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PINK1 overexpression suppresses p38 MAPK/NF‑κB signaling to attenuate chondrocyte senescence in osteoarthritis

  • Authors:
    • Lishi Jie
    • Yuanhui Zhang
    • Jiangyu Liu
    • Houyu Fu
    • Zaishi Zhu
    • Zeling Huang
    • Xiaoqing Shi
    • Peimin Wang
    • Songjiang Yin
  • View Affiliations / Copyright

    Affiliations: Department of Orthopedics, The Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210029, P.R. China, The First College of Clinical Medicine, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, P.R. China, Department of Orthopaedics and Traumatology, Suzhou Hospital of Traditional Chinese Medicine, Suzhou, Jiangsu 215031, P.R. China
    Copyright: © Jie et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 224
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    Published online on: October 15, 2025
       https://doi.org/10.3892/ijmm.2025.5665
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Abstract

PTEN‑induced putative kinase 1 (PINK1), a master regulator of mitophagy, is implicated in mitochondrial homeostasis, yet its role in knee osteoarthritis (OA) pathogenesis remains unclear. The present study investigated the mechanisms by which PINK1 modulates chondrocyte senescence during OA progression. Utilizing a destabilization of the medial meniscus‑induced OA murine model, decreased PINK1 expression, impaired mitochondrial function and suppressed mitophagy were observed in OA cartilage. In vitro, lipopolysaccharide‑induced chondrocyte senescence was exacerbated by PINK1 knockdown but mitigated by PINK1 overexpression, which restored mitophagy and reduced senescence‑associated β‑galactosidase activity, reactive oxygen species accumulation and mitochondrial membrane potential collapse. RNA sequencing and mechanistic studies identified the p38 MAPK/NF‑κB pathway as a downstream target; PINK1 knockdown amplified the phosphorylation of p38 MAPK/NF‑κB, promoting mitochondrial dysfunction and senescence. By contrast, pharmacological inhibition of p38 MAPK/NF‑κB rescued these effects in PINK1‑deficient chondrocytes. Collectively, PINK1 attenuated OA progression by suppressing chondrocyte senescence via inhibition of the p38 MAPK/NF‑κB pathway, highlighting its potential as a therapeutic target for OA management.
View Figures

Figure 1

Impaired mitochondrial autophagy and
enhanced cell senescence in KOA cartilage. (A) Representative
microcomputed tomography 3D reconstructions of knee joints from
sham-operated and destabilization of the Medial Meniscus)induced
KOA mice 14 days post-surgery. (B) Histopathological evaluation and
OARSI scoring of knee joint sections (n=6), stained with HE and SF.
Scale bar, 50 μm. (C) Western blot analysis and densitometry
of senescence-associated markers (MMP3, iNOS, p21, p16) in
cartilage, normalized to GAPDH (n=3). (D) Serum levels of
proinflammatory cytokines (TNF-α, IL-6, IL-1β) measured by ELISA
using samples collected via cardiac puncture (n=6). (E) Western
blot analysis and densitometry of mitophagy-related proteins
(PINK1, LC3B ratio, p62, BNIP3L, TUFm) in cartilage, normalized to
GAPDH (n=3). ***P<0.001 vs. sham. KOA, knee
osteoarthritis; HE, hematoxylin and eosin; TNF-α, tumor necrosis
factor-α; PINK1, PTEN-induced putative kinase 1; TUFm, Tu
translation elongation factor; OARSI, Osteoarthritis Research
Society International; SO/FG, Safranine O-Fast Green; iNOS,
inducible nitric oxide synthase; BNIP3L, BCL2-interacting Protein 3
Like; SMI, structure model index of trabeculae; BV/TV, bone volume
fration.

