Biochanin A attenuates LPS-induced pro-inflammatory responses and inhibits the activation of the MAPK pathway in BV2 microglial cells

Wu,Wang-Yang; Wu,Yang-Yang; Huang,Huan; He,Can; Li,Wei‑Zu; Wang,Hui-Li; Chen,Han-Qing; Yin,Yan-Yan

doi:10.3892/ijmm.2014.2020

Biochanin A attenuates LPS-induced pro-inflammatory responses and inhibits the activation of the MAPK pathway in BV2 microglial cells

Authors:
- Wang-Yang Wu
- Yang-Yang Wu
- Huan Huang
- Can He
- Wei‑Zu Li
- Hui-Li Wang
- Han-Qing Chen
- Yan-Yan Yin
View Affiliations

Affiliations: Department of Pharmacology, Anhui Medical University, Hefei, Anhui 230032, P.R. China, School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei, Anhui 230009, P.R. China
Published online on: December 2, 2014 https://doi.org/10.3892/ijmm.2014.2020
Pages: 391-398

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Inflammation in the brain, characterized by the activation of microglia, is believed to participate in the pathogenesis of Parkinson's disease. Biochanin A, an O-methylated isoflavone, is a natural organic compound and is classified as a phytoestrogen. In this study, using murine BV2 microglial cells, we investigated the anti-inflammatory effects of biochanin A and the possible mechanisms involved. BV2 microglial cells were treated with lipopolysaccharide (LPS) to induce pro-inflammatory responses and the cells were then treated with biochanin A. Cell viability was examined by MTT assay. The production of nitric oxide (NO) was examined using Griess reagent and intracellular reactive oxygen species (ROS production) was measured by DCFH-DA assay. The mRNA expression of interleukin-1β (IL-1β), inducible nitric oxide synthase (iNOS) and tumor necrosis factor-α (TNF-α) was examined by RT-PCR. The expression of p-ERK, p-JNK, p-p38 and iNOS was measured by western blot analysis. In addition, the protein and mRNA and phosphorylation levels of pro-inflammatory cytokines were determined by western blot analysis and RT-PCR, respectively. The results revealed that biochanin A attenuated LPS-induced microglial activation and the production of TNF‑α, IL-1β, nitric oxide and reactive oxygen species in a dose-dependent manner. Biochanin A significantly decreased the LPS-induced mRNA expression of TNF-α and IL-1β, and inhibited iNOS mRNA and protein expression. Furthermore, biochanin A significantly inhibited the LPS-induced phosphorylation of c-Jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38. These findings suggest that the inhibitory effects of biochanin A on LPS-induced proinflammatory responses may be associated with the inhibition of mitogen-activated protein kinase (MAPK) signaling pathways in BV2 microglial cells.

