Browsing by Author "Petersen, Ashlin Veruscka"

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    Investigating the effects of glucocorticoids and pro-inflammatory cytokines on oxidative stress in the HepG2 cell line
    (Stellenbosch : Stellenbosch University, 2023-03) Petersen, Ashlin Veruscka; Verhoog, Nicolette; Louw-Dutoit, Renate; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: Type 2 diabetes mellitus (T2DM) is characterized by hyperglycemia, hyperinsulinemia, and the inability of insulin-target tissues (the liver, adipose tissue, and skeletal muscle) to respond to insulin. If target tissues are unable to respond to insulin to maintain glucose homeostasis, insulin resistance develops, resulting in the onset of T2DM. Furthermore, oxidative stress is defined as an imbalance of reactive oxygen species (ROS) and antioxidants linked to obesity, cancer, and T2DM. Organelles such as mitochondria and transmembrane proteins such as NADPH oxidases (NOX) generate ROS. NADPH oxidase 3 (NOX3)-derived ROS has been linked to insulin resistance in the liver, which was investigated further in this project. In contrast, the antioxidant system is important in eliminating ROS and nuclear factor erythroid- 2 related factor 2 (NRF-2) was investigated as it is the most abundant antioxidant transcription factor in the liver. Glucocorticoids, which are associated with the stress response, and inflammatory mediators such as pro-inflammatory cytokines impair insulin signalling. However, the underlying cause of this impairment is not fully understood and therefore the primary aim of this study was to examine if oxidative stress contributes to stress- and inflammation-induced insulin resistance, either via NOX3-derived ROS or by affecting the antioxidant system through NRF-2 regulation. The findings showed that the synthetic glucocorticoid dexamethasone and the pro-inflammatory cytokines tumour necrosis factor- alpha (TNF-α) and interleukin-6 (IL-6) impair insulin signalling at different exposure times. Dexamethasone and TNF-α both increased ROS production (in the absence and presence of insulin). While IL-6 only induced ROS production in the presence of insulin only. The increase in ROS production was not accompanied by an increase in NOX3 mRNA or protein levels. The downregulation of NOX3 mRNA expression was observed in response to TNF-α in the absence and presence of dexamethasone. Similarly, IL-6 together with dexamethasone in the presence of insulin upregulated NOX3 mRNA expression. Additionally, NRF-2 expression, which is inhibited by glycogen synthase kinase-3 beta (GSK-3β), was not affected by any of the treatment conditions. To conclude the findings of this study, indicate that although glucocorticoids and pro-inflammatory cytokines cause hepatic oxidative stress, whether this ROS is produced via NOX3 or if the antioxidant system via NRF-2 plays a role is unclear and remains to be elucidated.