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Apigenin: Is It a Pro- or Anti-Inflammatory Agent?

      To the Editor-in Chief:
      We read with special interest the elegant article by Sawatzky and colleagues
      • Sawatzky D
      • Willoughby D
      • Colville-Nash P
      • Rossi A
      The involvement of the apoptosis-modulating proteins Erk 1/2, Bcl-xL, and Bax in the resolution of acute inflammation in vivo.
      regarding the involvement of apoptosis-modulating proteins in the resolution of acute inflammation in vivo. One of the findings described by the authors was that in contrast to specific inhibitors of ERK-1/2 proteins, the flavonoid apigenin, which is a nonspecific inhibitor of ERK and Cox-2, actually exacerbated inflammation. Apigenin is a phytopolyphenol widely distributed in the human diet. Like many other flavonoids, apigenin has been reported to exert anti-inflammatory effects such as lowering oxidative stress and preventing the expression of several inflammatory factors, as confirmed by Sawatzky and collaborators.
      • Sawatzky D
      • Willoughby D
      • Colville-Nash P
      • Rossi A
      The involvement of the apoptosis-modulating proteins Erk 1/2, Bcl-xL, and Bax in the resolution of acute inflammation in vivo.
      We would like to draw attention to the fact that flavonoids are potent anti-inflammatory factors, even when administered after inflammation is settled. Our group has investigated the effect of xanthohumol, a prenylated flavonoid, in nude mice inoculated with human breast cancer MCF7 cells (Soares et al

      Soares R, Monteiro R, Guerreiro S, Incio J, Lopes R, Oliveira e Silva A, Gärtner F, Calhau C: Modulation of angiogenesis in breast cancer by dietary zantho-humol. Sociedade Portuguesa de Farmacologia 2005, (Book of Abstracts): C33

