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Citrate and hydroxycinnamate derivatives from Mume Fructus protect LPS-injured intestinal epithelial cells by
Zhihua Liu1, Zhengxu Zhang1, Xiaonan Chen1
1School of Pharmacy, Shanghai Jiao Tong University, Shanghai, 800 Dongchuan Road, Shanghai, 200240, People's Republic of China.
Ethnopharmacological Relevance:
Mume Fructus (MF) is processed from the near-ripe fruit of Prunus mume (Siebold) Siebold & Zucc by drying at low temperature until the color turns black. MF is often used in Chinese medicine for the treatment of chronic diarrhea and dysentery. Previous studies have shown that the active components of MF against Crohn's disease (CD) are mainly citrate and hydroxycinnamate derivatives, which can alleviate the CD-induced inflammatory response and intestinal barrier damage. However, their molecular mechanisms on CD still need further elucidation.
Aim Of The Study:
To investigate the protective effects and underlying mechanisms of citrate and hydroxycinnamate derivatives in MF on intestinal epithelial injury.
Materials And Methods:
Network pharmacology technology was used to predict the anti-CD targets and molecular mechanisms of 4 citrate and 11 hydroxycinnamate derivative prototypes and 5 hydroxycinnamate derivative metabolites in the 40% ethanol fraction of MF (MFE40), the active anti-CD ingredient group of MF. Lipopolysaccharide (LPS)-treated IEC-6 cells were used to investigate the effects of the above components on the proliferation of damaged IEC-6 cells and to verify the molecular mechanism of their regulation on the FAK/PI3K/AKT signaling pathways for the promotion of the proliferation of IEC-6 cells.
Results:
A "compound-target-pathway" network was constructed based on network pharmacology analysis, including 20 citrate and hydroxycinnamate derivatives that target 316 core proteins and 36 CD-related pathways, of which PI3K-AKT pathway and focal adhesion were the most enriched pathways. Further cell validation experiments showed that 1 citric acid (CA) compound and 10 hydroxycinnamate derivatives, including 3-O-caffeoylquinic acid (3CQA), 4-O-caffeoylquinic acid (4CQA), 5-O-caffeoylquinic acid (5CQA), caffeic acid (CFA), p-coumaric acid (PCMA), m-coumaric acid (MCMA), ferulic acid (FUA), isoferulic acid (IFUA), 3-hydroxyphenylpropionic acid (3HPPA) and hippuric acid (HPP), could promote the proliferation of IEC-6 cells and inhibit the damage of LPS to IEC-6 cells. Ethyl caffeate (ECFA), a hydroxycinnamic acid derivative, had no effect on promoting the proliferation of IEC-6 cells and was weak in inhibiting the damage of IEC-6 cells caused by LPS. Further mechanistic verification experiments showed that 7 citrate and hydroxycinnamate derivatives (CA, CFA, 3CQA, MCMA, FUA, 3HPPA, and HPP) could upregulate the expression of p-FAK, p-PI3K, and p-AKT proteins. Among them, CA had the better effect on activating the FAK-PI3K-AKT signaling pathway.
Conclusions:
Citrate and hydroxycinnamate derivatives in MF can ameliorate LPS-induced intestinal epithelial cell injury to demonstrate potential for Crohn's disease alleviation. This protective effect can be achieved by upregulating FAK/PI3K/AKT pathway.
Insights
Citrate and hydroxycinnamate derivatives from Mume Fructus (MF) protect intestinal cells from injury. These compounds, particularly citric acid (CA), promote cell proliferation and repair by activating the FAK/PI3K/AKT pathway, offering potential for Crohn
Area of Science:
- Pharmacology and Traditional Medicine
- Gastroenterology and Inflammatory Bowel Disease Research
- Molecular Biology and Cell Signaling
Background:
- Mume Fructus (MF), a traditional Chinese medicine, contains citrate and hydroxycinnamate derivatives known to mitigate inflammatory responses and intestinal barrier damage.
- Previous research indicates these compounds are key active components in MF for managing Crohn's disease (CD), but their precise molecular mechanisms require further investigation.
- This study focuses on elucidating the protective effects and underlying mechanisms of these derivatives in MF on intestinal epithelial injury relevant to CD.
Purpose of the Study:
- To investigate the protective effects of citrate and hydroxycinnamate derivatives from Mume Fructus (MF) on intestinal epithelial injury.
- To elucidate the molecular mechanisms by which these MF derivatives exert their protective effects, particularly concerning the FAK/PI3K/AKT signaling pathway.
- To evaluate the potential of these compounds as therapeutic agents for conditions like Crohn's disease.
Main Methods:
- Network pharmacology was employed to predict anti-CD targets and molecular mechanisms of identified citrate and hydroxycinnamate derivatives within MF.
- Lipopolysaccharide (LPS)-induced IEC-6 cell injury model was used to assess the effects of these compounds on cell proliferation and damage.
- Western blotting was utilized to verify the regulation of the FAK/PI3K/AKT signaling pathway by the active compounds.
Main Results:
- Network pharmacology identified 20 citrate and hydroxycinnamate derivatives targeting 316 proteins across 36 CD-related pathways, with PI3K-AKT and focal adhesion pathways being most enriched.
- Cell experiments confirmed that citric acid (CA) and 10 hydroxycinnamate derivatives promoted IEC-6 cell proliferation and inhibited LPS-induced damage, while ethyl caffeate showed limited efficacy.
- Seven compounds, including CA, caffeic acid (CFA), and 3-O-caffeoylquinic acid (3CQA), upregulated p-FAK, p-PI3K, and p-AKT, with CA demonstrating the strongest activation of the FAK/PI3K/AKT pathway.
Conclusions:
- Citrate and hydroxycinnamate derivatives in Mume Fructus (MF) effectively ameliorate LPS-induced intestinal epithelial injury.
- These protective effects are mediated through the upregulation of the FAK/PI3K/AKT signaling pathway.
- The findings support the potential of MF derivatives as a therapeutic strategy for alleviating symptoms associated with Crohn's disease.
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