Establishment of a High Content Image Platform to Measure NF-κB Nuclear Translocation
in LPS-induced RAW264.7 Macrophages for Screening Anti-inflammatory
Drug Candidates

Yan-Yu      Zhang; Yun-Da      Yao; Qi-Qing      Cheng; Yu-Feng      Huang; Hua      Zhou

doi:10.2174/1389200223666220411121614

Abstract

Background: High Content Image (HCI), an automatic imaging and analysis system, provides a fast drug screening method by detecting the subcellular distribution of protein in intact cells.

Objective: This study established the first standardized HCI platform for lipopolysaccharide (LPS)-induced RAW264.7 macrophages to screen anti-inflammatory compounds by measuring nuclear factor-κB (NF-κB) nuclear translocation.

Methods: The influence of the cell passages, cell density, LPS induction time and concentration, antibody dilution, serum, dimethyl sulfoxide, and analysis parameters on NF-κB nuclear translocation and HCI data quality was optimized. The BAY-11-7085, the positive control for inhibiting NF-κB, and the Western blot assay were separately employed to verify the stability and reliability of the platform. Lastly, the effect of BHA on NO release, iNOS expression, IL-1β, IL-6, and TNF-α mRNA in LPS-induced RAW264.7 cells was detected.

Results: The optimal conditions for measuring NF-κB translocation in LPS-induced RAW264.7 cells by HCI were established. Cells that do not exceed 22 passages were seeded at a density of 10 k cells/well and pretreated with compounds following 200 ng/mL LPS for 40 min. Parameters including the nuclear area of 65 μm2, cell area of 80 μm2, collar of 0.9 μm, and sensitivity of 25% were recommended for image segmentation algorithms in the analysis workstation. Benzoylhypaconine from aconite was screened for the first time as an anti-inflammatory candidate by the established HCI platform. The inhibitory effect of benzoylhypaconine on NF-κB translocation was verified by Western blot. Furthermore, benzoylhypaconine reduced the release of NO, inhibited the expression of iNOS, and decreased the mRNA levels of IL-1β, IL-6, and TNF-α.

Conclusion: The established HCI platform could be applied to screen anti-inflammatory compounds by measuring the NF-κB nuclear translocation in LPS-induced RAW264.7 cells.

Keywords: High content image, lipopolysaccharide, RAW264.7 macrophages, nuclear factor-κB nuclear translocation, anti-inflammation, BAY 11-7085, benzoylhypaconine.

« Previous Next »

