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Role of riboflavin in cell fate: identification of determinants and effectors

Reference	BB/C518206/1
Principal Investigator / Supervisor	Professor Hilary Powers
Co-Investigators / Co-Supervisors	Professor Bernard Corfe, Professor Harry Moore, Dr Stuart Riley
Institution	University of Sheffield
Department	Human Nutrition Unit
Funding type	Research
Value (£)	358,031
Status	Completed
Type	Research Grant
Start date	01/06/2005
End date	31/12/2008
Duration	43 months

Abstract

Riboflavin is a water soluble vitamin with a well-established role in redox processes and important to growth and development. National studies in the United Kingdom have revealed low intakes and biochemical deficiency to riboflavin in substantial numbers in some groups, especially in young adults. There is therefore a strong public health argument for understanding the functional effects of low intakes of riboflavin. Feeding a diet low in riboflavin to young rats leads to a disruption of the normal development of the small intestine and the changes observed are compatible with a role for riboflavin deficiency in dysregulation of apoptosis and or proliferation. We hypothesise that riboflavin has multiple roles in the enterocyte important to the regulation of cell division, differentiation and death. We wish to determine the cellular effects of riboflavin in this context and to map the precise cellular effects in terms of alterations in gene expression and in the proteome. We will use two approaches: the culture of human intestinal cells exposed to different concentrations of riboflavin and the study of intestinal tissue in humans before and after riboflavin supplementation. 1) we will culture Caco-2 cells and determine effects of different concentrations of riboflavin on cell proliferation, apoptosis and cell differentiation. Effects of riboflavin will be mediated by changes in gene and protein expression and the nature of these changes will reflect mechanistic pathways. We will use the human Affymetrix Gene-Chip Microarray system to identify a group of genes regulated by riboflavin. The transcriptomics will be supported by proteomic analysis of cells, using 2d gel electrophoresis. These approaches will yield a panel of proteins that may be the key determinants of or biomarkers for the cellular response to riboflavin. Using QI-TOF-MS we aim to identify the key proteins of interest. We have conducted preliminary experiments on Caco-2 cells in culture that confirm a role for riboflavin in cell survival, and have also demonstrated effects or low riboflavin availability on the expressed proteome. 2) The in vitro work will inform the strategy to determine effects of riboflavin repletion on intestinal structure and cellular function, in human subjects. We will conduct a randomised controlled trial of riboflavin supplementation in two patient groups from whom we can collect intestinal biopsy material from the small and the large intestine. We will determine changes in intestinal morphology (supporting earlier animal studies) and proliferative and apoptotic activity (supporting the in vitro observations). In addition we will use the transcriptomics and proteomics data from the cell studies to inform a study of the effects of riboflavin supplementation on protein expression, using 1d electrophoresis and Western blotting, and, where necessary, RT-PCR. These approaches will provide an insight into the mechanisms whereby riboflavin helps to determine whether a cell divides, differentiates or dies, and thus how riboflavin directs normal development of the intestinal tract.

Summary

unavailable

Committee	Closed Committee - Agri-food (AF)
Research Topics	Ageing, Diet and Health
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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