Inhibition of cc chemokine receptor 10 ameliorates osteoarthritis via inhibition of the phosphoinositide-3-kinase/Akt/mammalian target of rapamycin pathway

Background
Osteoarthritis (OA) is a joint disorder characterized by inflammation and the progressive breakdown of cartilage. A key pathological feature of OA is the apoptosis of chondrocytes. Interleukin-1β (IL-1β), a pro-inflammatory cytokine, plays a significant role in promoting cartilage degradation and is widely used to stimulate primary human chondrocytes in vitro, establishing an OA model. Additionally, IL-1β contributes to OA pathogenesis by activating the phosphoinositide-3-kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK) signaling pathways.

CCR10, a G-protein-coupled receptor involved in various cancers, has an unclear role in OA progression. This study aimed to explore the protective role of CCR10 in IL-1β-stimulated CHON-001 cells and uncover the underlying molecular mechanisms.

Methods
CHON-001 cells were transfected with either control small interfering RNA (siRNA) or CCR10-siRNA for 24 hours, followed by stimulation with 10 ng/mL IL-1β for 12 hours to create an in vitro OA model. Quantitative reverse transcription PCR and western blotting were used to measure the expression levels of CCR10, cleaved-caspase-3, MMP-3, MMP-13, Collagen II, Aggrecan, phosphorylated-PI3K (p-PI3K), PI3K, phosphorylated-Akt (p-Akt), Akt, phosphorylated-mTOR (p-mTOR), and mTOR. Cell viability, cytotoxicity, and apoptosis were evaluated using the MTT assay, lactate dehydrogenase (LDH) release assay, and flow cytometry, respectively. Levels of inflammatory cytokines (TNF-α, IL-6, and IL-8) were measured by enzyme-linked immunosorbent assay (ELISA).

Results
IL-1β stimulation significantly increased CCR10 levels in CHON-001 cells compared to the control group, but CCR10 expression was effectively reduced in CCR10-siRNA-transfected cells. Suppression of CCR10 mitigated IL-1β-induced inflammatory damage, as evidenced by improved cell viability, reduced LDH release, lower apoptotic cell counts, and decreased cleaved-caspase-3 PDGFR 740Y-P expression. Furthermore, CCR10 inhibition reversed the IL-1β-induced elevation of TNF-α, IL-6, IL-8, MMP-3, and MMP-13, while restoring the levels of Collagen II and Aggrecan. These protective effects were negated upon treatment with the PI3K agonist 740Y-P.

Moreover, IL-1β stimulation activated the PI3K/Akt/mTOR signaling pathway, but this activation was suppressed by CCR10-siRNA. Treatment with 740Y-P restored the pathway’s activation despite CCR10 inhibition.

Conclusion
CCR10 inhibition alleviates IL-1β-induced chondrocyte injury by suppressing the PI3K/Akt/mTOR signaling pathway, highlighting CCR10 as a potential therapeutic target for OA treatment.