Fig 2: Lkb1 discriminates regulatory from inflammatory transcriptional programmes. a GSEA of transcriptional profiles of DCs from mice treated with LPS compared with that of Cd11cCreLkb1f/f mice. The upper part shows the list of gene sets related to inflammation; the lower part shows a representative plot of the gene set; –, no enrichment. b Scatter plot of Log2 counts of the genes expressed in splenic DCs from Lkb1f/f mice treated with LPS (n = 1) compared with those from Cd11cCreLkb1f/f mice (n = 3). Grey dots mark genes with less than two-fold change in any group. Red and blue dots mark the positively correlated genes with more than two-fold change from the two groups, while negatively correlated genes are indicated in green. The names of pro-inflammatory genes only upregulated in LPS-treated DCs are indicated in blue characters, and Ox40l, upregulated in both groups, is indicated in red characters. c mRNA levels of Cxcl11, Il1a, Il27, Cxcl9, Il15, Cd80, Il1f9, Cxcl10 and Ccl5 in splenic DCs from Lkb1f/f mice, Lkb1f/f mice treated with LPS, and Cd11cCreLkb1f/f mice. d mRNA level of Ox40l in DCs sorted from mice treated with or without LPS. e, f Flow cytometric analysis (e) and quantification of relative MFI (f) of OX40L protein expression on splenic DCs from mice treated with or without LPS (1.5 mg kg−1). The results are presented as the mean ± S.E.M., *P < 0.05, **P < 0.01, ***P < 0.001, by Student's t-test (c, d, f). Data are pooled from c, d, f or are representative of e at least two independent experiments with similar results

Fig 3: MC-mediated immune stimulatory properties of fibroblasts and keratinocytes.(A) NIH3T3 or (B) MPEK stable cells lines expressing either GFP alone (neg. control), OVA or MC.OVA were incubated for 60 hours. MC.OVA +rim had 10 nM rim present in the culture. After 60 hours, cells were harvested and MHC-I expression was analyzed by flow cytometry. (C-E) MPEK stable cells surface expression of CD80, CD86, and CD40. MFI values of live cells are reported relative to the mean MFI of negative control cells, transduced with an empty-vector, in each respective sample. (F) CD3+CD8+ OT-1 or WT T cells were purified by MACs and labeled with CellTrace violet (CTV) dye. Then 5 x 104 CD3+CD8+ CTV dye-labeled OT-1 or WT T cells were incubated with 5 x 104 B6-MPEK cells for 60 hours. By flow cytometry, live CTV+ cells were analyzed for CTV dye dilution as a measure of cell proliferation. (G) Representative histograms of live CTV+ cells. Percentages represent the percent of cells proliferating. n = 3, *p<0.05, One-way ANOVA, Tukey correction for multiple comparisons.

Fig 4: Tr1 cells suppress immune responses via IL-10 signaling.(A) Representative histogram of surface MHC-II, CD40, CD80, and CD86 on BMDCs in each group (n = 4 for each group). (B) Quantitative data of the MFI of surface MHC-II, CD40, CD80, and CD86 on BMDCs in each group (n = 4 for each group). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001. (C) Representative histogram of CFSE dilution of CD8+ T cells in each group (n = 4 for each group). (D) Quantitative data of the proportion of CFSEdim proliferating CD8+ T cells in each group (n = 4 for each group). Values are shown as the mean ± SEM; ***P < 0.001 and ****P < 0.0001.

Fig 5: PQ/PB-Gel for triggering antitumor innate and adaptive immune response after B16F10 melanoma surgery. (A and B) Representative flow cytometric images (left) and relative quantitative images (right) of M2-like TAMs (CD206hi) and M1-like TAMs (CD80hi) gating on F4/80+CD11b+CD45+ cells. Data are presented as means ± SD (n = 5). (C) Representative flow cytometric images (left) and relative quantification images (right) of CD80+CD86hi DCs gating on CD11c+ cells. Data are presented as means ± SD (n = 5). (D) Relative quantitative images of CD11c+ cells. Data are presented as means ± SD (n = 5). (E) Relative quantification of CD3+ cells within the tumor. Data are presented as means ± SD (n = 5). (F and G) The quantification results of CD4+, CD8+ cells within the tumor in different groups. Data are presented as means ± SD (n = 5). (H) Representative flow cytometric analysis of T cells infiltration within the tumor gating on CD3+ CD45+ cells. (I) The ratio of CD8 to CD4 cells within the tumor in different groups. Data are presented as means ± SD (n = 5). *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001 by one-way analysis of variance.