Effect Of Neonatal BCG Vaccination On Splenic And Lung Dendritic Cell Development Of BALB/c Mice

Background and Objective Bronchial asthma is one of the most common chronic diseases in childhood."Hygiene hypothesis"suggests that the improved hygiene in industrialized countries and the use of vaccines and antibiotics cause the decline of many infectious diseases that would normally stimulate Th1 development in some way that mitigates against asthma. However, more and more results indicated that Th1/Th2 imbalance could not interpret all of asthma pathogensis. Our previous study suggested that neonatal BCG vaccination induced Th1/Tr1 development in mice spleen and attenuated airway inflammation and airway hyperreactivity (AHR) without Th1/Th2 cytokines change from bronchoalveolar lavage fluid (BALF) in mice which mechanism is not clarified. Recently the impact of dendritic cells (DCs) and regulatory T cells (Tr) on the pathogensis of asthma have been investigated over the past decades. As the professional antigen presenting cells, DCs not only prime immune response directing Th0 cells toward different T subtypes but also induce immune tolerance. To further identify the mechanism of neonatal BCG vaccination on T cell subsets polarization, we investigated the effect of neonatal BCG vaccination on splenic and lung DCs development and lung T cells of BALB/c mice.Methods Neonatal SPF BALB/c murine were divided into BCG-treated intraperitonally group and BCG-treated subcutaneously group and control group, simultaneously adult SPF BALB/c murine were set up as control group. Post BCG vaccination for four weeks, spleen and lung cells were isolated. With flow cytometry, phenotype of splenic and lung DC were analysed. Moreover, cells were stimulated by LPS or BCG in vitro for 18 or 48 hours and cytokines concentration was detected by ELISA. Furthermore, mRNA expression of T cell transcription factor of lung cells was assayed by RT-PCR.Results Part one: Effect of neonatal BCG vaccination on phenotype and function of splenic DC of BALB/c mice: (1) In comparison with the control group, the percent of CD8α+DCs was strikingly increased and that of CD8α-DCs was significantly declined in neonatal BCG-treated mice, furthermore, expression of costimulatory molecules on DC was upregulated and concentration of IL-12p70 induced by LPS and IL-10 induced by BCG from spleen cells culture supernatant was noticeably elevated in neonatal BCG-treated mice, but IL-6 did not. (2) In contrast, the percent of CD8α-DCs was markedly increased and that of CD8α+DCs was noticeably reduced in adult-vaccinated ones. Compared with the control group, expression of CD40 and MHC-Ⅱmolecules on CD8α- and CD8α+DC was significantly higher and that of CD86 was strikingly lower in adult-vaccinated ones. (3) The percent of CD8α-DC was significantly higher and that of CD8α+DC was strikingly lower in adult-vaccinated mice than that of neonatal-vaccinated ones. The expression of costimulatory molecules on DC had no difference between neonatal and adult BCG vaccination mice. (4) There were no difference in DC subtypes and expression of costimulatory molecules on DC between neonatal BCG-vaccinated intraperitonally mice and subcutaneously ones.Part two: Effect of neonatal BCG vaccination on lung DC and T cells of BALB/c mice: (1) CD8α-DCs was the main DCs in lung from BALB/c mice, with low expression of MHC-Ⅱmolecules; The percent of CD8α-DC and CD8α+DC and concentration of IL-12p70 and IL-6 from lung cells culture supernatant was the same between neonatal BCG-vaccinated mice and control ones. In comparison with control group, the expression of MHC-Ⅱmolecules on DC was upregulated in neonatal-vaccinated group. (2) The percent of CD4+CD25+T cells and expression of Foxp3,T-bet,GATA-3 mRNA of lung cells was the same between neonatal BCG-vaccinated mice and control ones. (3) The percent of CD8α-DC was significantly higher and that of CD8α+DC was strikingly lower from lung in adult-vaccinated mice than that of adult control ones. The expression of costimulatory molecules on DC was significantly upregulated in adult BCG-vaccinated mice. (4) There was no difference in DC phenotype and expression of Foxp3,T-bet,GATA-3 mRNA of T cells from lung between neonatal-vaccinated mice and adult-vaccinated ones. (5) The percent of DC subtypes and expression of Foxp3,T-bet,GATA-3 mRNA of T cells from lung was the same between neonatal-vaccinated intraperitoneally mice and neonatal-vaccinated subcutaneously ones. The expression of MHC-Ⅱmolecules on lung DC was significantly higher in neonatal-vaccinated intraperitoneally mice than that of neonatal-vaccinated subcutaneously ones.Conclusion (1) By up-regulating splenic CD8α+DCs and inducing IL-12p70 and IL-10 production in BALB/c mice, neonatal BCG vaccination promoted Th1/Tr1 development.(2) Neonatal BCG vaccination promoted lung DC maturation, but had no effect on lung T cell subsets. It is supposed that the effect of neonatal BCG vaccination to inhibit AHR and lung inflammation is not mediated by T cell subsets polarization.(3) The effect of BCG vaccination on DC in BALB/c mice is different between neonates and adult.