      ). Administration of xanthohumol in the mouse's beverage for 2 months starting 24 hours after inoculation, resulted in dramatic decreases in inflammatory cells, both polymorphonuclear cells and lymphocytes, as compared to vehicle-treated mice. These findings imply that flavonoids prevent inflammation even when administered after the onset of disease.
      Corroborating our findings, studies using apigenin and other flavonoids after inflammation has been initiated
      • Kowalski J
      • Samojedny A
      • Paul M
      • Pietsz G
      • Wilczok T
      Effect of apigenin, kaempferol and resveratrol on the expression of interleukin-1beta and tumor necrosis factor-alpha genes in J774.2 macrophages.
      • Hendriks JJ
      • Alblas J
      • van der Pol SM
      • van Tol EA
      • Dijkstra CD
      • de Vries HE
      Flavonoids influence monocytic GTPase activity and are protective in experimental allergic encephalitis.
      have reported anti-inflammatory effects of these flavonoids. Hendriks and colleagues
      • Hendriks JJ
      • Alblas J
      • van der Pol SM
      • van Tol EA
      • Dijkstra CD
      • de Vries HE
      Flavonoids influence monocytic GTPase activity and are protective in experimental allergic encephalitis.
      examined the effect of another flavonoid compound, luteolin, in lesions of the central nervous system; they observed reduced inflammation and axonal damage when luteolin was administered either before or after disease onset.
      Sawatzky and colleagues
      • Sawatzky D
      • Willoughby D
      • Colville-Nash P
      • Rossi A
      The involvement of the apoptosis-modulating proteins Erk 1/2, Bcl-xL, and Bax in the resolution of acute inflammation in vivo.
      argued that the increase in inflammation they observed after administration of apigenin could be due to inhibition of Cox-2, since Cox-2 inhibitors given at the peak of inflammation resulted in a prolonged inflammatory response. We think that another hypothesis may be applied: flavonoids are potent phytoestrogens and are capable of inducing estrogen-dependent gene transcription, resulting in activation of several tyrosine kinases.
      • Stroheker T
      • Pinnert MF
      • Picard K
      • Chagnon MC
      • Canivenc-Lavier MC
      Estrogenic effects of apigenin, kaempferol, and bisphenol A in immature Wistar female rats and in MCF-7 cells.
      Estrogens are also able to activate several tyrosine kinase receptors, such as epidermal growth factor receptor and transforming growth factor-β.
      • Soares R
      • Reis-Filho JS
      • Gartner F
      • Schmitt FC
      Vascular endothelial growth factor, transforming growth factor-alpha, and estrogen receptors: possible cross-talks and interactions.
      • Soares R
      • Guo S
      • Gartner F
      • Schmitt FC
      • Russo J
      17beta-estradiol-mediated vessel assembly and stabilization in tumor angiogenesis requires TGFbeta and EGFR crosstalk.
      Therefore, it is quite possible that flavonoids interact with these signaling pathways as well.
      In conclusion, the complexity of the inflammatory process and the variety of flavonoid effects may establish some unpredictability in their interactions. The potential relevance of flavonoids in health justifies the pursuit of this path of investigation.
      Authors' Reply:
      We thank Drs. Soares and Azevedo for their special interest in our study and for highlighting the diverse biological effects and possible mechanisms of action of the dietary plant flavonoid apigenin. In our recent study
      • Sawatzky DA
      • Willoughby DA
      • Colville-Nash PR
      • Rossi AG
      The involvement of the apoptosis modulating proteins ERK 1/2, Bcl-xL, and Bax in the resolution of acute inflammation in vivo.
      we described that by modulating signaling pathways by specific pharmacological inhibitors, we can influence the resolution of established inflammation in an acute model of pleurisy in the rat. In addition to using specific inhibitors of the mitogen-activated protein kinase (MAPK) pathway and the pro-apoptotic Bcl-2 family member Bax, we used another pharmacological reagent, the phytopolyphenol apigenin, which is a flavonoid that has been reported to have wide-ranging anti-inflammatory and anti-cancer properties.
      • Sarkar FH
      • Li Y
      Cell signaling pathways altered by natural chemopreventive agents.
      ,
      • Ross JA
      • Kasum CM
      Dietary flavonoids: bioavailability, metabolic effects, and safety.
      The precise mechanisms by which apigenin and other flavonoids exert their anti-inflammatory properties, however, remain to be elucidated. Apigenin has been shown to affect a number of key signaling pathways, including MAPK, protein kinase C, phosphatidylinositol-3 kinase, nuclear factor-κB and cyclooxygenase (COX),
      • Sarkar FH
      • Li Y
      Cell signaling pathways altered by natural chemopreventive agents.
      ,
      • Ross JA
      • Kasum CM
      Dietary flavonoids: bioavailability, metabolic effects, and safety.
      that likely lead to its diverse and complex biological profile in a wide variety of cell types and tissues. Thus, these effects are likely to influence, in a complex manner, all features of the inflammatory response, including the processes of initiation, progression, and resolution of inflammation.
      • Gilroy DW
      • Lawrence T
      • Perretti M
      • Rossi AG
      Inflammatory resolution: new opportunities for drug discovery.
      In our study
      • Sawatzky DA
      • Willoughby DA
      • Colville-Nash PR
      • Rossi AG
      The involvement of the apoptosis modulating proteins ERK 1/2, Bcl-xL, and Bax in the resolution of acute inflammation in vivo.
      we found that apigenin, perhaps surprisingly given its better known anti-inflammatory properties, administered intrapleurally at the peak of inflammation in the rat model of carrageenan-induced pleurisy augmented inflammation, presumably by slowing the resolution phase of the inflammatory response. We suggested that a possible mechanism for its effects might be that the flavonoid could influence COX-2 expression and/or activity, possibly by inhibiting important signaling pathways. Our rationale for this explanation is that selective COX-2 inhibition by NS-398 and traditional nonsteroidal anti-inflammatory drugs, when administered at the peak of carrageenan pleurisy, cause a prolonged inflammatory response.
      • Gilroy DW
      • Colville-Nash PR
      • Willis D
      • Chivers J
      • Paul-Clark MJ
      • Willoughby DA
      Inducible cyclooxygenase may have anti-inflammatory properties.
      This is likely attributable to inhibition of the anti-inflammatory cyclopentenone prostaglandins, 15deoxyΔ12,14 PGJ2 and PGD2 synthesis, prostanoids that directly promote apoptosis of neutrophils and macrophages,
      • Gilroy DW
      • Colville-Nash PR
      • McMaster S
      • Sawatzky DA
      • Willoughby DA
      • Lawrence T
      Inducible cyclooxygenase-derived 15-deoxyΔ12–14PGJ2 brings about acute inflammatory resolution in rat pleurisy by inducing neutrophil and macrophage apoptosis.
      ,
      • Ward C
      • Dransfield I
      • Murray J
      • Farrow SN
      • Haslett C
      • Rossi AG
      Prostaglandin D2 and its metabolites induce caspase-dependent granulocyte apoptosis that is mediated via inhibition of IκBα degradation using a peroxisome proliferator-activated receptor-γ-independent mechanism.
      apoptosis being a fundamental process for the successful resolution of inflammation.
      • Gilroy DW
      • Lawrence T
      • Perretti M
      • Rossi AG
      Inflammatory resolution: new opportunities for drug discovery.
      ,
      • Riley NA
      • Ward C
      • Sawatzky DA
      • Sheldrake TA
      • Dransfield I
      • Haslett C
      • Rossi AG
      Granulocyte apoptosis and macrophage clearance of apoptotic cells as targets for pharmacological intervention in inflammatory diseases.
      We have preliminary unpublished evidence that apigenin inhibits COX-2 expression in pleural inflammatory cells when administered 24 hours after established carrageenan-induced inflammation; however, given the recent interest in flavonoids, this finding merits further investigation.
      Drs. Soares and Azevedo correctly highlight another interesting possible explanation. They have brought to our attention their recent preliminary study that another flavonoid, xanthohumol, given orally for 2 months starting 24 hours after nude mice had been inoculated with human breast cancer MCF cells reduced inflammatory cell recruitment.

