Evaluating the neuroprotective effect of curcumin on a PINK1 cell model of Parkinson's disease

Thesis (MSc)--University of Stellenbosch University, 2022.ENGLISH ABSTRACT: Parkinson’s disease (PD) is a neurodegenerative disorder characterized by a loss of neurons producing the neurotransmitter dopamine. Notably, despite extensive studies that have revealed numerous dysregulated processes associated with PD, fundamental gaps still exist in our knowledge of the disease pathophysiology. Our understanding of the disease includes processes such as mitochondrial dysfunction causing increased oxidative stress and energy failure, as well as misfolded protein accumulation in large inclusions. However, the spatiotemporal sequence of events leading to PD and, importantly, the initial factors that trigger disease onset remain elusive. This challenges the development of effective therapeutic strategies to prevent and cure PD. Moreover, the various side effects associated with conventional pharmaceuticals currently used to treat this disease motivate the search for a natural treatment that can avoid augmenting the suffering associated with an already debilitating disease. It is evident that mitochondrial dysfunction and oxidative stress are likely to be involved in the disease pathogenesis, therefore studying these phenomena in PD may lead to the development of more effective therapeutic strategies. Curcumin is a plant-based polyphenol that has been observed to have antioxidant properties, increase cell viability, and enhance mitochondrial function. Consequently, the aim of the present study was to create a PD cellular model and evaluate the potential protective effects of curcumin. Therefore, we sought to establish an appropriate PD model by transfecting SH-SY5Y cells with wild-type (WT) or G309D mutant PINK1 cDNA, of which the latter has been shown to upregulate dopamine and lead to cytotoxicity. Additionally, since several neurotoxins have been shown to trigger PD, the toxic herbicide paraquat was administered to complement the model with cellular damage and mitochondrial dysfunction. To confirm the model, we performed RT-qPCR to measure gene expression levels of PINK1 and tyrosine hydroxylase (TH), an enzyme in the dopamine synthesis pathway. PINK1 was significantly upregulated in the mutant, however, no significant difference in TH gene expression was observed between groups. We then sought to measure the levels of dopamine in transfected cells using liquid chromatography-mass spectrometry (LC-MS). Although no dopamine was detected using LC-MS, higher levels of phenylalanine, a precursor of dopamine, were observed in the mutant. Using this model, we sought to test the protective effects of curcumin using assays that measure cellular and mitochondrial health. A toxic paraquat concentration of 1.7 mM was chosen to elicit a 50 % decrease in cell viability for the model, while a curcumin concentration of 2.5 μM was chosen as it exhibited no toxic effects. Following the establishment of the model, four treatment groups were established for all experiments thereon: untreated control, curcumin only treatment, paraquat only treatment, and pre-treatment (curcumin treatment followed by paraquat treatment). We found that curcumin was unable to significantly rescue the paraquat-induced reduction in cell viability and mitochondrial membrane potential. The latter was significantly reduced in PINK1 transfected groups, more so in the G309D mutant, indicating the toxic effects of the mutation. Thereafter, the effects of curcumin and polycaprolactone encapsulated nanocurcumin on cell viability were compared. Formulations of curcumin including nanocurcumin are postulated to improve the stability and efficacy of curcumin. Interestingly, curcumin had a greater protective effect, whereas nanocurcumin as well as the empty nanoparticles elicited toxicity. In fact, p re-treatment with the nanocurcumin prior to paraquat treatment caused a 30 % greater loss in cell viability compared to the paraquat treatment alone. Finally, a literature review was published, exploring the potential of consistent dietary consumption of curcumin as an alternative or supplement to existing therapies. We speculate that curcumin binds to α- synuclein protein (found to accumulate in PD) and that this complex is subsequently excreted from the body via the large intestine. In this view, replacing some of the PD drugs in an individual’s treatment regime with a nutraceutical, or ‘functional food’, like curcumin may improve therapeutic benefits with fewer side effects. Considering these results and the published evidence for curcumin as a dietary ‘nutraceutical’, further studies are required to optimize curcumin treatment before advocation of its widespread use as a PD therapeutic agent. Study limitations include the use of an unverified WT plasmid and an undifferentiated cell line, which can be addressed in future work. The findings in this study are of importance as they may contribute to advancing the development of novel plant-based therapies to treat and potentially prevent this detrimental disease.AFRIKAANSE OPSOMMING: Parkinson se siekte (PS) is 'n neurodegeneratiewe versteuring wat gekenmerk word deur 'n verlies aan neurone wat die