Abstract
Small intestine damage caused by diclofenac is called diclofenac enteropathy. Berberine (BBR), a class of isoquinoline alkaloids derived from Berberis vulgaris and Phellodendron amurense, is widely used in intestinal diseases. The present study evaluated the protective effect of BBR on the intestinal mucosal mechanical barrier in diclofenac enteropathy and its possible action mechanism. The in vitro animal experiment revealed that BBR downregulated the expression of long non-coding RNA H19 (lncRNA H19) in the small intestine and exosomes. In the co-culture experiment involving exosomes and intestinal epithelial cell-6 (IEC-6) cells, the results of qRT-PCR, western blotting, and immunofluorescence assays demonstrated that the elevated expression of lncRNA H19 in the small intestine, conveyed via exosomes derived from the diclofenac group, suppressed the expression levels of autophagy-associated protein 5 (Atg 5) and light chain 3 (LC 3), as well as and the tight junction (TJ) proteins zonula occludens-1 (ZO-1), claudin-1, and occluding, relative to the control group. BBR treatment attenuated exosomal lncRNA H19 levels, upregulated the expression of Atg5 and LC3 expression, enhanced TJ protein expression, and increased the light chain 3 (LC3)-II/LC3-I ratio. These findings significantly elucidated that BBR promoted the restoration of autophagy in IECs by inhibiting exosomal lncRNA H19, thereby mitigating the impairment of the intestinal mucosal mechanical barrier function in diclofenac enteropathy. The process involving exosomal lncRNA H19 regulating autophagy, thereby affecting the intestinal mucosal mechanical barrier, offers a novel perspective for the application of BBR in the treatment of diclofenac enteropathy.
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Data availability
Data supporting the findings of this study are available from the corresponding author [Shanshan Chen] on request.
Abbreviations
- BBR:
-
Berberine
- LncRNA H19:
-
Long non-coding RNA H19
- IEC:
-
Intestinal epithelial cell
- Atg 5:
-
Autophagy-associated protein 5
- TJ:
-
Tight junction
- ZO-1:
-
Zonula occludens-1
- LC 3:
-
Light chain 3
- TLR4:
-
Toll-like receptor 4
- NF-κ B:
-
Nuclear factor kappa B
- MTORC:
-
Mammalian target of rapamycin complex
- ADA:
-
Adenosine deaminase
- HPLC:
-
High-performance liquid chromatography
- SD:
-
Sprague Dawley
- H&E:
-
Hematoxylin and eosin
- PBS:
-
Phosphate-buffered saline
- DTT:
-
Dithiothreitol
- BCA:
-
Bicinchoninic acid
- SDS-PAGE:
-
Sodium dodecyl sulfate–polyacrylamide gel electrophoresis
- PVDF:
-
Polyvinylidene fluoride
- BSA:
-
Blocking buffer
- ECL:
-
Electrochemiluminescent
- DMEM:
-
Dulbecco’s modified Eagle’s medium
- FBS:
-
Fetal bovine serum
- shRNA:
-
Short hairpin RNA
- IF:
-
Immunofluorescence
- qRT-PCR:
-
Quantitative real-time polymerase chain reaction
- DAPI:
-
4′6-Diamidino-2-phenylindole
- ANOVA:
-
Analysis of variance
- SD:
-
Standard deviation
- LSD:
-
Least significant difference
- PPIs:
-
Proton pump inhibitors
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Acknowledgements
We thank the Zhejiang Chinese Medical University for its experimental support and research facility.
Funding
This work was supported by the Zhejiang Medicine and Health Science and Technology Plan Project (2023KY850, 2021KY834), Zhejiang Traditional Chinese Medicine Administration (2023ZL417), National Natural Science Foundation (82074214) and Leading Talents of Scientific and Technological Innovation in the “ten thousand talents plan” of Zhejiang Province [2020R52024].
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Ruonan He conducted the experiment, performed data analysis, and writing—original draft, review and editing. Ying Li and Yi He performed data analysis and writing—review and editing. Qianqian Wang conducted the experiment and performed data analysis. Shanshan Chen and Shuo Zhang conceived and designed the experiments, writing—review and editing, and project administration. All authors read and approved the final manuscript.
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The animal study was approved by the Institutional Animal Care and Use Committee of Zhejiang Chinese medical university (Approval No. IACUC-20210712-10). All animal care (including the euthanasia procedure) was done with the guidelines of the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC) and the Institutional Animal Care and Use Committee (IACUC).
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He, R., Li, Y., He, Y. et al. Berberine mitigates diclofenac-induced intestinal mucosal mechanical barrier dysfunction through the restoration of autophagy by inhibiting exosome-mediated lncRNA H19. Inflammopharmacol (2024). https://doi.org/10.1007/s10787-024-01487-y
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DOI: https://doi.org/10.1007/s10787-024-01487-y