Browsing by Author "Shaw, Klarissa"
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- ItemQuantification of the insulin response in rat L6 skeletal muscle cells(Stellenbosch : Stellenbosch University, 2020-04) Shaw, Klarissa; Snoep, Jacob Leendert; Van Niekerk, David Douglas; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.ENGLISH ABSTRACT: Mathematical models can help us to understand complex molecular mechanisms underlying diseases. There is a lack of these types of models describing the cellular processes involved in conditions such as insulin resistance and Type 2 diabetes (T2D). Therefore, this study aimed to quantify the reference state of the insulin signalling pathway for the construction of a mathematical model describing the dynamics of the pathway intermediates. This model will serve as a comparison for a model describing the diabetic state. Western blot analysis was used to quantify the phosphorylation states of Akt Ser473 and Thr308 at varying concentrations of insulin and over time, with or without insulin. The glucose transporter, GLUT4, activity was also investigated at deferent concentrations of insulin. These data were used in the construction of a model for the reference state. Ordinary deferential equations (ODEs) were derived to describe the dynamics of Akt over time. Steady state constraints were used to _t the dose response for Akt and made it possible to estimate a dephosphorylation/phosphorylation ratio. The estimated ratio was used to determine if the model could describe the time course data. A similar analysis was done for GLUT4. The model was able to accurately describe the phosphorylation and dephosphorylation dynamics of the Akt Ser473 and Thr308 site. In order to determine the link between insulin signalling and glucose transport, transport activity was plotted against Akt phosphorylation and the model was fitted to this data. The _t for both the Ser473 and Thr308 site did not differ. We found that the model predictions correlated quite well with the experimental data despite the lack of data on intermediates between insulin and Akt.