neurotransmitter dopamien produseer. Merkwaardig, ten spyte van omrykende studies wat talle gedisreguleerde prosesse verwant aan PS geopenbaar het, bestaan fundamentele gapings steeds in ons kennis van die siektepatofisiologie. Ons begrip van die siekte sluit prosesse soos mitochondriale disfunksie in wat verhoogde oksidatiewe stres en energiemislukking veroorsaak, asook verkeerde proteïenophoping in groot insluitings. Die tydruimtelike volgorde van gebeure wat tot PS lei en, belangriker, die aanvanklike faktore wat siekte-aanvang veroorsaak, bly egter ontwykend. Dit daag die ontwikkeling van effektiewe terapeutiese strategieë uit om PS te voorkom en te genees. Bowendien, het die huidige konvensionele farmaseutiese behandelinge vir PS tale newe- effekte. Dit dien as motivering vir soektogte na ‘n natuurlike behandeling wat die aanvullende leiding kan verminder van ‘n reeds afbrekende siekte. Dit is duidelik dat mitochondriale disfunksie en oksidatiewe stres betrokke is by die siekte se patologie, daarom kan die bestudering van hierdie verskynsels in PS lei tot die ontwikkeling van meer effektiewe terapeutiese strategieë. Kurkumien is 'n plantgebaseerde polifenol wat waargeneem is om antioksidant eienskappe te toon, verhoog sel lewenskragtigheid en verbeter mitochondriale funksie. Gevolglik was die doel van die huidige studie om 'n PS-sellulêre model te skep en die potensiële beskermende effekte van kurkumien te evalueer. Daarom het ons probeer om 'n toepaslike PS-model te vestig deur SH-SY5Y-selle met wilde-tipe (WT) of G309D muteerde PINK1 cDNA te transreguleer, waarvan laasgenoemde dopamien op gereguleer het wat gelei het tot sitotoksiteit. Daarbenewens, aangesien verskeie neurotoksiene getoon is om PS te aktiveer, is die giftige onkruiddoder, parakwat, toegedien om die model aan te vul met sellulêre skade en mitochondriale disfunksie. Om die model te bevestig, het ons RT-qPCR uitgevoer om geen-uitdrukkingsvlakke van PINK1 en tyrosine hydroxylase (TH), 'n ensiem betrokke by die chemiese samestelling van dopamien, kwantitatief te meet. PINK1 was aansienlik opgereguleer in die muteerde, maar geen beduidende verskil was opgemerk in TH-geenuitdrukking tussen groepe nie. Ons het toe probeer om die vlakke van dopamien in transfekte selle te meet met behulp van vloeibare chromatografie-massaspektrometrie (VC-MS). Alhoewel geen dopamien opgespoor was met behulp van VC-MS nie, was hoër vlakke van fenylalanien, 'n voorloper van dopamien, in die muteerde waargeneem. Met behulp van hierdie model het ons probeer om die beskermende effekte van kurkumien te toets met behulp van eksperimente wat sellulêre en mitochondriale gesondheid kan bepaal. 'n Giftige parakwat konsentrasie van 1.7 mM was gekies om 'n 50% afname in sel lewensvatbaarheid in die model te ontlok, terwyl 'n kurkumien konsentrasie van 2.5 μM gekies was, aangesien dit geen giftige effekte getoon het nie. Na die bevestiging van die model was vier behandelingsgroepe gestig vir alle eksperimente wat volg daarna, dit sluit in: onbehandelde kontrole, kurkumien alleenlik, parakwat alleenlik, en voor-behandeling (kurkumien behandeling gevolg deur parakwat behandeling). Ons het gevind dat kurkumien nie merkwaardig die selle kon beskerm teen die verlaagde sel lewenskragtigheid en mitochondirale membraanpotentiaal wat geïnduseer was deur parakwat nie. Die mitochondriale membraanpotentiaal was aansienlik verminder in die PINK1-getransfekteerde groepe, meer so as in die G309D-muteerde, dit dui die giftige effekte van die mutasie aan. Daarna is die effek van kurkumien en policaprolactone ingeslote nanokurkumien op sel lewensvatbaarheid vergelyk. Formulerings van kurkumien, insluitend nanokurcumin, word gepostuleer om die stabiliteit en doeltreffendheid van kurkumien te verbeter. Interessant genoeg, kurkumien het 'n groter beskermende effek gehad, terwyl nanokurcumin sowel as die leë nanopartikels toksisiteit getoon het. In werklikheid, het die voorbehandeling met nanokurkumien (voor parakwat behandeling) ‘n 30% groter verlies in sel lewenskragtigheid veroorsaak as die selle met slegs parakwat behandeling. Ten slotte is 'n literatuuroorsig gepubliseer, wat die potensiaal van konsekwente dieet verbruik van kurkumien ondersoek as 'n alternatief of aanvulling tot bestaande terapieë. Ons spekuleer dat kurkumien aan α- sinukleien proteien bind (gevind om te versamel in PS) en dat hierdie kompleks daarna uit die liggaam uitgeskei word deur die dikderm. Deur dit in ag te neem, kan die vervanging van sommige van die huidige PS-behandelinge met nutraceutiese produkte, of ‘funksionele kos’, soos kurkumien, voordelig wees en minder newe-effekte hê. Samevattend met hierdie resultate en die gepubliseerde bewyse vir kurkumien as ‘n dieet ‘nutraceutical’, is verdere studies nodig om kurkumien behandeling te optimiseer voordat die wydverspreide gebruik daarvan as 'n PS terapeutiese middel kan gebeur. Studie beperkings sluit in die gebruik van 'n ongeverifieerde WT-plasmied en 'n ongedifferensieerde sellyn, wat in toekomstige werk aangespreek kan word. Die bevindings in hierdie studie is van belang, aangesien dit kan bydra tot die bevordering van die ontwikkeling van nuwe plantgebaseerde terapieë om hierdie nadelige siekte te behandel en moontlik te voorkom