Figure 2

PINK1 overexpression decreases
senescence and promotes mitophagy in chondrocytes. (A) Western blot
analysis and densitometric quantification of mitophagy markers
(PINK1, LC3B, p62, BNIP3L, TUFm) in primary mouse chondrocytes
following transduction with Lv-PINK1, followed by LPS treatment (1
μg/ml, 24 h). Protein signals were normalized to GAPDH. (B)
Mitochondrial membrane potential was assessed using JC-1 staining.
Red, healthy mitochondria (J-aggregates); green, depolarized
mitochondria (J-monomers). (C) Levels of intracellular ROS were
detected using the DCFH-DA probe. Green fluorescence intensity was
quantified in ≥100 cells/group. Scale bar, 20 μm. (D)
Cellular senescence (blue) was evaluated by SA-β-gal staining. The
percentage of SA-β-gal-positive cells was quantified (scale bar, 50
μm). (E) Western blot quantification of
senescence-associated secretory phenotype markers (MMP3, iNOS) and
cell cycle inhibitors (p21, p16). Protein signals were normalized
to GAPDH. n=3. ***P<0.001 vs. NC;
∆∆∆P<0.001 vs. LPS. PINK1, PTEN-induced putative
kinase 1; TUFm, Tu translation elongation factor; ROS, reactive
oxygen species; SA-β-gal, senescence-associated β-galactosidase;
LPS, lipopolysaccharide; BNIP3L, BCL2-interacting Protein 3 Like;
Lv, Lentiviral Vector; iNOS, inducible nitric oxide synthase; NC,
negative Control.

Figure 3

Transcriptomic analysis of PINK1
overexpression in OA chondrocytes. (A) Correlation analysis, (B)
PCA and (C) number and (D) heatmap of differentially expressed
genes between groups. Top 10 differentially expressed genes in (E)
KOA vs. con and (F) Lv-PINK1 vs. KOA group. KEGG enrichment
analysis (G) donut chart and (H) bubble plot of differentially
expressed genes in the KOA vs. con group. (I) GSEA of the MAPK
signaling pathway in the KOA vs. con group. KEGG enrichment
analysis (J) donut chart and (K) bubble plot of differentially
expressed genes in the Lv-PINK1 vs. KOA group. (L) GSEA of the MAPK
signaling pathway in the Lv-PINK1 vs. KOA group. (M) Changes in
MAPK signaling pathway-related target genes in the KOA vs. con and
(N) Changes in MAPK signaling pathway-related target genes in the
Lv-PINK1 vs. KOA group. PINK1, PTEN-induced putative kinase 1; PCA,
principal component analysis; KEGG, Kyoto Encyclopedia of Genes and
Genomes; GSEA, gene set enrichment analysis; KOA, knee
osteoarthritis; Lv, lentiviral vector; con, control.

Figure 4

Activation of the p38 MAPK/NF-κB
pathway induces senescence and mitochondrial damage in LPS-treated
chondrocytes. (A) ELISA was performed to detect the levels of TNF-α
and IL-6 in the culture medium of immortalized human chondrocytes.
(B) Western blotting was performed to detect the protein expression
of MMP3, iNOS, p21 and p16 in immortalized human chondrocytes.
Representative (C) JC-1 staining showing mitochondrial membrane
potential in immortalized human chondrocytes. (D) DCFH-DA staining
showing ROS levels in immortalized human chondrocytes. (E) Western
blotting was performed to detect the protein expression of PINK1,
LC3B, p62, BINPL and TUFm in immortalized human chondrocytes. (F)
Representative SA-β-Gal staining showing senescence levels in each
group of immortalized human chondrocytes. (G) Western blotting was
performed to detect the protein expression levels of p-p65, p65,
p-p38 and p38. n=3. **P<0.01,
***P<0.001 vs. sham; ∆∆∆P<0.001 vs.
LPS. LPS, lipopolysaccharide; ROS, reactive oxygen species; PINK1,
PTEN-induced putative kinase 1; TUFm, Tu translation elongation
factor; SA-β-gal, senescence-associated β-galactosidase; iNOS,
inducible nitric oxide synthase; p-, phosphorylated; BINP3L, BCL2
Interacting Protein 3 Like.

