rs100a8  (ProSpec)

Journal: Theranostics

Article Title: CEBPB-high dormant tumor cells drive immune evasion via S100A8 orchestrated tumor-associated macrophages reprogramming

doi: 10.7150/thno.124789

Figure Lengend Snippet: CEBPB transcriptionally activates S100A8 to promote M2 macrophage polarization. A , Venn diagram of three datasets identifying four common secretory factors: differentially expressed genes from CEBPB-overexpressing 4T07 cells bulk RNA-seq, CEBPB binding peaks in 4T07 cells from anti-CEBPB ChIP-seq, and the profile of mouse secretory proteins. B , CEBPB binding peaks for S100A8 and Cxcl1 from anti-CEBPB ChIP-seq in 4T07 cells. C , ChIP-qPCR showing the CEBPB binding regions in the S100a8 promoter or upstream regions of Cxcl1 of 4T07 and EMT6 cells. D , Relative luciferase activity in CEBPB-overexpressing HEK293T cells with the S100a8 promoter WT or MUT, respectively. E-F , mRNA expression of S100a8 or Cxcl1 in 4T07 cells with CEBPB overexpression (E) or knockdown (F). G , Protein expression of S100A8 in 4T07 cells with or without CEBPB overexpression. H , Representative multiplex immunohistochemistry images of Ki67, CEBPB, and S100A8 staining in a TMA of TNBC patient tumor smaples ( n = 80). I , mRNA expression of S100a8 in eGFP -/lo and eGFP +/hi tumor cells sorted from 4T07-TetOn or EMT6-TetOn tumor masses. J-K , Percentages of CD206 (J) and MHCII (K) expressing BMDMs with the addition of rS100A8 (100 or 500 ng/mL) or rCXCL1 (100 or 500 ng/mL) for 48 hours. L-N , Tumor volume (L), image (M) and weight (N) of CEBPB-overexpressing with or without S100A8 knockdown EMT6 allografts in BALB/c mice ( n = 5). O-R , Flow cytometry analysis of CD8 + T cells (O), CD11b + F4/80 + cells (P), CD206 + macrophages (Q), and MHCII + macrophages (R) infiltrated in tumors developed in (L). n.s., not significant, *, P < 0.05, **, P < 0.01, ***, P < 0.001.

Article Snippet: BMDMs or PBMC-M φ cells were co-cultured in conditional medium supplemented with 10% FBS and 1% penicillin-streptomycin, or in complete DMEM or 1640 medium supplemented with rS100A8 (MCE, #HY- P73671 for BMDMs and #HY- P70531 for PBMC-M φ ) or rCXCL1 (MCE, #HY-P7188 for BMDMs) for 48 hours.

Techniques: RNA Sequencing, Binding Assay, ChIP-sequencing, ChIP-qPCR, Luciferase, Activity Assay, Expressing, Over Expression, Knockdown, Multiplex Assay, Immunohistochemistry, Staining, Flow Cytometry

Journal: Theranostics

Article Title: CEBPB-high dormant tumor cells drive immune evasion via S100A8 orchestrated tumor-associated macrophages reprogramming

doi: 10.7150/thno.124789

Figure Lengend Snippet: The S100A8 inhibitor remodels TME and improves ICB efficacy. A-B , Percentages of CD206 expressing in BMDMs (A) or PBMC-Mφ (B) with the addition of paquinimod (PAQ, 5 µg/mL) and/or rS100A8 (500 ng/mL) for 48 hours. C , Migration assay of BMDMs in DMEM with the addition of PAQ (5 µg/mL) and/or rS100A8 (500 ng/mL) for 24 hours. D-E , Percentages of CD206 expressing in BMDMs (D) or PBMC-Mφ (E) co-cultured with CM and/or PAQ (5 µg/mL) for 48 hours, including CM from 4T07 (D) or MDA-MB-231 (E) with or without CEBPB overexpression. F-H , Tumor volume (F), image (G) and weight (H) of EMT6 allografts in BALB/c mice ( n = 5). Tumor-bearing mice were randomly divided into four groups until tumor volume reached approximately 100 mm 3 . I-L , Flow cytometry analysis of CD8 + T cells (I), CD11b + F4/80 + macrophages (J), CD206 + macrophages (K) MHCII + macrophages (L) infiltrated in tumors developed of the indicated groups. M , Kaplan-Meier analysis of S100A8 expression in a public cohort with 238 patients receiving anti-PD-1 therapy. n.s., not significant, *, P < 0.05, **, P < 0.01, ***, P < 0.001 .

Article Snippet: BMDMs or PBMC-M φ cells were co-cultured in conditional medium supplemented with 10% FBS and 1% penicillin-streptomycin, or in complete DMEM or 1640 medium supplemented with rS100A8 (MCE, #HY- P73671 for BMDMs and #HY- P70531 for PBMC-M φ ) or rCXCL1 (MCE, #HY-P7188 for BMDMs) for 48 hours.

Techniques: Expressing, Migration, Cell Culture, Over Expression, Flow Cytometry

Journal: Shock (Augusta, Ga.)

Article Title: TARGETING S100A9-TLR2 AXIS CONTROLS MACROPHAGE NLRP3 INFLAMMASOME ACTIVATION IN FATTY LIVER ISCHEMIA REPERFUSION INJURY

doi: 10.1097/SHK.0000000000002470

Figure Lengend Snippet: S100A9 induces inflammatory injury via macrophage TLR2 activation BMMs were isolated from WT mice and incubated with rS100A9 or rS100A8 respectively for 24 h. (A) Western-assisted analysis of TLR2 and ATF4 in rS100A9-stressed macrophages. (B) Western-assisted analysis of TLR2 and ATF4 in rS100A8-stressed macrophages; BMMs were transfected with the TLR2-siRNA or control vector followed by incubation with rS100A9. (C) Detection of cytokines IL-1β, CXCL1, CXCL-2, and IL-6 by qRT-PCR in BMMs. (D) Western-assisted analysis of ATF4 in nuclear extracts. (E) Immunofluorescence staining for ATF4 distribution in macrophages. DAPI was used to visualize nuclei. Scale bars: 50 μm, 20 μm. N = 3–6/group. All data represent the mean ± SD, * P < 0.05, ** P < 0.01, *** P < 0.001. BBM, bone-derived macrophage. WT, wild type; qRT-PCR, quantitative real-time PCR.

Article Snippet: After a period of 24–48 h, the cells were treated with recombinant S100A9 (rS100A9) or rS100A8 (Prospec, Rehovot, Israel) for additional 24 h.

Techniques: Activation Assay, Isolation, Incubation, Western Blot, Transfection, Control, Plasmid Preparation, Quantitative RT-PCR, Immunofluorescence, Staining, Derivative Assay, Real-time Polymerase Chain Reaction