Periodic Reporting for period 2 - EarlyLife (Gut epithelial dynamics and function at the nexus of early life infection and long-term health)
Berichtszeitraum: 2023-02-01 bis 2024-07-31
Early life is now recognized as a critical and non-redundant time period during which the mucosal tissue and immune system are primed and the enteric microbiota is established. Three factors: the enteric microbiota, the mucosal immune system and the epithelial barrier cooperate to establish intestinal host-microbial homeostasis after birth. They also determine the risk to develop prevalent inflammatory, immune-mediated, and metabolic diseases. Maturation of the mucosal immune system and establishment of the enteric microbiota have been extensively studied. In contrast, postnatal evolvement of epithelial cell type heterogeneity and functional specialization and the influence of enteric infection on this process have not been explored. Therefore, the analysis of the cellular heterogeneity of the intestinal epithelium and its functional specialization during healthy postnatal development but also after early life challenge is the primary aim of the EarlyLife project. Chronic inflammatory, immune-mediated, and metabolic diseases account for significant morbidity in the society and contribute to reduced quality of life, labor force, and increased health care costs. Studies during the last decades suggested that host-microbial interaction at mucosal body sites and here in particular the epithelial surface of the gastrointestinal tract represents a key aspect of immune priming that influences the pathogenesis of prevalent human inflammatory, immune-mediated, and metabolic diseases. In addition, infections of the gastrointestinal tract remain a major cause of childhood mortality and morbidity worldwide. The main clinical symptoms are diarrhea, fluid loss and electrolyte disturbance but gastrointestinal infections during infancy have also been associated with indirect consequences such as reduced growth, body weight gain and delayed intellectual development suggesting a lasting influence on mucosal tissue homeostasis. Thus, a better understanding of the developmental and adaptive processes that determine intestinal epithelial cell type heterogeneity and function both during postnatal development in the healthy individual and after transient challenge for example by enteropathogenic microorganisms is expected to provide novel targets for preventive and therapeutic strategies to reduce the societal burden of chronic diseases. EarlyLife employs innovative, multiscale technical approaches and analytical protocols in combination with novel in vivo models to generate the first comprehensive map of postnatal epithelial cell type and subtype differentiation and analyze the impact of early life infection by important human bacterial, viral, and parasitic pathogens. Effects will be functionally studied using epigenetic profiling, microbiota-transfer experiments, stem cell organoid culture, and genetic models. Identified mechanisms will be confirmed using for example human stem cell organoids. As a result, EarlyLife is expected to identify mechanisms of enhanced infection susceptibility of the neonate, decipher the critical and non-redundant influence of the postnatal period for mucosal homeostasis and explain the role of early life imprinting for long term immune-mediated, inflammatory, and metabolic diseases.
So far, we have characterized the cellular specialization and functional characterization across the postnatal period until postweaning and adulthood in mice (Torow, Hand and Hornef, 2023). This analysis has been performed using a multidimensional approach including global but also single cell transcriptomics, proteomics, epigenetics, and metabolomics. In addition, certain features were studied in greater detail using stem cell organoids and multi-color immunostaining of tissue sections. These analyses facilitated the generation of an intestinal epithelial cell atlas that depicts the dynamic age-dependent changes in cellular heterogeneity and function. They illustrated the particular situation of the neonate host and allowed insight in significant age-dependent changes in the digestive and substrate transport capacity, tissue remodeling, maturation of antimicrobial functions, and the regenerative capacity of the intestinal epithelium and are consistent with the particular role of the postnatal period. In addition, we have comparatively analyzed mice bred under germ-free conditions and conventional breeding conditions and exposed neonatal mice to the important human enteropathogens Giardia lamblia, Salmonella and enteropathogenic E. coli. The results show a core epithelial response to pathogen exposure but also pathogen-specific responses. Studies are underway to perform transient early life infection (i.e. restrict the course of the infection to the immediate neonatal period) and analyse long-term consequences of this early life challenge. Here, we focus on immune priming and its influence on the long-term susceptibility to immune-mediated diseases such as asthma or atopic enteritis. Finally, we have established methods to also address the situation in humans. Here, ethical concerns limit access to cells and tissue for further analysis in healthy human babies and infants. Therefore, we have established and validated a method to obtain total RNA from human fecal material that contains epithelial cells that have been exfoliated as part of their normal life cycle. The results are promising and indicate that this approach may allow the analysis of epithelial gene expression in healthy and diseased human infant.
By comparing the intestinal epithelium of neonate and adult mice both under physiological and challenged conditions, we have made a number of unexpected observations that discovered differences in the host-microbe interaction and antimicrobial host response between neonates and adult mice illustrating the particular situation of the neonate host. For example, we noted enhanced macromolecule translocation by neonatal intestinal epithelial cells consistent with the concept of gut closure. Strikingly, this increase in translocation was not due to immaturity but rather functional differences associated with the metabolic requirements of the neonatal host (Schwentker et al., submitted manuscript). Also, we observed delayed maturation of M cells in the dome of Peyer’s patches after birth and this delayed maturation was shown to set the pace of adaptive immune maturation. Strikingly, M cell maturation was influenced by exposure to microbial stimuli for example leading to an earlier M cell appearance and adaptive immune maturation following infection with Salmonella (Torow et al., Immunity, 2023). Moreover, we identified the uptake of infected enterocytes by neighboring epithelial cells in the infected neonate host (Zhang et al., J. Exp. Med., 2024). This was in striking contrast to adult animals that remove infected epithelial cells by exfoliation into the gut lumen. Finally, we noted that early life infection drives the postnatal cell differentiation program leading to accelerated maturation of different cell types such as Paneth cells and absorptive enterocytes (Schlößer et al., submitted manuscript). Until the end of the project period, we expect a detailed cell atlas of postnatal cell type maturation, heterogeneity, and function, a characterization of early life infection on immune priming and susceptibility to immune-mediated diseases as well as further striking examples of differences in the host-microbial interaction of the neonatal host.