Journal: International Journal of Molecular Sciences

Article Title: Inflammation-Driven Secretion Potential Is Upregulated in Osteoarthritic Fibroblast-Like Synoviocytes

doi: 10.3390/ijms231911817

Figure Lengend Snippet: Growth factor secretion levels by HFLS and HFLS-OA stimulated with TNFα or LPS measured using the ELISA method. Secretion of growth factors measured in media using the ELISA method. Cells were stimulated for 24 h with 10 ng/mL of TNFα ( A , C , E ) or LPS ( B , D , F ). Afterward, the media were collected, and ANGPTL1 ( A , B ), FGF5 ( C , D ), and IGF2 ( E , F ) levels were determined. Data are presented as pg or ng/mL ± SEM and analyzed with two-way ANOVA followed by Tukey’s post hoc test. *** p < 0.001 vs. control group. ## 0.01 > p > 0.001; ### p < 0.001 HFLS vs. HFLS-OA stimulated groups. HFLS, human fibroblast-like synoviocytes; HFLS-OA, osteoarthritic human fibroblast-like synoviocytes; TNFα, tumor necrosis factor alpha; LPS, lipopolysaccharide; ELISA, enzyme linked immunosorbent assay; ANGPTL1, angiopoietin like 1; FGF5, fibroblast growth factor 5; IGF2, insulin growth factor 2.

Article Snippet: The protein levels of the CXCL6 (Human GCP2 (Granulocyte Chemotactic Protein 2) ELISA Kit, ELK Biotechnology, Wuhan, China), CXCL10 (Human IP-10/CXCL10 ELISA Kit, Elabscience, Wuhan, China), CXCL16 (Human CXCL16 ELISA Kit, Elabscience, Wuhan, China), ANGPTL1 (Human ANGPTL1 ELISA Kit, ELK Biotechnology, Wuhan, China), FGF5 (Human FGF5 ELISA Kit, ELK Biotechnology, Wuhan, China) and IGF-2 (Human IGF-2 ELISA Kit, Elabscience, Wuhan, China) were measured using commercially available enzyme-linked immunosorbent assay kits according to the manufacturers’ instructions.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Cell Death & Disease

Article Title: Cancer-associated fibroblasts secrete FGF5 to inhibit ferroptosis to decrease cisplatin sensitivity in nasopharyngeal carcinoma through binding to FGFR2

doi: 10.1038/s41419-024-06671-0

Figure Lengend Snippet: Human CAFs and NFs were isolated from NPC tissues and the adjacent normal tissues, respectively. A The expression levels of α-SMA and FAP in CAFs were detected by immunofluorescence. Green, α-SMA. Red, FAP. Blue, DAPI. Scale bar: 100 μm. B The protein levels of α-SMA, vimentin and FAP in NFs and CAFs were detected by western blot. C The protein level of FGF5 in fibroblasts was detected by western blot. D The secretion of FGF5 in fibroblasts was assessed by ELISA assay. NPC/HK1 and C666-1 cells were divided into following groups: control, DDP (5 μg/mL), DDP (5 μg/mL)+CM and DDP (5 μg/mL)+anti-FGF5 antibody. E Cell viability was monitored by CCK-8 assay. F The protein levels of GPX4 and SLC7A11 were detected by western blot. G – I The levels of MDA ( G ), Fe 2+ ( H ) and GSH ( I ) in NPC cells were measured using commercial kits. J NPC cells were stained with BODIPY C11 and lipid peroxidation was assessed by flow cytometry. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: The protein level of FGF5 in culture medium secreted from CAFs or NFs was detected using Human FGF5 ELISA Kit (EH191RB, Invitrogen) according to the protocol.

Techniques: Isolation, Expressing, Immunofluorescence, Western Blot, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Staining, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Cancer-associated fibroblasts secrete FGF5 to inhibit ferroptosis to decrease cisplatin sensitivity in nasopharyngeal carcinoma through binding to FGFR2

doi: 10.1038/s41419-024-06671-0

Figure Lengend Snippet: A The mRNA level of Nrf2 in transfected NPC cells were detected by qRT-PCR. NPC/HK1 and C666-1 cells were divided into six groups: control, DDP (5 μg/mL), DDP (5 μg/mL) + Vector-CM, DDP (5 μg/mL)+FGF5-CM, DDP (5 μg/mL)+FGF5-CM+shNC and DDP (5 μg/mL) + FGF5-CM + shNrf2#2. B The protein levels of Keap1, Nrf2 and HO-1 in NPC cells were detected by western blot. C Cell viability was monitored by CCK-8 assay. D – F The levels of MDA ( D ), Fe 2+ ( E ) and GSH ( F ) in NPC cells were measured using commercial kits. G NPC cells were stained with BODIPY C11 and lipid peroxidation was assessed by flow cytometry. H The protein levels of GPX4 and SLC7A11 were detected by western blot. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: The protein level of FGF5 in culture medium secreted from CAFs or NFs was detected using Human FGF5 ELISA Kit (EH191RB, Invitrogen) according to the protocol.