Author Correction: The IPDGC/GP2 Hackathon - an open science event for training in data science, genomics, and collaboration using Parkinson’s disease data

Correction to: npj Parkinson’s Disease https://doi.org/10.1038/s41531-023-00472-6, published online 04 March 2023// In this article the affiliation details for Alastair J Noyce, Jonggeol Jeff Kim, Isabelle Francesca Foote, Sumit Dey were incorrectly given as ‘Department of Genetics and Genomic Sciences and Mindich Child Health and Development Institute, Icahn School of Medicine at Mount, Hess Center for Science and Medicine, New York, NY 10029, USA,’ but should have been ‘Preventive Neurology Unit, Wolfson Institute of Population Health, Queen Mary University of London, London, UK’.// The affiliation details for Prabhjyot Saini were incorrectly given as ‘Preventive Neurology Unit, Wolfson Institute of Population Health, Queen Mary University of London, London, UK’ but should have been ‘The Neuro (Montreal Neurological Institute-Hospital), McGill University, Montreal, QC, Canada’. The original article has been corrected

The IPDGC/GP2 Hackathon - an open science event for training in data science, genomics, and collaboration using Parkinson’s disease data

Open science and collaboration are necessary to facilitate the advancement of Parkinson's disease (PD) research. Hackathons are collaborative events that bring together people with different skill sets and backgrounds to generate resources and creative solutions to problems. These events can be used as training and networking opportunities, thus we coordinated a virtual 3-day hackathon event, during which 49 early-career scientists from 12 countries built tools and pipelines with a focus on PD. Resources were created with the goal of helping scientists accelerate their own research by having access to the necessary code and tools. Each team was allocated one of nine different projects, each with a different goal. These included developing post-genome-wide association studies (GWAS) analysis pipelines, downstream analysis of genetic variation pipelines, and various visualization tools. Hackathons are a valuable approach to inspire creative thinking, supplement training in data science, and foster collaborative scientific relationships, which are foundational practices for early-career researchers. The resources generated can be used to accelerate research on the genetics of PD

The IPDGC/GP2 Hackathon - an open science event for training in data science, genomics, and collaboration using Parkinson's disease data

Open science and collaboration are necessary to facilitate the advancement of Parkinson's disease (PD) research. Hackathons are collaborative events that bring together people with different skill sets and backgrounds to generate resources and creative solutions to problems. These events can be used as training and networking opportunities, thus we coordinated a virtual 3-day hackathon event, during which 49 early-career scientists from 12 countries built tools and pipelines with a focus on PD. Resources were created with the goal of helping scientists accelerate their own research by having access to the necessary code and tools. Each team was allocated one of nine different projects, each with a different goal. These included developing post-genome-wide association studies (GWAS) analysis pipelines, downstream analysis of genetic variation pipelines, and various visualization tools. Hackathons are a valuable approach to inspire creative thinking, supplement training in data science, and foster collaborative scientific relationships, which are foundational practices for early-career researchers. The resources generated can be used to accelerate research on the genetics of PD.This project was supported by the Global Parkinson’s Genetics Program (GP2). GP2 is funded by the Aligning Science Against Parkinson’s (ASAP) initiative and implemented by The Michael J. Fox Foundation for Parkinson’s Research (https://gp2.org).Open Access funding provided by the National Institutes of Health (NIH).This research was supported in part by the Intramural Research Program of the NIH, National Institute on Aging (NIA), National Institutes of Health, Department of Health and Human Services; project numbers ZO1 AG000535 and ZO1 AG000949, as well as the National Institute of Neurological Disorders and StrokePeer reviewe

