Bibian van der Voorn

195 GENERAL DISCUSSION AND SUMMARY In chapter 8 , we showed that poor intrauterine and postnatal growth was associated with early life alterations in HPA axis activity in children born preterm, while at age 8 years HPA axis settings seemed to be recovered, as these did not differ from their well grown equivalent anymore. Since adaptation of the HPA axis is thought to underlie these changes, this could contribute to increased health risks later in life. Moreover with the data presented in chapter 9 , it can be made plausible that development of the HPA axis takes place in a gender-specific manner, with a gender-specific evolution induced by puberty, thereby contributing to a gender-specific DOHaD. STRENGTHS AND LIMITATIONS OF THIS THESIS One of the strengths of this thesis is the development of new methodologies, specifically the LC-MS/MS assay to measure glucocorticoids in human milk. Although many studies have focused on the beneficial effects of nutritional contents of early life feeding, nutritive and non-nutritive, bioactive components of milk cannot be seen separately 8 . Accordingly, our data gave some more insights into the possible role of milk glucocorticoids, by showing the existence of a diurnal rhythm which correlated highly with maternal HPA axis activity. We believe that more attention should be paid to the non-nutritive signaling components of milk, in addition to the nutritional factors, and facilitated that by developing this new assay. All studies that were presented in this thesis were observational, and therefore could be hampered by selection, performance and detection biases. Animal studies have the advantage that stress can be randomly assigned, environmental circumstances strictly controlled and in vivo effects determined at any time. Still, human studies are necessary to translate the hypotheses that were gained from those experiments to the very preterm newborn. When interpreting our observational data, it is important to take into consideration that we had to deal with uncontrollable factors like the amount of stress exposure ( chapter 3, 7 and 8 ), timing of outpatient visit appointments ( chapter 2 and 3 ), necessity to adhere to feeding times ( chapter 5 ), selection by survival ( chapter 7 ), attrition at follow-up ( chapter 7 and 8 ), and the lack of standardization of study protocols and presentation of data ( chapter 9 and 10 ). Among these biases, selection bias is the most difficult one. While the causes of preterm birth are complex, there might be an overlap between factors that cause individual variations in stress responses, and factors that lie in the causal pathway of being born preterm or surviving preterm birth. Many researchers therefore advocate to include a full term control group when studying (very) preterm newborns. Some