Figure 5

PINK1 gene knockdown enhances p38
MAPK/NF-κB phosphorylation and exacerbates chondrocyte senescence.
(A) Adenoviral infection efficiency of immortalized human
chondrocytes observed under a fluorescence microscope. Western blot
analysis of (B) PINK1, LC3B, p62, BINPL and TUFm and (C) p-p65,
p65, p-p38 and p38 protein expression. Representative (D) JC-1
staining showing mitochondrial membrane potential and (E) DCFH-DA
staining showing ROS levels in immortalized human chondrocytes. (F)
Western blot analysis of MMP3, iNOS, p21 and p16 protein expression
in immortalized human chondrocytes. (G) Representative SA-β-Gal
staining showing senescence levels in immortalized human
chondrocytes. n=3. **P<0.01, ***P<0.001
vs. sham; ∆P<0.05, ∆∆P<0.01,
∆∆∆P<0.001 vs. LPS. PINK1, PTEN-induced putative
kinase 1; TUFm, Tu translation elongation factor; ROS, reactive
oxygen species; SA-β-Gal, senescence-associated β-galactosidase;
BINP3L, BCL2-interacting Protein 3 Like; p-, Phosphorylation; iNOS,
inducible nitric oxide synthase; LPS, lipopolysaccharide; NC,
Negative Control; sh, short hairpin; BF, Bright field.

Figure 6

Inhibition of p38 MAPK/NF-κB pathway
ameliorates senescence and mitochondrial damage in PINK1-silenced
chondrocytes. (A) Western blot analysis of protein expression of
p-p65, p65, p-p38 and p38. (B) Representative JC-1 assay showing
mitochondrial membrane potential and (C) DCFH-DA staining showing
ROS levels, (D) Western blot analysis of protein expression of
MMP3, iNOS, p21 and p16 and (E) representative SA-β-Gal staining
showing senescence levels in immortalized human chondrocytes. n=3.
**P<0.01, ***P<0.001 vs. sham;
#P<0.05, ##P<0.01,
###P<0.001 vs. LPS; ∆P<0.05,
∆∆P<0.01, ∆∆∆P<0.001 vs. diprovocim.
ROS, reactive oxygen species; PINK1, PTEN-induced putative kinase
1; SA-β-gal, senescence-associated β-galactosidase; p-,
Phosphorylation; iNOS, inducible nitric oxide synthase; sh, short
hairpin; NC, Negative Control; LPS, Lipopolysaccharide.
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Copy and paste a formatted citation
Spandidos Publications style
Jie L, Zhang Y, Liu J, Fu H, Zhu Z, Huang Z, Shi X, Wang P and Yin S: PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis. Int J Mol Med 56: 224, 2025.
APA
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z. ... Yin, S. (2025). PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis. International Journal of Molecular Medicine, 56, 224. https://doi.org/10.3892/ijmm.2025.5665
MLA
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z., Shi, X., Wang, P., Yin, S."PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis". International Journal of Molecular Medicine 56.6 (2025): 224.
Chicago
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z., Shi, X., Wang, P., Yin, S."PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis". International Journal of Molecular Medicine 56, no. 6 (2025): 224. https://doi.org/10.3892/ijmm.2025.5665
Copy and paste a formatted citation
x
Spandidos Publications style
Jie L, Zhang Y, Liu J, Fu H, Zhu Z, Huang Z, Shi X, Wang P and Yin S: PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis. Int J Mol Med 56: 224, 2025.
APA
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z. ... Yin, S. (2025). PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis. International Journal of Molecular Medicine, 56, 224. https://doi.org/10.3892/ijmm.2025.5665
MLA
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z., Shi, X., Wang, P., Yin, S."PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis". International Journal of Molecular Medicine 56.6 (2025): 224.
Chicago
Jie, L., Zhang, Y., Liu, J., Fu, H., Zhu, Z., Huang, Z., Shi, X., Wang, P., Yin, S."PINK1 overexpression suppresses p38 MAPK/NF‑&kappa;B signaling to attenuate chondrocyte senescence in osteoarthritis". International Journal of Molecular Medicine 56, no. 6 (2025): 224. https://doi.org/10.3892/ijmm.2025.5665
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