Metabolic and Immune System Responses to a Mixed Meal

The aim of this research is to investigate differences in metabolism and inflammation in adipose tissue and the circulation in response to a meal with increasing adiposity. It is hypothesised that participants with increased fat mass, will have a more pronounced inflammatory response to a meal compared to participants with a 'healthy' weight. The inflammatory response will be assessed by measuring changes in gene expression and protein secretion in the adipose tissue and by measuring markers of metabolism, insulin resistance, cardiovascular disease and inflammation in the blood.

Following advertisement of the study, interested potential participants will be asked to contact the Chief Investigator for further information via email/telephone correspondence. There will be an initial assessment of eligibility based on inclusion/exclusion criteria and, if these requirements are met, the potential participant will be invited to a meeting to further discuss the trial. After they have read the participant information sheet and seen the flowchart outlining the timeline for the study, if they would like to take part, they will be asked to sign a consent form. Dates will then be scheduled for some preliminary anthropometric measurements to be taken, 9 days of physical activity monitoring and the trial date (1 full day in the laboratory only).

Preliminary measurements:

Preliminary testing will include anthropometric measurements of height, weight, waist and hip circumferences, sagittal waist height and blood pressure. These measurements will take place at the University of Bath Physiology Laboratories.

Physical activity monitoring:

For 9 days, participants will be fitted with a physical activity monitor (Actiheart™) and asked to record a corresponding diary of their physical activity during this period to aid its interpretation and allow a more accurate calculation of average daily activity energy expenditure. Participants should not make any conscious changes to their normal lifestyle habits/routines during this period. The first 2 days of activity monitoring will be excluded from analysis to account for reactivity.

Main trial day:

In the 3 days prior to the main trial day, participants will be asked to record their food and drink intake and 48 hours before they should refrain from performing any strenuous physical activity. In the 24 hours prior participants should also refrain from consuming any alcohol or caffeine.

Participants should arrive at the Sports Training Village, University of Bath in the morning following a 10 hour fast where they will have their body composition precisely assessed using a dual-energy Xray absorptiometer (DEXA). They will then be taken to the Physiology Resting Laboratory where baseline expired gases will be collected for assessment of resting metabolic rate (RMR). A cannula will be inserted into a forearm vein and baseline blood sample(s) taken for analysis of metabolic/inflammatory markers and isolation of T cells. A fat sample will also be obtained using a needle aspiration technique.

The participant will then be asked to consume a breakfast type meal that is high in carbohydrate and fat contents consisting of brioche, jam, butter, and milkshake which has been adapted from a meal used by Rijkelijkhuizen J. et al, 2008, which induced stronger beta cell responses (i.e. of insulin) compared to a glucose load and showed differences with insulin sensitivity, so this meal type should identify differences in metabolism based on adiposity.

Furthermore, there is some evidence that high starch/low fibre foods may induce greater inflammation (Manning, Sutherland et al. 2008). The meal composition for all individuals will be identical but the energy content from carbohydrate will be calculated as a portion of the individuals' resting metabolic rate. This allows standardisation of energy intake for differences body weight/composition and will reflect energy requirements of the individual whilst at rest.

The meal should be consumed within 15 minutes and blood samples will be taken from the cannula at 15, 30, 60, and 90 minutes and then at every hour for 6 hours after consumption of the meal to monitor changes in metabolic and inflammatory markers. Additional blood samples will be obtained at 2 and 6 hours post meal by venepuncture for analysis of inflammatory markers as there is evidence that the cannula can stimulate local inflammation which interferes with the true result.

Expired gas samples for RMR will also be taken hourly and indirect calorimetry will be used to estimate relative fat and carbohydrate metabolism in response to the meal.

A second fat sample will be taken at 6 hours to examine inflammatory and metabolic responses to the meal within adipose tissue. A second blood sample for T cell isolation will also be obtained at this time point to examine corresponding metabolic/inflammatory responses in T cells.

Analysis:

On the day of the trial whole blood will be analysed for white blood cell count, glucose and lactate. Plasma and serum samples will be extracted from the whole blood via centrifugation and stored at -80. Monocytes and T lymphocytes (populations of white blood cells) will also be isolated from whole blood for later assessment of insulin sensitivity and culture experiments.

In each fat sample, separate portions of either adipose tissue or adipocytes (isolated from the other cells within the adipose tissue using a digestion method) will be cultured and media collected for later investigations. mRNA expression will also be examined in adipose tissue and isolated adipocytes. The remaining cells from the adipose tissue (SVF) will also be stored for later analysis of cell populations using flow cytometry, and expression/secretion analysis.