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Published on: July 26, 2017
Demethyleneberberine alleviated the inflammatory response by targeting MD-2 to inhibit the TLR4 signaling
Yaxing Zhao1,2, Peng Liu3, Haofan Luan1
1State Key Laboratory of Natural Medicines, Department of Biochemistry, China Pharmaceutical University, Nanjing, China.
Introduction:
The colitis induced by trinitrobenzenesulfonic acid (TNBS) is a chronic and systemic inflammatory disease that leads to intestinal barrier dysfunction and autoimmunedisorders. However, the existing treatments of colitis are associated with poor outcomes, and the current strategies remain deep and long-time remission and the prevention of complications. Recently, demethyleneberberine (DMB) has been reported to be a potential candidate for the treatment of inflammatory response that relied on multiple pharmacological activities, including anti-oxidation and antiinflammation. However, the target and potential mechanism of DMB in inflammatory response have not been fully elucidated.
Methods:
This study employed a TNBS-induced colitis model and acute sepsis mice to screen and identify the potential targets and molecular mechanisms of DMB in vitro and in vivo. The purity and structure of DMB were quantitatively analyzed by high-performance liquid chromatography (HPLC), mass spectrometry (MS), Hydrogen nuclear magnetic resonance spectroscopy (1H-NMR), and infrared spectroscopy (IR), respectively. The rats were induced by a rubber hose inserted approximately 8 cm through their anus to be injected with TNBS. Acute sepsis was induced by injection with LPS via the tail vein for 60 h. These animals with inflammation were orally administrated with DMB, berberine (BBR), or curcumin (Curc), respectively. The eukaryotic and prokaryotic expression system of myeloid differentiation protein-2 (MD-2) and its mutants were used to evaluate the target of DMB in inflammatory response.
Resluts:
DMB had two free phenolic hydroxyl groups, and the purity exceeded 99% in HPLC. DMB alleviated colitis and suppressed the activation of TLR4 signaling in TNBS-induced colitis rats and LPS-induced RAW264.7 cells. DMB significantly blocked TLR4 signaling in both an MyD88-dependent and an MyD88-independent manner by embedding into the hydrophobic pocket of the MD-2 protein with non-covalent bonding to phenylalanine at position 76 in a pi-pi T-shaped interaction. DMB rescued mice from sepsis shock induced by LPS through targeting the TLR4-MD-2 complex.
Conclusion:
Taken together, DMB is a promising inhibitor of the MD-2 protein to suppress the hyperactivated TLR4 signaling in inflammatory response.
Insights
Demethyleneberberine (DMB) effectively treats colitis and sepsis by inhibiting the Toll-like receptor 4 (TLR4)-myeloid differentiation protein-2 (MD-2) complex. This study elucidates DMB
Area of Science:
- Pharmacology and Toxicology
- Immunology
- Gastroenterology
Background:
- Inflammatory diseases like colitis and sepsis pose significant health challenges with limited treatment options.
- Trinitrobenzenesulfonic acid (TNBS)-induced colitis models chronic inflammation and intestinal barrier dysfunction.
- Demethyleneberberine (DMB) shows potential anti-inflammatory and anti-oxidative properties, but its mechanism is unclear.
Purpose of the Study:
- To identify the molecular targets and mechanisms of demethyleneberberine (DMB) in inflammatory responses.
- To evaluate DMB's efficacy in preclinical models of colitis and sepsis.
- To elucidate DMB's interaction with the Toll-like receptor 4 (TLR4) signaling pathway.
Main Methods:
- Utilized a trinitrobenzenesulfonic acid (TNBS)-induced colitis rat model and an acute sepsis mouse model.
- Administered DMB, berberine (BBR), or curcumin (Curc) orally to evaluate therapeutic effects.
- Employed in vitro and in vivo systems, including myeloid differentiation protein-2 (MD-2) expression systems, to determine DMB's target.
Main Results:
- Demethyleneberberine (DMB) significantly alleviated colitis and protected against sepsis shock.
- DMB suppressed Toll-like receptor 4 (TLR4) signaling via both MyD88-dependent and independent pathways.
- DMB non-covalently bound to the MD-2 protein's hydrophobic pocket, specifically interacting with phenylalanine at position 76.
Conclusions:
- Demethyleneberberine (DMB) acts as a potent inhibitor of the MD-2 protein.
- DMB effectively suppresses hyperactivated TLR4 signaling, offering a novel therapeutic strategy.
- DMB demonstrates significant potential for treating inflammatory conditions like colitis and sepsis.