      Soares R, Monteiro R, Guerreiro S, Incio J, Lopes R, Oliveira e Silva A, Gärtner F, Calhau C: Modulation of angiogenesis in breast cancer by dietary xantho-humol. Sociedade Portuguesa de Farmacologia 2005(Book of Abstracts): C33

      An interesting hypothesis that has been postulated is that flavonoids, being potent phytoestrogens,
      • Ross JA
      • Kasum CM
      Dietary flavonoids: bioavailability, metabolic effects, and safety.
      may be capable of inducing estrogen-dependent gene transcription thereby influencing certain tyrosine kinase-linked receptors.
      • Soares R
      • Reis-Filho JS
      • Gartner F
      • Schmitt FC
      Vascular endothelial growth factor, transforming growth factor-α, and estrogen receptors: possible cross-talks and interactions.
      ,
      • Soares R
      • Guo S
      • Gartner F
      • Schmitt FC
      • Russo J
      17β-estradiol-mediated vessel assembly and stabilization in tumor angiogenesis requires TGFβ and EGFR crosstalk.
      Although there is good evidence that estrogens can modulate carrageenan-induced pleurisy in the rat,
      • Esposito E
      • Iacono A
      • Raso GM
      • Pacilio M
      • Coppola A
      • Di Carlo R
      • Meli R
      Raloxifene, a selective estrogen receptor modulator, reduces carrageenan-induced acute inflammation in normal and ovariectomized rats.
      ,
      • Cozzocrea S
      • Mazzou E
      • Sautebin L
      • Serraino I
      • Dugo L
      • Calabro G
      • Caputi AP
      • Maggi A
      The protective role of endogenous estrogen in carageenan-induced lung injury in the rat.
      ,
      • Cuzzocrea S
      • Santagati S
      • Sautebin L
      • Mazzon E
      • Calabro G
      • Serraino I
      • Caputi AP
      • Maggi A
      17β-estradiol anti-inflammatory activity in carrageenan-induced pleurisy.
      whether flavonoids such as apigenin influence the resolution of inflammation in the acute pleurisy model described in our study by modulating estrogen-dependent pathways requires direct investigation. Given the many reported biological effects of flavonoids, the structural diversity of this family (more than 5000 flavonoids have been described) and the relatively unknown pharmacological and pharmacokinetic properties of these compounds, further research in our opinion is warranted.

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