Alzheimer’s disease (AD) is characterized by a progressive cognitive decline and is one of the most common neurodegenerative diseases in elderly. Given the aging population and predicted prevalence of AD in the future, it has become very important to identify populations at risk for AD in an early stage, in order to try to prevent the disorder and benefit from the potential (development of) targeted therapy in the future. There is evidence that environmental and lifestyle factors exert a modulatory role in the age-of-onset and progression of AD. Stress is also considered a relevant lifestyle factor that alters the risk to develop AD and the effects of stress can be more pronounced when they occur during a critical developmental time window in early-life. Indeed, stress during the sensitive perinatal period elicits lasting effects on the structure and function of the adult brain and might further enhance the risk for age-related cognitive decline, as well as AD itself.

In this thesis, we aimed to extend our knowledge on the role of early-life stress (ES) exposure in the progression of AD features, including cognitive deficits with aging, neuropathology levels and different forms of brain plasticity, like hippocampal neurogenesis. We further set out to study whether such ES-induced consequences might be associated with changes in the immune cells of the brain, microglia, and in neuroinflammatory regulation.

In our studies, we make use of an established mouse model for chronic ES. The mice are placed in a cage with minimal nesting material from postnatal day (P)2 to P9. This leads to well-described alterations in the adult hippocampus and also to cognitive deficits several months later. In addition, we use naturally aged wild type mice as well as the APPswe/PS1dE9 and APP.V717I AD mouse models, that develop genetically driven expression of β-amyloid (Aβ) neuropathology.

In Chapter 1, we review the impact of different forms of early-life adversity, i.e. stress, nutritional changes and immune activation, on later hippocampal structure and function. These different forms of adversity affect the later neuroendocrine stress profile, the nutritional profile and the immunological profile in the brain and we argue that these different systems are in fact highly interactive. In addition, these different forms of early-life adversity are all known to impair hippocampal neurogenesis and cognitive functioning. Therefore, we hypothesize that the later-life outcome after ES exposure and other adversities is determined by the interactive characteristic of the stress, nutritional and immunological profiles, that together might lead to a synergistic activation of these systems.

In Chapter 2, we followed up on this hypothesis and subjected APP/PS1 male mice and their wild type littermates to chronic ES and studied if ES affected Aβ neuropathology and neuroinflammation. We showed that ES altered Aβ neuropathological hallmarks in a differential manner at two ages; ES firstly reduced Aβ levels in the hippocampus at 4 months of age, while the Aβ deposits were ultimately aggravated at 10 months of age. We further observed that ES affected neuroinflammatory factors in the hippocampus of wild type mice, directly after stress exposure at postnatal day 9, as well as in 4-month-old adult mice. In addition, while the 4-month-old APP/PS1 mice had reduced Aβ levels after ES exposure, they also showed a mild, but enhanced expression of microglial markers by ES at this same age. In contrast, the aggravated Aβ plaque load in the ES-exposed APP/PS1 mice was accompanied by reduced clustering of microglial Iba1 density around Aβ deposits at 10 months of age, and by a reduced expression of microglial markers. These findings indicate that ES differentially affected Aβ levels at 4 and 10 months, while the neuroinflammatory profile showed an opposite phenotype in the ES-exposed APP/PS1 mice.

In Chapter 3, we investigated microglial functioning and neuroinflammatory dysregulation after ES exposure. We showed that, relative to control cultures, an acute neuroinflammatory challenge exacerbated the pro-inflammatory response in primary hippocampal glial cultures from P9 ES-exposed mice. In addition, the phagocytic response of microglia in culture was enhanced by ES exposure; the microglia in ES-derived cultured internalized more beads, although internalization of aggregated Aβ1-42 was not significantly altered by ES. These results are preliminary and might require a larger sample-size. In addition to these data on microglial functioning after acute, in vitro challenges, ES altered the expression of various regulators of microglial functioning in the hippocampus at P9 and at 10 months of age. These alterations included factors that mediate the interaction between microglia and neurons. This might therefore, in part regulate the neuroinflammatory phenotypes at these ages. In addition, APP/PS1 mice had an elevated expression of various neuroinflammatory mediators, that was dampened in ES-exposed APP/PS1 mice. Similarly, proliferation of microglia in 10-month-old APP/PS1 mice was reduced by ES exposure. With these findings, we expand on previously described later-life consequences of ES for neuroinflammation and show that ES might sensitize the acute inflammatory response in P9 cultures and that ES dampens the activation upon continuous Aβ exposure in adult APP/PS1 mice.