Movement of prion-like α-synuclein along the gut–brain axis in Parkinson's disease : a potential target of curcumin treatment

A pathological hallmark of the neurodegenerative disorder, Parkinson's disease (PD), is aggregation of toxic forms of the presynaptic protein, α-synuclein in structures known as Lewy bodies. α-Synuclein pathology is found in both the brain and gastrointestinal tracts of affected individuals, possibly due to the movement of this protein along the vagus nerve that connects the brain to the gut. In this review, we discuss current insights into the spread of α-synuclein pathology along the gut–brain axis, which could be targeted for therapeutic interventions. The prion-like propagation of α-synuclein, and the clinical manifestations of gastrointestinal dysfunction in individuals living with PD, are discussed. There is currently insufficient evidence that surgical alteration of the vagus nerve, or removal of gut-associated lymphoid tissues, such as the appendix and tonsils, are protective against PD. Furthermore, we propose curcumin as a potential candidate to prevent the spread of α-synuclein pathology in the body by curcumin binding to α-synuclein's non-amyloid β-component (NAC) domain. Curcumin is an active component of the food spice turmeric and is known for its antioxidant, anti-inflammatory, and potentially neuroprotective properties. We hypothesize that once α-synuclein is bound to curcumin, both molecules are subsequently excreted from the body. Therefore, dietary supplementation with curcumin over one's lifetime has potential as a novel approach to complement existing PD treatment and/or prevention strategies. Future studies are required to validate this hypothesis, but if successful, this could represent a significant step towards improved nutrient-based therapeutic interventions and preventative strategies for this debilitating and currently incurable disorder.South African National Research Foundation and South African Medical Research Council.http://www.wileyonlinelibrary.com/journal/ejn2022-05-27hj2022Neurolog

Movement of Prion‐Like α‐Synuclein along the Gut‐Brain Axis in Parkinson’s Disease: A Potential Target of Curcumin Treatment

A pathological hallmark of the neurodegenerative disorder, Parkinson's disease (PD), is aggregation of toxic forms of the presynaptic protein, α-synuclein in structures known as Lewy bodies. α-Synuclein pathology is found in both the brain and gastrointestinal tracts of affected individuals, possibly due to the movement of this protein along the vagus nerve that connects the brain to the gut. In this review, we discuss current insights into the spread of α-synuclein pathology along the gut–brain axis, which could be targeted for therapeutic interventions. The prion-like propagation of α-synuclein, and the clinical manifestations of gastrointestinal dysfunction in individuals living with PD, are discussed. There is currently insufficient evidence that surgical alteration of the vagus nerve, or removal of gut-associated lymphoid tissues, such as the appendix and tonsils, are protective against PD. Furthermore, we propose curcumin as a potential candidate to prevent the spread of α-synuclein pathology in the body by curcumin binding to α-synuclein's non-amyloid β-component (NAC) domain. Curcumin is an active component of the food spice turmeric and is known for its antioxidant, anti-inflammatory, and potentially neuroprotective properties. We hypothesize that once α-synuclein is bound to curcumin, both molecules are subsequently excreted from the body. Therefore, dietary supplementation with curcumin over one's lifetime has potential as a novel approach to complement existing PD treatment and/or prevention strategies. Future studies are required to validate this hypothesis, but if successful, this could represent a significant step towards improved nutrient-based therapeutic interventions and preventative strategies for this debilitating and currently incurable disorder.South African National Research Foundation and South African Medical Research Council.http://www.wileyonlinelibrary.com/journal/ejn2022-05-27hj2022Neurolog

The IPDGC/GP2 Hackathon - an open science event for training in data science, genomics, and collaboration using Parkinson’s disease data

Abstract Open science and collaboration are necessary to facilitate the advancement of Parkinson’s disease (PD) research. Hackathons are collaborative events that bring together people with different skill sets and backgrounds to generate resources and creative solutions to problems. These events can be used as training and networking opportunities, thus we coordinated a virtual 3-day hackathon event, during which 49 early-career scientists from 12 countries built tools and pipelines with a focus on PD. Resources were created with the goal of helping scientists accelerate their own research by having access to the necessary code and tools. Each team was allocated one of nine different projects, each with a different goal. These included developing post-genome-wide association studies (GWAS) analysis pipelines, downstream analysis of genetic variation pipelines, and various visualization tools. Hackathons are a valuable approach to inspire creative thinking, supplement training in data science, and foster collaborative scientific relationships, which are foundational practices for early-career researchers. The resources generated can be used to accelerate research on the genetics of PD