anti vav1  (Cell Signaling Technology Inc)

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A Adv TrkA mice experimental procedure. B IF of CGRP (red) in Adv TrkA mice injected with Veh or TMX at 14 days after Modeling. ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. C H&E and Masson staining of peritendinous tissue from Adv TrkA mice ( n = 6). Scale bars: 300 μm. D Analysis of ROM of Adv TrkA mice ( n = 6). E IHC of Col1 and Col3 expression of peritendinous tissue from Adv TrkA mice ( n = 6). Scale bars: 100 μm. F Protein expressions of Col 1 and Col 3 in fibrosis tissue from Adv TrkA mice ( n = 3). G IF of Prrx1 (red) from Adv TrkA mice ( n = 6). Scale bars: 50 μm. H Double IF of αSMA (green) and Col1 (red) of peritendinous tissue from Adv TrkA mice ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. I Adv DTA mice experimental procedure. J Analysis of ROM of Adv DTA mice ( n = 6). K IHC of Col1 and Col3 expression of peritendinous tissue from Adv DTA mice. Scale bars: 100 μm. L Protein expressions of Col 1 and Col 3 in fibrosis tissue of Adv DTA mice. M IF of αSMA (green) of peritendinous tissue from Adv DTA mice. ( n = 6). Scale bars: 50 μm. Yellow dashed line shows space between tendon and surrounding tissues. Black or white dashed line shows space occupied by fibrosis tissues. T represents tendon; AD represents fibrosis tissue; M represents muscle (mean ± SD; B – H / J – M : unpaired t-test; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; A/I created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Injection, Staining, Expressing

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A Schematic diagram of cholera toxin B (CTB) retrograde neuronal tracing. B Representative NeuN/CTB staining of DRG neurons ( n = 6). C Relative mRNA expression of NGF, NPY, SP, and CGRP in WT mice at normal, 7, 14, and 28 days ( n = 6). D Relative mRNA expression of NGF at the fibrosis site and DRG at 14 days ( n = 6). E NGF protein concentration at fibrotic sites in WT and Adv TrkA mice (pg/mg protein, n = 6). F Double IF of NGF (green) and NeuN (red) after modeling on 14 day in DRG from normal, Adv TrkA mice injected with Veh or TMX ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. G IF of NGF (green) after modeling on 14 day in fibrosis site from normal, Adv TrkA mice injected with Veh or TMX ( n = 6). Scale bars: 50 μm. H Experimental workflow of intrathecal AAV-GFP-siNGF injection. I H&E and Masson staining of WT mice with intrathecal injection of AAV-GFP-siNGF or NC at 14 days. Scale bars: 300 μm. J Adv NGF mice experimental procedure. K CGRP (red) staining in Adv NGF mice injected with Veh or TMX at 14 days after Modeling ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. L Analysis of ROM of Adv NGF mice ( n = 6). M Double IF of αSMA (green) and Col1 (red) of Adv NGF mice. ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm.Yellow dashed line shows space between tendon and surrounding tissues. Black or white dashed line shows space occupied by fibrosis tissues. T represents tendon; AD represents fibrosis tissue; M represents muscle(mean ± SD; F , G : ANOVA with multiple comparisons, I / K – M : unpaired t-test; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; A / H (mouse, injector and intrathecal space)/ J created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed; H (tendon and fibrosis tissue) created in BioRender. Author, li, y. (2025)) https://BioRender.com/0hlz1hf ).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Staining, Expressing, Protein Concentration, Injection

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A Prrx1 TrkA mice experimental procedure. B Analysis of ROM of Prrx1 TrkA mice injected with Veh or TMX at 14 and 28 days after modeling ( n = 6). C , D Double IF of CGRP (red) in Prrx1 TrkA mice ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. E Double IF of αSMA (green) and Col1 (red) in Prrx1 TrkA mice ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. F IHC of Col1 and Col3 expression of peritendinous tissue from Prrx1 TrkA mice injected with Veh or TMX at 14 and 28 days after modeling ( n = 6). Scale bars: 100 μm. G RNA-Sequencing experimental procedure of Prrx1 TrkA mice. H GO analysis of Prrx1 TrkA mice. ( I ) KEGG analysis of Prrx1 TrkA mice. J Heatmap of key genes expression in GO and KEGG analysis of Prrx1 TrkA mice. ( K ) Heatmap of EMT related gene set expression of Prrx1 TrkA mice. L IF of Engrailed-1 (green) in WT, Adv TrkA , Adv NGF , Prrx1 TrkA mice injected with TMX at 14 days after modeling. ( n = 6). Scale bars: 50 μm.Yellow dashed line shows space between tendon and surrounding tissues. Black or white dashed line shows space occupied by fibrosis tissues. T represents tendon; AD represents fibrosis tissue; M represents muscle; S represents skin. (mean ± SD; B / D – F : unpaired t-test, H / I / L : ANOVA with multiple comparisons; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; A (mouse)/G(mouse) created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed; G (tendon, fibrosis tissue and MSCs) created in BioRender. Author, li, y. (2025) https://BioRender.com/0hlz1hf ; G (Mechanical and enzymatic digestion, FACS) created in BioRender. Author, li, y. (2025) https://BioRender.com/1qycjy7 ).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Injection, Expressing, RNA Sequencing

