Immune maturation and allergy development during childhood in relation to early-life microbial exposuresBabies are born with an immature immune system, which gradually matures during their first year(s) of life. This maturation is dependent on environmental exposures such as the establishment of a gut flora and exposure to various infections, but also on exposure to microbial compounds. These types of interactions between the surrounding environment and the neonate will provide the child with an immune system which knows how to react and also to what antigens it should respond. In contrast, microbial deprivation, especially during early years of life, could result in poor immune maturation and/or altered immune balance and later also to immune mediated diseases, like allergy. Our research aims to understand how early-life microbial exposures (bacterial and viral) influence immune maturation and allergy development in children.

Monocyte and natural killer cell function and maturation in children
Among innate cells, monocytes are the ones with the highest density of toll like receptors on their cell surface. Accordingly, they are efficient responders to several different classes of microbial stimuli. As they differentiate into dendritic cells they can greatly influence immune maturation and adaptive immunity. Monocytes are further divided into different subsets with different characteristics and behaviour, stressing the heterogeneity of this cell type. We have shown that up to the age of 2 years, maternal allergic heredity is associated with altered anti-microbial responses (Amoudruz et al 2005, Saghafian-Hedengren et al 2008). A 5 years of age however, monocyte function is primarily influenced by allergic disease and not allergic heredity (Amoudruz et al 2009).
We investigate innate immunity in early life, in relation to both maternal disease and allergic outcomes. To understand the role of maternal disease for neonatal innate immune function, the phenotype, function and regulation of both placental and peripheral monocytes/macrophages from healthy women and women with pregnancy-associated complications are investigated together with another important innate immune actor– the natural killer cell. We have previously shown that pregnancy-associated complications can influence maternal placental and peripheral natural killer cells and placental pro-inflammatory capacity (Bachmayer et al 2006 and 2009, Holmlund et al 2007).
To explore innate immune maturation in children, we study monocytes and their different subtypes at different ages. With flow cytometry-based techniques, we investigate phenotypes, the activity of intra-cellular signalling pathways as well as secreted substances, both at basal levels and following activation with different microbial compounds. But as immune responses to allergens are of an adaptive nature, we also explore T cell responses following microbial stimulation. For example we investigate transcription factors which correspond to different types of T cell immunity with real time RT-PcR and the production of cytokines by ELISA-based techniques. We will then be able to see how microbes influence adaptive responses and if this depends on allergy.

The commensal gut microbiota, breast feeding and allergy development
We have shown that children, who develop allergy during their first 5 years of life, have a reduced biodiversity and altered gut flora composition in very early fecal samples (Sjögren et al 2009). In addition we have demonstrated that the early life gut microbiota composition influences both mucosal and systemic immune responses during childhood (Sjögren et al 2009).
We investigate how these microbes interact with the immune system and how breast-milk components influence these interactions. Gut microbiota – gut epithelium - immune cell interactions are studied in vitro. The bacterial species are selected based on their rich prevalence in early life, different prevalence in faeces from allergic and non-allergic children and/or for their demonstrated immunomodulatory effects. We further investigate how these bacteria influence immune cells of different maturity (= age).
Most neonates are breast-fed, at least for the very first months in their life. The breast milk is very rich in (immunological) factors, which are likely to influence the gut colonization and immune maturation. In addition, the breast milk composition depends on genetics (Sjögren et al 2007) but also on maternal exposures, such as country of birth, previous pregnancies etc (Amoudruz et al 2009). Therefore, we also study how breast milk influences this microbe - epithelium - immune interaction in the experimental set-ups described above. Main methods are real-time PcR, cell culture, ELISA-based techniques and flow cytometry.

Early-life viral infections, immune maturation and allergy development
Infections with herpesviruses like Epstein Barr virus (EBV) is most common in early life and are then usually asymptomatic or associated with mild symptoms. Early control of viral infections depends on interferon-g release by natural killer cells, cells which generally require the presence of accessory cells, like monocytes. In mice, herpesvirus latency causes systemic innate activation and protection from bacterial co-infection. In humans however, the situation is different. We have showed that an early EBV seroconversion influences both adaptive and innate immunity in 2-year old children (Nilssson et al 2009, Saghafian-Hedengren et al 2009). Further, an early EBV infection is associated with a reduced risk of persistent IgE-sensitization (Saghafian-Hedengren et al 2009).
We continue to analyse how these viruses modulate the immune system at different ages and how the time-point for seroconversion influences immunity. Much is known regarding how EBV influences B cells upon infection, as they are the major target cells for the virus. Much less is known about how EBV influences the innate players of the immune system, like monocytes and natural killer cells. We perform experiments on material from children participating in a large prospective allergy study, allowing us to relate experimental data to allergy outcomes at 2, 5 and 10 years of age. Cell culture, and flow cytometry-based techniques are key methods in this project.