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Title: Investigation of phosphatidylinositol 5-phosphate's role in insulin-stimulated glucose uptake in a skeletal muscle cell line
Author: Grainger, Deborah
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2011
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Phosphatidylinositol 5-phosphate (PtdIns5P) is the least well-characterised member of the phosphoinositide family of essential regulatory phospholipids. PtdIns5P levels are altered within cells in response to a number of stimuli and evidence is accumulating to suggest that it possesses important functions in cellular signalling. However, the physiological role of this lipid remains imperfectly understood. Previous studies have shown that PtdIns5P is elevated in adipocytes in response to insulin, and microinjection of PtdIns5P into these cells promotes plasma membrane insertion of the insulin-regulated glucose transporter GLUT4 (Sbrissa et al., 2004). This finding suggests a potential role of PtdIns5P as a mediator in insulin-stimulated glucose uptake, a process essential for efficient glucose homeostasis. As approximately 75% of postprandial glucose disposal is carried out by skeletal muscle, it is important to investigate the role of PtdIns5P in the response of this tissue to insulin. Therefore, this work has used differentiated myotubes of the rat muscle cell line, L6, to explore the effects of altered PtdIns5P levels on insulin-stimulated glucose uptake. This cell model had not been previously used in the laboratory so it first required characterisation. Here insulin is shown to stimulate a transient increase of PtdIns5P in L6 myotubes, indicative of a signalling role in response to insulin. This project developed several tools to further investigate this potential role for PtdIns5P in the insulin response of myotubes. One such development was the successful overexpression of the PtdIns5P 4-kinase PIP4KIIalpha in these cells, which was able to abolish the insulin-stimulated PtdIns5P rise. This correlated with a loss of insulin-stimulated glucose uptake (upon PIP4KIIalpha expression). Interestingly, artificial elevation of PtdIns5P in L6 myotubes increases glucose uptake in the absence of stimulation. This phenomenon appears to result from the activation of PI3-kinase signalling, as it is abolished by the PI3-kinase inhibitor wortmannin, and involves activation of the PI3-kinase effector Akt. These results are consistent with the idea that insulin-stimulated PtdIns5P production contributes to the robust PI3-kinase/Akt activation necessary for insulin-stimulated glucose uptake in muscle.
Supervisor: Dunne, Mark ; Hinchliffe, Katherine Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: skeletal muscle cell line ; L6 ; phosphatidylinositol 5-phosphate ; PtdIns5P ; Insulin ; GLUT4 ; Glucose uptake