An investigation of the neuroprotective properties of curcumin by monitoring autophagy and apoptosis
| dc.contributor.advisor | Bardien, Soraya | en_ZA |
| dc.contributor.advisor | Loos, Ben | en_ZA |
| dc.contributor.advisor | Abrahams, Shameemah | en_ZA |
| dc.contributor.author | Bekker, Minke | en_ZA |
| dc.contributor.other | Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Molecular Biology and Molecular Biology and Human Genetics. | en_ZA |
| dc.date.accessioned | 2021-04-30T07:57:06Z | |
| dc.date.available | 2021-04-30T07:57:06Z | |
| dc.date.issued | 2021-03 | |
| dc.description | Thesis (MSc)--Stellenbosch University, 2021. | en_ZA |
| dc.description.abstract | ENGLISH ABSTRACT: Parkinson’s disease (PD) is a neurodegenerative movement disorder, with a rapidly increasing prevalence and incidence throughout the global population. As current PD therapies only rely on symptomatic treatments, there is an urgent need for the development of neuroprotective therapies, to slow or halt progressive neuronal loss. This strategy, however, is dependent on a better understanding of the pathobiology and pathways underlying PD. Multiple causative factors have been postulated to be involved in the pathobiology of PD, with defective autophagy and the subsequent upregulation of apoptotic cascades hypothesised as main contributing factors. Although the crosstalk between these pathways has become evident, the exact role of autophagy and apoptosis regarding the neuronal fate in PD remains controversial. Curcumin is a polyphenolic plant compound that has been observed to mediate autophagy and apoptosis. Consequently, the aim of the present study was to investigate the potential of curcumin as a PD therapy, and its effect on autophagy and apoptosis in a PD model. Study objectives were set out to achieve this aim, with the first objective being to better understand the interplay between autophagy and apoptosis, by performing a literature review to identify molecular components involved in both pathways. The second objective was to establish an appropriate PD model for the experimental part of the study. The cellular model selected was treatment with a neurotoxin, paraquat, in a commercially available SH-SY5Y neuroblastoma cell line. This objective was executed through performing 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyl-2H-tetrazolium bromide and CyQUANT assays to determine appropriate concentrations for curcumin and paraquat treatment. The third objective was to assess the effect of curcumin in the autophagic pathway, through performing western blotting to measure levels of the autophagic LC3-II, p62 and LAMP2 proteins. Lastly, the fourth objective was to assess the effect of curcumin in the apoptotic pathway in an autophagy-inhibited model. This was executed through performing western blotting to measure the levels of the autophagic Beclin-1 and apoptotic Bax proteins. The literature review pinpointed Bcl-2, JNK, p38 and Akt as proteins with the potential to mediate the activation and/or inhibition of both pathways. It was concluded that novel PD therapies could target these connecting molecular components to mediate the balance between autophagy and apoptosis. Using our PD cellular model, our findings revealed a trend for curcumin treatment to increase the overall abundance of LC3-II protein levels, while also sustaining the protein levels of LAMP2 in the presence of the autophagic inhibitor bafilomycin A1. Curcumin treatment also facilitated the efficient clearance of p62 protein, in comparison to a blunted clearance of p62 protein observed upon paraquat treatment. These findings reflect the potential of curcumin to induce autophagy while also maintaining the balance of autophagic flux. Additionally, a trend for curcumin to increase Beclin- 1 and decrease Bax protein levels in the presence of the autophagic inhibitor, 3-methyladenine, was observed. Curcumin therefore exhibited the potential to counteract autophagic inhibition while also demonstrating anti-apoptotic properties, independently from autophagy. Considering these results and published evidence of curcumin-mediated changes in the expression of Bcl-2, JNK and, Akt, it is suggested that curcumin treatment exhibits the potential to facilitate the interplay between autophagy and apoptosis. Study limitations include the challenge of interpreting autophagic flux through western blotting and the use of an undifferentiated cell line and can be addressed in future work. The findings in this study are of importance, as they may contibute to a better understanding of the pathobiology of PD which could advance the development of novel therapies, to potentially ameliorate the detrimental nature of this disorder. | en_ZA |
