Effect of an obesogenic diet on the epigenetic profile of livers from Wistar rats and treatment with an aspalathin-rich rooibos extract
| dc.contributor.advisor | Pheiffer, Carmen | en_ZA |
| dc.contributor.advisor | Johnson, Rabia | en_ZA |
| dc.contributor.advisor | Samodien, Mugammad Ebrahim | en_ZA |
| dc.contributor.advisor | Windvogel, Shantal | en_ZA |
| dc.contributor.author | Abrahams, Yoonus | en_ZA |
| dc.contributor.other | Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Medical Physiology. | en_ZA |
| dc.date.accessioned | 2021-10-06T10:42:31Z | |
| dc.date.accessioned | 2021-12-22T14:15:28Z | |
| dc.date.available | 2021-10-06T10:42:31Z | |
| dc.date.available | 2021-12-22T14:15:28Z | |
| dc.date.issued | 2021-12 | |
| dc.description | Thesis (PhD)--Stellenbosch University, 2021. | en_ZA |
| dc.description.abstract | Background There is a scarcity of knowledge about obesity and metabolic syndrome development and progression in females, who are often under-represented in preclinical research. The aims of this study were thus to investigate the effects of high fat, high sugar (HFHS) feeding on gene expression and DNA methylation patterns in the livers of female and male Wistar rats and to determine whether an Aspalathin-rich rooibos extract, Afriplex® GRT, could prevent HFHS diet-induced metabolic dysregulation. Methods This study employed different animal models. Female and/or male Wistar rats were fed a HFHS diet for 12, 24 or 36 weeks. The effectiveness of Afriplex® GRT was investigated by daily supplementation with 60 mg/kg of bodyweight co-administered with diets for 36 weeks. Food and water intake, bodyweight, fasting blood glucose concentrations and glucose tolerance were measured regularly. After termination, organ weights, liver histology, fasting serum insulin, high- and low-density lipoprotein and cholesterol concentrations were measured. Gene expression was assessed using RT2 Profiler Rat Fatty Liver PCR arrays and TaqMan® gene expression assays. DNA methylation was quantified by pyrosequencing. The effects of DNA methylation were further investigated in the HepG2/C3A hepatocarcinoma cell line. Cells were treated with 2.5 μM of the DNA methyl transferase inhibitor, 5-aza-2’-deoxycytidine for 24 hours to induce DNA hypomethylation. Thereafter, steatosis and insulin resistance were induced by exposing cells to a 1:1 mixture of 0.5 mM palmitic and oleic acids for 24 hours. Cell viability, mitochondrial function, mitochondrial membrane potential, lipid accumulation, reactive oxygen species accumulation and gene expression were assessed. Results The HFHS diet increased visceral adiposity and hypertriglyceridaemia in both female and male rats, while hyperinsulinaemia and sustained obesity were observed in males only. Males fed the HFHS diet for 12 weeks showed reduced peroxisome proliferator-activated receptor gamma coactivator-1 alpha (Pgc1a) expression and hypermethylation of the Pgc1a promoter, but this was not observed in males fed the same diet for 36 weeks. Females fed HFHS showed a sex-specific reduction in peroxisome proliferator-activated receptor gamma (Pparg) expression. Afriplex® GRT did not prevent obesity or metabolic dysregulation. In vitro analysis demonstrated that DNA hypomethylation reduced lipid accumulation but did not affect insulin sensitivity. Steatosis and insulin resistance in the presence of DNA hypomethylation was associated with increased expression of uncoupling protein 2 (UCP2) and CCAAT/enhancer-binding protein beta (C/EBPß). Conclusion The HFHS diet elicited different responses in female and male Wistar rats. Male rats exhibited obesity, hyperinsulinaemia, glucose intolerance and altered methylation of Pgc1a. However, females exhibited a dampened metabolic response, potentially mediated through sex-specific downregulation of hepatic Pparg. Prophylactic daily supplementation with 60 mg/kg Afriplex® GRT did not prevent diet-induced obesity or metabolic dysregulation. For the first time, we show that DNA hypomethylation in HepG2 hepatocarcinoma cells decreases lipid accumulation in vitro and show that hypermethylation of a conserved CpG site in the promoter of Pgc1a is associated with decreased expression in the livers of male Wistar rats. We observed sex-specific hepatic differences between females and males which may have important implications for the development of therapeutic targets. In future, this may allow for the development of effective treatments free of sex-biased adverse effects and which confer protection against metabolic disease, reduce mortality and improve the quality of life of affected persons globally. | en_ZA |
