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Background: IgE‐mediated sensitisation to egg is common in infants. In some cases, the processes leading to egg sensitisation are established in early life, even before introduction to solid foods. The underlying mechanisms remain poorly understood. Methods: We performed detailed immune cell phenotyping of peripheral blood mononuclear cells and determined in vitro cytokine responses following allergen specific and non‐specific immune stimulation. To determine if unique immune profiles were linked to early‐life egg sensitisation, we compared 92 infants at 4–6 months of age, with (EggCAP+, n = 41) and without (EggCAP−, n = 51) early egg sensitisation. Additionally, 47 cord blood samples were analysed. For a subset of participants (n = 39), matching cord blood mononuclear cells were assessed by flow cytometry to establish the impact of IgE sensitisation on immune developmental trajectories. Results: EggCAP+ infants were found to exhibit a unique immune phenotype characterised by increased levels of circulating CD4+ T regulatory cells (Treg), CD4+ effector memory (EM) Treg and increased expression of the IgE receptor, FcεR1, on basophils. The increased CD4+ EM Treg profiles were already present in cord blood samples from EggCAP+ infants. A general Th2‐skewing of the immune system was observed based on increased IL‐13 production following phytohemagglutinin stimulation and by comparing immune developmental trajectories, EggCAP+ infants displayed an expansion of basophils and reduced levels of CD4− T cells compared to EggCAP− infants. Conclusions: Immunological profiles associated with egg sensitisation are detectable in infant circulation at 4–6 months of age and at birth. Understanding the immune mechanisms underlying early‐life sensitisation could provide important insights for future food allergy prevention strategies. [ABSTRACT FROM AUTHOR]

Background: Low vitamin D levels have been associated with allergic diseases. Vitamin D has potent immunomodulatory properties, but the mechanisms remain unclear. We have investigated the effect of oral vitamin D supplementation on circulating immune cell phenotypes in infants. Method: A double‐blinded randomised controlled trial was conducted to investigate the effect of oral vitamin D supplementation (400 IU/d) on eczema and immune development. A subset of 78 infants was included in this analysis. Phenotypic analysis of immune cell subsets was performed using flow cytometry. Results: Vitamin D supplementation resulted in median 25(OH)D levels of 80.5 vs 59.5 nmol/L in the placebo group at 3 months of age (P =.002) and 87.5 vs 77 nmol/L at 6 months of age (P =.08). We observed significant changes in immune cell composition from birth (cord blood) to 6 months of age. Vitamin D supplementation did not impact these changes, nor did immune cell composition correlate with plasma 25(OH)D levels. Through exploratory analysis, we identified possible associations with eczema development and increased abundance of naïve CD4‐ T cells at birth, as well as associations with basophils, iNKT and central memory CD4+ T cells, and altered expression patterns of IgE receptor (FcεR1) on monocytes and dendritic cells with eczema at 6 months. Conclusions: Vitamin D supplementation in infants who were vitamin D sufficient at birth did not affect developmental changes in immune cells during the first 6 months of life. However, immune cell profiles at birth and at 6 months of age were associated with early life eczema. [ABSTRACT FROM AUTHOR]

Dendritic cells (DCs) play a central role in regulating innate and adaptive immune responses. It is well accepted that their regulatory functions change over the life course. In order to study DCs function during early life it is important to characterize the function of neonatal DCs. However, the availability of neonatal DCs is limited due to ethical reasons or relative small samples of cord blood making it difficult to perform large-scale experiments. Our aim was to establish a robust protocol for the generation of neonatal DCs from cord blood derived CD34+ hematopoietic stem cells. For the expansion of DC precursor cells we used a cytokine cocktail containing Flt-3 L, SCF, TPO, IL-3 and IL-6. The presence of IL-3 and IL-6 in the first 2 weeks of expansion culture was essential for the proliferation of DC precursor cells expressing CD14. After 4 weeks in culture, CD14+ precursor cells were selected and functional DCs were generated in the presence of GM-CSF and IL-4. Neonatal DCs were then stimulated with Poly(I:C) and LPS to mimic viral or bacterial infections, respectively. Poly(I:C) induced a higher expression of the maturation markers CD80, CD86 and CD40 compared to LPS. In line with literature data using cord blood DCs, our Poly(I:C) matured neonatal DCs cells showed a higher release of IL-12p70 compared to LPS matured neonatal DCs. Additionally, we demonstrated a higher release of IFN-γ, TNF-α, IL-1β and IL-6, but lower release of IL-10 in Poly(I:C) matured compared to LPS matured neonatal DCs derived from cord blood CD34+ hematopoietic stem cells. In summary, we established a robust protocol for the generation of large numbers of functional neonatal DCs. In line with previous studies, we showed that neonatal DCs generated form CD34+ cord blood progenitors have a higher inflammatory potential when exposed to viral than bacterial related stimuli. • A robust protocol for the generation of high numbers of neonatal dendritic cells. • IL-3 and IL-6 are crucial for the proliferation of cord blood CD34+ progenitors. • Neonatal DCs have a higher inflammatory potential when exposed to viral stimuli. • LPS induces higher release of IL-10 in neonatal DCs compared to Poly(I:C). [ABSTRACT FROM AUTHOR]