Journal: bioRxiv

Article Title: Alirocumab attenuated plaque inflammation and PCSK9-induced proinflammatory signalling in M1 macrophages independently of lipid lowering

doi: 10.1101/2025.11.20.689631

Figure Lengend Snippet: By qPCR, levels of proinflammatory CXCL9 , CXCL10, TNF-α, IL-1β and IL-6 and anti-inflammatory CCL17, TGM2, IL-10 and TGF-β1 cytokines in M1 macrophages stimulated with PCSK9. TNF-α was used as a positive control. *p<0.05, **p<0.01, and ***p<0.001 vs. unstimulated cells, by One-sample t test, control value set to 1. N=4-5 independent biological replicates.

Article Snippet: The human taqman assay IDs were as follows: CXCL9 (Hs00171065_m1), CXCL10 (Hs00171042_m1), IL-1β (Hs01555410_m1), TNF-α (Hs00174128_m1), IL-6 (Hs0174131_m1), CCL17 (Hs00171074_m1), TGM2 (Hs01096681_m1), IL-10 (Hs00961622_m1), TGF-β1 (Hs00998133_m1), and TLR4 (Hs00152939_m1) serving 18S rRNA (Hs99999901_s1) as the endogenous control.

Techniques: Positive Control, Control

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: CCL17 was elevated in atherosclerotic model. The murine AS model was established by high-fat diet (HFD) to ApoE mice. A HE and Oil Red O analysis of aorta root tissues. B The levels of total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) in serum were measured by ELISA. C The ( C ) RNA levels of pro-inflammatory factors VCAM-1, MCP-1, TNF-α, IL-1β, and IL-6 was detected by qPCR assay. D Serum CCL17 level was detected by ELISA. E Immunofluorescence detection of co-localization of CCL17 and macrophages in plaque tissue of AS mice. F The level of CCL17 mRNA in mice plaque tissue was detected by qPCR. In vitro AS model was established by stimulating primary macrophage macrophages with ox-LDL. G The mRNA levels of inflammatory factors (IL-1β and IL-6) were detected by qPCR. H The level of CCL17 in cell supernatant was detected by ELISA. I The level of CCL17 in macrophages was detected by qPCR. * p < 0.05, *** p < 0.01, *** p < 0.001. N = 6

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Enzyme-linked Immunosorbent Assay, Immunofluorescence, In Vitro

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: Silencing CCL17 improves atherosclerosis in mice. After 4 weeks of AS modeling, sh-CCL17 encapsulated by AAV virus was injected into the tail vein of mice to knock down the expression of CCL17 in mice. A Western blot was used to detect the protein expression of CCL17 in mice plaque. B HE and oil red O staining were used to observe the pathological changes of arterial tissue in mice. C The levels of TC, TG and LDL-C in AS mice were measured by ELISA. D qPCR was used to detect the mRNA level of inflammatory factors (IL-1β, and IL-6). E The expression of lipid genes FASN, CD36, SCD1 and SREBP1 was detected by qPCR. * p < 0.05, *** p < 0.01, *** p < 0.001. N = 6

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Virus, Injection, Knockdown, Expressing, Western Blot, Staining, Enzyme-linked Immunosorbent Assay

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: Silencing CCL17 promotes the recruitment of Treg in AS plaque tissue. A The total CD4+ cells (CD3, CD4) in plaque tissue were detected by flow cytometry, and the proportion of cells in the upper right quadrant was counted. B The proportion of Treg cells (FOXP3, CD25) in CD4 cells was detected by flow cytometry. C The expression of FOXP3 in mouse plaque was detected by qPCR. D The expression of CD25 in mouse plaque was detected by immunofluorescence. * p <0.05, ** p <0.01, *** p <0.001. N =6

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Flow Cytometry, Expressing, Immunofluorescence

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: Macrophages stimulated by oxLDL affect the chemotaxis of Treg through CCL17 and CCL22 competition. Macrophages were stimulated with oxLDL, and Treg cells were co-cultured with macrophage supernatant. A Immunofluorescence detection of co-localization of CCL22 and macrophages (CD68) in plaque tissue of AS mice. B The content of CCL22 in the supernatant of macrophages was detected by ELISA. C Transwell detected the effect of macrophages on the chemotaxis of Treg. ( D - E ) Transwell detected the migration of Treg cells. * p < 0.05, ** p < 0.01, *** p < 0.001. N = 3

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Chemotaxis Assay, Cell Culture, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Migration

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: The stability of CCL17 protein is regulated by SENP3-mediated de-SUMOylation. A Western blot detected the level of SENP3 in macrophages induced by oxLDL. B SENP3 was overexpressed in macrophages, and the expressions of SENP3 and CCL17 were detected by western blot. C Bioinformatics on-line prediction of sumoylation site of CCL17 protein. D IP detection of the effect of SENP3 on the DeSUMOylation of CCL17. E & ( F ) The effect of SENP3 on the stability of CCL17 protein was detected by CHX protein degradation experiment. G Co-IP experiment verified the binding relationship between SENP3 and CCL17. H The DeSUMOylation of mutant CCL17 of K115R was detected by IP. I The binding relationship between SENP3 and CCL17 was verified through GST Pull-down. * p < 0.05, ** p < 0.01, *** p < 0.001. N = 3

