Grb10 and developmental programming: evaluation of a maternal diet restriction model during gestation

Sammanfattning: This study was conducted at the University of Bath, UK, from September to December 2014 as part of an ongoing research project aimed at elucidating how the Grb10 gene might act as a mediator of long-term health effects (such as predisposition to obesity, diabetes, coronary heart disease and hypertension) caused by environmental factors during development. This phenomenon is known as developmental programming. The purpose of this thesis was not primarily to answer any of the broader questions posed by this research project at large since this would require much more data than is reasonable to acquire over the course of a few months, but rather to evaluate the methods used in this project and reveal whether they are working the way they are presumed to. In the research project, which employs mice with the Grb10 gene knocked out as well as wild-type control mice, a dietary restriction model is utilized during gestation, which is supposed to generate offspring with lower birth weight and subsequent detrimental health effects in adulthood. This is supposed to be achieved by restricting the protein content of the pregnant mothers’ diet to 9 % (as opposed to the control diet of 20 %) throughout gestation. It is pivotal that the maternal dietary restriction model is functioning properly in order to gain developmental programming effects and perform worthwhile further testing later on in this project, such as blood pressure measurements, glucose tolerance tests and body composition analyzing. Any effects of the dietary restriction of the mothers during gestation are supposed to be shunted directly to the offspring and not to be lost by ”leaking off” and eliciting any adverse effects on the mothers themselves. Although the maternal diets differ in protein content, they are isocalorific and therefore not supposed to have any substantial effect on maternal growth during pregnancy. To validate this, 27 pregnant mice were weighed daily throughout gestation depending on genotype and diet. Genotype was determined via PCR of ear or tail clips of the mice. Food intake was also recorded. This set-up also allowed to record any hitherto unknown effects of Grb10 on growth induced by metabolic and endocrinological changes in adulthood such as pregnancy. An early indication of the dietary restriction effects on the offspring was also provided by weighing 60 pups at embryonic day 18.5 (e.g. one day before birth). The study revealed that there was no significant difference in the growth of pregnant mice according to genotype and diet, nor was the food intake affected by the dietary regime. However, it also gave a preliminary indication that there could potentially be problems with not achieving the desired lower birth weight effects on the offspring. Additionally, a deviation from the expected 50/50 birth ratio of maternal Grb10 knock-out offspring versus wild-type offspring when breeding a wild-type father with a Grb10 knock-out mother was discovered. The cause of the deviation is not known, but the findings indicate a perinatal mortality due to suffocation in maternal Grb10 knock-outs.