Study About The Whole Peptidoglycan Of Bifidobacterium Effect On Bone Marrow-Derived Dendritic Cells From Asthma Model Mice

【Objective】To explore in vitro the immunoregulation effect of the Whole Peptidoglycan of Bifidobacterium (WPG) and on bone marrow-derived dendritic cells (DC) from normal and asthmatic model mice by co-culturing with DC and observe if the immunoregulation effect is dependent on the change of time, revealing the mechanism of probiotics preventing and curing the allergic disease【Methods】16 Balb/c mice were randomly divided equally into normal control group and asthma group, bone marrow mononuclear cells were separated from normal and asthma mice, then induced to become immature DC with GM-CSF and IL-4 on day 7. WPG was added into the experimental group, Lipopolysaccharide (LPS) were added into the positive control group, culture medium was added into the negative control group. The changes of the shape of DC were observed continuously with phase-contrast microscope and electron microscope.24 hours later, the expression of CD11c,CD86 and MHC-Ⅱwas detected with flow cytometry, the capability of DC to stimulate the proliferation of allogeneic T lymphocytes was examined with MTT assay, the level of IL-12p70 and IL-10 in the culture supernatant was detected by enzyme linked immunosorbent assay (ELISA). To observe the level of IL-12 and IL-10 excreted by BMDC after interfering with WPG at different time points, the culture supernatant of BMDC from normal mice was collected at 0h,12h,24h,36h,48h,60h and 72h respectively after interfering with WPG, and the level of IL-12p70 and IL-10 in the culture supernatant was detected by ELISA.【Results】1. There was no significant difference between the negative control group of normal and asthma mice in the MLR test and the ability to secrete cytokine IL-12 and IL-10 (P>0.05), and the normal positive group was more competent than that of asthma positive group (P<0.05).For WPG group of normal mice, the marker CD86 and MHC-Ⅱwas the same with positive group on the whole, and it was more competent than the normal negative control group but less than the normal positive control group in the MLR test and the ability to secrete cytokine IL-12 and IL-10 (P<0.05).For asthma mice, the marker CD86 of DC in WPG group was lower than positive group, and it was more competent than the asthma negative control group and the asthma positive control group in the MLR test and the ability to secrete cytokine IL-12 and IL-10 ,but not so competent as that of WPG group and positive group of normal mice(P<0.05).3. After co-culturing WPG and BMDC, the level of IL-12 and IL-10 in the culture supernatant was increased gradually in 24 hours, and reached the peak at 24 hours then decreased 24 hours later.【Conclusions】1. There was mature and functional impairment in BMDC from asthma mice. WPG could regulate its function to some extent and promote Th1/Th2 balance but the dendritic cells couldn't recover to normal state, which suggested the immunoregulation effect of WPG was limited.2. The immunological regulation effect of WPG was dependent on the change of time, and it suggested that Probiotics regulated the function of BMDC only at a certain range of time.