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A , B experimental procedure and phenotypic diagram of Prrx1 Hif1a mice and Prrx1 VHL mice. C H&E and Masson staining and double IF of αSMA (green) and Col1 (red) of peritendinous tissue from WT mice injected with Veh or TPX0005, Prrx1 Hif1 α mice injected with Veh or NGF, Prrx1 VHL mice injected with TPX0005 at 14 days after modeling ( n = 6), and investigation analysis of ROM ( n = 6). Scale bars of of H&E and Masson staining: 300 μm. Scale bars of double-immunofluorescent staining: 100 μm. D , E Analysis of fibrosis percentage and ROM of WT mice, Prrx1 Hif1 α mice and Prrx1 VHL mice ( n = 6). F Analysis of Col1 + αSMA + cells of WT mice, Prrx1 Hif1 α mice and Prrx1 VHL mice ( n = 6). G IF of Prrx1 (red) cells and α-SMA (green) cells from four groups in vitro. Scale bars: 20 μm. H The diagram of sorting Prrx1 + MSCs from fibrosis tissues of four types of mice. I , J Protein expressions of Hif1α in fibrosis tissue of Prrx1 TrkA mice and Adv NGF mice ( n = 3). K , L Protein expressions of Twist1 in fibrosis tissue of WT mice, Prrx1 Hif1a mice, Prrx1 TrkA mice and Adv NGF mice ( n = 3). M Adv NGF mice, Prrx1 TrkA mice and Prrx1 Hif1a mice experimental procedure and phenotypic diagram in a skin wound fibrotic healing model. N , O Representative macroscopic illustration and macroscopic quantification of individual wound areas at D0-14 in Adv NGF mice, Prrx1 TrkA mice and Prrx1 Hif1a mice ( n = 6). P H&E stained sections of wound areas on D14 in Adv NGF mice, Prrx1 TrkA mice and Prrx1 Hif1a mice used for morphometric analysis of percentage of ( Q ) wound closure (length of newly formed epithelium (NFE)/length of NFE + length of gap between edges of wound epithelium (black dotted line) × 100), ( R ) area of HPE, ( S ) wound contraction (distance between wound border HFs (blue dotted line)), and ( T ) re-epithelialisation (length of NFE) ( n = 6). Yellow dashed line shows space between tendon and surrounding tissues. Black or white dashed line shows space occupied by fibrosis tissues. T represents tendon; AD represents fibrosis tissue; M represents muscle(mean ± SD; D – F : ANOVA with multiple comparisons, J / L / O / Q – T : unpaired t-test; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; A (mouse)/ B (mouse)/ H (mouse)/ M created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed; A (tendon, fibrosis tissue, MSCs and myofibroblasts)/ B (tendon, fibrosis tissue, MSCs and myofibroblasts)/ G (MSCs)/ H (tendon, fibrosis tissue and MSCs) created in BioRender. Author, li, y. (2025)) https://BioRender.com/0hlz1hf ).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Staining, Injection, In Vitro

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A Experimental procedure of WT mice injected with Veh or TPX-0005. B Representative macroscopic illustration of wound healing at D0-D14 in WT mice. C H&E analysis of D HPE area, E re-epithelialization, and F contraction ( n = 6). Boxed regions in granulation tissue (delineated by dotted lines) are shown at higher magnification in insets. ( n = 6). Scale bars: low power image is 500 μm, and the high power image is 100 μm. G H&E and Masson staining of peritendinous tissue from WT mice and investigation analysis of ROM ( n = 6). Scale bars: 300 μm. H Histological fibrosis percentage of peritendinous tissue from WT mice ( n = 6). I Quantitative analysis of ROM ( n = 6). J , K IF of Prrx1 (red) and α-SMA(green) cells in WT mice ( n = 6). Scale bars: 100 μm. L Relative mRNA expression of Col1 and Col3 in peritendinous tissue of WT mice ( n = 6). M Quantitative analysis of maximum load and stiffness of repaired tendons in normal WT mice ( n = 6). N Double IF of αSMA (green) and Col1 (red) in fibrosis tissue of Prrx1 TrkA mice and Nes TrkA mice ( n = 6). Scale bars: low power image is 100 μm, and the high power image is 50 μm. O Investigation analysis of ROM of Prrx1 TrkA mice and Nes TrkA mice ( n = 6). Black or white dashed line shows space occupied by fibrosis tissues. T represents tendon; AD represents fibrosis tissue; M represents muscle(mean ± SD; 8 D – F / H / I / K – O : unpaired t-test; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001; A (mouse) created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed; A (tendon) created in BioRender. Author, li, y. (2025)) https://BioRender.com/0hlz1hf ).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Injection, Staining, Expressing

