Abstract
Apigenin, a natural flavonoid, found in several plants, fruits, vegetables, herbs, and spices, is known to have anti-oxidant and anti-inflammatory properties that are evident in the use of these substances for centuries as medicinal approaches to treat asthma, insomnia, Parkinson’s disease, neuralgia, and shingles. However, there is a considerable dearth of information regarding its effect on immune cells, especially dendritic cells (DC) that maintain the critical balance between an immunogenic and tolerogenic immune response, in an immunospecialized location like the central nervous system (CNS). In this paper we looked at the anti-inflammatory properties of Apigenin in restoration of immune function and the resultant decrease in neuroinflammation. In vivo, a significant reduction in severity of experimental autoimmune encephalomyelitis (EAE) progression and relapse was observed in C57BL/6 (progressive) and SJL/J (relapse-remitting) mouse models of multiple sclerosis upon treatment with Apigenin. Apigenin treated EAE mice show decreased expression of α4 integrin and CLEC12A on splenic DCs and an increased retention of immune cells in the periphery compared to untreated EAE mice. This correlated consequently with immunohistochemistry findings of decreased immune cell infiltration and reduced demyelination in the CNS. These results indicate a protective role of Apigenin against the neurodegenerative effects resulting from the entry of DC stimulated pathogenic T cells into the CNS thus implicating a potential therapy for neuroinflammatory disease.
Similar content being viewed by others
References
Ali R, Nicholas RS, Muraro PA (2013) Drugs in development for relapsing multiple sclerosis. Drugs 73:625–650
Anandasabapathy N, Victora GD, Meredith M, Feder R, Dong B, Kluger C, Yao K, Dustin ML, Nussenzweig MC, Steinman RM, Liu K (2011) Flt3L controls the development of radiosensitive dendritic cells in the meninges and choroid plexus of the steady-state mouse brain. J Exp Med 208:1695–1705
Axtell RC, Webb MS, Barnum SR, Raman C (2004) Cutting edge: critical role for CD5 in experimental autoimmune encephalomyelitis: inhibition of engagement reverses disease in mice. J Immunol 173:2928–2932
Fletcher JM, Lonergan R, Costelloe L, Kinsella K, Moran B, O’Farrelly C, Tubridy N, Mills KH (2009) CD39+Foxp3+ regulatory T Cells suppress pathogenic Th17 cells and are impaired in multiple sclerosis. J Immunol 183:7602–7610
Gross CC, Jonuleit H, Wiendl H (2012) Fulfilling the dream: tolerogenic dendritic cells to treat multiple sclerosis. Eur J Immunol 42:569–572
Ha SK, Lee P, Park JA, Oh HR, Lee SY, Park JH, Lee EH, Ryu JH, Lee KR, Kim SY (2008) Apigenin inhibits the production of NO and PGE2 in microglia and inhibits neuronal cell death in a middle cerebral artery occlusion-induced focal ischemia mice model. Neurochem Int 52:878–886
Jager AK, Saaby L (2011) Flavonoids and the CNS. Molecules 16:1471–1485
Jeong GS, Lee SH, Jeong SN, Kim YC, Kim EC (2009) Anti-inflammatory effects of apigenin on nicotine- and lipopolysaccharide-stimulated human periodontal ligament cells via heme oxygenase-1. Int Immunopharmacol 9:1374–1380
Kang HK, Ecklund D, Liu M, Datta SK (2009) Apigenin, a non-mutagenic dietary flavonoid, suppresses lupus by inhibiting autoantigen presentation for expansion of autoreactive Th1 and Th17 cells. Arthritis Res Ther 11:R59
Lassmann H, van Horssen J (2011) The molecular basis of neurodegeneration in multiple sclerosis. FEBS Lett 585:3715–3723
Lee JH, Zhou HY, Cho SY, Kim YS, Lee YS, Jeong CS (2007) Anti-inflammatory mechanisms of apigenin: inhibition of cyclooxygenase-2 expression, adhesion of monocytes to human umbilical vein endothelial cells, and expression of cellular adhesion molecules. Arch Pharm Res 30:1318–1327
Lefort EC, Blay J (2013) Apigenin and its impact on gastrointestinal cancers. Mol Nutr Food Res 57:126–144
Liang YC, Tsai SH, Tsai DC, Lin-Shiau SY, Lin JK (2001) Suppression of inducible cyclooxygenase and nitric oxide synthase through activation of peroxisome proliferator-activated receptor-gamma by flavonoids in mouse macrophages. FEBS Lett 496:12–18
Liu K, Nussenzweig MC (2010) Origin and development of dendritic cells. Immunol Rev 234:45–54
Manuel SL, Rahman S, Wigdahl B, Khan ZK, Jain P (2007) Dendritic cells in autoimmune diseases and neuroinflammatory disorders. Front Biosci 12:4315–4335
