nitzschie grew decreased MDA level in blood samples. After 24-hour exposure of blood cells to medium in which P. delicatissima (PN1) was grown, a significant decrease in MDA concentration compared to the control group was observed (p ≤ 0.05). 4- and 24-hour exposure of blood cells to medium in which PN1, PN2 and PN3 were grown led to a decrease in PC level, however, the decrease was not significant compared to control. The observed increase in GSH level in blood samples exposed to the medium in which Pseudo-nitzschie grew indicate that in blood cells activation of the defence against oxidative stress occurred. Decrease in MDA and PC level may be explained with increased GSH level that prevented oxidative damage to lipids and proteins. Therefore, it can be concluded that the tested Pseudo-nitzschie species into the nutrient medium secrete metabolite(s), which can induce oxidative stress in human blood cells.

• Keywords: Pseudo-nitzschia; domoična kiselina; oksidacijski stres; glutation; malondialdehid; proteinski karbonili
• Keywords (english): Pseudo-nitzschia; oxidative stress; glutathione; malondialdehyde; protein carbonyls
• Language: croatian
• URN:NBN: urn:nbn:hr:163:568386
• Study programme: Title: Pharmacy; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra farmacije (magistar/magistra farmacije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2022-07-05 12:13:15