| dc.description.abstract | AFRIKAANSE OPSOMMING: Parkinson se siekte (PS) is ʼn degeneratiewe, en bewegingsverwante, neurologiese siekte waarvan die voorkoms in gevalle spoedig toeneem in die globale populasie. Met huidige PS terapieë wat slegs staatmaak op die behandeling van PS simptome, word die ontwikkeling van ‘n terapie met die vermoë om progressiewe neurodegenerasie te vertraag, dringend benodig. Hierdie strategie is egter afhanklik van die verbetering van kennis rakende die onderliggende patobiologie en padweë teenwoordig in PS. Meervoudige faktore word gepostuleer om betrokke te wees in the die patobiologie van PS, waarvan defektiewe autofagie en die daaropvolgende opregulering van die apoptotiese kaskade as hoof veroorsakende faktore gehipotiseer word. Hoewel ‘n duidelike interaksie tussen die padweë bekend staan, bly die presiese rol van autofagie en apoptose, rakende die neurale noodlot in PS, steeds kontroversieel. Curcumin is ‘n polifenoliese plantsamestelling met die waargeneemde vermoë om autofagie en apoptose te reguleer. Gevolglik was die hoofdoel van die huidige studie om die terapeutiese potensiaal van curcumin as ‘n terapie vir PS te ondersoek deur die effek daarvan op autofage en apoptose te bestudeer. Die eerste doelwit om die hoofdoel te voltooi, was om ‘n verbeterde kennis rakende die wisselwerking tussen autofagie en apoptose op te doen. ‘n Literatuurstudie om die molekulêre komponente te identifiseer wat by altwee padweë betrokke is, is bewerkstelling. Die tweede doelwit het die vestiging van ‘n aanvaarbare PS model vir die eksperiment afdeling van die studie behels. Die verkose sellulêre model het die behandeling van ‘n kommersieel beskikbare SH-SY5Y neuroblastoma sellyn met die neurologiese toksien, paraquat, behels. Die doelwit was uitgevoer deur 3-(4,5-dimetieltiasol-2-iel)-2,5-difeniel-2H-tetrasolium bromide en CyQUANT toetse om gepaste konsentrasies van curcumin en paraquat behandelings te bepaal. Die derde doelwit was om die effek van curcumin in die autofagiese padweg te ondersoek deur die vlakke van die autofagiese LC3-II, p62 en LAMP2 proteÏene deur Westerse kladtegnieke te bestudeer. Laastens was die vierde doelwit om die effek van curcumin in die apoptose padweg, in ‘n autofagies-geÏnhibeerde model te ondersoek. Dit is uitgevoer deur die vlakke van die autofagiese Beclin-1 en apoptotiese Bax proteÏene, deur Westerse kladtegnieke, te bestudeer. In die literatuurstudie is Bcl-2, JNK, p38 en Akt as proteÏene, met die potensiaal om die aktivering en/of inhibisie van beide padweë te reguleer, uitgewys. Die gevolgtrekking was dat nuwe PS terapieë hierdie verwisselnde molekulêre komponente kan teiken, om sodoende die balans tussen autofagie en apoptose te reguleer. Deur gebruik te maak van ons sellulêre model, het ons bevindings uitgewys dat curcumin behandeling ‘n verhoogde neiging in die algehele vlakke van LC3-II tot effek gehad het, terwyl dit ook die vlakke van LAMP2, in die teenwoordigheid van die autofagiese inhibitor, bafilomycin A1, konstant gehou het. Curcumin behandeling het ook die effektiewe proteÏenopruiming van p62 gefasilitieer in vergelyking met die afgestompte proteÏenopruiming waargeneem vir die paraquat behandeling. Hierdie bevindings verteenwoordig die potensiaal van curcumin om autofagie te aktiveer terwyl dit ook die balans in die autofagiese vloed te onderhou. Daarbenewens het curcumin behandeling ook gelei tot die onderskeie toenemende en dalende neiging in Beclin-1 en Bax proteÏen vlakke in die teenwoordigheid van die autofagiese inhibitor, 3-metieladenien. Hierdeur was die potensiaal van curcumin, om autofagiese inhibisie teë te werk, uitgewys. Terselfdertyd het curcumin ook, onafhanklik van autofagie, anti-apoptotiese eienskappe gedemonstreer. Deur hierdie bevindings, asook die gepubliseerde bewyse van curcumin bemiddelde veranderings in Bcl-2, JNK en Akt vlakke, word dit voorgestel dat curcumin behandeling die vermoë het om die wisselwerking tussen autofagie en apoptose te reguleer. Die studie was beperk deur die uitdagende interpretasie van die autofagiese vloed, uitgevoer deur Westerse kladtegnieke, asook die gebruik van ‘n ongedifferensieerde sellyn. Verdere navorsing word benodig om hierdie beperkings aan te spreek. Die bevindings in hierdie studie word as belangrik geag, aangesien dit kan bydra tot ‘n verbeterde kennis in die patobiologie van PS, wat kan lei tot die ontwikkeling van nuwe terapieë om die nadelige natuur van PS teë te werk. | af_ZA |
| dc.description.version | Masters | en_ZA |
| dc.format.extent | 155 pages | en_ZA |
| dc.identifier.uri | http://hdl.handle.net/10019.1/110429 | |
| dc.language.iso | en_ZA | en_ZA |
| dc.publisher | Stellenbosch : Stellenbosch University | en_ZA |
| dc.rights.holder | Stellenbosch University | en_ZA |
| dc.subject | Nervous system -- Degeneration | en_ZA |
| dc.subject | UCTD | en_ZA |
| dc.subject | Parkinson’s disease (PD) | en_ZA |
| dc.subject | Curcumin as a PD therapy | en_ZA |
| dc.subject | Neuroprotective agents | en_ZA |
| dc.title | An investigation of the neuroprotective properties of curcumin by monitoring autophagy and apoptosis | en_ZA |
| dc.type | Thesis | en_ZA |