Fig 1: Ablating KCs prevents the hepatic response to atherogenic dyslipidemia.a, Volcano plot of genes downregulated in female D374Y mice treated with clodronate liposome during 10-d post-tamoxifen administration while maintained on a chow diet (n = 3 versus n = 3; P value from DESeq2 two-sided Wald test). b,c, Effects of clodronate liposomes on the expression of core identity genes of KCs (b), LAMs, monocytes and CV and capsular macrophages (c) in the liver of D374Y mice. d, Effect of clodronate liposomes on the day 10 conserved gene expression in both APOE cKO and D374Y mice. e, IL18BP plasma levels in dyslipidemic APOE cKO and D374Y mice given clodronate liposomes or Dil liposomes controls (ng ml−1, APOE cKO n = 3 and D374Y n = 4). f, Volcano plot and heat map indicating minimal response of the liver to dyslipidemia when comparing littermate control versus D374Y with both treated with clodronate liposomes, as determined by mRNA-seq (n = 3 versus n = 3; P value from DESeq2 two-sided Wald test). g, Total plasma (mg dl−1) and liver (µg mg−1 of protein) cholesterol and triglyceride measurements in dyslipidemic APOE cKO and D374Y mice given clodronate liposomes or Dil liposomes controls with respective littermate controls also administered clodronate liposomes (APOE cKO n = 4 versus n = 3 versus n = 4 and D374Y n = 3 versus n = 4 versus n = 3). h, Plasma CD5l (µg ml−1) concentrations as determined by ELISA in D374Y and littermate control mice 10 d after tamoxifen administration (n = 9 versus n = 9). i, Plasma CD5l (µg ml−1) concentrations in dyslipidemic D374Y mice and littermate control mice given clodronate liposomes and dyslipidemic D374Y mice administered Dil liposomes controls (n = 3 versus n = 4 versus n = 3). All experiments were conducted on day 10 after tamoxifen treatment, and mice were maintained on a normal chow diet. All plots are ±s.e.m. Two-sided t-test (e,h) or one-way ANOVA (g,i).
Fig 2: Conserved and divergent responses to sustained atherogenic dyslipidemia.a, Plasma levels of cholesterol in dyslipidemic APOE cKO and D374Y mice with respective littermate controls, maintained on a high fat diet for 20 weeks. (APOE cKO n = 16 vs 11; and D374Y n = 7 vs 3). Plots are ± SEM, two-sided t-test b, Representative liver sections stained with ORO in APOE cKO and D374Y mice after 20 weeks on a high fat diet (scale bars = 100 μm, n = 2 vs 2 for D374Y and n = 3 vs 3 for APOE cKO, where n indicates a different mouse). c, Representative liver sections stained with H&E in APOE cKO and D374Y mice after 20 weeks on a high fat diet (scale bars = 100 μm, n = 2 vs 2 in each group where n indicates a different mouse).d, Unique and common upregulated genes in both strains at 8, 12, and 20 weeks on a high fat diet. Unique genes for each strain are indicated at the bottom. e, Flow cytometry of CLEC4F levels on Kupffer cells from female D374Y versus littermate control after 8 weeks of high fat diet. f, Heatmap of the 72 genes upregulated at all time points in both strains, expressed in 76 human single cell types. g, Flow cytometry plots showing percentages of blood and liver monocytes, and liver Kupffer cells in dyslipidemic D374Y mice given clodronate liposome or Dil liposomes control for 8 weeks while on a high fat diet. h, Immunohistochemistry of liver to detect CD5L production in control or clodronate treated D374Y mice after 8 weeks of high fat diet (scale bars = 100 μm). i, The 14 genes not affected by clodronate-induced Kupffer cells depletion, expressed in liver myeloid cell clusters. j, Correlation between plasma IL18BP and liver PCSK9 expression in humans.
Fig 3: PCA analysis of serum cytokines and CBC in C57BL6 mice with PBI/BM5 and/or IL-18BP treatment. (A) PC score plot of non-irradiated mice with d2, d3, and d7 irradiated vehicle/IL-18BP treatment mice. Each dot represents one animal. (B) Loading score plot of (A). n = 4, 6, 4, 6, 6 and 4 for the 0 Gy, d2 vehicle, d2 IL-18BP, d3 vehicle, d3 IL-18BP, and d7 vehicle groups, respectively. The d7 IL-18BP group had only one survivor, and therefore was not included for analysis here.
Fig 4: CBC changes in the C57BL6 mice after PBI/BM5. The mice were exposed to 14.48 Gy PBI/BM5, and then treated with a vehicle or 2.0 mg/kg IL-18BP at d1 after radiation. The serum samples were collected on d2, d3, and d7 after radiation. The blood samples from the IL-18 wild-type and knockout mice were included here too (d14 blood samples after 14.73 Gy PBI/BM5). The CBC differences between the different groups were analyzed using one-way ANOVA, and a Tukey post-test. The post-test showed no difference between the vehicle or IL-18BP treatment on the same day, or wild-type d14 vs. IL-18KO d14. Therefore, the differences shown here were all compared to the 0 Gy group. n = 4, 6, 4, 6, 6, 4, 1, 2, and 6 mice in the 0 Gy, Veh d2, BP d2, Veh d3, BP d3, Veh d7, BP d7, WT d14, and IL-18KO d14 groups, respectively. V, BP, WT, and KO stand for vehicle group, IL-18BP treatment group, wild-type, and IL-18 knockout. The time of the tissue collection and radiation doses are shown at the bottom. Each dot represents one animal. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001.
Fig 5: Serum IL-18 and IL-18BP levels in C57BL6 mice after PBI/BM5. The mice were exposed to 14.48 Gy PBI/BM5, and then treated with a vehicle or 2.0 mg/kg IL-18BP on d1 after radiation. Serum samples were collected on d2, d3, and d7 after radiation exposure. The blood samples from the IL-18 WT and KO mice were included here too (d14 blood samples after 14.73 Gy PBI/BM5). The IL-18 and IL-18BP differences between the different groups were analyzed using one-way ANOVA, and a Tukey post-test. The post-test showed no significant difference between the vehicle or IL-18BP treatment on the same day, or wild-type d14 vs. IL-18KO d14. Therefore, the differences shown here were all compared to the 0 Gy group. n = 4, 6, 4, 6, 6, 4, 1, 2, and 6 mice in the 0 Gy, Veh d2, BP d2, Veh d3, BP d3, Veh d7, BP d7, WT d14, and IL-18KO d14 groups, respectively. The BP d7 sample was lost in the IL-18BP assay. V, BP, WT, and KO stand for vehicle group, IL-18BP treatment group, wild-type, and IL-18 knockout. The time of the tissue collection and radiation doses are shown at the bottom. Each dot represents one animal. *, p < 0.05.
Supplier Page from Abcam for Mouse IL-18BP ELISA Kit