Reversing/delaying the age-related switch leading to β-cell quiescence
1 Purpose: Previously we developed a mouse model of monogenic diabetes, carrying HNF1a inducible mutation, which showed a premature accumulation of senescence markers, leading to hyperglycemia and thus, diabetes onset (experiments performed under FOTS 10785 and breeding application no. 12015). By following our experimental mice we also observed a difference in hyperglycemia level, which might be connected either with stress or with diet. Caloric restriction was previously shown to influence cell senescence. This project aims at delaying aging of insulin-producing cells lacking HNF1a (our monogenic diabetes model) by reducing caloric input, thus analyzing the impact of diet in diabetes mutation carriers as well as diabetic mice. We also want to assess the quality of diet on premature diabetes onset due to high fat food intake, and further analyze the differences between female and male mutation-carriers. We showed that HFD worsens the diabetic phenotype, leading to rapid weight gain and increased hyperglycemia, which can be decreased to almost normal levels by restricting the food intake. We are currently investigating the molecular players involved in this process.
2 Distress: Caloric restriction which is calculated at 60% of normal food intake up to 6 months; The mice are followed daily and weighted regularly, thus identifying quickly any factors affecting their welfare. The mice show a moderate initial weight loss, which stabilizes after 2 weeks.
Mice are housed 2 per cage containing a separator wall in order to administrate precise measurement of food intake and also to avoid detrimental behaviours related to competition for food.
3 Expected benefit: Monogenic diabetes like MODY (maturity onset of diabetes of the young) shows a high variability related to the age of the disease onset. This may be correlated with diet, or with the early senescence of the insulin-producing cells. Identifying ways to keep insulin-producing cells healthy and functional, especially by simple measures as caloric restriction, would greatly benefit people carrying the disease mutation but not yet exhibiting the disease signs. Our study identified potential benefits of caloric reduction before and after diabetes onset. We plan to study further the cellular and molecular mechanisms involved in diabetes onset, which could also lead to novel targets to be used in diabetes treatment.
4 Number of animals, and what kind: We will use transgenic mice with inducible mutation in HNF1a (floxed) or HNF1a (humanized – carrying the human mutation), under the control of RIPcre or RIPcreERT, specific promoters that lead to the selective expression of the mutation only in insulin producing cells. These mice are also carrying a florescent marker (YFP), which will label the cells carrying the mutated protein. We would like to add also a mouse strain carrying the HNF1 mutation in all endocrine islet cells (expression under the control of Ngn3cre), as our studies showed the involvement of other islet cells aside beta-cells, in diabetes development.
5 How to adhere to 3R: These experiments require a living organism due to its target on systemic metabolism, studying several organs’ interactions (pancreas-liver-intestine). We will optimize the number of mice necessary for our projects by using common experimental groups, where the same set of collected samples will be processed separately for replying to distinct questions, hence halving the number of required animals. For establishing the experimental setups we screened the literature in order to establish a proven protocol, thus eliminating the need to pretesting the experimental conditions.
2 Distress: Caloric restriction which is calculated at 60% of normal food intake up to 6 months; The mice are followed daily and weighted regularly, thus identifying quickly any factors affecting their welfare. The mice show a moderate initial weight loss, which stabilizes after 2 weeks.
Mice are housed 2 per cage containing a separator wall in order to administrate precise measurement of food intake and also to avoid detrimental behaviours related to competition for food.
3 Expected benefit: Monogenic diabetes like MODY (maturity onset of diabetes of the young) shows a high variability related to the age of the disease onset. This may be correlated with diet, or with the early senescence of the insulin-producing cells. Identifying ways to keep insulin-producing cells healthy and functional, especially by simple measures as caloric restriction, would greatly benefit people carrying the disease mutation but not yet exhibiting the disease signs. Our study identified potential benefits of caloric reduction before and after diabetes onset. We plan to study further the cellular and molecular mechanisms involved in diabetes onset, which could also lead to novel targets to be used in diabetes treatment.
4 Number of animals, and what kind: We will use transgenic mice with inducible mutation in HNF1a (floxed) or HNF1a (humanized – carrying the human mutation), under the control of RIPcre or RIPcreERT, specific promoters that lead to the selective expression of the mutation only in insulin producing cells. These mice are also carrying a florescent marker (YFP), which will label the cells carrying the mutated protein. We would like to add also a mouse strain carrying the HNF1 mutation in all endocrine islet cells (expression under the control of Ngn3cre), as our studies showed the involvement of other islet cells aside beta-cells, in diabetes development.
5 How to adhere to 3R: These experiments require a living organism due to its target on systemic metabolism, studying several organs’ interactions (pancreas-liver-intestine). We will optimize the number of mice necessary for our projects by using common experimental groups, where the same set of collected samples will be processed separately for replying to distinct questions, hence halving the number of required animals. For establishing the experimental setups we screened the literature in order to establish a proven protocol, thus eliminating the need to pretesting the experimental conditions.