Figure 5.

Mechano-stress initiates the positive loop of NF-κB and periostin via PIEZO1

(A) Immunoblotting results of three control and three degenerated human NP tissues. C, control; D, degenerated. (B) Quantitation of Ca²⁺ influx changes in hNPCs after 5 μM Yoda1 stimulation (left panel), and representative fluorescence images at indicated time points (right panel). Yoda1 was administrated at 10 s. (C) Representative images of Fluo-8 AM in hNPCs cultured on different hydrogels. Scale bar = 100 μm. (D) Representative β-Gal staining images of hNPCs treated with vehicle or Yoda1 for 24 h (left panel), and quantitation of β-Gal⁺ senescent cells (right panel). n = 3 biologically independent samples. Scale bar, 100 μm. (E–G) qPCR results of senescence (E), SASP (F), and matrix metabolism (G) markers in hNPCs treated with vehicle or Yoda1 for 24 h. n = 3 biologically independent samples. (H) Representative immunostaining images of NF-κB p65 (red) in hNPCs under indicated treatments for 30 min. DAPI (blue) indicates nuclei. (I) Quantification of p65 localization in (H), presented as mean ± SD. In total, 237 cells from the vehicle group and 232 cells from the Yoda1 group were analyzed. ∗Represents the statistical analysis was conducted in terms of the proportion of N > C between the two groups and the p < 0.05. n = 3 biologically independent samples. Scale bar = 25 μm. (J) Immunoblotting results of hNPCs treated with vehicle or Yoda1 for 48 h. (K and L) Representative β-Gal staining images of hNPCs under indicated conditions for 24 h (K), and quantitation of β-Gal⁺ senescent cells (L). n = 3 biologically independent samples. Scale bar, 100 μm. (M) Relative mRNA expression of the matrix metabolism and senescence markers and POSTN in hNPCs under indicated conditions. n = 3 biologically independent samples. (N and O) Representative T2WI image (N) and MRI Pfirrmann grade (O) of rat tail IVDs with indicated administration. n = 9 discs. Data are presented as the mean ± SD. ∗p < 0.05.