Fig 2: PS3D1@DMXAA elicits TAA-specific CD8+ T cell priming.(A) Schematic illustrating the process of DC activation in the TME and DC migration to the tdLN for antigen presentation and activation of T cells. Mice bearing B16.F10 tumors were treated with PBS (1), free DMXAA (2), PS3D1 (3), a physical mixture of empty PS3D1 and DMXAA (4), or PS3D1@DMXAA (5) intravenously for five times, 2 days apart. (B) Percentages of CD103+CD11c+ DCs of CD45+ cells in tumors were measured by flow cytometry on day 14 (n = 4). (C and D) Representative flow cytometric analysis (C) and the corresponding quantification (D) of CD103+CD11c+ DCs of CD45+ cells in tdLN on day 14 (n = 4). (E) Intravenous treatment scheme for mice with established B16-OVA tumor. Mice bearing B16-OVA tumors were treated as in (A). Then, the mice were sacrificed on day 7 for different analysis. (F and G) Percentages of OVA (SIINFEKL)–specific CD8+ T cells in TME (F) and spleen (G) were measured by flow cytometry (n = 3). (H) ELISpot analysis of IFN-? spot-forming cells among splenocytes after ex vivo restimulation with OVA (SIINFEKL) peptide on day 7. (I) Flow cytometric quantification of granzyme B+CD8+ T cells among splenocytes of mice after ex vivo restimulation with SIINFEKL (OVA) peptide. (J) Granzyme B protein levels in the supernatant of splenocytes after ex vivo restimulation with OVA (SIINFEKL) peptide for 24 hours on day 7 (n = 4). (K) Flow cytometric quantification of CD107a+CD8+ T cells among splenocytes after ex vivo restimulation with SIINFEKL (OVA) peptide on day 7. Data are means ± SD, and statistical significance was calculated by one-way ANOVA with Tukey’s test. ***P < 0.001 and **P < 0.01.

Fig 3: Gemcitabine-cisplatin chemotherapy promotes an overwhelmingly high release of inhibitory DAMP to attenuate DC maturation.a Western blot analysis of COX-2 expression in G69 cell lysates treated with either vehicle or GC for 48 h. GAPDH expression was used as loading control (n = 3 biologically independent experiments). b ELISA for PGE2 release in the culture medium of G69 cells treated with either vehicle control or GC for 48 h (n = 4 independent biological samples). Data are presented as mean values ± SEM. (Two-tailed, unpaired t-test **p = 0.0016). c COX-2 expression in G69-tumor tissue lysates treated with either vehicle or GC for two chemotherapy cycles. GAPDH expression was used as a loading control (n = 3 biologically independent experiments). d ELISA for PGE2 release in the serum of G69-tumor-bearing mice treated with either vehicle or GC for 2 cycles (n = 5 and n = 4 biologically independent samples for vehicle and GC, respectively). Data are presented as mean values ± SEM. (Two-tailed, unpaired t-test ****p < 0.0001). e Immunofluorescence analysis of COX-2 expression in G69-tumor tissues treated with either vehicle or GC for two chemotherapy cycles (Scale bar: 100 µm; Images representative of n = 3 independent experiments). f Schematic representation of BMDC treatment with conditioned media in the presence or absence of iDAMP blockade. g Flow cytometric analysis of CD40, CD86, and MHCI expression in BMDCs treated with G69-conditioned media with or without iDAMP blockade. h Geometric mean fluorescence intensity (gMFI) of CD40, CD86, and MHCI molecule expression in BMDCs upon treatment with condition media as in g (n = 2 biologically independent samples). Data are presented as mean values. i Schematic representation of footpad vaccination assay. j Flow cytometric analysis of CD40, CD86 and MHCI expression in vaccination-draining lymph node CD11c + MHC2 + DCs upon injection of G69 cells treated with GC in the presence or absence of iDAMP blockade (n = 5 biologically independent samples). Non-vaccination-draining lymph node DCs were used as control (n = 3 biologically independent samples). Data are presented as mean values ± SEM. (One-way ANOVA-Tukey’s multiple comparisons test. CD40: GC vs GC+ Celex, p = 0.045; GC vs GC+ aPGE2, p = 0.036; CD86: GC vs GC+ Celex, p = 0.046; GC vs GC+ aPGE2, p = 0.0007; H2-K: GC vs GC+ Celex, p = 0.0004; GC vs GC+ aPGE2, p = 0.005). Source data are provided as a Source data file. ns not significant.

Fig 4: Cell-type-specific role of caspase-11 in antibacterial host defense(A) Survival of Casp11fl/fl (n = 8), Casp11-/- (n = 8), LyzM-cre+Casp11fl/fl (n = 8), MRP8-cre+Casp11fl/fl (n = 8), Cd11c-cre+Casp11fl/fl (n = 8), and Villin-cre+ Casp11fl/fl mice (n = 8) infected i.p. with B. thailandensis (103 CFUs).(B and C) Bacterial loads in the spleen (B) and liver (C) of Casp11fl/fl (n = 8), Casp11-/- (n = 8), LyzM-cre+Casp11fl/fl (n = 8), MRP8-cre+Casp11fl/fl (n = 8), and Cd11c-cre+Casp11fl/fl mice (n = 8) infected i.p. with B. thailandensis (103 CFUs) on 3 days post-infection. Data from two independent experiments are shown. Each circle represents a mouse, and the horizontal lines represent the mean.(D) Summary of cell-type-specific roles of cytosolic LPS sensing in LPS shock and antibacterial host defense against B. thailandensis. Plus sign (+) indicates the magnitude of contribution (+++ and + indicate strong and weak contributions, respectively); -, no role; NA, not assessed. ***p = 0.0002 (adjusted p value; Mantel-Cox test).

Fig 5: Mono-therapeutic interventions with tacrolimus suppressed aberrantly activated ovarian F4/80+ CD11c+ macrophages in treated mice. Detection of M1 activated F4/80+ CD11c+ macrophages in the tacrolimus- and metformin-treated mice and their untreated and control groups at postcoital/gd 4.5 by confocal microscope (A) and by flow-cytometry (B). Red fluorescence in the upper two horizontal panels in (A) indicates the presence of CD11c+ and the green fluorescence in the third horizontal panel is the surface staining for F4/80+ macrophages, respectively. The merged fluorescence images in the lower horizontal panel in (A) (orange) further identify the classically activated F4/80+ CD11c+ macrophages. As opposed to the NFD-NONcNZO and those receiving tacrolimus or metformin, infiltration of the vascular granulosa cells layer with CD11c+ macrophages was pathognomonic feature of uncoordinated follicular growth among untreated HFD-dNONcNZO mice. Figures in the second horizontal panel in A are high magnifications of the corresponding framed boxes shown in the first horizontal panel. Bars in A = 100 µm. The peri-conceptional differential expression profiles of the activated ovarian M1 (CD11c+ CD206+) macrophages during postcoital days 2.5, 4.5 and 6.5 in all experimental groups are represented in bar graphs in (B). Nuclei (blue fluorescence) were counterstained with DAPI.