goat anti human igg  (R&D Systems)

Journal: bioRxiv

Article Title: Mechano-activated chondroprogenitors (MACs) drive intrinsic cartilage repair; a process that is arrested in osteoarthritis

doi: 10.1101/2025.08.21.671507

Figure Lengend Snippet: Low density porcine chondrocytes were stimulated with injury CM in the presence or absence of FGFRi (250nM SB402451) or anti-FGF2 neutralising antibody (4 μg/mL) and imaged using the IncuCyte imaging system. A , Representative images of cells after 96h culture. B , C , Quantitation of cell density assessed by IncuCyte imaging (at 2h intervals) over 96h. Movie showing change in cell morphology and proliferation for each condition are presented in supplementary video data file S5. D , Yap-Tead GFP reporter assay after stimulation of cells with IL-6/IL-6R (positive control), injury CM at indicated dilutions, with or without FGFRi (250 nM). Graph shows mean ± SD. Data points indicate independent experiments. One-way ANOVA with Tukey’s post-test, n = 6. Quantification E , and representative images F , of in vivo murine cartilage repair model, induced at 10 weeks of age and assessed after 8 weeks, in FGF2 -/- and DBA1 control mice. Histological sections were scored using a modified Pineda score. Mean ± SEM; Mann-Whitney U test; *p>0.05, **p>0.01, n = 12 biological replicates.

Article Snippet: Human recombinant FGF2 was purchased from Peprotech EC Ltd. Recombinant human Activin A (MAB3381), TGF-βR3 (QK054), FGF2 neutralizing antibody (AF-233-NA) and Activin A neutralizing antibody (MAB3381) were obtained from R&D Systems.

Techniques: Imaging, Quantitation Assay, Reporter Assay, Positive Control, In Vivo, Control, Modification, MANN-WHITNEY

Journal: bioRxiv

Article Title: Mechano-activated chondroprogenitors (MACs) drive intrinsic cartilage repair; a process that is arrested in osteoarthritis

doi: 10.1101/2025.08.21.671507

Figure Lengend Snippet: A , Illustration of SJD. B, Volcano plot showing mean difference (log) in protein expression against adjusted p-values, pre and post SJD in paired samples (n = 16, 5471 proteins, padj ≤ 0.05). Top regulated proteins labelled. C, Line plots for a selection of regulated proteins. INHBA, inhibin βA; ACTIVIN A, inhibin βA homodimer; ACTIVIN AC, Inhibin βA:Inhibin βC heterodimer; BMP, bone morphogenetic proteins-5 & 6. SOX-9, SRY box transcription factor-9; COL2A1, collagen type II a1 chain; COL10A1, collagen type 10 a1 chain; HPLN1, link protein; HSPG2, perlecan. D , Top 10 pathways after gene enrichment analysis (Reactome). E , Cell confluency by IncuCyte imaging of primary chondrocytes stimulated with injury CM in the presence of anti-activin A. F , Chondrogenesis assay of MACs (generated by 72h treatment with injury CM) in the presence of recombinant FGF2 or activin A. Expression of chondrogenic marker genes measured by qPCR at day 3. G , Schematic of the intrinsic repair cycle of articular cartilage. A and G created using BioRender.

Article Snippet: Human recombinant FGF2 was purchased from Peprotech EC Ltd. Recombinant human Activin A (MAB3381), TGF-βR3 (QK054), FGF2 neutralizing antibody (AF-233-NA) and Activin A neutralizing antibody (MAB3381) were obtained from R&D Systems.

Techniques: Expressing, Selection, Imaging, Generated, Recombinant, Marker

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Flow chart of the experimental design and verification of FGF2 deletion. a Diagram of experimental design. Bone marrow precursor cells were collected from the tibia and femur of WT and FGF2 KO mice aged 8–12 weeks. They were differentiated into BMDM for 7 days using 100ng/ml M-CSF. Macrophages in C57BL/6 male mice were depleted using clodronate liposomes. BMDM from WT and KO mice were injected via the tail vein into mice to reconstitute macrophages after 2 days. Mice were treated with Sham or CLP, and the samples were analyzed 24 h later. b Serum FGF2 protein levels were assessed by ELISA ( n = 3–5). c FGF2 gene expression levels were measured in the lungs, spleen, and liver using real-time PCR ( n = 3). d Western blot analysis of FGF2 protein levels in lung tissue extractions ( n = 3). e Immunofluorescence staining of FGF2 in BMDM from WT and FGF2 KO mice ( n = 3). f The levels of FGF2 gene expression were quantified in the lung tissue of mice subjected to elimination-reconstruction procedures. Bar is 50 μm. * p < 0.05 vs. WT or vs. Clo + WT

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Liposomes, Injection, Enzyme-linked Immunosorbent Assay, Gene Expression, Real-time Polymerase Chain Reaction, Western Blot, Immunofluorescence, Staining

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Effect of FGF2 deficiency on BMDM apoptosis and polarization. a – c FGF2 deletion increased BMDM apoptosis. a Apoptosis in BMDM deprived of FBS for 24 h was assessed by flow cytometry ( n = 4). b - c Percentage of PI + Annexin V + and PI- Annexin V + BMDM after starvation. d - k FGF2 deletion in BMDM promoted M1 polarization. d - g Flow cytometric analysis of macrophage markers in BMDM treated with LPS or IL4, including CD86, iNOS, CD206, and Arg1 ( n = 3). h - k The levels of CD86, iNOS, CD206 and Arg1 in BMDM after treatment with LPS or IL4. N represents no treatment; * p < 0.05, vs. WT; Ψ p < 0.05, vs. N + WT; Ω p < 0.05, vs. N + FGF2 KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Flow Cytometry