Graphical Abstract

[1] 
Yeung, Y.T.; Aziz, F.; Guerrero-Castilla, A.; Arguelles, S. Signaling pathways in inflammation and anti-inflammatory therapies. Curr. Pharm. Des.,  2018, 24(14), 1449-1484.
[http://dx.doi.org/10.2174/1381612824666180327165604] [PMID:  29589535] 
[2] 
Ivanenkov, Y.A.; Balakin, K.V.; Lavrovsky, Y. Small molecule inhibitors of NF-kB and JAK/STAT signal transduction pathways as promis-ing anti-inflammatory therapeutics. Mini Rev. Med. Chem.,  2011, 11(1), 55-78.
[http://dx.doi.org/10.2174/138955711793564079] [PMID:  21034406] 
[3] 
Yenmis, G.; Yaprak Sarac, E.; Besli, N.; Soydas, T.; Tastan, C.; Dilek Kancagi, D.; Yilanci, M.; Senol, K.; Karagulle, O.O.; Ekmekci, C.G.; Ovali, E.; Tuncdemir, M.; Ulutin, T.; Kanigur Sultuybek, G. Anti-cancer effect of metformin on the metastasis and invasion of primary breast cancer cells through mediating NF-kB activity. Acta Histochem.,  2021, 123(4), 151709.
[http://dx.doi.org/10.1016/j.acthis.2021.151709] [PMID:  33711726] 
[4] 
Hunto, S.T.; Kim, H.G.; Baek, K.S.; Jeong, D.; Kim, E.; Kim, J.H.; Cho, J.Y. Loratadine, an antihistamine drug, exhibits anti-inflammatory activity through suppression of the NF-kB pathway. Biochem. Pharmacol.,  2020, 177, 113949.
[http://dx.doi.org/10.1016/j.bcp.2020.113949] [PMID:  32251678] 
[5] 
Lai, J.L.; Liu, Y.H.; Liu, C.; Qi, M.P.; Liu, R.N.; Zhu, X.F.; Zhou, Q.G.; Chen, Y.Y.; Guo, A.Z.; Hu, C.M. Indirubin inhibits LPS-induced inflammation via TLR4 abrogation mediated by the NF-kB and MAPK signaling pathways. Inflammation,  2017, 40(1), 1-12.
[http://dx.doi.org/10.1007/s10753-016-0447-7] [PMID:  27718095] 
[6] 
Park, J.S.; Park, M.Y.; Cho, Y.J.; Lee, J.H.; Yoo, C.G.; Lee, C.T.; Lee, S.M. Anti-inflammatory effect of erdosteine in lipopolysaccharide-stimulated RAW 264.7 cells. Inflammation,  2016, 39(4), 1573-1581.
[http://dx.doi.org/10.1007/s10753-016-0393-4] [PMID:  27317418] 
[7] 
Nabel, G.; Baltimore, D. An inducible transcription factor activates expression of human immunodeficiency virus in T cells. Nature,  1987, 326(6114), 711-713.
[http://dx.doi.org/10.1038/326711a0] [PMID:  3031512] 
[8] 
Mitchell, J.P. Carmody, R.J. NF-κB and the transcriptional control of inflammation. Int. Rev. Cell Mol. Biol.,  2018, 335, 41-84.
[http://dx.doi.org/10.1016/bs.ircmb.2017.07.007] [PMID:  29305014] 
[9] 
Zhang, Q.; Lenardo, M.J.; Baltimore, D. 30 Years of NF-κB: A blossoming of relevance to human pathobiology. Cell,  2017, 168(1-2), 37-57.
[http://dx.doi.org/10.1016/j.cell.2016.12.012] [PMID:  28086098] 
[10] 
Chen, F.E.; Huang, D.B.; Chen, Y.Q.; Ghosh, G. Crystal structure of p50/p65 heterodimer of transcription factor NF-kappaB bound to DNA. Nature,  1998, 391(6665), 410-413.
[http://dx.doi.org/10.1038/34956] [PMID:  9450761] 
[11] 
Neurath, M.F.; Becker, C.; Barbulescu, K. Role of NF-kappaB in immune and inflammatory responses in the gut. Gut,  1998, 43(6), 856-860.
[http://dx.doi.org/10.1136/gut.43.6.856] [PMID:  9824616] 
[12] 
Bakkar, N.; Guttridge, D.C. NF-kappaB signaling: A tale of two pathways in skeletal myogenesis. Physiol. Rev.,  2010, 90(2), 495-511.
[http://dx.doi.org/10.1152/physrev.00040.2009] [PMID:  20393192] 