In Chapter 4, we determined whether ES also accelerated or aggravated the behavioral deficits and hippocampal neurogenesis alterations in APP/PS1 mice. ES wild type mice appeared to show an earlier cognitive impairment than ES APP/PS1 mice at a young adult (3 months) age. This indicates thus a rather protective phenotype in ES APP/PS1 mice, than the hypothesized acceleration of cognitive decline. At 10 months of age, ES did also not further impair the learning behavior of APP/PS1 mice. While APP/PS1 overexpression induced alterations in hippocampal neurogenesis at both 4 and 10 months of age, these alterations were not modulated by previous exposure to ES at either age. We can thus conclude that, even though these same mice were previously described to exhibit an altered Aβ neuropathological and neuroinflammatory profile (Chapter 2), the exposure to ES did not accelerate or aggravate the cognitive decline and neurogenesis impairments in the APP/PS1 mice at the ages indicated.

In Chapter 5, we study how the presence of Aβ neuropathology affected adult hippocampal neurogenesis and neuroinflammation in middle-aged (10 months) to aged (27 months) APP.V717I mice. This is a milder single transgenic model for Aβ neuropathological progression, when compared to the double transgenic APP/PS1 mice. We showed that with increasing age, the Aβ accumulation induced parallel microglial activation in the APP.V717I mice. We further showed that hippocampal neurogenesis was reduced with age in both wild type and APP.V717I mice, but this decline was not affected by the Aβ and microglial changes at any age. Thus, the slower development of the AD-related neuropathological hallmarks in APP.V717I mice did not induce any subsequent response in hippocampal neurogenesis. We and others showed that a faster-progressing pathological accumulation of Aβ at younger ages did impact neurogenesis (Chapter 4). Together, these findings indicate that in view of its endogenous reduction with age, adult hippocampal neurogenesis might be less responsive to the emergence of neuropathological hallmarks particularly in middle-aged to old mice.

In Chapter 6, we investigate the consequences of ES for age-related cognitive decline, adult hippocampal neurogenesis and neuroinflammation in 20-month-old C57BL/6J mice. ES exposure had a tendency to enhance the variance in inter-individual performance and this seemed to lead to an amplification of the individual extremes within the ES population, in comparison to the control group. Although the ES-exposed mice showed a lasting reduction in hippocampal volume, other age-related changes in hippocampal neurogenesis were not affected by ES. Also, the expression of microglial priming-related factors and other inflammatory markers were not differentially affected followed ES exposure. These findings lead to the conclusion that mice that underwent ES exposure in the first week of their life do not show an aggravation in their age-related alterations in cognitive decline and hippocampal parameters when they reached 20 months of age.

Chapter 7 provides a general discussion of the data presented in this thesis in the context of the recent literature. Both positive and negative early-life experiences have been shown to alter Aβ neuropathology, while the effects of ES on tau neuropathology have been hardly studied to date. Several other studies have furthermore shown that these AD-related neuropathological alterations were associated with both alterations in cognitive decline, as well as in neuroplasticity in the hippocampus of various AD transgenic models. In addition, ES exposure altered cognitive decline in naturally aged rodents, which resulted in either an enhanced cognitive decline or higher inter-individual variance. However, little direct evidence has so far been provided for the underlying mechanisms and we discuss how stress-related, neuroinflammatory and metabolic factors could potentially mediate the altered AD progression following ES exposure. We bring forward a model of ES-induced vulnerability for AD development and progression and discuss the outstanding questions that can increase our understanding of this research field.

In conclusion, the findings in this thesis provide evidence for ES mediated modulation of Aβ neuropathological progression, while cognitive decline and neurogenesis were not aggravated by ES exposure in aging or in APP/PS1 mice. Furthermore, ES affected the microglial cells in the hippocampus and the expression of various neuroinflammatory factors in both the wild type and APP/PS1 mice. Such modulation potentially contributes to a higher pro-inflammatory response upon immune activation, and to an ES-enhanced progression of AD neuropathology. These hypotheses require further experimental investigation. Together, the findings presented in this thesis provide some first indications for a modulation of neuroinflammation and Aβ neuropathology by chronic ES. As such, it highlights the importance of future pre-clinical as well as clinical studies to further elucidate the relationship between these different factors and their underlying mechanisms.