Miean KH, Mohamed S (2001) Flavonoid (myricetin, quercetin, kaempferol, luteolin, and apigenin) content of edible tropical plants. J Agric Food Chem 49:3106–3112
Naves R, Singh SP, Cashman KS, Rowse AL, Axtell RC, Steinman L, Mountz JD, Steele C, De Sarno P, Raman C (2013) The interdependent, overlapping, and differential roles of type I and II IFNs in the pathogenesis of experimental autoimmune encephalomyelitis. J Immunol 191:2967–2977
Ni HT, Spellman SR, Jean WC, Hall WA, Low WC (2001) Immunization with dendritic cells pulsed with tumor extract increases survival of mice bearing intracranial gliomas. J Neurooncol 51:1–9
Nicholas C, Batra S, Vargo MA, Voss OH, Gavrilin MA, Wewers MD, Guttridge DC, Grotewold E, Doseff AI (2007) Apigenin blocks lipopolysaccharide-induced lethality in vivo and proinflammatory cytokines expression by inactivating NF-kappaB through the suppression of p65 phosphorylation. J Immunol 179:7121–7127
Nico B, Quondamatteo F, Ribatti D, Bertossi M, Russo G, Herken R, Roncali L (1998) Ultrastructural localization of lectin binding sites in the developing brain microvasculature. Anat Embryol (Berl) 197:305–315
Nijveldt RJ, van Nood E, van Hoorn DE, Boelens PG, van Norren K, van Leeuwen PA (2001) Flavonoids: a review of probable mechanisms of action and potential applications. Am J Clin Nutr 74:418–425
Oh J, O’Connor PW (2015) Established disease-modifying treatments in relapsing-remitting multiple sclerosis. Curr Opin Neurol 28:220–229
Pashenkov M, Huang YM, Kostulas V, Haglund M, Soderstrom M, Link H (2001) Two subsets of dendritic cells are present in human cerebrospinal fluid. Brain 124:480–492
Patel D, Shukla S, Gupta S (2007) Apigenin and cancer chemoprevention: progress, potential and promise (review). Int J Oncol 30:233–245
Plattner VE, Germann B, Neuhaus W, Noe CR, Gabor F, Wirth M (2010) Characterization of two blood-brain barrier mimicking cell lines: distribution of lectin-binding sites and perspectives for drug delivery. Int J Pharm 387:34–41
Raiotach-Regue D, Grau-Lopez L, Naranjo-Gomez M, Ramo-Tello C, Pujol-Borrell R, Martinez-Caceres E, Borras FE (2012) Stable antigen-specific T-cell hyporesponsiveness induced by tolerogenic dendritic cells from multiple sclerosis patients. Eur J Immunol 42:771–782
Renno T, Lin JY, Piccirillo C, Antel J, Owens T (1994) Cytokine production by cells in cerebrospinal fluid during experimental allergic encephalomyelitis in SJL/J mice. J Neuroimmunol 49:1–7
Rice GP, Hartung HP, Calabresi PA (2005) Anti-alpha4 integrin therapy for multiple sclerosis: mechanisms and rationale. Neurology 64:1336–1342
Rice-Evans C (2001) Flavonoid antioxidants. Curr Med Chem 8:797–807
Romanova D, Vachalkova A, Cipak L, Ovesna Z, Rauko P (2001) Study of antioxidant effect of apigenin, luteolin and quercetin by DNA protective method. Neoplasma 48:104–107
Ross JA, Kasum CM (2002) Dietary flavonoids: bioavailability, metabolic effects, and safety. Annu Rev Nutr 22:19–34
Rouse M, Singh NP, Nagarkatti PS, Nagarkatti M (2013) Indoles mitigate the development of experimental autoimmune encephalomyelitis by induction of reciprocal differentiation of regulatory T cells and Th17 cells. Br J Pharmacol 169:1305–1321
Sagar D, Foss C, El Baz R, Pomper MG, Khan ZK, Jain P (2012a) Mechanisms of dendritic cell trafficking across the blood-brain barrier. J Neuroimmune Pharmacol 7:74–94
Sagar D, Lamontagne A, Foss CA, Khan ZK, Pomper MG, Jain P (2012b) Dendritic cell CNS recruitment correlates with disease severity in EAE via CCL2 chemotaxis at the blood-brain barrier through paracellular transmigration and ERK activation. J Neuroinflammation 9:245
Shukla S, Gupta S (2010) Apigenin: a promising molecule for cancer prevention. Pharm Res 27:962–978
Singh NP, Hegde VL, Hofseth LJ, Nagarkatti M, Nagarkatti P (2007) Resveratrol (trans-3,5,4′-trihydroxystilbene) ameliorates experimental allergic encephalomyelitis, primarily via induction of apoptosis in T cells involving activation of aryl hydrocarbon receptor and estrogen receptor. Mol Pharmacol 72:1508–1521
Steinman L (2012) The discovery of natalizumab, a potent therapeutic for multiple sclerosis. J Cell Biol 199:413–416
Steinman RM, Cohn ZA (1973) Identification of a novel cell type in peripheral lymphoid organs of mice. I. Morphology, quantitation, tissue distribution. J Exp Med 137:1142–1162
Steinman RM, Hemmi H (2006) Dendritic cells: translating innate to adaptive immunity. Curr Top Microbiol Immunol 311:17–58
