Mathematical modelling of glycolysis in differentiated mouse adipocytes 3T3-L1

De Villiers, William (2021-03)

Thesis (MSc)--Stellenbosch University, 2021.

Thesis

ENGLISH ABSTRACT: Diabetes mellitus is a metabolic disease characterised by high blood sugar levels, primarily caused by insulin resistance and the resulting inability of the body to process glucose. Any tissue or cell that utilizes glucose is a target of diabetes research. Obesity is closely related to the development of diabetes due to the swelling of adipocytes caused by triacylglycerol formation. As glucose and its breakdown by glycolysis are directly responsible for supplying triacylglycerol formation, the focus of this work was on glycolysis in 3T3-L1 adipocytes. A bottom-up modelling approach was used in which all enzymes within glycolysis and the initial lipogenic enzyme G3PDH were characterised in order to create a kinetic model for glycolysis in 3T3-L1 adipocytes. Nondiabetic adipocyte cell extracts were utilized due to the need for a reference state model. Validation was performed using an HPLC analysis of glycolytic intermediates and co-factors. The validation was partially successful with glucose consumption and several glycolytic intermediate dynamics well described. Lactate production was underestimated in the model due to the high G3PDH activity within cell extracts which caused a signi cant fraction of the glucose to be converted via the glycerol branch, which was not seen to the same degree in the HPLC analysis. Being the rst model of its kind to be constructed for adipocytes, this work has laid the groundwork for further modelling of adipocyte glucose metabolism to better understand the diabetic condition in adipose tissue.

AFRIKAANSE OPSOMMING: Diabetes mellitus is 'n metaboliese siekte wat gekenmerk word deur hoë bloedsuikervlakke, hoofsaaklik veroorsaak deur insulienweerstandigheid en die gevolglike onvermoë van die liggaam om glukose te verwerk. Enige weefsel of sel wat glukose gebruik, is 'n teiken vir navorsing oor diabetes. Vetsug hou nou verband met die ontwikkeling van diabetes as gevolg van die swelling van adiposiete wat veroorsaak word deur die vorming van triasielgliserol. Aangesien glukose en die afbreek daarvan deur glikolise direk verantwoordelik is vir die verska ng van triasielgliserolvorming, was die fokus van hierdie werk op glikolise in 3T3-L1-adiposiete. 'n Benaderingsmodel benadering is gebruik waarin alle ensieme binne glikolise en die aanvanklike lipogene ensiem G3PDH gekarakteriseer is om 'n kinetiese model vir glikolise in 3T3-L1 adiposiete te skep. Nie-diabetiese adiposiet selekstrakte is gebruik as gevolg van die behoefte aan 'n verwysingstoestandmodel. Validasie is uitgevoer met behulp van 'n HPLC-analise van glikolitiese tussenprodukte en medefaktore. Die validering was gedeeltelik suksesvol met glukoseverbruik en verskeie glikolitiese intermedi êre dinamika wat goed beskryf is. Laktaatproduksie is in die model onderskat as gevolg van die hoë G3PDH-aktiwiteit in selekstrakte, wat veroorsaak het dat 'n beduidende fraksie van die glukose via die gliserolvertakking omgeskakel is, wat nie in dieselfde mate in die HPLC-analise gesien is nie. Aangesien dit die eerste model in sy soort is wat vir adiposiete vervaardig is, het dit die basis gelê vir verdere modellering van metabolisme van adiposiete glukose om die diabetiese toestand in vetweefsel beter te verstaan.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/110283
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