Adoptive transfer of LPS-treated BMDCs influences the number and function of pulmonary memory CD4+ T cells in OVA-induced asthmatic mice
It is widely believed that Th1/Th2 ratio imbalances may lead to the pathogenesis of allergic asthma, in which memory T cells play a crucial role in the capacity of initiating Th2 response under repeated allergen challenges21. Memory T cells can be divided into two distinct subsets, central memory T cells (TCM cells) and effector memory T cells (TEM cells), according to their homing characteristics and effector functions22, 23. In order to assess whether DClps could suppress the Th2 immune recall response in airways of treated mice, we sought to examine the number and function of memory CD4+ T cells in lung tissues. We thus established asthma model mice and treated these mice with diverse DCs, as described above; however, instead of sacrificing the mice on day 19, we rechallenged the animals with OVA from days 35-39 (Fig. 4A).
On day 40, mice were sacrificed and lungs were removed to obtain single-cell suspensions for the isolation of memory CD4+ T cells. We confirmed that both DC10 and DClps observably decreased the pulmonary CD4+CD44high memory T cells, whereas the difference in DCia was negligible compared to the OVA group (Fig. 4B). On the other hand, we next identified CD4+TCM cells that were CD62L+CCR7+ double positive and CD4+ TEM cells that were CD62L-CCR7- double negative by flow cytometry (Fig. 4B). Surprisingly, we saw no discernible difference in the proportion of TCM and TEM cells among mice treated with DClps, DC10 or DCia. To gain insight into this phenomenon, we assessed whether the adoptive transfer of DClps affected the proliferation of pulmonary memory CD4+ T cells with an MLR assay. After MLR, the proliferation activity of pulmonary memory CD4+ T cells was determined by CFSE. Consistent with the above results, it was readily apparent the proliferation activity of pulmonary memory CD4+ T cells isolated from mice in the DC10 and DClps groups was obviously suppressed, compared with the DCia and OVA groups, especially on day 3 and day 5 (Fig. 5). Meanwhile, we collected the cell supernatant of MLR to assess IL-4 and IFN-γ secretion by memory CD4+ T cells. We found that compared with cells in the OVA group, both TEM and TCM cells exhibited a notable decline in the secretion of IL-4 in the DClps and DC10 groups (Fig. 6A), whereas no treatment effect was detectable in the DCia group (Fig. 6B). Interestingly, in the DClps and DC10 groups, the level of IFN-γ dramatically reduced in only TEM cells but not in TCM cells (Fig. 6B). Taken together, these data indicate that the adoptive transfer of DClps may inhibit the allergen recall response by suppressing memory CD4+ T cells. Moreover, the reduction in memory CD4+ T cells may well reduce the conversion of Th2 cells, which further results in the downregulation of Th2 cytokines and improves the Th1/Th2 imbalance in the immune response.