Swanborg RH (2001) Experimental autoimmune encephalomyelitis in the rat: lessons in T-cell immunology and autoreactivity. Immunol Rev 184:129–135
Verbeek R, Plomp AC, van Tol EA, van Noort JM (2004) The flavones luteolin and apigenin inhibit in vitro antigen-specific proliferation and interferon-gamma production by murine and human autoimmune T cells. Biochem Pharmacol 68:621–629
Verbeek R, van Tol EA, van Noort JM (2005) Oral flavonoids delay recovery from experimental autoimmune encephalomyelitis in SJL mice. Biochem Pharmacol 70:220–228
Wu GF, Laufer TM (2007) The role of dendritic cells in multiple sclerosis. Curr Neurol Neurosci Rep 7:245–252
Yednock TA, Cannon C, Fritz LC, Sanchez-Madrid F, Steinman L, Karin N (1992) Prevention of experimental autoimmune encephalomyelitis by antibodies against alpha 4 beta 1 integrin. Nature 356:63–66
Yoon MS, Lee JS, Choi BM, Jeong YI, Lee CM, Park JH, Moon Y, Sung SC, Lee SK, Chang YH, Chung HY, Park YM (2006) Apigenin inhibits immunostimulatory function of dendritic cells: implication of immunotherapeutic adjuvant. Mol Pharmacol 70:1033–1044
Acknowledgments
The authors wish to acknowledge US Public Health Service/National Institutes of Health grants: R01CA054559 and R56AI077414 to PJ.
Conflict of interest
The authors declare that they have no competing interests.
Author information
Authors and Affiliations
Corresponding authors
Electronic supplementary material
Below is the link to the electronic supplementary material.
Supplementary Figure 1
Myeloid cell quantification and characterization within the blood and lymph nodes of mice induced with relapse-remitting type of EAE. Cells isolated from the blood and lymph nodes of the EAE + vehicle and EAE + Apigenin groups in SJL/J mice were pooled in each group (n=5) and run in triplicates for cell quantification. The cells were stained for CD11c, CD8α, CD11b, CD68, CD45R, and CD4 immune cell markers. Plots represent absolute cell counts of each of the cell types from animals in each group. *p<0.05 (GIF 184 kb)
Supplementary Figure 2
Impact of Apigenin on DC stimulation and antigen presentation and T cell adhesion in SJL/J mice. Splenocytes from both the EAE + vehicle and EAE + Apigenin groups of SJL/J mice were stimulated with PLP323-339 peptides for 3 days followed by activation with PMA, ionomycin and brefeldin A for 5 hours. These cells were subsequently stained with antibodies against CD11c, CD8α, MHC II, and CD86, α4, and CLEC12A. Data represents CD11c+ dendritic cells from mice expressing MHCII+CD86+(top), and CD4+ T cells expressing α4 integrin (middle), and CLEC12A (bottom) upon stimulation with PLP323-339. Each bar is representative of the mean percentage for every marker per group. Histogram plots are representative of one animal per group. *p<0.05 (GIF 298 kb)
Supplementary Figure 3
Tregs up-regulation and Th17 down-modulation in SJL/J mice upon Apigenin treatment. Flow cytometry analysis representing CD4+ cells from lymphocytes of the SJL/J mice expressing IL-17A (top) and CD25+FOXP3 (bottom) upon stimulation with PLP323-339 peptides respectively for 3 days followed by activation with PMA, ionomycin and brefeldin A for 5 hours. Each bar is representative of the mean percentage for every marker per group. Contour plots representative of one animal per group are shown on the left. *p<0.05 (GIF 326 kb)
Supplementary Figure 4
Effect of Apigenin on IFN-γ production. Flow cytometry analysis representing CD4+ cells expressing IFNγ in SJL/J mice upon ex-vivo stimulation of lymphocytes with PLP323-339 and activation with PMA, ionomycin and brefeldin. Lymphocytes were pooled from mice in each group (n=5) and run for quantifying and stimulating procedures. (GIF 188 kb)
Supplementary Figure 5
Apigenin treatment reduces immune cell migration into the CNS. Spinal cord tissues from RR-EAE (SJL/J) mice were sectioned. Spinal cord tissue from SJL/J mice was subjected to LFB and H&E staining showing areas of myelination (blue) and cellular infiltration (reddish brown). (GIF 654 kb)
Rights and permissions
About this article
Cite this article
Ginwala, R., McTish, E., Raman, C. et al. Apigenin, a Natural Flavonoid, Attenuates EAE Severity Through the Modulation of Dendritic Cell and Other Immune Cell Functions. J Neuroimmune Pharmacol 11, 36–47 (2016). https://doi.org/10.1007/s11481-015-9617-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11481-015-9617-x