[13] 
Gilmore, T.D. Introduction to NF-kappaB: Players, pathways, perspectives. Oncogene,  2006, 25(51), 6680-6684.
[http://dx.doi.org/10.1038/sj.onc.1209954] [PMID:  17072321] 
[14] 
Gloire, G.; Legrand-Poels, S.; Piette, J. NF-kappaB activation by reactive oxygen species: Fifteen years later. Biochem. Pharmacol.,  2006, 72(11), 1493-1505.
[http://dx.doi.org/10.1016/j.bcp.2006.04.011] [PMID:  16723122] 
[15] 
Ewins, B.A.; Vassiliadou, M.; Minihane, A.M.; Rimbach, G.H.; Weinberg, P.D. Techniques for quantifying effects of dietary antioxidants on transcription factor translocation and nitric oxide production in cultured cells. Genes Nutr.,  2006, 1(2), 125-131.
[http://dx.doi.org/10.1007/BF02829954] [PMID:  18850206] 
[16] 
Malo, N.; Hanley, J.A.; Cerquozzi, S.; Pelletier, J.; Nadon, R. Statistical practice in high-throughput screening data analysis. Nat. Biotechnol.,  2006, 24(2), 167-175.
[http://dx.doi.org/10.1038/nbt1186] [PMID:  16465162] 
[17] 
Hill, A.A.; LaPan, P.; Li, Y.; Haney, S. Impact of image segmentation on high-content screening data quality for SK-BR-3 cells. BMC Bioinformatics,  2007, 8(1), 340.
[http://dx.doi.org/10.1186/1471-2105-8-340] [PMID:  17868449] 
[18] 
Boutros, M.; Heigwer, F.; Laufer, C. Microscopy-based high-content screening. Cell,  2015, 163(6), 1314-1325.
[http://dx.doi.org/10.1016/j.cell.2015.11.007] [PMID:  26638068] 
[19] 
Iannetti, E.F.; Willems, P.H.; Pellegrini, M.; Beyrath, J.; Smeitink, J.A.; Blanchet, L.; Koopman, W.J. Toward high-content screening of mito-chondrial morphology and membrane potential in living cells. Int. J. Biochem. Cell Biol.,  2015, 63, 66-70.
[http://dx.doi.org/10.1016/j.biocel.2015.01.020] [PMID:  25668473] 
[20] 
Kajihara, D. High content cell profiling as a tool for early drug safety testing using GE healthcare cardiomyocytes and IN Cell Analyzer 2000. In: Annual Meeting of the Japanese Society of Toxicology; , 2012; p. P-65.
[21] 
Trask, O.J. Nuclear factor kappa B (NF-κB) translocation assay development
and validation for high content screening. In: Markossian, S.;
Grossman, A.; Brimacombe, K.; et al., Eds. Assay Guidance Manual, Bethesda (MD): Eli Lilly & Company and the National Center for Advancing
Translational Sciences. 2004.
[22] 
Kelly, C.; Lawlor, C.; Burke, C.; Barlow, J.W.; Ramsey, J.M.; Jefferies, C.; Cryan, S.A. High-throughput methods for screening liposome-macrophage cell interaction. J. Liposome Res.,  2015, 25(3), 211-221.
[http://dx.doi.org/10.3109/08982104.2014.987785] [PMID:  25547801] 
[23] 
Njikan, S.; Manning, A.J.; Ovechkina, Y.; Awasthi, D.; Parish, T. High content, high-throughput screening for small molecule inducers of NF-κB translocation. PLoS One,  2018, 13(6), e0199966-e0199966.
[http://dx.doi.org/10.1371/journal.pone.0199966] [PMID:  29953522] 
[24] 
Koch, P.D.; Miller, H.R.; Yu, G.; Tallarico, J.A.; Sorger, P.K.; Wang, Y.; Feng, Y.; Thomas, J.R.; Ross, N.T.; Mitchison, T. A high content screen in macrophages identifies small molecule modulators of STING-IRF3 and NFkB signaling. ACS Chem. Biol.,  2018, 13(4), 1066-1081.
[http://dx.doi.org/10.1021/acschembio.7b01060] [PMID:  29553248] 
[25] 
