Fig 1: Anti-IGFBP7 treatment rescues IFN-γ–induced endothelial glycocalyx destruction and T cell adhesion.(A) GSEA revealing the signaling pathways enriched in IGFBP7hi psoriatic ECs compared with IGFBP7lo psoriatic ECs. (B) Relative mRNA expression assessed by quantitative real-time PCR of IGFBP7 in HMEC-1 cells (n = 3/group) treated with PBS or IFN-γ. (C and D) Representative staining and MFI quantification of IGFBP7 in HMEC-1 cells (n = 6/group) treated with PBS or IFN-γ. (E) HMEC-1 cells stimulated with different conditioned media (CM) were treated with anti-IGFBP7 or control IgG. (F and G) Representative staining and MFI quantification of HA and HS (n = 6/group). (H and I) 3D reconstruction and volume quantification of HA and HS (n = 6/group). (J) HMEC-1 cells stimulated with different conditioned media were first treated with anti-IGFBP7 or control IgG and then cocultured with CD4+ T cells. (K and L) Representative images and quantification of adherent CD4+ T cells in the coculture assay (n = 6/group). Data are represented as mean ± SD. Data in B and D were analyzed using unpaired Student’s t test. Data in G, I, and L were analyzed using 1-way ANOVA with Tukey’s post hoc test. Labeled P values in G and I represent the differences between the corresponding group and the control group.
Fig 2: IGFBP7 induces endothelial glycocalyx degradation and promotes T cell adhesion.(A and B) Representative staining and MFIs of HA and HS in HMEC-1 cells (n = 6/group) treated with PBS, endothelial glycocalyx degradation, or rhIGFBP7. (C and D) 3D reconstruction and quantification of HA and HS volume (n = 6/group). (E and F) Representative images and quantification of adherent CD4+ T cells after coculturing with HMEC-1 cells, which were pretreated with PBS, endothelial glycocalyx degradation, or rhIGFBP7 (n = 6/group). (G) Snapshots of CD4+ T cells flowing on HMEC-1 cells at different time points (n = 6/group). (H) The rolling velocity of CD4+ T cells in G (n = 20 T cells/group). (I) The number of adherent CD4+ T cells (n = 9 views/group) at the end of the flow assay in G. Data are represented as mean ± SD. All data were analyzed using 1-way ANOVA with Tukey’s post hoc test. Labeled P values in B and D represent the differences between the corresponding group and the PBS group.
Fig 3: Elevated IGFBP7 leads to endothelial dysfunction and skin inflammation in IMQ mice.(A) TEM of skin endothelial glycocalyx in control mice (n = 6 mice/group) that received rmIGFBP7 or vehicle i.v. The endothelial glycocalyx is highlighted by the yellow dotted lines. (B and C) Endothelial glycocalyx thickness and coverage were quantified (n = 30 vessels of 6 mice/group). (D) Dermatoscopy of IMQ mice (n = 6 mice/group) with or without rmIGFBP7 injection. Yellow arrowheads indicate dilated skin capillaries; white arrowheads indicate psoriatic scales. (E) Quantification of dilated capillaries in D (n = 30 views of 6 mice/group). (F) Skin histology of IMQ mice (n = 6 mice/group) treated with or without rmIGFBP7. Green arrowheads indicate erythrocyte exocytosis. (G) Quantification of erythrocyte exocytosis in F (n = 30 views of 6 mice/group). (H) Immunofluorescence staining for Ki67 (green) in dorsal skin from IMQ mice (n = 6 mice/group) with or without rmIGFBP7 injection. Dashed white lines mark the interface between the epidermis and dermis. (I and J) Epidermis thickness and the percentage of Ki67+ cells in the basal layer in H were quantified (n = 30 views of 6 mice/group). (K) Inflammatory gene expression in skin tissues from IMQ mice treated with or without rmIGFBP7 (n = 6 mice/group). (L) Whole-mount immunofluorescence staining for skin blood vessels (red) and CD3 (cyan) in IMQ mice (n = 6 mice/group) treated with or without rmIGFBP7. (M and N) 3D surface rendering of blood vessels and CD3 based on whole-mount immunofluorescence staining. (O–R) Quantifications of blood vessel diameter, infiltrated T cells, extravascular T cells, and intravascular T cells (n = 30 views of 6 mice/group). Data are represented as mean ± SD. Significance was calculated using unpaired Student’s t test. *P < 0.05; **P < 0.01.
Fig 4: IGFBP7hi ECs are increased in psoriatic skin.(A) Pseudotime trajectory of healthy and psoriatic ECs. (B) Volcano plot of DEGs between the psoriatic ECs (circled by dashed line) and the rest of the psoriatic ECs (see A right panel). Genes with log (fold change) > 0 are upregulated in psoriatic ECs (circled by dashed line). (C) IGFBP7 expression in psoriatic ECs. (D) GO analysis showing the pathways enriched in psoriatic IGFBP7hi ECs compared with psoriatic IGFBP7lo ECs. The x axis represents the rich factor. Bar colors indicate adjusted P value. (E) Regulon activity of IGFBP7hi and IGFBP7lo ECs. (F) Pseudotime trajectory of each cluster from psoriatic ECs. (G) Cluster composition of psoriatic IGFBP7hi ECs.
Fig 5: Anti-IGFBP7 treatment restores the endothelial glycocalyx and alleviates skin inflammation in IMQ mice.(A) Dermatoscopy of dorsal skin appearances in IMQ mice (n = 6 mice/group) with or without anti-IGFBP7 treatment. Yellow arrowheads indicate dilated skin capillaries; white arrowheads indicate psoriatic scales. (B) Quantification of dilated skin capillaries in A (n = 30 views of 6 mice/group). (C) Skin histology in IMQ mice (n = 6 mice/group) treated with or without anti-IGFBP7. Green arrowheads indicate erythrocyte exocytosis. (D and E) Quantification of epidermal thickness and erythrocyte exocytosis in C (n = 30 views of 6 mice/group). (F) Immunofluorescence for Ki67 (green) and Hoechst (blue) in dorsal skin tissue from IMQ mice (n = 6 mice/group) treated with or without anti-IGFBP7. The white dashed lines mark the interface between the epidermis and dermis. (G) Percentages of Ki67+ cells in the basal layer in F were quantified (n = 30 views of 6 mice/group). (H) mRNA levels of inflammatory genes in skin tissues (n = 6 mice/group). (I) TEM of skin endothelial glycocalyx in IMQ mice that received anti-IGFBP7 or control IgG (n = 6 mice/group). The vessel structure is highlighted by the yellow dotted line. (J and K) Endothelial glycocalyx thickness and coverage were quantified (n = 30 vessels of 6 mice/group). (L) Whole-mount immunofluorescence for skin blood vessels (red) and CD3 (cyan) (n = 6 mice/group). (M and N) 3D surface rendering of blood vessels and CD3 (cyan) based on whole-mount immunofluorescence. (O and P) Blood vessel diameter and infiltrated T cells in L were examined (n = 30 views/group). (Q and R) Quantification of extravascular and intravascular T cells in N (n = 30 views/group). Data are represented as mean ± SD. Significance was calculated using unpaired Student’s t test. *P < 0.05, ***P < 0.001, ****P < 0.0001).
Supplier Page from Sino Biological, Inc. for Human IGFBP7 / IBP-7 Protein (His Tag)