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Ca² + and endothelin-1 (ET-1) contribute to Piezo1-dependent activation of hypoxia-inducible factor 1-alpha (HIF-1α). a) and b) Gene set enrichment analysis (GSEA) of epidural scar tissue from mice after laminectomy. c) Ca² + influx measured by Fluo-4 AM in the blank, type I rat tail collagen-encapsulated, and collagen-encapsulated with GsMTX4 groups. d) Western blot analysis of NIH/3T3 fibroblasts treated with type I rat tail collagen and the calcium influx blocker Nimodipine (n = 3). e) Western blot analysis of NIH/3T3 fibroblasts treated with Yoda1 and Nimodipine (n = 3). f) Quantitative analysis of relative Edn1 messenger RNA (mRNA) expression in blank and collagen-encapsulated NIH/3T3 fibroblasts. g) and h) Western blot analysis of NIH/3T3 fibroblasts treated with type I rat tail collagen and the ET-1 inhibitor Bosentan (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001, independent-samples t -test (two groups) or analysis of variance (multiple groups).

Journal: Bone & Joint Research

Article Title: Piezo1 drives fibroblast activation in epidural fibrotic remodelling via the ET-1/HIF-1α pathway

doi: 10.1302/2046-3758.158.BJR-2025-0662.R1

Figure Lengend Snippet: Ca² + and endothelin-1 (ET-1) contribute to Piezo1-dependent activation of hypoxia-inducible factor 1-alpha (HIF-1α). a) and b) Gene set enrichment analysis (GSEA) of epidural scar tissue from mice after laminectomy. c) Ca² + influx measured by Fluo-4 AM in the blank, type I rat tail collagen-encapsulated, and collagen-encapsulated with GsMTX4 groups. d) Western blot analysis of NIH/3T3 fibroblasts treated with type I rat tail collagen and the calcium influx blocker Nimodipine (n = 3). e) Western blot analysis of NIH/3T3 fibroblasts treated with Yoda1 and Nimodipine (n = 3). f) Quantitative analysis of relative Edn1 messenger RNA (mRNA) expression in blank and collagen-encapsulated NIH/3T3 fibroblasts. g) and h) Western blot analysis of NIH/3T3 fibroblasts treated with type I rat tail collagen and the ET-1 inhibitor Bosentan (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001, independent-samples t -test (two groups) or analysis of variance (multiple groups).

Article Snippet: Alternatively, the following drugs were used to stimulate NIH/3T3 cells: Yoda1 (25 μM; HY-P1410), a chemical activator of Piezo1; BAY 87-2243 (1 μM; HY-15836), an inhibitor of hypoxia-inducible factor 1-alpha (HIF-1α); SB-431542 (10 μM; HY-10431), an inhibitor of TGF-β1; SIS 3 (10 μM; HY-13013), an inhibitor of Smad3; GsMTX4 (2.5 μM; HY-P1410); Nimodipine (60 μM; HY-B0265), a calcium channel blocker; and Bosentan (1 μM; HY-A0013, all MedChemExpress), an endothelin-1 (ET-1) receptor antagonist.

Techniques: Activation Assay, Western Blot, Expressing