Vitexin reduces neutrophil migration to inflammatory focus by down-regulating pro-inflammatory mediators via inhibition of p38, ERK1/2 and JNK pathway

Abstract

Background: Vitexin is a flavonoid found in plants of different genus such as Vitex spp. and Crataegus spp. Despite being an important molecule present in phytomedicines and nutraceuticals, the mechanisms supporting its use as anti-inflammatory remains unclear.

Purpose: To investigate the cellular and molecular mechanisms involved in acute anti-inflammatory effect of vitexin with regard to neutrophil recruitment and macrophages activation.

Methods: Anti-inflammatory properties of vitexin were evaluated in four models of neutrophil recruitment. The regulation of inflammatory mediators release was assessed in vivo and in vitro. Vitexin (5, 15 and 30 mg/kg p.o) effects on leukocytes migration to peritoneal cavity induced by zymosan (ZY), carrageenan (CG), n-formyl-methionyl-leucyl-phenylalanine (fMLP) and lipopolysaccharide (LPS) were evaluated in Swiss-Webster mice and the effects on the levels of TNF-α, IL-1β and IL-10 cytokines, and NO concentration were in the LPS-peritonitis. RAW 264.7 macrophages viability were determined by Alamar Blue assay as well as the capacity of vitexin in directly reducing the concentrations of TNF-α, IL-1β, IL-10, NO and PGE2. Additionally, vitexin effects upon the transcriptional factors p-p38, p-ERK1/2 and p-JNK were evaluated by western blotting in cells activated with LPS.

Results: Vitexin was not cytotoxic (IC50 > 200 µg/ml) in RAW 264.7 and at all doses tested it effectively reduced leukocyte migration in vivo, particularly neutrophils in the peritoneal lavage, independently of the inflammatory stimulus used. It also reduced TNF-α, IL-1β and NO releases in the peritoneal cavity of LPS-challenged mice. Vitexin had low cytotoxicity and was able to reduce the releases of TNF-α, IL-1β, NO, PGE2 and increase in IL-10 release by LPS activated RAW 264.7 cells. Vitexin was also able to regulate transcriptional factors for pro-inflammatory mediators, reducing the expression of p-p38, p-ERK1/2 and p-JNK in LPS-elicited cells.

Conclusions: Vitexin presented no in vitro cytotoxicity. Inhibition of neutrophil migration and pro-inflammatory mediators release contributes to the anti-inflammatory activity of vitexin. These effects are associated with the inactivation of important signaling pathways such as p38, ERK1/2 and JNK, which act on transcription factors for eliciting induction of inflammatory response.

Keywords: Cytotoxicity; Flavonoid; Immunomodulation; Leukocyte recruitment.