Techniques: Transfection, Quantitative RT-PCR, Plasmid Preparation, Western Blot, CCK-8 Assay, Staining, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Cancer-associated fibroblasts secrete FGF5 to inhibit ferroptosis to decrease cisplatin sensitivity in nasopharyngeal carcinoma through binding to FGFR2

doi: 10.1038/s41419-024-06671-0

Figure Lengend Snippet: A The interactions between FGF5 and FGFRs in NPC tissues were detected by Co-IP assay. B The mRNA level of FGFR2 in N69, NPC/HK1 and C666-1 cells was detected by qRT-PCR. C The protein level of FGFR2 in N69, NPC/HK1 and C666-1 cells was detected by western blot. D The expression of FGFR2 in nasal polyp and NPC tissues was determined by tissue microarray. The first column: nasal polyp tissues. 2–9 columns: NPC tissues. E The immunoreactivities of FGF5 and FGFR2 in NPC tissues were detected by IHC analysis. Scale bar: 20 μm. * P < 0.05 and *** P < 0.001.

Article Snippet: The protein level of FGF5 in culture medium secreted from CAFs or NFs was detected using Human FGF5 ELISA Kit (EH191RB, Invitrogen) according to the protocol.

Techniques: Co-Immunoprecipitation Assay, Quantitative RT-PCR, Western Blot, Expressing, Microarray

Journal: Cell Death & Disease

Article Title: Cancer-associated fibroblasts secrete FGF5 to inhibit ferroptosis to decrease cisplatin sensitivity in nasopharyngeal carcinoma through binding to FGFR2

doi: 10.1038/s41419-024-06671-0

Figure Lengend Snippet: A The mRNA level of FGFR2 in transfected NPC cells were detected by qRT-PCR. B The protein level of FGFR2 in transfected NPC cells were detected by western blot. NPC/HK1 and C666-1 cells were divided into five groups: control, DDP (5 μg/mL), DDP (5 μg/mL)+FGF5-CM, DDP (5 μg/mL) + FGF5-CM + shNC and DDP (5 μg/mL) + FGF5-CM + shFGFR2#1. C The protein levels of FGFR2, Keap1, Nrf2 and HO-1 in NPC cells were detected by western blot. D Cell viability was monitored by CCK-8 assay. E – G The levels of MDA ( E ), Fe 2+ ( F ) and GSH ( G ) in NPC cells were measured using commercial kits. H NPC cells were stained with BODIPY C11 and lipid peroxidation was assessed by flow cytometry. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: The protein level of FGF5 in culture medium secreted from CAFs or NFs was detected using Human FGF5 ELISA Kit (EH191RB, Invitrogen) according to the protocol.

Techniques: Transfection, Quantitative RT-PCR, Western Blot, CCK-8 Assay, Staining, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Cancer-associated fibroblasts secrete FGF5 to inhibit ferroptosis to decrease cisplatin sensitivity in nasopharyngeal carcinoma through binding to FGFR2

doi: 10.1038/s41419-024-06671-0

Figure Lengend Snippet: Male BALB/c nude mice were randomly divided into three groups: control, DDP (4 mg/kg) and DDP (4 mg/kg)+FGF5 recombinant protein groups. A Representative photos of xenograft tumors. B , C Quantitative analyses of tumor volume ( B ) and weight ( C ). D The immunoreactivity of 4-HNE in tumors was detected by IHC analysis. Scale bar: 5 μm. E – G The levels of MDA ( E ), Fe 2+ ( F ) and GSH ( G ) in xenograft tumors were measured using commercial kits. H The protein levels of GPX4, SLC7A11, FGFR2, Keap1, Nrf2 and HO-1 were detected by western blot. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: The protein level of FGF5 in culture medium secreted from CAFs or NFs was detected using Human FGF5 ELISA Kit (EH191RB, Invitrogen) according to the protocol.