Raschke, W.C.; Baird, S.; Ralph, P.; Nakoinz, I. Functional macrophage cell lines transformed by Abelson leukemia virus. Cell,  1978, 15(1), 261-267.
[http://dx.doi.org/10.1016/0092-8674(78)90101-0] [PMID:  212198] 
[26] 
Lio, C.K.; Luo, J.F.; Shen, X.Y.; Dai, Y.; Machado, J.; Xie, Y.; Yao, Y.D.; Yu, Y.; Liu, J.X.; Yao, X.S.; Luo, P.; Zhou, H. Nardosinanone N suppresses LPS-induced macrophage activation by modulating the Nrf2 pathway and mPGES-1. Biochem. Pharmacol.,  2020, 173, 113639.
[http://dx.doi.org/10.1016/j.bcp.2019.113639] [PMID:  31536727] 
[27] 
Zhou, H.; Liu, J.X.; Luo, J.F.; Cheng, C.S.; Leung, E.L.; Li, Y.; Su, X.H.; Liu, Z.Q.; Chen, T.B.; Duan, F.G.; Dong, Y.; Zuo, Y.H.; Li, C.; Lio, C.K.; Li, T.; Luo, P.; Xie, Y.; Yao, X.J.; Wang, P.X.; Liu, L. Suppressing mPGES-1 expression by sinomenine ameliorates inflammation and arthritis. Biochem. Pharmacol.,  2017, 142, 133-144.
[http://dx.doi.org/10.1016/j.bcp.2017.07.010] [PMID:  28711625] 
[28] 
Cui, F.; Jiang, L.; Qian, L.; Sun, W.; Tao, T.; Zan, X.; Yang, Y.; Wu, D.; Zhao, X. A macromolecular α-glucan from fruiting bodies of Volvariella volvacea activating RAW264. 7 macrophages through MAPKs pathway. Carbohydr. Polym.,  2020, 230, 115674.
[http://dx.doi.org/10.1016/j.carbpol.2019.115674] [PMID:  31887864] 
[29] 
Tabarsa, M.; Dabaghian, E.H.; You, S.; Yelithao, K.; Palanisamy, S.; Prabhu, N.M.; Li, C. Inducing inflammatory response in RAW264.7 and NK-92 cells by an arabinogalactan isolated from Ferula gummosa via NF-κB and MAPK signaling pathways. Carbohydr. Polym.,  2020, 241, 116358.
[http://dx.doi.org/10.1016/j.carbpol.2020.116358] [PMID:  32507213] 
[30] 
Davatelis, G.; Tekamp-Olson, P.; Wolpe, S.D.; Hermsen, K.; Luedke, C.; Gallegos, C.; Coit, D.; Merryweather, J.; Cerami, A. Cloning and characterization of a cDNA for murine Macrophage Inflammatory Protein (MIP), a novel monokine with inflammatory and chemokinetic properties. J. Exp. Med.,  1988, 167(6), 1939-1944.
[http://dx.doi.org/10.1084/jem.167.6.1939] [PMID:  3290382] 
[31] 
Park, S.; Shin, H.J.; Shah, M.; Cho, H.Y.; Anwar, M.A.; Achek, A.; Kwon, H.K.; Lee, B.; Yoo, T.H.; Choi, S. TLR4/MD2 specific peptides stalled in vivo LPS-induced immune exacerbation. Biomaterials,  2017, 126, 49-60.
[http://dx.doi.org/10.1016/j.biomaterials.2017.02.023] [PMID:  28254693] 
[32] 
Yang, S.; Yu, Z.; Wang, L.; Yuan, T.; Wang, X.; Zhang, X.; Wang, J.; Lv, Y.; Du, G. The natural product bergenin ameliorates lipopolysaccha-ride-induced acute lung injury by inhibiting NF-kappaB activition. J. Ethnopharmacol.,  2017, 200, 147-155.
[http://dx.doi.org/10.1016/j.jep.2017.02.013] [PMID:  28192201] 
[33] 
Zhang, X.; Yang, L.; Liu, Y.; Song, Z.; Zhao, J.; Chen, D.; Yu, H.; Li, R.; Wang, Y.; Yang, K.; Chen, Y.; Xia, M.; Zhang, L.W. Detection of nanocarrier potentiation on drug induced phospholipidosis in cultured cells and primary hepatocyte spheroids by high content imaging and analysis. Toxicol. Appl. Pharmacol.,  2018, 348, 54-66.
[http://dx.doi.org/10.1016/j.taap.2018.04.016] [PMID:  29678448] 
[34] 
Lv, X.; Chen, D.; Yang, L.; Zhu, N.; Li, J.; Zhao, J.; Hu, Z.; Wang, F.J.; Zhang, L.W. Comparative studies on the immunoregulatory effects of three polysaccharides using high content imaging system. Int. J. Biol. Macromol.,  2016, 86, 28-42.