Journal: Nature Communications

Article Title: Nociceptive sensory neuron-derived NGF orchestrates a fibrotic mesenchymal stromal cell neurogenic niche to drive tendon pathological fibrosis

doi: 10.1038/s41467-025-67396-y

Figure Lengend Snippet: A Nociceptive sensory nerves infiltrate the peripheral trauma area, mediating the biased repair response due to the formation of fibrotic mesenchymal stromal cells (MSCs) neurogenic niche. Mechanistically, nociceptive sensory neuron-derived nerve growth factor (NGF) activates the TrkA receptors in MSCs and triggers the Hif1α-Twist1 pathway, driving the differentiation of MSCs into myofibroblasts to mediate fibrosis. And TPX-0005 exerts therapeutic effects by inhibiting TrkA to reverse fibrosis (A(hand, DRG, nerve, microenvirenment) created in Adobe. Author, Yanhao Li. (2025). Contact the author for publication if needed; A(tendon, fibrosis tissue and other cells) created in BioRender. Author, li, y. (2025))ve sensory nerves infiltrate the peripheral trauma area, mediating the biased repair response due to the formation of fibrotic mesenchymal stromal cells (MSCs) neurogenic niche. Nociceptive sensory neuron-derived nerve growth factor activates the TrkA receptors in MSCs and triggers the Hif1α-Twist1 pathway, driving the differentiation of MSCs into myofibroblasts to mediate fibrosis. TPX-0005 exerts therapeutic effects by inhibiting TrkA to reverse fibrosis. https://BioRender.com/0hlz1hf ).

Article Snippet: The adhesion grading scale and healing grading scale were assessed as before. (Supplementary Fig. ; Supplementary Fig. ) The primary antibodies used in this section were as follows: α-smooth muscle actin (αSMA, Cell Signaling, 19245 and 48938, 1:200); calcitonin gene-related peptide (CGRP, Cell Signaling, 14959, 1:200); nerve growth factor (NGF, Abcam, 52918, 1:150); neuron nucleus (NeuN, Cell Signaling, 94403 s, 1:100); Nestin (Aves Labs, NES, 1:200); TrkA (BIOSS, bs-0193R, 1:200); and Hif1α (NOVUS, H1alpha67, 1:300); Prrx1(NOVUS, NBP1-06067, 1:200); Collagen 1(Abcam, 138492, 1:200 for IF, 1:500 for WB); Collagen 3(Abcam, 184993, 1:200 for IF, 1:500 for WB); PGP9.5(Cell Signaling, 60702, 1:200); VHL(Abcam, 140989, 1:500); Engrailed-1(Sigma, AB5732, 1:200).

Techniques: Derivative Assay

Journal: bioRxiv

Article Title: Induction of Human Pruriceptors from Pluripotent Stem Cells via Transcription Factors

doi: 10.1101/2025.06.11.659208

Figure Lengend Snippet: (A) Heatmap analysis on the samples from 2-step N1 and N1-I, showing the expression of indicated markers. Transcript expression values (TPM) were log10-transformed, and Z-scores were calculated. (B) Representative images showing neuronal morphology at different stages of differentiation under the 2-step N1-I, namely iPruriceptor, protocol on day12, 17, 21. Scale bars indicate 100μm. (C) Immunofluorescence staining of indicated markers on the induced neurons under iPruriceptor protocol on day 28. Scale bars indicate 50μm. The table shows the proportion of induced neurons under iPruriceptor protocol expressing the indicated markers on day 28. The percentages of TRPV1-, TrkA-, RET-, IL31ra-, or OSMR-positive cells among the β3 tubulin–positive population, which were stained simultaneously with each described marker, were shown.

Article Snippet: The cells were then incubated overnight at 4°C with the following primary antibodies diluted in blocking buffer: [anti-Tubb3 (Biolegend, 801201, 1:1000), anti-MAP2 (Abcam, AB5392,1:20000), anti-TRPV1 (Almone Lab, ACC-030,1:200), anti- Trka (Almone Lab, ANT-018, 1:200), anti-IL31RA (Abcam, AB113498, 1:200), anti- OSMR (LSBio, LS-B11477-50, 1:200), anti-RET (Novus biologicals, AF1485, 1:200)].

Techniques: Expressing, Transformation Assay, Immunofluorescence, Staining, Marker