Quantitative analysis of key protein phosphorylation events as pathway biomarkers of metabolic disease in human cells.

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A combination of a more sedentary lifestyle and diets rich in fat has resulted in an explosion of health problems associated with metabolic dysfunction. For example, there are 2.3 million people in the UK with Type 2 diabetes, while one in ten women of child bearing age will experience polycystic ovarian syndrome. Indeed, simply being overweight increases the chances of developing these diseases. Type 2 diabetes alone costs the NHS around #2 billion pounds per year, while diabetes diagnosis is expected to double in the next 10 years. Type 2 diabetes is a major cause of heart disease, blindness, amputation and stroke, so reduces quality of life for individuals for many years as well as reducing life expectancy by around 10 years. One of the key features of Type 2 diabetes is poor response to the hormone insulin. All evidence points to this ?insulin resistance? occurring prior to the development of Type 2 diabetes, but it is technically impossible at present to screen for insulin resistance in the general population. We strongly believe that earlier diagnosis and more targeted therapy would prevent progression to Type 2 diabetes, hence preventing the devastating complications of the disease. In addition, insulin resistance is strongly associated with all obesity related diseases, including polycystic ovarian syndrome, hypertension and fatty liver disease. Therefore the same diagnostic approach, and potentially the same therapy could be applied to all these ?metabolic diseases?.In this proposal we aim to develop novel methodology (using state-of the-art Mass Spectrometry techniques) to accurately measure multiple biomarkers of insulin action inside cells. Insulin induces modifications of many proteins inside cells, and these modifications do not occur in the same pattern when cells or tissues are insulin resistant. Therefore accurate measurement of these biomarkers gives three things; firstly the pattern of modification after exposure to insulin provides the insulin sensitivity of the cells or tissues, secondly, an abnormal pattern provides the specific location of the molecular defect in the cells or tissues, and finally identification of the molecular defect allows classification of different forms of insulin resistance in order to identify the correct treatment for that subgroup.The project will be split into two phases, the development of the methodology, and validation of the new methodology using cells and carefully collected muscle biopsies from volunteers with different amounts of body fat (obesity), and therefore a range of different levels of insulin resistance.