[http://dx.doi.org/10.1016/j.ijbiomac.2016.01.048] [PMID:  26783639] 
[35] 
Syam, S. Bustamam, A.; Abdullah, R.; Sukari, M.A.; Hashim, N.M.; Mohan, S.; Looi, C.Y.; Wong, W.F.; Yahayu, M.A.; Abdelwahab, S.I. β Mangostin suppress LPS-induced inflammatory response in RAW 264.7 macrophages in vitro and carrageenan-induced peritonitis in vivo. J. Ethnopharmacol.,  2014, 153(2), 435-445.
[http://dx.doi.org/10.1016/j.jep.2014.02.051] [PMID:  24607509] 
[36] 
Schmidt, H.H.; Warner, T.D.; Nakane, M.; Förstermann, U.; Murad, F. Regulation and subcellular location of nitrogen oxide synthases in RAW264.7 macrophages. Mol. Pharmacol.,  1992, 41(4), 615-624.
[PMID: 1373797] 
[37] 
Harrill, J.A.; Robinette, B.L.; Mundy, W.R. Use of high content image analysis to detect chemical-induced changes in synaptogenesis in vitro. Toxicol. In Vitro,  2011, 25(1), 368-387.
[http://dx.doi.org/10.1016/j.tiv.2010.10.011] [PMID:  20969947] 
[38] 
Lin, S.; Schorpp, K.; Rothenaigner, I.; Hadian, K. Image-based high-content screening in drug discovery. Drug Discov. Today,  2020, 25(8), 1348-1361.
[http://dx.doi.org/10.1016/j.drudis.2020.06.001] [PMID:  32561299] 
[39] 
Bray, M.A.; Singh, S.; Han, H.; Davis, C.T.; Borgeson, B.; Hartland, C.; Kost-Alimova, M.; Gustafsdottir, S.M.; Gibson, C.C.; Carpenter, A.E. Cell Painting, a high-content image-based assay for morphological profiling using multiplexed fluorescent dyes. Nat. Protoc.,  2016, 11(9), 1757-1774.
[http://dx.doi.org/10.1038/nprot.2016.105] [PMID:  27560178] 
[40] 
Hussain, S.; Anees, A.; Das, A.; Nguyen, B.P.; Marzuki, M.; Lin, S.; Wright, G.; Singhal, A. High-content image generation for drug discovery using generative adversarial networks. Neural Netw.,  2020, 132, 353-363.
[http://dx.doi.org/10.1016/j.neunet.2020.09.007] [PMID:  32977280] 
[41] 
Mendes, T.; Herledan, A.; Leroux, F.; Deprez, B.; Lambert, J.C.; Kilinc, D. High-content screening for protein-protein interaction modulators using proximity ligation assay in primary neurons. Curr. Protoc. Cell Biol.,  2020, 86(1), e100.
[http://dx.doi.org/10.1002/cpcb.100] [PMID:  31876395] 
[42] 
Sero, J.E.; Sailem, H.Z.; Ardy, R.C.; Almuttaqi, H.; Zhang, T.; Bakal, C. Cell shape and the microenvironment regulate nuclear translocation of NF-κB in breast epithelial and tumor cells. Mol. Syst. Biol.,  2015, 11(3), 790.
[http://dx.doi.org/10.15252/msb.20145644] [PMID:  26148352] 
[43] 
Xiong, Q.; Zhang, L.; Xin, L.; Gao, Y.; Peng, Y.; Tang, P.; Ge, W. Proteomic study of different culture medium serum volume fractions on RANKL-dependent RAW264.7 cells differentiating into osteoclasts. Proteome Sci.,  2015, 13(1), 16.
[http://dx.doi.org/10.1186/s12953-015-0073-6] [PMID:  25969670] 
[44] 
Toma, L.; Stancu, C.S.; Botez, G.M.; Sima, A.V.; Simionescu, M. Irreversibly glycated LDL induce oxidative and inflammatory state in hu-man endothelial cells; added effect of high glucose. Biochem. Biophys. Res. Commun.,  2009, 390(3), 877-882.
[http://dx.doi.org/10.1016/j.bbrc.2009.10.066] [PMID:  19850013] 
[45] 
