Biochemistry and Microbiology

Permanent URI for this collection

https://hdl.handle.net/10530/475

Browse

Browsing Biochemistry and Microbiology by Subject "aspalathin"

Now showing 1 - 1 of 1
  • Thumbnail Image
    Item
    A study of the protective effects of an aspalathin enriched green rooibos extract on experimentally induced hepatic steatosis
    (2019) Khuboni, Noxolo Thobile
    Background: Non-alcoholic fatty liver disease (NAFLD) is a hepatic manifestation of the metabolic syndrome, and is strongly associated with insulin resistance, obesity and type 2 diabetes (T2D). Growing evidence shows NAFLD as a multi-system disease, affecting several extra hepatic organs and regulatory pathways (Byrne et al., 2015). The prevalence of NAFLD is expected to escalate dramatically with the upsurge in metabolic diseases, particularly obesity and type 2 diabetes. However, an optimal treatment regime to cure NAFLD has not been established. Rooibos is gaining increasing recognition for its ability to improve health and has demonstrated hepatoprotective effects against carbon tetrachloride (CCl4) and tert-butyl hydroperoxide induced hepatotoxicity in rodents. This study will establish if an aspalathin rich green rooibos extract (Afriplex GRT™) is able to modulate or prevent the excessive fat accumulation in experimental models of NAFLD. Aim: The aim of the study was to assess the modulatory effect of GRT extract in in vitro and in vivo models of experimentally induced NAFLD. Methods: The study used immortalized C3A liver cells and oleic acid to induce steatosis. Assessment of the effect of GRT on oleic acid induced steatosis was determined using different concentrations of aspalathin-enriched rooibos (GRT) extract. Cells not exposed to oleic acid were included as controls. Pioglitazone was used as a positive control. Steatosis of C3A cells was induced using 1 mM oleic acid for 24 hours. Treatment with GRT and pioglitazone for 24 hours, either commenced in the presence of oleic acid (simultaneous treated group) or 24 hours after oleic acid induced steatosis (post-treated group). MTT assay was performed to determine cell viability, while lipid content was measured using oil red O staining normalized for cell number by crystal violet staining. The effect of GRT extract on mRNA relevant to lipogenic genes (PPAR-α, SREBP, FASN and CPT1), inflammation (TNF-α) and glucose metabolism (GCK and ChREBP) were assessed. Further investigation was done using leptin-receptor deficient C57BL6/J db/db mice (n=8/group) and their lean littermates (db/+) (n=8/group). The mice received standard ii rodent chow ad libitum. Treatments were given daily with powdered chow containing a calculated dose equal to 74 mg/kg BW and 740 mg/kg BW GRT or 10 mg/kg BW pioglitazone. Food intake, water intake, body weight and fasting glucose was monitored weekly for the duration of the study. A week prior to termination of the study, oral glucose tolerance tests were performed. At termination the livers were collected and either fixed in formalin or snap frozen for storage at -80oC. Gene and protein expression were performed using qPCR and Western blot, respectively, for the genes and proteins related to the development of NAFLD. The effect of GRT extract on the liver was confirmed by histology. The data was analysed using one-way analysis of variance (ANOVA) followed by Dunnet multiple comparison post hoc test. Statistical significance was set at p ˂ 0.05. Results: This study showed that 1 mM oleic acid for 24 hrs induced lipid accumulation in C3A cells and produced a good in vitro model for studying NAFLD. GRT extract displayed protective effects against NAFLD in oleic acid induce steatosis and decreased lipid content by < 80% in steatotic C3A cells. The reduction was comparable to pioglitazone (control drug). mRNA analysis revealed that GRT extract regulated fatty acid synthesis by downregulating genes involved in lipogenesis (SREBP 1c and FASN) and glucose metabolism (GCK and ChREBP) as well as inflammation (TNF-α). In db/db mice, GRT improved glucose tolerance and reduced the liver weight to body weight ratio. In terms of genes and proteins that are involved in NAFLD, GRT increased key lipolytic genes PPAR-α and CPT 1 involved in the regulation of beta-oxidation of lipids and reduced HMG-CoA synthase 1 (HMGCS1), the gene regulating cholesterol synthesis. Histology confirmed increased steatosis limited to zone 3 of the liver acinus synonymous with NAFLD. Treatment with GRT did not significantly affect the steatotic severity score of the db/db mice. Conclusion: GRT was effective at reducing oleic acid induced steatosis in C3A cells by regulating key genes involved in fatty acid and cholesterol synthesis, and inflammation. In db/db mice GRT extract modulated glucose tolerance and increased the expression of effector proteins involved with beta-oxidation in the liver but was not effective at reducing NAFLD
University of Zululand