[27] Stimulation by TLR has been shown to involve the activation of MAPK signalling pathways in human monocytes,[9, 28] macrophages,[29] eosinophils[30] and CB progenitor cells.[21] In relation to progenitor cells, we have previously shown that IL-5-stimulated or GM-CSF-stimulated peripheral blood progenitor cells undergo rapid phosphorylation of p38 MAPK within 1–5 min using phospho-ELISA.[17] Although not in a kinetic study, Kim et al.[21] also

showed that in CB progenitors stimulated with TLR-9 agonists there is up-regulation of both p38 MAPK and ERK 1/2. Our findings therefore complement and extend the latter study, showing that significant phosphorylation of p38 MAPK is also detected in CB CD34+ APO866 cells stimulated with other TLR (LPS) agonists (Fig. 7). While others have reported that BM-derived CD34+ cells respond to TLR stimulation with the production of cytokines including GM-CSF,[6-8] the potential mechanism(s) of this secretion were not investigated. Our demonstration Veliparib mw that blocking p38 MAPK signalling

in CB CD34+ cells suppresses LPS-induced GM-CSF secretion is therefore novel. Related to this, Kim et al.[21] have demonstrated that TLR9 stimulation of CB CD34+ cells activates the p38 MAPK and ERK 1/2 pathways involved in IL-8 secretion. Our data show for the first time that LPS-induced GM-CSF production, which facilitates Eo/B CFU, directly involves TLR4/p38 MAPK signal transduction in CB CD34+ cells. In this way, LPS is only one component of this autocrine effect, a co-factor in Eo/B CFU formation, which uses the production of GM-CSF

through MAPK signalling pathways to induce Eo/B differentiation from CB CD34+ cells. This is in line with studies that have shown that p38 MAPK is an integral part of the TLR4 axis of signal transduction.[31] We have previously shown that CB progenitor cells from high-atopic risk infants have reduced capacity for Eo/B CFU formation after LPS stimulation.[12] It has recently been shown that children of atopic mothers have reduced TLR-dependent p38 MAPK signalling in their blood monocytes up to the age of 2 years.[32, 33] In light Orotic acid of our current results, we hypothesize that reduced CB Eo/B differentiation after LPS stimulation in high-atopic risk infants[12] may be the result of reduced p38 MAPK-induced GM-CSF production by CD34+ cells, possibly related to epigenetic effects on p38 MAPK expression in utero. Along these lines, prenatal exposure to bacterial microflora (Acinetobacter lowffii F78) has been shown to prevent the development of allergy in offspring[34] through microbial-induced epigenetic regulation of the IFN-γ promoter.[35] Although the assessment of atopy was not the objective of this study because we were interested solely in the biological implications of LPS stimulation on human CB CD34+ cells, we are now in position to examine this hypothesis in prospective birth cohorts.