Long non-coding RNA LINC-PINT: emerging clinical significance and molecular mechanisms in knee osteoarthritis

The underlying pathophysiological mechanisms of knee osteoarthritis (KOA) remain incompletely understood. This study aimed to investigate the clinical significance of long non-coding RNA LINC-PINT in KOA and elucidate its molecular mechanism in regulating chondrocyte function via the miR-324-3p/GPX4 axis.

100 KOA patient samples and 40 normal knee cartilage tissue samples were collected, and the relevant gene expression was quantified using RT-qPCR. C28/I2 was cultured in vitro to establish an IL-1β-induced KOA cellular model. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were performed to validate the targeted molecular regulatory interactions. Cell proliferation and apoptosis, as well as inflammatory factors levels, were assessed using cell counting kit-8 (CCK-8), flow cytometry, and enzyme-linked immunosorbent assay (ELISA), respectively. Oxidative stress-related indicators and Fe²⁺ concentration were evaluated using commercial assay kits. Western blotting was performed to detect protein expression.

In KOA tissues, LINC-PINT and glutathione peroxidase 4 (GPX4) were significantly downregulated, whereas miR-324-3p was markedly upregulated. LINC-PINT demonstrated the potential diagnostic discrimination ability and was identified as an independent protective factor in KOA. Overexpression of LINC-PINT effectively counteracted interleukin-1 beta (IL-1β)-induced suppression of chondrocyte proliferation, reduced apoptosis, and attenuated the secretion of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), while significantly alleviating cartilage matrix degradation, as indicated by increased expression of collagen II and aggrecan and decreased matrix metallopeptidase 13 (MMP13) levels. Moreover, LINC-PINT directly bound to miR-324-3p, which in turn directly targeted GPX4. Functional rescue assays further confirmed that transfection with a miR-324-3p mimic reversed the protective effects of LINC-PINT overexpression on chondrocytes, whereas GPX4 overexpression mitigated the detrimental effects induced by the miR-324-3p mimic. IL-1β disrupted the cellular redox balance by increasing the levels of Fe²⁺, reactive oxygen species (ROS), and (malondialdehyde) MDA and by inhibiting superoxide dismutase (SOD) activity and the GSH/GSSG ratio. Overexpression of LINC-PINT can counteract these effects, and the protective effect was enhanced when combined with Per-1. Transfection with miR-mimic reversed the protective effect of LINC-PINT, suggesting that LINC-PINT may act as a competitive endogenous RNA (ceRNA) to capture this pro-oxidative miR-324-3p. Finally, overexpression of GPX4 could protect cells from miRNA-induced damage, highlighting its crucial role in alleviating iron-dependent oxidative stress and iron-overloaded apoptosis.

LINC-PINT demonstrated the potential diagnostic discrimination ability and exerted a protective effect in KOA by modulating the miR-324-3p/GPX4 axis.