A neolignan enantiomer from Piper hancei with anti-neuroinflammatory effect by attenuating NF-κB signaling pathway.

Background: In the traditional "Yao" ethno-medicine system, Piper hancei Maxim. is used to treat rheumatism, wind-cold, and inflammation. Previous studies indicate that lignans obtained from P. hancei stems have anti-neuroinflammatory potential in LPS-stimulated microglial cells. However, identification of the lignan enantiomers and the precise mechanism by which they work to reduce inflammation is yet to be explored.

Purpose: To identify the active anti-neuroinflammatory lignan enantiomers isolated from P. hancei stems and to elucidate the mechanism of action both in vitro and in vivo.

Methods: The lignan enantiomers from P. hancei stems were isolated and elucidated using various chromatographic and spectroscopic methods. The anti-neuroinflammatory potential of all the compounds was initially screened by measuring nitric oxide (NO) inhibition in LPS-stimulated BV-2 microglial cells. Then anti-neuroinflammatory efficacy of the most active compound was assessed with LPS-stimulated microglial cell model, microglia-induced neuronal injury SH-SY5Y cell model, and LPS-intracerebroventricular injection neuroinflammation mouse model. The underlying mechanism was further explored by qRT-PCR analysis, Western blot analysis, and immunofluorescence staining experiments to understand the intervention pathway.

Results: Phytochemical analysis of P. hancei stems resulted in the isolation of 13 pairs of neolignan enantiomers (1-13), including 4 new pairs named piperhancin D-G (1-4). All right-handed (+) and left-handed (-) enantiomers of each pair (1-13) were isolated successfully. Notably, (+)-futoquinol (5) demonstrated significant anti-neuroinflammatory activity without cytotoxicity, unlike its inactive enantiomer (-)-5 in LPS-stimulated microglial cells. The representative compound (+)-5 effectively suppressed pro-inflammatory cytokines in LPS stimulated BV-2 cells and mouse brains, and alleviated microglia-induced neuronal damage in SH-SY5Y cells. Behavioral tests showed that (+)-5 alleviated the LPS-induced cognitive dysfunction in mice. Furthermore, the compound was able to reduce LPS-induced neuronal damage and microglial activation in mouse brains. A mechanistic study demonstrated that (+)-5 hindered the nuclear translocation of NF-κB p65 and downregulated the pro-inflammatory mediators to relieve neuroinflammation.

Conclusion: This is the first example of both in vitro and in vivo study on the anti-neuroinflammatory effects and underlying mechanism of the neolignan enantiomers isolated from P. hancei. Notably, (+)-futoquinol (5) emerged as a potential lead for further drug development to treat neurodegenerative diseases.