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: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 overexpression prevents SCI in mice. (A) Mice were exposed to Ctrl or SCI surgery, and FGF5 mRNA level in the spinal cord was detected at indicating times. (B) FGF5 level was detected using an ELISA kit. (C, D) Mice were intraspinally injected with 2 μL lentivirus carrying FGF5 to overexpress FGF5 in the spinal cord, and FGF5 mRNA and protein levels were detected 2 weeks post‐injection. (E) Mice were overexpressed with FGF5 using lentiviral vectors and then were exposed to SCI or sham surgery 2 weeks post‐injection. BMS score, from 0 (no ankle movement) to 9 (normal gait), was determined at indicating times. (F, G) Sensitivities to mechanical and thermal stimulation were determined 28 days after SCI. (H) Extravasation of Evans blue dye was determined 28 days after SCI. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Over Expression, Enzyme-linked Immunosorbent Assay, Injection, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 knockdown exacerbates SCI in mice. (A, B) Mice were intraspinally injected with 2 μL lentivirus carrying shFGF5 to knockdown FGF5 in the spinal cord, and FGF5 mRNA and protein levels were detected 2 weeks post‐injection. (C) Mice were injected with shFGF5, and then were exposed to SCI or sham surgery 2 weeks post‐injection. BMS score, from 0 (no ankle movement) to 9 (normal gait), was determined at indicating times. (D, E) Sensitivities to mechanical and thermal stimulation were determined 28 days after SCI. (F) Extravasation of Evans blue dye was determined 28 days after SCI. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Knockdown, Injection, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 overexpression reduces inflammation and oxidative stress in SCI mice. (A) Mice were overexpressed with FGF5 using lentiviral vectors, and then were exposed to SCI or sham surgery 2 weeks post‐injection. IL‐6 and TNF‐α levels in the spinal cord were detected. (B) MPO activity in the spinal cord. (C) Western blot images and quantification of p65 phosphorylation. (D) Relative NF‐κB transcription activity. (E, F) Relative levels of NRF2 protein and transcription activity. (G) ROS level detected by DCFH‐DA probe. (H) Quantification of hydrogen peroxide and superoxide anion in spinal cord. (I) The levels of MDA, 3‐NT and 8‐OHdG. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Over Expression, Injection, Activity Assay, Western Blot, Phospho-proteomics, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 knockdown elevates inflammation and oxidative stress in SCI mice. (A) Mice were injected with shFGF5, and then were exposed to SCI or sham surgery 2 weeks post‐injection. IL‐6 and TNF‐α levels in the spinal cord were detected. (B) MPO activity in the spinal cord. (C) Western blot images and quantification of p65 phosphorylation. (D) Relative NF‐κB transcription activity. (E, F) Relative levels of NRF2 protein and transcription activity. (G) ROS level detected by DCFH‐DA probe. (H) Quantification of hydrogen peroxide and superoxide anion in spinal cord. (I) The levels of MDA, 3‐NT and 8‐OHdG. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Knockdown, Injection, Activity Assay, Western Blot, Phospho-proteomics, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 overexpression reduces inflammation, oxidative stress and SCI through activating AMPK. (A) Mice were overexpressed with FGF5 using lentiviral vectors, and then were exposed to SCI or sham surgery 2 weeks post‐injection. Western blot images and quantification of AMPK phosphorylation in the spinal cord were detected. (B) To inhibit AMPK, mice with or without FGF5 overexpression were intraperitoneally injected with CC every 2 days 1 week pre‐SCI. IL‐6 and TNF‐α levels in the spinal cord were detected. (C) ROS level detected by DCFH‐DA probe. (D) The levels of MDA, 3‐NT and 8‐OHdG. (E) BMS score, from 0 (no ankle movement) to 9 (normal gait), was determined at indicating times. (F, G) Sensitivities to mechanical and thermal stimulation were determined 28 days after SCI. (H) Extravasation of Evans blue dye was determined 28 days after SCI. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Over Expression, Injection, Western Blot, Phospho-proteomics, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: FGF5 overexpression activates AMPK through cAMP/PKA pathway. (A, B) Mice were intraspinally injected with 2 μL lentivirus carrying FGF5 to overexpress FGF5 in the spinal cord, and cAMP level and PKA activity were detected 2 weeks post‐injection. (C) To inhibit PKA, mice with or without FGF5 overexpression were intraperitoneally injected with H89 every 2 days 1 week pre‐SCI. Western blot images and quantification of AMPK phosphorylation in the spinal cord were detected. (D) ROS level detected by DCFH‐DA probe. (E) The levels of MDA, 3‐NT and 8‐OHdG. (F) IL‐6 and TNF‐α levels in the spinal cord were detected. (G) BMS score, from 0 (no ankle movement) to 9 (normal gait), was determined at indicating times. (H, I) Sensitivities to mechanical and thermal stimulation were determined 28 days after SCI. (J) Extravasation of Evans blue dye was determined 28 days after SCI. n = 6 for each groups. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Over Expression, Injection, Activity Assay, Western Blot, Phospho-proteomics, Standard Deviation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Fibroblast growth factor 5 protects against spinal cord injury through activating AMPK pathway

doi: 10.1111/jcmm.17934

Figure Lengend Snippet: Serum FGF5 level positively correlates with the sensory and motor function in SCI patients. (A) Serum levels of FGF5 in Ctrl and SCI patients. (B, C) SCI patients were divided into four groups according to serum FGF5 levels, and ASIA sensation and motor scores were measured at indicating groups. n = 56 for Ctrl group and n = 92 for SCI group. Data are expressed as the mean ± standard deviation and p < 0.05 was considered statistically significant. * p < 0.05.

Article Snippet: FGF5 in mouse spinal cord and human serum samples were detected using Mouse FGF5 ELISA Kit (#CSB‐EL008632MO; CUSABIO) or Human FGF5 ELISA Kit (#CSB‐EL008632HU; CUSABIO), respectively according to the manufacturer's instructions.

Techniques: Standard Deviation

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