Cord Blood (CB) Progenitor Toll- Like Receptor (TLR) Expression: An Alternate Innate Immune Pathway In The Development Of Atopy

RATIONALE: Atopy in early life may be modulated by the expression and stimulation of TLRs on immunocompetent cells. Infants at high risk for developing atopy demonstrate phenotypic alterations of their CB progenitor cell hemopoietic cytokine receptors (HCR). Since hemopoietic mechanisms are involved in atopic development and maintenance, we investigated the co-expression of TLR and HCR on CB progenitors. METHODS: Fresh CB, enriched for CD34+ cells through magnetic cell separation techniques, were stimulated with 10 μL lipopolysaccharide (LPS) overnight and stained for surface and intracellular expression of TLR-2, TLR-4, TLR-9, IL-5R, IL-3R and GM-CSFR. Median fluorescence intensity (MFI) and mean percent expression were calculated. RESULTS: Prior to stimulation, mean expression and MFI were: TLR-2 (5.1 ± 5%, 1.8+/-0), TLR-4 (1.8 ± 0.1%, 1.6+/-0.2), TLR-9 (86 ± 2%, 3.8+/-0.5), IL-5R (18.2 ± 10%, 3.2+/-1.2), IL-3R (4.1 ± 0.08%, 1.2+/-0.1), GM-CSFR (17 ± 13%, 2.4+/-0.1). After stimulation, mean expression of TLR-2 (1.6 ± 0.7%) decreased (p = 0.0009, n = 4); TLR-9 (94.9 ± 0.8%) increased (p = 0.0001, n = 4), while mean expression of IL-5R (10.1 ± 4.3%), IL-3R (0.6 ± 0.4%) and GM-CSFR (8.9 ± 3.7%) decreased (p = 0.01, n = 4). CONCLUSIONS: We have demonstrated that CB progenitor cells have significant TLR expression and TLR stimulation directly affects both TLR and HCR expression. These alterations may have functional consequences for CB progenitor cell differentiation and maturation of pro-inflammatory cells previously shown to be important in allergic inflammation (e.g. eosinophils). This may indicate an alternate innate immune pathway of microbial influence on development of atopy in early life.