Journal: Frontiers in Immunology

Article Title: Nocardia rubra cell wall skeleton-induced MARCO expression: implications for improved phagocytosis and cytokine secretion in tumor-associated macrophages

doi: 10.3389/fimmu.2026.1611476

Figure Lengend Snippet: Nr-CWS affects MARCO expression through TLR4. The expression of TLR4 significantly increased after Nr-CWS treatment [ (D-G) , scale bars = 100 μm], while the expression TLR2 was unchanged (A-C) . Subsequent inhibition of TLR4 expression resulted in decreased levels of MARCO [ (H-J) , scale bars = 100 μm]. n = 3, * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following commercial antibodies (vendor, catalog number and dilution) utilized for western blot and immunohistochemical staining were employed in accordance with the manufacturers’ guidelines: mouse anti-CD68 antibody (Abcam, ab201340, 1:200), rabbit anti-MARCO antibody (Abcam, ab231046, 1:1000 for western blot and 1:100 for immunohistochemistry), rabbit anti-CD163 antibody (Abways Technology, CY6845, 1:500), mouse anti-CD86 antibody (Proteintech, 68674-2-Ig, 1:5000), mouse anti-TLR4 antibody (Santa Cruz Biotechnology, sc-293072, 1:1000 for western blot and 1:200 for immunohistochemistry), rabbit anti-TLR2 antibody(Abways Technology, CY5102, 1:2000), mouse anti-GAPDH antibody (Proteintech, 6004-1-Ig, 1:50000), HRP-goat anti-mouse recombinant secondary antibody (H+L)(Proteintech, RGAM001, 1:5000), goat anti-mouse IgG (H+L) Alexa Fluor 594 (Abways Technology, AB0152, 1:300), HRP-goat anti-rabbit recombinant secondary antibody (H+L) (Proteintech, RGAR001, 1:5000), goat anti-rabbit IgG (H+L) secondary antibody DyLightTM 594 (Report Biotech, S7002, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 488 (Report Biotech, S6001, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 594 (Abways Technology, AB0152, 1:500).

Techniques: Expressing, Inhibition

Journal: International Journal of Molecular Sciences

Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

doi: 10.3390/ijms262412024

Figure Lengend Snippet: Biglycan (BGN) promotes proliferation and migration of esophageal squamous cell carcinoma (ESCC) cells through its receptor TLR4. ( A ) Expression of TLR4 in ESCC cells was confirmed by Western Blotting. ( B ) Double immunofluorescence staining for BGN (green) and TLR4 (red) was performed in ESCC cells treated with recombinant human BGN (rhBGN; 100 ng/mL). Nuclei were counterstained with DAPI (blue). ( C , D ) MTS assays ( C ) and Transwell migration assays ( D ) were conducted to evaluate changes in the proliferation ( C ) and migration ( D ) of TE-9, TE-10, and TE-15 cells following treatment with rhBGN (100 ng/mL) in the presence of a TLR4-neutralizing antibody (1 μg/mL) or control immunoglobulin G (IgG; 1 μg/mL). Migrated cells were counted in five representative microscopic fields after 48 h, and representative images are shown beneath the graphs ( D ). The data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( C , D ). ** p < 0.01, *** p < 0.001. Scale bars: 20 μm ( B ) and 100 μm ( D ).

Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

Techniques: Migration, Expressing, Western Blot, Double Immunofluorescence Staining, Recombinant, Control

Journal: International Journal of Molecular Sciences

Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

doi: 10.3390/ijms262412024

Figure Lengend Snippet: Biglycan (BGN) promotes proliferation, migration, and activation of macrophages and fibroblasts. ( A ) Cell–cell communication network analysis of BGN -expressing cancer-associated fibroblasts (CAFs) in esophageal squamous cell carcinoma (ESCC) tissues. Fibroblasts in single-cell RNA sequencing (scRNAseq) datasets from ESCC tissues were stratified into CAF_ BGN _High (scaled expression > 3) and CAF_ BGN _Low (≤3) groups based on BGN expression levels. Cell–cell communication inferred by CellChat revealed that CAF_ BGN _High cells exhibited strong outgoing signaling toward epithelial and myeloid cells, as well as prominent autocrine signaling within the CAF_ BGN _High population. ( B , C ) MTS assay ( B ) and Transwell migration assay ( C ) showing increased proliferation and migration of mesenchymal stem cells (MSCs) treated with recombinant human BGN (rhBGN; 100 ng/mL), respectively. ( D ) Western Blot analysis showing that fibroblast activation protein (FAP) and α-smooth muscle actin (αSMA) (CAF markers) were upregulated in MSCs treated with rhBGN for 24 h, whereas interleukin-6 (IL6) expression remained unchanged. TLR4 expression was detected; however, phosphorylation of NF-κB and Erk was not increased by rhBGN. ( E , F ) MTS assay ( E ) and Transwell migration assay ( F ) showing that rhBGN-induced proliferation and migration of MSCs were not suppressed by a TLR4-neutralizing antibody (1 μg/mL). ( G , H ) MTS assay ( G ) and Transwell migration assay ( H ) showing increased proliferation and migration of macrophages treated with rhBGN (100 ng/mL), respectively. ( I ) Western Blot analysis showing that rhBGN treatment upregulated the expression of CD163 and CD206 (M2 macrophage markers) and increased NF-κB phosphorylation, while TLR4 expression was also detected, but phosphorylated Erk was not changed. ( J , K ) MTS assay ( J ) and Transwell migration assay ( K ) showing that rhBGN-induced proliferation and migration of macrophages were attenuated by treatment with a TLR4-neutralizing antibody (1 μg/mL), respectively. ( L , M ) MTS assay ( L ) and Transwell migration assay ( M ) showing that treatment with the NF-κB pathway inhibitor Bay 11-7082 (1 μM) attenuated rhBGN-induced proliferation and migration to a similar extent, respectively. Data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( B , C , E – H , J – M ). * p < 0.05, ** p < 0.01, *** p < 0.001. N.S., not significant. Scale bars: 100 μm ( C , F , H , K , M ).

Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

Techniques: Migration, Activation Assay, Expressing, RNA Sequencing, MTS Assay, Transwell Migration Assay, Recombinant, Western Blot, Phospho-proteomics

Journal: International Journal of Molecular Sciences

Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

doi: 10.3390/ijms262412024

Figure Lengend Snippet: A schematic illustration of the role of BGN in the tumor–stromal interactions among ESCC cells, CAFs, and macrophages. BGN secreted from MSCs that have undergone CAF transition upon direct contact with ESCC cells promotes ESCC cell proliferation and migration through the TLR4–Erk/NF-κB signaling pathways. In addition, BGN enhances the proliferation and migration of MSCs and induces their differentiation into CAFs. Furthermore, BGN promotes the proliferation and migration of macrophages and drives their polarization toward the M2 phenotype through the TLR4–NF-κB signaling pathways.

Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

Techniques: Migration, Protein-Protein interactions