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Western Blot, Co-Immunoprecipitation Assay, Binding Assay, Mutagenesis

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: Knocking down SENP3 improved the progress of AS. After 4 weeks of AS modeling, sh-SENP3 encapsulated by AAV virus was injected into the tail vein of mice to knock down the level of SENP3 in mice. A Western blot detected the expression of SENP3 in mice plaque tissue. B HE staining and oil red O staining were used to observe the pathological changes of arterial tissue in mice. C The contents of TC, TG and LDL-C in mice were detected by ELISA. D The levels of inflammatory factors in mice were detected by ELISA. E qPCR detected the expression level of lipid-related genes. F Flow detection of Treg cells in mice plaque tissue. G The co-localization of CCL17/CD68 in mice plaque tissues was detected by immunofluorescence. H The level of CD25 in the mice plaque was detected by immunofluorescence. * p < 0.05, ** p < 0.01, *** p < 0.001. N = 6

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques: Virus, Injection, Knockdown, Western Blot, Expressing, Staining, Enzyme-linked Immunosorbent Assay, Immunofluorescence

Journal: Cell Biology and Toxicology

Article Title: SENP3 mediated DeSUMOylation of macrophage derived CCL17 accelerates atherosclerosis via regulation of Treg

doi: 10.1007/s10565-025-10099-3

Figure Lengend Snippet: SENP3-mediated regulation of CCL17 in atherosclerosis. In atherosclerosis, SENP3 levels are elevated in macrophages, where it mediates the DeSUMOylation of CCL17, stabilizing its protein levels. Increased CCL17 secretion subsequently competitively inhibits Treg cell recruitment, exacerbating atherosclerosis progression

Article Snippet: Prior to the transwell migration assay, the induced Tregs were treated with recombinant mouse CCL17 (MCE, HY-P71891A) or recombinant mouse CCL22 (MCE, HY-P7248) or anti-CCL17 (R&D, Catalog #: MAB529) and anti-CCL22 antibodies (R&D, Catalog #: MAB529).

Techniques:

Journal: The EMBO Journal

Article Title: DNGR-1 signalling limits dendritic cell activation for optimal antigen cross-presentation

doi: 10.1038/s44318-025-00620-z

Figure Lengend Snippet: ( A , B ) Flow cytometric analyses of surface proteins and ELISA of indicated proteins released in culture supernatants from ( A ) RAW 264.7 cells ectopically expressing indicated receptors or transduced with EV stimulated overnight ± Zym-D or ( B ) C9 KO MuTuDCs reconstituted with indicated receptors stimulated overnight ± DNGR-1L. Data shown as mean ± SEM from biological triplicates. Note that the TNF-α data in ( A ) are plotted in two different ways (absolute versus relative concentration to untreated controls) to emphasise the similarity between C7 and C9(I6G)::C7. ( C ) Uptake of DNGR-1L-coupled beads by C9 KO MuTuDCs or those reconstituted with indicated receptors assessed by flow cytometry. Plotted as phagocytic index (% internalised beads x bead MFI of bead + cells/arbitrary unit) mean ± SD from biological duplicates. ( D , E ) C9 KO MuTuDCs reconstituted with C7::C9 or C7(G14I)::C9 receptors co-cultured with DNGR-1L-OVA coupled beads. Single internalised bead + cells were subsequently sorted and co-cultured with OT-I T E cells. ( D ) Schematic of experiment (left) and representative flow plots from pre- and post-sort enrichment (right). ( E ) ELISA for IFN-γ released from OT-I T E cells co-cultured with MuTuDCs from ( D ) or MuTuDCs loaded with exogenous SIINFEKL peptide. Mean ± SD from biological duplicates is plotted. Data are representative of two ( D , E ) or ≥ three ( A – C ) independent experiments. Data were analysed using Tukey-corrected two-way ANOVA ( A – C , E ). Significant values comparing against untreated controls ( A , B ) or between KO/C7(G14I)::C9 ( E ) are plotted. ( A ) H2-D d ** P = 0.0043, *** P = 0.0006, **** P < 0.0001; CD83 * P = 0.0374, *** P = 0.0002, **** P < 0.0001; TNF-α ** P = 0.0017 (C7), P = 0.0067 (C9(Y7F)::C7, *** P = 0.0004 (C7(G14I)), P = 0.0006 (C9::C7), **** P < 0.0001, ( B ) CD86 * P = 0.0469, **** P < 0.0001; CCR7 ** P = 0.0097, *** P = 0.0004, **** P < 0.0001; CCL17 **** P < 0.0001; CCL22 * P = 0.0217, ** P = 0.0018, **** P < 0.0001, ( C , E ) **** P < 0.0001. See also Fig. . .

Article Snippet: Taqman Ccl17 , Thermo Fisher Scientific , Assay ID: Mm00516136_m1.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Transduction, Concentration Assay, Flow Cytometry, Cell Culture