Anti-Neuroinflammatory effects of the extract of Achillea fragrantissima

Anat Elmann,Alona Telerman,Miriam Rindner,Hilla Erlank,Sharon Mordechay,Rivka Ofir

doi:10.1186/1472-6882-11-98

Abstract

BackgroundThe neuroinflammatory process plays a central role in the initiation and progression of neurodegenerative diseases such as Parkinson's and Alzheimer's diseases, and involves the activation of brain microglial cells. During the neuroinflammatory process, microglial cells release proinflammatory mediators such as cytokines, matrix metalloproteinases (MMP), Reactive oxygen species (ROS) and nitric oxide (NO). In the present study, extracts from 66 different desert plants were tested for their effect on lipopolysaccharide (LPS) - induced production of NO by primary microglial cells. The extract of Achillea fragrantissima (Af), which is a desert plant that has been used for many years in traditional medicine for the treatment of various diseases, was the most efficient extract, and was further studied for additional anti-neuroinflammatory effects in these cells.MethodsIn the present study, the ethanolic extract prepared from Af was tested for its anti-inflammatory effects on lipopolysaccharide (LPS)-activated primary cultures of brain microglial cells. The levels of the proinflammatory cytokines interleukin1β (IL-1β) and tumor necrosis factor-α (TNFα) secreted by the cells were determined by reverse transcriptase-PCR and Enzyme-linked immunosorbent assay (ELISA), respectively. NO levels secreted by the activate cells were measured using Griess reagent, ROS levels were measured by 2'7'-dichlorofluorescein diacetate (DCF-DA), MMP-9 activity was measured using gel zymography, and the protein levels of the proinflammatory enzymes cyclooxygenase-2 (COX-2) and induced nitric oxide synthase (iNOS) were measured by Western blot analysis. Cell viability was assessed using Lactate dehydrogenase (LDH) activity in the media conditioned by the cells or by the crystal violet cell staining.ResultsWe have found that out of the 66 desert plants tested, the extract of Af was the most efficient extract and inhibited ~70% of the NO produced by the LPS-activated microglial cells, without affecting cell viability. In addition, this extract inhibited the LPS - elicited expression of the proinflammatory mediators IL-1β, TNFα, MMP-9, COX-2 and iNOS in these cells.ConclusionsThus, phytochemicals present in the Af extract could be beneficial in preventing/treating neurodegenerative diseases in which neuroinflammation is part of the pathophysiology.

Highlights

The neuroinflammatory process plays a central role in the initiation and progression of neurodegenerative diseases such as Parkinson’s and Alzheimer’s diseases, and involves the activation of brain microglial cells
The excessive amounts of nitric oxide (NO), a free radical produced by induced NO-synthase (iNOS), and of prostaglandin E, an arachidonic acid metabolite produced by COX-2, which are secreted by activated microglial cells during the neuroinflammatory process, cause nitrosative stress and brain cell death [5,6]
Reagents Dulbecco’s modified Eagle’s medium (DMEM), RPMI1640, Leibovitz-15 medium, glutamine, antibiotics (10,000 IU/ml penicillin and 10,000 μg/ml streptomycin), soybean trypsin inhibitor, fetal bovine serum (FBS) and Dulbecco’s phosphate buffered saline (PBS) were purchased from Biological Industries (Beit Haemek, Israel); Griess reagent and rabbit anti COX-2 polyclonal antibody were obtained from Cayman chemical, Ml, USA; DreamTaq Green PCR master Mix (2x) and ReverAid First Strand cDNA Synthesis Kit were purchased from Fermentas life sciences

Summary

Introduction

The neuroinflammatory process plays a central role in the initiation and progression of neurodegenerative diseases such as Parkinson’s and Alzheimer’s diseases, and involves the activation of brain microglial cells. The increase in the life span of populations in the Western world has been accompanied by an elevation in the frequencies of neurodegenerative diseases, e.g., Alzheimer’s and Parkinson’s diseases. In these diseases, a gradual and progressive neuronal cell death occurs, or to the lipopolysaccharide (LPS) excreted during bacterial infection. The excessive amounts of NO, a free radical produced by iNOS, and of prostaglandin E, an arachidonic acid metabolite produced by COX-2, which are secreted by activated microglial cells during the neuroinflammatory process, cause nitrosative stress and brain cell death [5,6]. Co-induction and co-regulation of iNOS and COX-2 have been demonstrated in a number of cell culture studies and in inflammatory animal model systems [8]

Methods

Results

Discussion

Conclusion