| dc.description.abstract | Agtergrond Kennis is skaars oor vetsug en metaboliese sindroomontwikkeling en progressie by vroue, wat dikwels onderverteenwoordig is in prekliniese navorsing. Die doel van hierdie studie was om die gevolge van hoë vet, hoë suiker (HFHS) voeding op geenuitdrukking en DNA-metilerings- patrone in die lewers van vroulike en manlike Wistar-rotte te ondersoek en om te bepaal of 'n Aspalathin-ryke rooibos-uittreksel, Afriplex®GRT, kan voorkom dat die HFHS-dieët metaboliese disregulering veroorsaak. Metodes Hierdie studie het verskillende dieremodelle gebruik. Vroulike en / of manlike Wistar-rotte is 12, 24 of 36 weke lank 'n HFHS-dieet gevoer. Die effektiwiteit van Afriplex®GRT is ondersoek deur daaglikse aanvulling met 60 mg/kg liggaamsgewig wat saam met diëte vir 36 weke toegedien is. Voedsel- en waterinname, liggaamsgewig, vastende bloedglukosekonsentrasies en glukosetoleransie is gereeld gemeet. Na beëindiging is orgaangewigte, lewerhistologie, vastende seruminsulien, hoë- en lae-digtheid lipoproteïen en cholesterolkonsentrasies gemeet. Geenuitdrukking is beoordeel aan die hand van RT2 Profileerder Rot Vet Lever PCR-skikkings en TaqMan®-geenuitdrukkingstoetse. DNA-metilering is gekwantifiseer deur pirovolgordebepaling. Die effekte van DNA-metilering is verder ondersoek in die HepG2/C3A-lewer-sellyn. Selle is 24 uur lank behandel met 2.5 μM van die DNA- metieltransferase-inhibitor 5-aza-2'-deoxycytidine om DNA-hipometilering te veroorsaak. Daarna is steatose en insulien-weerstandigheid veroorsaak, deur selle aan 'n 1: 1-mengsel van 0,5 mM-palmities- en oleïensure vir 24 uur. Sel lewensvatbaarheid, mitochondriale funksie, mitochondriale membraanpotensiaal, lipiedophoping, reaktiewe suurstofspesie-ophoping en geenuitdrukking is assesseer. Resultate Die HFHS-dieet het visserale adipositeit en hipertrigliseridemie by vroulike, sowel as manlike rotte verhoog, terwyl hiperinsulinemie en volgehoue vetsug slegs by manlike rotte waargeneem is. Mannetjies wat die HFHS-dieet vir 12 weke gevoer was, het 'n verminderde peroksisoom- proliferatorgeaktiveerde reseptor gamma-koaktiveerder-1 alfa (Pgc1a) uitdrukking en hipermetilering van die Pgc1a-promotor, maar dit was nie waargeneem by mannetjies wat dieselfde dieet gedurende 36 weke gevoer was nie. Wyfies wat die HFHS gevoer was, het 'n geslag-spesifieke vermindering in peroksisoom-proliferatorgeaktiveerde gamma (Pparg) uitdrukking getoon. Afriplex®GRT het nie vetsug of metaboliese disregulering verhoed nie. In vitro-analise het getoon dat DNA-hipometilering die opeenhoping van lipied verminder het, maar nie die insulien sensitiwiteit beïnvloed het nie. Steatose en insulien-weerstandigheid in die teenwoordigheid van DNA-hipometilering was geassosieer met 'n verhoogde uitdrukking van ontkoppeling proteïen 2 (UCP2) en CCAAT / enhancer-bindende proteïen beta (C/EBPß). Gevolgtrekking Die HFHS-dieet het verskillende metaboliese reaksies, geenuitdrukking en DNA- metileringspatrone by vroulike en manlike Wistar-rotte ontlok. Mannetjies het vetsug, hiperinsulinemie, glukose-onverdraagsaamheid en veranderde metilering van Pgc1a getoon. Wyfies het egter 'n gedempte metaboliese respons getoon, wat moontlik bemiddel kan word deur geslagspesifieke onderregulering van lewer Pparg in wyfies. Profilaktiese daaglikse aanvulling met 60 mg/kg Afriplex®GRT het nie dieet-geïnduseerde vetsug of metaboliese disregulasie verhoed nie. Vir die eerste keer wys ons dat DNA-hipometilering in HepG2- hepatokarsinoomselle die opeenhoping van lipiede in vitro verminder. Verder, wys ons dat hipermetilering van 'n behoue CpG-setel in die promotor van Pgc1a geassosieer word met 'n verminderde uitdrukking in die lewers van manlike Wistar-rotte. Ons het geslagspesifieke lewerskille tussen vroulike en manlike rotte waargeneem, wat belangrike implikasies vir die ontwikkeling van terapeutiese teikens kan hê. In die toekoms kan dit voorsiening maak vir die ontwikkeling van effektiewe behandelings vry van seksbevooroordeelde nadelige effekte en wat beskerming teen metaboliese siekte verleen, sterftes verminder en die lewensgehalte van geaffekteerde persone wêreldwyd verbeter. | af_ZA |
| dc.format.extent | 207 pages | en_ZA |
| dc.identifier.uri | http://hdl.handle.net/10019.1/123678 | |
| dc.language.iso | en_ZA | en_ZA |
| dc.publisher | Stellenbosch : Stellenbosch University | en_ZA |
| dc.rights.holder | Stellenbosch University | en_ZA |
| dc.subject | Obesity | en_ZA |
| dc.subject | Nonalcoholic fatty liver disease | en_ZA |
| dc.subject | DNA -- Methylation | en_ZA |
| dc.subject | Rooibos extract | en_ZA |
| dc.subject | Wistar rats | en_ZA |
| dc.subject | Fatty liver | en_ZA |
| dc.subject | Liver -- Diseases | en_ZA |
| dc.subject | Epigenetics | en_ZA |
| dc.subject | UCTD | en_ZA |
| dc.title | Effect of an obesogenic diet on the epigenetic profile of livers from Wistar rats and treatment with an aspalathin-rich rooibos extract | en_ZA |
| dc.type | Thesis | en_ZA |