Abstract
Background
Liver fibrosis can progress to cirrhosis and hepatic carcinoma without treatment. CircDCBLD2 was found to be downregulated in liver fibrosis. However, the precise underlying mechanism requires further investigation.
Methods
qRT-PCR, Western blot, and immunohistochemistry assays were used to detect the related molecule levels. HE, Masson’s trichrome, and Sirius Red staining were used to assess the pathological changes in mice’s liver tissues. Flow cytometric analysis and commercial kit were used to assess the levels of lipid reactive oxygen species (ROS), malonaldehyde (MDA), glutathione (GSH), and iron. Cell viability was assessed by MTT. Immunoprecipitation was used to study the ubiquitination of PARK7. Mitophagy was determined by immunostaining and confocal imaging. RIP and Co-IP assays were used to assess the interactions of circDCBLD2/HuR, HuR/STUB1, and STUB1/PARK7. Fluorescence in situ hybridization and immunofluorescence staining were used to assess the co-localization of circDCBLD2 and HuR.
Results
CircDCBLD2 was downregulated, whereas PARK7 was upregulated in liver fibrosis. Ferroptosis activators increased circDCBLD2 while decreasing PARK7 in hepatic stellate cells (HSCs) and mice with liver fibrosis. CircDCBLD2 overexpression reduced cell viability and GSH, PARK7, and GPX4 expression in erastin-treated HSCs while increasing MDA and iron levels, whereas circDCBLD2 knockdown had the opposite effect. CircDCBLD2 overexpression increased STUB1-mediated PARK7 ubiquitination by promoting HuR-STUB1 binding and thus increasing STUB1 mRNA stability. PARK7 overexpression or HuR knockdown reversed the effects of circDCBLD2 overexpression on HSC activation and ferroptosis. CircDCBLD2 reduced liver fibrosis in mice by inhibiting PARK7.
Conclusion
CircDCBLD2 overexpression increased PARK7 ubiquitination degradation by upregulating STUB1 through its interaction with HuR, inhibiting HSC activation and promoting HSC ferroptosis, ultimately enhancing liver fibrosis.
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Abbreviations
- HSCs:
-
Hepatic stellate cells
- GSH:
-
Glutathione
- GPX4:
-
Glutathione peroxidase 4
- ROS:
-
Reactive oxygen species
- PARK7:
-
Parkinson disease 7
- STUB1:
-
STIP1 homology and U-Box containing protein 1
- circRNAs:
-
Circular RNAs
- FBS:
-
Fetal bovine serum
- shRNA:
-
Short hairpin RNA
- act-D:
-
Actinomycin D
- MTT:
-
3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide
- MDA:
-
Malondialdehyde
- RIP:
-
RNA immunoprecipitation
- CHX:
-
Cycloheximide
- Co-IP:
-
Co-immunoprecipitation
- CCl4 :
-
Carbon tetrachloride
- H&E:
-
Hematoxylin and eosin
- IHC:
-
Immunohistochemistry
- ALT:
-
Alanine aminotransferase
- AST:
-
Aspartate aminotransferase
- HYP:
-
Hydroxyproline
- qRT-PCR:
-
Quantitative real-time PCR
- PVDF:
-
Polyvinylidene difluoride
- HO-1:
-
Heme oxygenase-1
- IRP2:
-
Iron-regulatory protein 2
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Funding
This study was funded by grants from the National Natural Science Foundation of China (No. 82170639), the National Natural Science Foundation of China Youth Fund (No. 82200671), and the Hunan Provincial Natural Science Foundation (No. 2022JJ70068, 2022JJ70070).
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JW: conceptualization; methodology; HZ: visualization; supervision; LC: validation; formal analysis; KF: investigation; resources; YY: data curation; writing–original draft; ZL: writing—review and editing; project administration; funding acquisition.
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The Animal Care and Use Committee of Central South University’s Third **angya Hospital approved all mouse-related experiments.
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Wang, J., Zhang, H., Chen, L. et al. CircDCBLD2 alleviates liver fibrosis by regulating ferroptosis via facilitating STUB1-mediated PARK7 ubiquitination degradation. J Gastroenterol 59, 229–249 (2024). https://doi.org/10.1007/s00535-023-02068-6
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DOI: https://doi.org/10.1007/s00535-023-02068-6