Abstract
Purpose : To evaluate the anti-neuroinflammatory effects of Houttuynia cordata extract ( H. cordata ) in lipopolysaccharide (LPS)-stimulated BV-2 microglial cells, and its anti-oxidant properties. Methods : Anti-oxidant properties were evaluated by 1, 1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenging assay. Cell viability was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5- diphenyl-tetrazolium bromide (MTT) assay. LPS was used to stimulate BV-2 cells. Nitric oxide (NO) levels were measured using Griess assay. Inducible NO synthase (iNOS) expression, interleukin (IL)-6 expressional level were determined by enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Results : Ethyl actetae (HC-EA) extract of H. cordata significantly scavenged DPPH free radicals in a concentration-dependent fashion. The increased levels of NO, iNOS and IL-6 in LPS-stimulated BV-2 microglial cells were also suppressed by HC-EA extract in a concentration-dependent manner. Conclusion: The result indicate that the HC-EA extract exhibited strong anti-oxidant properties and inhibited the excessive production of pro-inflammatory mediators, including NO, iNOS and IL-6, in LPSstimulated BV-2 cells. The anti-oxidant phenolic compounds present in HC-EA extract might play an important role in ameliorating neuroinflammatory processes in LPS-stimulated BV-2 microglial cells. Keywords: Houttuynia cordata , DPPH radicals, antioxidant, neuroinflammation, BV-2 cells, iNOS, COX-2, IL-6.
Highlights
Inflammation in brain disorders is characterized by activation of glial cells further expressing key inflammatory mediators as well as neurotoxic free radicals
Pre-treatment with HC-ethyl acetate (EA) extract significantly suppressed the LPS-stimulated increased nitric oxide (NO) release in BV-2 cells in a concentration-dependent manner compared to LPS cells
Western blot analysis showed that the increased protein expression of inducible NO synthase (iNOS) in the LPS-stimulated BV-2 cells was suppressed in a concentration-dependent manner (Fig 4)
Summary
Inflammation in brain disorders is characterized by activation of glial cells (mainly microglia and astrocytes) further expressing key inflammatory mediators as well as neurotoxic free radicals. The anti-neuroinflammatory activity of H. cordata has not been studied in activated BV2 microglial cells. We sought to determine whether H. cordata extract possesses anti-neuroinflammatory activity in LPS-stimulated BV-2 microglial cells. Cells were pre-treated with the indicated concentrations of HC-EA) for 1 hr before the addition of LPS (1 μg/ml, SigmaAldrich, St. Louis, MO, USA) in serum free DMEM. The anti-oxidant property of the HC-EA extract was determined using the stable radical 2, 2diphenyl-1-picrylhydrazyl (DPPH, Sigma-Aldrich, St. Louis, MO, USA). Treatment with HCEA at various concentrations (0.1 μg/ml to 100 μg/ml) did not affect the overall cell viability nor did they exhibit any significant cytotoxicity in BV2 microglia (Fig 2). BV-2 microglia cells (1 x 105 cells/well) were cultured on 96 well plates and treated with HCEA extract at indicated dosages and LPS (1μg/ml).
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