Ha et al. Chemistry Central Journal (2018) 12:14 https://doi.org/10.1186/s13065-018-0386-5 RESEARCH ARTICLE Open Access Anti‑infammatory efect of oligostilbenoids from Vitis heyneana in LPS‑stimulated RAW 264.7 macrophages via suppressing the NF‑κB activation Do Thi Ha1*, Phung Thanh Long1, Tran Thi Hien2,3, Dao Trong Tuan4, Nguyen Thi Thuy An1,5, Nguyen Minh Khoi1, Ha Van Oanh6 and Tran Manh Hung7* Abstract Background: Vitis heyneana is widely distributed in the north of Vietnam, it has been used in Vietnamese traditional medicine as an agent for treatment of arthritis, bronchitis, carbuncles and infammatory conditions, and menstrual irregularities. However, this plant has not been investigated in phytochemical constituents and biological efects, especially in the anti-infammatory property. Results: Bioassay-guided fractionation of the EtOAc soluble fraction from the aerial part of Vitis heyneana resulted in the isolation of a series of oligostilbenoids as piceid (1), 2-r-viniferin (2), betulifol A (3), vitisinol C (4), (-)-trans-ε-viniferin (5), α-viniferin (6), shoreaketon (7), amurensin B (8), vitisinol B (9), and cis-vitisin B (10). Compound 5 showed the most potent inhibitory activities by suppressing LPS-induced COX-2 expression and PGE2 production. This compound exhibited signifcantly reduced LPS-induced nitric oxide (NO) release in a dose-dependent manner. These efects are accompanied with the inhibition of transcription factor NF-κB activation. Conclusion: The results suggested that trans-ε-viniferin exerts anti-infammatory efects via suppression the NF-κB activation in RAW 264.7 cells. Keywords: Vitis heyneana, (-)-Trans-ε-viniferin, COX-2, PGE2, NO, NF-κB Background resveratrol named stilbenoids [3–7]. To date, over 1000 Te genus Vitis (family Vitaceae) is in the major group stilbenoids have been identifed in various plant families with more than 66 species identifed and is distrib- such as Celastraceae, Cyperaceae, Fabaceae, Iritaceae, uted throughout the world [1]. In Vietnam, 6 species Moraceae, Paeoniaceae, and Vitaceae. Te Vitaceae of Vitis genus have been reported until now, including family includes about 900 species within 14–17 gen- V. larbusca L., V. heyneana Roem, & Schult., V. retordii era, primarily in tropical climate [8, 9]. Of these genera, Roman du Caill. Ex Planch. V. vinifera L., V. balansana only fve, the Ampelopsis, Cissus, Cyphostemma, Par- Planch., V. fexuosa Tunb [2]. Typical constituents thenocissus, and Vitis genera reported the presence of in Vitis genus have been reported to be oligomers of stilbenoids. However, chemical constituents of the spe- cies in Vitis genus have been studied the most. Stilbe- noids possessed various pharmacological activities such *Correspondence: [email protected]; [email protected] as antioxidative, anti-infammatory, and antimicrobial 1 Vietnam National Institute of Medicinal Materials, 3B Quangtrung, Hanoi, Vietnam activities, as well as having cardioprotective, hepatopro- 7 Department of Biomedical Sciences, Institute for Research and Executive tective, and neuroprotective efects [10–14]. Although Education (VNUK), The University of Danang, 41 Le Duan, Haichau district, approximately 100 stilbenoid monomers, dimers, and Danang 551000, Vietnam Full list of author information is available at the end of the article oligomers have been found in all Vitis species, research © The Author(s) 2018. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Ha et al. Chemistry Central Journal (2018) 12:14 Page 2 of 9 on the chemical compositions and biological activities of Vitis genus remains lacking [15]. Our screening anti- infammatory efect of an ethanol extract of 4 Vitis spe- cies including V. heyneana Roem. & Schult. (VH), Vitis vinifera L. (VV); Vitis balansana Planch. (VB), and Vitis labrusca L. (VL) collected in Vietnam via suppression of LPS-induced COX-2 expression found that 96% of ethanol extract of VH exhibited the most activity [16]. Besides, V. heyneana is widely distributed in northern Vietnam, for example in Cao Bang, Lang Son, and Lao Cai provinces. Te stems and roots of this species are traditionally used for the treatment of arthritis, bron- chitis, carbuncles and infammatory conditions, and menstrual irregularities in Vietnamese indigenous peo- ples [17]. So far, few studies have been done to investi- Fig. 1 COX-2 expression efects of V. heyneana extract (VH) and its gate the chemical constituents of VH. Currently, only a fractions (VHH: n-hexane, VHE: ethyl acetate, VHW: water fraction; all 50 µg/mL). Eighteen hours after treating