Acrylamide induces human chondrocyte cell death by initiating autophagy‑dependent ferroptosis

Wang,Hui; Tang,Zizheng; Liu,Shasha; Xie,Kangqi; Zhang,Hua

doi:10.3892/etm.2023.11945

Acrylamide induces human chondrocyte cell death by initiating autophagy‑dependent ferroptosis

Authors:
- Hui Wang
- Zizheng Tang
- Shasha Liu
- Kangqi Xie
- Hua Zhang
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Affiliations: Department of Rheumatology, The Fourth Hospital of Jinan, Jinan, Shandong 250031, P.R. China, Department of Rheumatology, Zaozhuang Municipal Hospital (Affiliated Hospital of Jining Medical College), Zaozhuang, Shandong 277102, P.R. China
Published online on: April 12, 2023 https://doi.org/10.3892/etm.2023.11945
Article Number: 246

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Abstract

Acrylamide (ACR) is formed during heat treatment of foodstuffs and ACR may serve as a probable malignant neoplastic disease agent in all organs and tissues of the human body. However, it is unknown if ACR is associated with ankylosing spondylitis (AS) pathogenesis. Cell viability and proliferation were determined using CCK‑8 assay and EdU staining. Flow cytometry was used to determine cell death and cell cycle arrest. Intracellular lipid reactive oxygen species, Fe2+ and mitochondrial membrane potential (MMP) were analyzed using a C11‑BODIPY581/591 fluorescent probe, FerroOrange staining and a JC‑1 MMP Assay kit, respectively. The present study showed that ACR decreased chondrocyte cell viability in a dose‑dependent manner and that ACR significantly promoted chondrocyte senescence. ACR also elevated the expression of cell cycle arrest‑associated proteins, including p53, cyclin‑dependent kinase inhibitor 1 and cyclin‑dependent kinase inhibitor protein, in human chondrocytes. Similarly, DNA damage was also enhanced following ACR treatment in chondrocytes. In addition, the ferroptosis‑specific inhibitor ferrostatin‑1 (Fer‑1) and the autophagy inhibitor 3‑methyladenine abolished ACR‑induced cell death in chondrocytes. ACR was shown to activate autophagic flux and induce mitochondrial dysfunction by increasing the MMP. Western blot analysis of ferroptosis‑related proteins demonstrated that ACR decreased the expression of glutathione peroxidase 4, solute carrier family 7 member 11, transferrin receptor protein 1 and ferritin heavy chain 1 in chondrocytes whereas Fer‑1 abolished these effects. ACR treatment significantly elevated the phosphorylation levels of AMP‑activated protein kinase (AMPK) and serine/threonine‑protein kinase ULK1 in human chondrocytes. Notably, the effect of ACR was diminished by knockdown of AMPK, as evidenced by reduced lipid reactive oxygen species accumulation and Fe²⁺ levels. Hence, ACR inhibited cell proliferation and contributed to cell death by inducing autophagy‑dependent ferroptosis while promoting autophagy by activating AMPK‑ULK1‑mTOR signaling in human chondrocytes. It was hypothesized that the presence of ACR in foodstuffs may increase the risk of AS and that decreasing ACR in food products is of importance.

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