View Figures

View References

1	Liu B and Hong JS: Role of microglia in inflammation-mediated neurodegenerative diseases: mechanisms and strategies for therapeutic intervention. J Pharmacol Exp Ther. 304:1–7. 2003. View Article : Google Scholar
2	McGeer PL, Itagaki S, Boyes BE and McGeer EG: Reactive microglia are positive for HLA-DR in the substantia nigra of Parkinson’s and Alzheimer’s disease brains. Neurology. 38:1285–1291. 1988. View Article : Google Scholar : PubMed/NCBI
3	Kreutzberg GW: Microglia: a sensor for pathological events in the CNS. Trends Neurosci. 19:312–318. 1996. View Article : Google Scholar : PubMed/NCBI
4	Gao HM, Hong JS, Zhang W and Liu B: Distinct role for microglia in rotenone-induced degeneration of dopaminergic neurons. J Neurosci. 22:782–790. 2002.PubMed/NCBI
5	Gayle DA, Ling Z, Tong C, Landers T, Lipton JW and Carvey PM: Lipopolysaccharide (LPS)-induced dopamine cell loss in culture: roles of tumor necrosis factor-alpha, interleukin-1beta, and nitric oxide. Brain Res Dev Brain Res. 133:27–35. 2002. View Article : Google Scholar : PubMed/NCBI
6	Kim WG, Mohney RP, Wilson B, Jeohn GH, Liu B and Hong JS: Regional difference in susceptibility to lipopolysaccharide-induced neurotoxicity in the rat brain: role of microglia. J Neurosci. 20:6309–6316. 2000.PubMed/NCBI
7	Liu B, Du L and Hong JS: Naloxone protects rat dopaminergic neurons against inflammatory damage through inhibition of microglia activation and superoxide generation. J Pharmacol Exp Ther. 293:607–617. 2000.PubMed/NCBI
8	Liu B, Gao HM, Wang JY, Jeohn GH, Cooper CL and Hon JS: Role of nitric oxide in inflammation-mediated neurodegeneration. Ann NY Acad Sci. 962:318–331. 2002. View Article : Google Scholar : PubMed/NCBI
9	Jung WK, Ahn YW, Lee SH, Choi YH, Kim SK, Yea SS, Choi I, Park SG, Seo SK, Lee SW and Choi IW: Ecklonia cava ethanolic extracts inhibit lipopolysaccharide-induced cyclooxygenase-2 and inducible nitric oxide synthase expression in BV2 microglia via the MAP kinase and NF-kappaB pathways. Food Chem Toxicol. 47:410–417. 2009. View Article : Google Scholar
10	Lei M, Wang JG, Xiao DM, Fan M, Wang DP, Xiong JY, Chen Y, Ding Y and Liu SL: Resveratrol inhibits interleukin 1β-mediated inducible nitric oxide synthase expression in articular chondrocytes by activating SIRT1 and thereby suppressing nuclear factor-κB activity. Eur J Pharmacol. 674:73–79. 2012. View Article : Google Scholar
11	Chen HQ, Jin ZY and Li GH: Biochanin A protects dopaminergic neurons against lipopolysaccharide-induced damage through inhibition of microglia activation and proinflammatory factors generation. Neurosci Lett. 417:112–117. 2007. View Article : Google Scholar : PubMed/NCBI
12	Puthli A, Tiwari R and Mishra KP: Biochanin A enhances the radiotoxicity in colon tumor cells in vitro. J Environ Pathol Toxicol Oncol. 32:189–203. 2013. View Article : Google Scholar : PubMed/NCBI
13	Su SJ, Yeh YT and Shyu HW: The preventive effect of biochanin a on bone loss in ovariectomized rats: involvement in regulation of growth and activity of osteoblasts and osteoclasts. Evid Based Complement Alternat Med. 2013:5948572013.PubMed/NCBI
14	Green LC, Wagner DA, Glogowski J, Skipper PL, Wishnok JS and Tannenbaum SR: Analysis of nitrate, nitrite, and [15N]nitrate in biological fluids. Anal Biochem. 126:131–138. 1982. View Article : Google Scholar : PubMed/NCBI
15	Bruce-Keller AJ, Keeling JL, Keller JN, Huang FF, Camondola S and Mattson MP: Antiinflammatory effects of estrogen on microglial activation. Endocrinology. 141:3646–3656. 2000.PubMed/NCBI
16	Boje KM: Nitric oxide neurotoxicity in neurodegenerative diseases. Front Biosci. 9:763–776. 2004. View Article : Google Scholar : PubMed/NCBI
17	Han IO, Kim KW, Ryu JH and Kim WK: p38 mitogen-activated protein kinase mediates lipopolysaccharide, not interferon-gamma, -induced inducible nitric oxide synthase expression in mouse BV2 microglial cells. Neurosci Lett. 325:9–12. 2002. View Article : Google Scholar : PubMed/NCBI
18	Kim M, Li YX, Dewapriya P, Ryu B and Kim SK: Floridoside suppresses pro-inflammatory responses by blocking MAPK signaling in activated microglia. BMB Rep. 46:398–403. 2013. View Article : Google Scholar : PubMed/NCBI
19	Le W, Rowe D, Xie W, Ortiz I, He Y and Appel SH: Microglial activation and dopaminergic cell injury: an in vitro model relevant to Parkinson’s disease. J Neurosci. 21:8447–8455. 2001.PubMed/NCBI
20	Minghetti L and Levi G: Microglia as effector cells in brain damage and repair: focus on prostanoids and nitric oxide. Prog Neurobiol. 54:99–125. 1998. View Article : Google Scholar : PubMed/NCBI