Techniques: Recombinant, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Inflammation-Driven Secretion Potential Is Upregulated in Osteoarthritic Fibroblast-Like Synoviocytes

doi: 10.3390/ijms231911817

Figure Lengend Snippet: Growth factor secretion levels by HFLS and HFLS-OA stimulated with TNFα or LPS measured using the ELISA method. Secretion of growth factors measured in media using the ELISA method. Cells were stimulated for 24 h with 10 ng/mL of TNFα ( A , C , E ) or LPS ( B , D , F ). Afterward, the media were collected, and ANGPTL1 ( A , B ), FGF5 ( C , D ), and IGF2 ( E , F ) levels were determined. Data are presented as pg or ng/mL ± SEM and analyzed with two-way ANOVA followed by Tukey’s post hoc test. *** p < 0.001 vs. control group. ## 0.01 > p > 0.001; ### p < 0.001 HFLS vs. HFLS-OA stimulated groups. HFLS, human fibroblast-like synoviocytes; HFLS-OA, osteoarthritic human fibroblast-like synoviocytes; TNFα, tumor necrosis factor alpha; LPS, lipopolysaccharide; ELISA, enzyme linked immunosorbent assay; ANGPTL1, angiopoietin like 1; FGF5, fibroblast growth factor 5; IGF2, insulin growth factor 2.

Article Snippet: The protein levels of the CXCL6 (Human GCP2 (Granulocyte Chemotactic Protein 2) ELISA Kit, ELK Biotechnology, Wuhan, China), CXCL10 (Human IP-10/CXCL10 ELISA Kit, Elabscience, Wuhan, China), CXCL16 (Human CXCL16 ELISA Kit, Elabscience, Wuhan, China), ANGPTL1 (Human ANGPTL1 ELISA Kit, ELK Biotechnology, Wuhan, China), FGF5 (Human FGF5 ELISA Kit, ELK Biotechnology, Wuhan, China) and IGF-2 (Human IGF-2 ELISA Kit, Elabscience, Wuhan, China) were measured using commercially available enzyme-linked immunosorbent assay kits according to the manufacturers’ instructions.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Inflammation-Driven Secretion Potential Is Upregulated in Osteoarthritic Fibroblast-Like Synoviocytes

doi: 10.3390/ijms231911817

Figure Lengend Snippet: Chemokine secretion levels by HFLS and HFLS-OA stimulated with TNFα or LPS measured using the ELISA method. Upregulation of chemokine level secretion measured in media using the ELISA method. Cells were stimulated for 24 h with 10 ng/mL of TNFα ( A , C , E ) or LPS ( B , D , F ). Afterward, the media were collected, and CXCL6 ( A , B ), CXCL10 ( C , D ), and CXCL16 ( E , F ) levels were determined. Data are presented as pg/mL ± SEM and analyzed with two-way ANOVA followed by Tukey’s post hoc test. ** 0.01 > p > 0.001; *** p < 0.001 vs. control group. ## 0.01 > p > 0.001; ### p < 0.001 HFLS vs. HFLS-OA stimulated groups. HFLS, human fibroblast-like synoviocytes; HFLS-OA, osteoarthritic human fibroblast-like synoviocytes; TNFα, tumor necrosis factor alpha; LPS, lipopolysaccharide; ELISA, enzyme linked immunosorbent assay; CXCL6, chemokine (C-X-C motif) ligand 6; CXCL10, chemokine (C-X-C motif) ligand 10; CXCL16, chemokine (C-X-C motif) ligand 16.

Article Snippet: The protein levels of the CXCL6 (Human GCP2 (Granulocyte Chemotactic Protein 2) ELISA Kit, ELK Biotechnology, Wuhan, China), CXCL10 (Human IP-10/CXCL10 ELISA Kit, Elabscience, Wuhan, China), CXCL16 (Human CXCL16 ELISA Kit, Elabscience, Wuhan, China), ANGPTL1 (Human ANGPTL1 ELISA Kit, ELK Biotechnology, Wuhan, China), FGF5 (Human FGF5 ELISA Kit, ELK Biotechnology, Wuhan, China) and IGF-2 (Human IGF-2 ELISA Kit, Elabscience, Wuhan, China) were measured using commercially available enzyme-linked immunosorbent assay kits according to the manufacturers’ instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Control