Browsing by Author "Myburgh, Lauren"

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    Modelling the insulin signaling pathway and glucose transport induction in differentiated 3T3-L1 mouse adipocytes
    (Stellenbosch : Stellenbosch University, 2021-04) Myburgh, Lauren; Snoep, Jacob Leendert; Van Niekerk, David Douglas; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: Insulin is the principal controller of energy homeostasis in the human body. It lowers postprandial blood glucose and enforces a change in cellu- lar metabolism to maintain glucose homeostasis by stimulating glucose uptake into muscle and adipose tissue. Dysfunction in this response can lead to insulin resistance which is a manifestation of type 2 diabetes for which the molecular mechanisms are not well understood. Although adipose tissue contributes less to peripheral glucose utilization compared to muscle tissue, knockout studies have identified adipose tissue as a key component in the dysfunction of insulin signalling. With the prevalence of this disease increasing drastically world- wide and majority of these patients being overweight, adipose tissue can be regarded as unhealthy despite its many health benefits, which include regu- lating energy balance and communicating with other organs as it functions as an endocrine organ. Therefore, it is crucial to firstly understand the ref- erence state under which the insulin signalling network and glucose uptake occurs as there is difficulty in identifying key components that result in type 2 diabetes. The network originates with the activation of the insulin receptor (IR) by insulin before eliciting a wide range of cellular responses that together coordinate glucose uptake into adipose tissue. Therefore, the primary purpose of this study was to describe the interactions involved between insulin and the intermediates, IR and Protein Kinase B (Akt) in the insulin signalling network and the e ect thereof on glucose uptake and metabolism in adipose tissue under a reference state using a systems biology approach. Firstly, the adipose tissue used in this study was 3T3-L1 adipocytes. Secondly, the intermediates, IR and Akt, were analyzed using Western blot analysis and glucose transport activity was measured using radio-labelled glucose uptake using these di erentiated 3T3-L1 adipocytes. Finally, the ux through glycolysis was measured to determine whether the transport activity changes in the same degree to the ux. The response of the insulin signalling network intermediates and glucose transport activity in a dose-dependent and time responsive manner were determined. This resulted in a hyperbolic relationship between insulin and the IR over time, demonstrated by both the experimental results and the mathematical model used. Experimentally an overshoot response, which is de ned as transient peaks of phosphorylation followed by a lower steady state level, was observed between insulin and Akt. The lower steady state level was caused by a down-regulation of the signal while still in the presence of insulin. However, the model could not predict this response as the molecular mechanisms that result in the overshoot behaviour are not known and therefore were not included in the model restricting the system to a hyperbolic response. The model could to some extent, however, predict the time responsive phosphorylation and dephosphorylation dynamics of the interactions involved. Thus, the study was able to capture the dynamic behaviour of the system in question thereby allowing a better understanding of the interactions between insulin and its intermediates and its a ect on glucose transporter activity and metabolism within a reference state in 3T3-L1 adipocytes.