cells with LPS (5 µg/mL) with few studies have referred to the presence of stilbenoids or without fractions in RAW 264.7 cells. Samples were harvested and in VH [17–19]. However, evaluation of anti-infamma- lysated for COX-2 mRNA level by qPCR. Relative changes in the COX-2 tory activities of the VH species have not been studied mRNA expression (*signifcant as compared to control, *, p < 0.05; yet. Tis study reports the anti-infammatory efects of **, p < 0.01; ***, p < 0.001; #signifcant as compared to LPS group, n 4–5) VH extracts and its isolated oligostilbenoids via sup- = pressing LPS-induced COX-2 expression, PGE2 and NO productions, and NF-κB activation in RAW 264.7 macrophages. Screening inhibitory activities of oligostilbenoids by suppressing LPS‑induced COX‑2 expression and PGE2 Results and discussion production in RAW 264.7 macrophages Screening inhibitory activities Te efects of all isolated compounds at concentration To study the cytotoxic efects of extract and its fractions of 10 µM, and a reference agent, meloxicam (20 µM) on cell viability, the RAW 264.7 cells were incubated were further examined on LPS-induced COX-2 protein, and treated a concentration of all materials (50 µg/mL). mRNA expression and LPS-induced PGE2 production in Te results showed that all the extract and fractions that RAW 246.7 cells by reported method with slight modif- induced cell toxicities were negligible at the above con- cation. As the results in Fig. 3, western blotting and real centrations [16]. In order to examine candidate extract/ time-PCR assays revealed that, among the active metabo- fractions inhibiting COX-2 in RAW 264.7 cells, ethanol lites, trans-ε-viniferin (5; 10 µM) potently suppressed V. heyneana extract (VH) and its fractions (n-hexane- LPS-induced COX-2 expression in RAW 246.7 cells VHH, ethyl acetate-VHE and water-VHW) were tested (Fig. 3a, b). Furthermore, ELISA assay confrmed that 5 on COX-2 mRNA expression level by qPCR. As shown in most inhibited the level of LPS-induced PGE2 produc- Fig. 1, the inhibitory efect of COX-2 mRNA was mostly tion in RAW 264.7 cells (Fig. 3c). potently suppressed by VHE and VH. Efects of 5 on COX‑2, iNOS expression and PGE2 Structure identifcation of the isolated compounds production in RAW 264.7 macrophages To investigate the active components from the poten- Considering that the 5 was the most active metabolite, tial fraction, several chromatographic techniques were the efects of this compound was investigated on LPS- applied and ten compounds were obtained after puri- induced PGE2 production in RAW 264.7 cells using fcation. On the basis of NMR spectroscopic analysis, ELISA method with slight modifcation. As the results in and in comparison with the previous studies, the chemi- Fig. 4a show, stimulation of cells resulted in the increase cal structures of these compounds were identifed as of PGE2 production compared with unstimulated vehi- piceid (1), 2-r-viniferin (2), betulifol A (3), vitisinol C cle cells. In the administration of 5 (1–30 µM), the PGE2 (4), (-)-trans-ε-viniferin (5), α-viniferin (6), shoreaketon productions were remarkably reduced in a concentra- (7), amurensin B (8), vitisinol B (9), and cis-vitisin B (10) tion dependent manner (p < 0.05). We further investi- (Fig. 2) [17, 20–26]. gated whether the inhibition efect by the same range of concentration of 5 was related to the gene expression, Ha et al. Chemistry Central Journal (2018) 12:14 Page 3 of 9 4 HO O 11' OH 1 O 11' OH OH 3' 13' 7 6'' 2' OH H 13' 4'' 1 7 10' 5'' O Ha 4' 8 H HO 8 10' 2 1' 9' H O 1 HO H 9 9' HO 2'' 3 5' 4 H H 3'' OH 1'' 9 HO 8' 6' 8' OH 13 7' 4 6 Hb 10 7' 1' 4' 11 5 1' HO 13 11 O OH OH OH 13'' 1 4'' 3 H 3 HO 11'' OH 9'' 8'' 10'' O 3 4' H 8''' O OH 7''' 12' 7'' 1 1''' 11' 7 HO H HO 4'' H 5 11 10''' 9''' 10' 8 OH OH 1' HO 7' 9' 7' 13 4''' 13 11''' 1' 13' 9 8' 9 13''' OH 3' 8' 9' 10' OH 11 8 OH 2 OH 1 HO 5' O 7 4 3' HO 13' OH 4 11' 1' 5 O 7' OH 11 3 O 11' 13' O 3'' 13' 7'' 13 8' 1 7 HO HO OH 11' OH 10 9' 1'' 3 O OH 11' 5 8 10' 14' 8'' 5'' 1 7 13' 9 10' 9' HO 5' 3 9 1 8 8'' O 9'' 8 7 10'' 11 8' 5 10' 9' 3' HO HO 7' 13'' 9'' 7'' 1'' OH 9 5 O 1' 13 11'' 11 7' 11'' 3'' OH 1' HO 8' 13'' HO 13 O OH 3'' 5'' OH OH 4'' 3' 5' OH 6 8 7'' 1'' HO 8'' 13''' 9'' 13'' OH 8''' HO 9''' 3 HO 11''' 11'' 7''' O H 5 3''' 1''' HO 1 O OH 14 H 7 11' 8 OH 5''' HO 10 12 9 13' 3''' OH H H 9' 12''' 5 10 8' 1''' HO O H 7' 9''' H H H HO 7''' O 11' 2' 1' 8'' R 10''' 8''' O 3 1 7 OH 7'' H H 8 11'' 14 13' 4' 1'' 9'' HO 9 10' 9' OH O R= H 5'' 13'' 12 7' 1'' 3'' 10 8' 3'' OH HO OH 1' 5'' 7 3' 5' OHC HO 9 Fig.