21	Sawada M, Kondo N, Suzumura A and Marunouchi T: Production of tumor necrosis factor-alpha by microglia and astrocytes in culture. Brain Res. 491:394–397. 1989. View Article : Google Scholar : PubMed/NCBI
22	Akassoglou K, Bauer J, Kassiotis G, et al: Oligodendrocyte apoptosis and primary demyelination induced by local TNF/p55TNF receptor signaling in the central nervous system of transgenic mice: models for multiple sclerosis with primary oligodendrogliopathy. Am J Pathol. 153:801–813. 1998. View Article : Google Scholar : PubMed/NCBI
23	Botchkina GI, Meistrell ME III, Botchkina IL and Tracey KJ: Expression of TNF and TNF receptors (p55 and p75) in the rat brain after focal cerebral ischemia. Mol Med. 3:765–781. 1997.
24	Meda L, Cassatella MA, Szendrei GI, et al: Activation of microglial cells by beta-amyloid protein and interferon-gamma. Nature. 374:647–650. 1995. View Article : Google Scholar : PubMed/NCBI
25	Sriram K, Matheson JM, Benkovic SA, Miller DB, Luster MI and O’Callaghan JP: Mice deficient in TNF receptors are protected against dopaminergic neurotoxicity: implications for Parkinson’s disease. FASEB J. 16:1474–1476. 2002.PubMed/NCBI
26	Rothwell N, Allan S and Toulmond S: The role of interleukin 1 in acute neurodegeneration and stroke: pathophysiological and therapeutic implications. J Clin Invest. 100:2648–2652. 1997. View Article : Google Scholar
27	Chao CC, Hu S, Molitor TW, Shaskan EG and Peterson PK: Activated microglia mediate neuronal cell injury via a nitric oxide mechanism. J Immunol. 149:2736–2741. 1992.PubMed/NCBI
28	Jeohn GH, Kim WG and Hong JS: Time dependency of the action of nitric oxide in lipopolysaccharide-interferon-gamma-induced neuronal cell death in murine primary neuron-glia co-cultures. Brain Res. 880:173–177. 2000. View Article : Google Scholar : PubMed/NCBI
29	Iravani MM, Kashefi K, Mander P, Rose S and Jenner P: Involvement of inducible nitric oxide synthase in inflammation-induced dopaminergic neurodegeneration. Neuroscience. 110:49–58. 2002. View Article : Google Scholar : PubMed/NCBI
30	Li FQ, Wang T, Pei Z, Liu B and Hong JS: Inhibition of microglial activation by the herbal flavonoid baicalein attenuates inflammation-mediated degeneration of dopaminergic neurons. J Neural Transm. 112:331–347. 2005. View Article : Google Scholar
31	Dawson VL and Dawson TM: Nitric oxide in neuronal degeneration. Proc Soc Exp Biol Med. 211:33–40. 1996. View Article : Google Scholar : PubMed/NCBI
32	Qin L, Liu Y, Wang T, et al: NADPH oxidase mediates lipopolysaccharide-induced neurotoxicity and proinflammatory gene expression in activated microglia. J Biol Chem. 279:1415–1421. 2004. View Article : Google Scholar
33	Wang T, Qin L, Liu B, et al: Role of reactive oxygen species in LPS-induced production of prostaglandin E2 in microglia. J Neurochem. 88:939–947. 2004. View Article : Google Scholar : PubMed/NCBI
34	Farber JL: Mechanisms of cell injury by activated oxygen species. Environ Health Perspect. 102(Suppl 10): S17–S24. 1994. View Article : Google Scholar
35	Estevez AG and Jordan J: Nitric oxide and superoxide, a deadly cocktail. Ann NY Acad Sci. 962:207–211. 2002. View Article : Google Scholar : PubMed/NCBI
36	Xie Z, Wei M, Morgan TE, et al: Peroxynitrite mediates neurotoxicity of amyloid beta-peptide1–42- and lipopolysaccharide-activated microglia. J Neurosci. 22:3484–3492. 2002.PubMed/NCBI
37	Kim EK and Choi EJ: Pathological roles of MAPK signaling pathways in human diseases. Biochim Biophys Acta. 1802:396–405. 2010. View Article : Google Scholar : PubMed/NCBI
38	Bhat NR, Zhang P, Lee JC and Hogan EL: Extracellular signal-regulated kinase and p38 subgroups of mitogen-activated protein kinases regulate inducible nitric oxide synthase and tumor necrosis factor-alpha gene expression in endotoxin-stimulated primary glial cultures. J Neurosci. 18:1633–1641. 1998.PubMed/NCBI
39	Choi Y, Lee MK, Lim SY, Sung SH and Kim YC: Inhibition of inducible NO synthase, cyclooxygenase-2 and interleukin-1beta by torilin is mediated by mitogen-activated protein kinases in microglial BV2 cells. Br J Pharmacol. 156:933–940. 2009. View Article : Google Scholar : PubMed/NCBI
40	Johnson GL and Lapadat R: Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Science. 298:1911–1912. 2002. View Article : Google Scholar : PubMed/NCBI
41	Park JS, Woo MS, Kim DH, et al: Anti-inflammatory mechanisms of isoflavone metabolites in lipopolysaccharide-stimulated microglial cells. J Pharmacol Exp Ther. 320:1237–1245. 2007. View Article : Google Scholar
42	Song Y, Qu R, Zhu S, Zhang R and Ma S: Rhynchophylline attenuates LPS-induced pro-inflammatory responses through downregulation of MAPK/NF-kappaB signaling pathways in primary microglia. Phytother Res. 26:1528–1533. 2012.PubMed/NCBI