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Deficiency of FGF2 in BMDM resulted in the upregulation of M1 markers and proinflammatory cytokine expression and increased nuclear translocation of NF-KB p65. a - j BMDM were treated with LPS or IL4, and real-time PCR was used to determine the expression of M1 and M2 markers and cytokines ( n = 3). k , l P65 nuclear translocation was detected and quantitatively analyzed by immunofluorescence ( n = 3). m The expression of MMP9 in WT and FGF2 KO BMDM treated with LPS or IL4 were determined with real-time PCR. Bar in the first three rows is 50 μm, while bar in the fourth rows is 20 μm, * p < 0.05 vs. WT; Ψ p < 0.05 vs. N + WT; Ω p < 0.05 vs. N + KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Expressing, Translocation Assay, Real-time Polymerase Chain Reaction, Immunofluorescence

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Transcriptome sequencing was used to analyze BMDM from WT and FGF2 KO mice treated with or without LPS. a Heat map of DEGs in BMDM from WT and FGF2 KO mice stimulated with or without LPS (Average TPM each group, n = 3). b KEGG enrichment analysis identified the top 20 altered signaling pathways in the four groups (WT, FGF2 KO, WT + LPS, and FGF2 KO + LPS). c Construction of a regulatory network to modulate PPI involving FGF2 and LPS using DEGs. d KEGG pathway network based on similarity in gene expression profiles. e Volcano plot showing DEGs of WT + LPS and FGF2 KO + LPS. f Heat map of DEGs in BMDM of WT + LPS and FGF2 KO + LPS. g Top 20 KEGG pathways for DEGs in BMDM from WT + LPS and FGF2 KO + LPS. h KEGG pathway annotation of differentially expressed genes between WT + LPS and FGF2 KO + LPS. i , j Differently regulated pathways in the GSEA. k Inflammation and cytokine gene heat map for WT + LPS and FGF2 KO + LPS. l , m TPM changes in the gene groups compared to WT weights. * p < 0.05 vs. WT; Δ p < 0.05 vs. WT + LPS; # p < 0.05 vs. FGF2 KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Sequencing, Protein-Protein interactions, Gene Expression

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: FGF2 deletion in macrophages aggravates lung injury in CLP mice. a - c Clodronate liposomes were injected intravenously to deplete macrophages, followed by quantification of F4/80 + spleen cells using flow cytometry to measure macrophage clearance. * p < 0.05 vs. control. d - f Levels of the inflammatory cytokines IL1β, TNFα, and IL6 in bronchoalveolar lavage fluid (BALF) from the four groups were quantified via ELISA. The four groups were Clo + WT + Sham, Clo + WT + CLP, Clo + FGF2 KO + Sham, and Clo + FGF2 KO + CLP. g The lung wet-to-dry weight ratios were measured in the four groups. h - i The total cells and total protein in BALF were evaluated in the four groups. j The OD value of Evans blue in the lung tissues from the four groups. k - l HE staining of lung tissue from the four groups and their Smith score. (m-o) Blood gas analysis was performed using abdominal aortic blood from the four groups. Bar is 250 μm. * p < 0.05 vs. WT; Δ p < 0.05 vs. WT + LPS; # p < 0.05 vs. FGF2 KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Liposomes, Injection, Flow Cytometry, Control, Enzyme-linked Immunosorbent Assay, Staining

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Mice reconstituted with FGF2 KO macrophages and subjected to CLP demonstrate increased M1 polarization in lung tissue. a - f The presence and levels of CD206, CD86, and F4/80 markers on macrophages within lung tissue were identified and quantitatively assessed using immunofluorescence staining. Bar is 20 μm. * p < 0.05, vs. WT; Δ p < 0.05 vs. WT + LPS; # p < 0.05 vs. FGF2 KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Immunofluorescence, Staining

Journal: Molecular Biomedicine

Article Title: Deleting fibroblast growth factor 2 in macrophages aggravates septic acute lung injury by increasing M1 polarization and inflammatory cytokine secretion

doi: 10.1186/s43556-024-00203-0

Figure Lengend Snippet: Mice reconstituted with FGF2 KO macrophages and subjected to CLP exhibit increased proinflammatory gene expression and apoptosis. a - c The expression of the inflammatory factors CXCL1, IL1β, and IL6 in lung tissue was detected by real-time PCR ( n = 3 per group). d TUNEL staining was used to evaluate apoptosis in the lung tissue. e - f The apoptosis-associated genes BCL2 and Bax were identified by western blot analysis. Bar is 20 μm. * p < 0.05, vs. WT; Δ p < 0.05, vs. WT + LPS; # p < 0.05 vs. FGF2 KO

Article Snippet: Briefly, BMDM were stimulated with 10 ng/ml LPS or 10 ng/ml IL4 for 24 h. After fixation with 4% paraformaldehyde (Cat No.BL539A, Labgic, Beijing, China) for 10 min, the cells were blocked with normal goat serum (Cat No. A7007, Beyotime, Jiangsu, China) for 30 min at room temperature, followed by incubation with either FGF2 antibody (1:100, Cat No. MA00121, Boster, Wuhan, China) or P65 antibody (1:500, Cat No.8242, CST, USA) for 1 h at 37 °C.

Techniques: Gene Expression, Expressing, Real-time Polymerase Chain Reaction, TUNEL Assay, Staining, Western Blot