Extended Data Fig. 4: Characterisation of mCherry-APOB responsive cells.
From: Kupffer cells dictate hepatic responses to the atherogenic dyslipidemic insult
a, Schematic of targeting strategy of the Apob locus to produce mCherry-APOB. b, Confocal analysis of liver from D374Y and D374Y-APOB strains 10 days after tamoxifen treatment and maintained on a chow diet. Scale bars are 100μM, n = 2 D374Y-APOB and n = 4 D374Y, where n represet a different liver sections. c, Western blot of plasma from D374Y-APOB mice probed with anti-mCherry antibody. d, Plasma total cholesterol measurements from female mCherry-APOB and D374Y- mCherry-APOB after 4 weeks of high-fat diet following tamoxifen-induction (n = 4 D374Y and n = 3 littermate control mice). Plots are ± SEM. statistical analysis was performed with two-sided t-test. e, Heatmap of the top 5 marker genes for every cluster using Seurat analysis of scRNA-seq data from Live CD45+mCherry-APOB+ cells isolated from digested liver of female D374Y 10 days after tamoxifen administration and maintained on a chow diet. Yellow indicates high expression of a particular gene, and purple indicates low expression. f, Expression of receptors that are capable of APOB lipoproteins uptake in the cell types found in a mouse liver. g, Quantification of liver Kupffer cells (defined Live CD45 + CD3-CD19-Ly6G-CD64 + F4/80 + TIM4 + ) in male D374Y and littermate controls 10 days after tamoxifen administration and maintained on a chow diet. n = 6 D374Y and n = 8 littermate control mice. Data are shown as mean ± SEM. h, Representative gating strategy applied to Kupffer cells analysis by flow cytometry. i, Histogram representing the different cell populations responsible for LDL uptake in the liver of a dyslipidemic APOE cKO mouse injected i.v. with a solution containing purified LDL-AF-488. j, Histogram and mean fluorescent intensity for mCherry in the liver of a APOE cKO mouse injected i.v. with D374Y-mCherry plasma (red line), compared to a APOE cKO mouse injected with non-labelled D374Y plasma (black line) (n = 7 individual samples obtained from the liver of the injected APOE cKO mouse and n = 3 individual samples obtained from the liver of the injected littermate control mouse). Data are presented as mean ± SEM. statistical analysis was performed with two-sided t-test. In all cases, mice were administered tamoxifen 10 days prior and maintained on a chow diet.