Ryu, J.H.; Park, H-J.; Jeong, Y-Y.; Han, S.; Shin, J-H.; Lee, S.J.; Kang, M.J.; Sung, N-J.; Kang, D. Aged red garlic extract suppresses nitric oxide production in lipopolysaccharide-treated RAW 264.7 macrophages through inhibition of NF-κB. J. Med. Food,  2015, 18(4), 439-445.
[46] 
Zhao, D.; Wang, J.; Cui, Y.; Wu, X. Pharmacological effects of Chinese herb aconite (fuzi) on cardiovascular system. J. Tradit. Chin. Med.,  2012, 32(3), 308-313.
[http://dx.doi.org/10.1016/S0254-6272(13)60030-8] [PMID:  23297548] 
[47] 
Bisset, N.G. Arrow poisons in China. Part II. Aconitum-botany, chemistry, and pharmacology. J. Ethnopharmacol.,  1981, 4(3), 247-336.
[48] 
Nyirimigabo, E.; Xu, Y.; Li, Y.; Wang, Y.; Agyemang, K.; Zhang, Y. A review on phytochemistry, pharmacology and toxicology studies of Aconitum. J. Pharm. Pharmacol.,  2015, 67(1), 1-19.
[http://dx.doi.org/10.1111/jphp.12310] [PMID:  25244533] 
[49] 
Gao, W.; Liu, X.G.; Liu, L.; Li, P.; Yang, H. Targeted profiling and relative quantification of benzoyl diterpene alkaloids in Aconitum roots by using LC-MS/MS with precursor ion scan. J. Sep. Sci.,  2018, 41(18), 3515-3526.
[http://dx.doi.org/10.1002/jssc.201800149] [PMID:  29974648] 
[50] 
Zhi, M.; Liu, K.; Han, S.; Xu, J.; Li, W.; Li, F.; Han, X.; Tang, Y.; Liu, Z.; Wang, H.; Du, H. Influence of different dosage forms on pharma-cokinetics of 6 alkaloids in raw Aconiti kusnezoffii radix (Caowu) and Chebulae fructus- (Hezi-) processed Caowu by UPLC-MS/MS. BioMed Res. Int.,  2020, 2020, 1942849.
[http://dx.doi.org/10.1155/2020/1942849] [PMID:  33029492] 
[51] 
Zhang, N.; Song, Y.; Song, Q.; Shi, S.; Zhang, Q.; Zhao, Y.; Li, J.; Tu, P. Qualitative and quantitative assessments of Aconiti lateralis radix praeparata using high-performance liquid chromatography coupled with diode array detection and hybrid ion trap-time-of-flight mass spec-trometry. J. Chromatogr. Sci.,  2016, 54(6), 888-901.
[http://dx.doi.org/10.1093/chromsci/bmv245] [PMID:  27048641] 
[52] 
Sun, Q.; Cao, H.; Zhou, Y.; Wang, X.; Jiang, H.; Gong, L.; Yang, Y.; Rong, R. Qualitative and quantitative analysis of the chemical constitu-ents in Mahuang-Fuzi-Xixin decoction based on high performance liquid chromatography combined with time-of-flight mass spectrometry and triple quadrupole mass spectrometers. Biomed. Chromatogr.,  2016, 30(11), 1820-1834.
[http://dx.doi.org/10.1002/bmc.3758] [PMID:  27183898] 
[53] 
Liu, X.; Li, H.; Song, X.; Qin, K.; Guo, H.; Wu, L.; Cai, H.; Cai, B. Comparative pharmacokinetics studies of benzoylhypaconine, benzoylme-saconine, benzoylaconine and hypaconitine in rats by LC-MS method after administration of radix Aconiti lateralis praeparata extract and Dahuang fuzi decoction. Biomed. Chromatogr.,  2014, 28(7), 966-973.
[http://dx.doi.org/10.1002/bmc.3102] [PMID:  24343604] 
[54] 
Yang, H.; Gao, W.; Liu, L.; Liu, K.; Liu, E.H.; Qi, L.W.; Li, P. Discovery of characteristic chemical markers for classification of aconite herbs by chromatographic profile and probabilistic neural network. J. Pharm. Biomed. Anal.,  2015, 115, 10-19.
[http://dx.doi.org/10.1016/j.jpba.2015.06.021] [PMID:  26142559] 
[55] 
Guzik, T.J.; Korbut, R.; Adamek-Guzik, T. Nitric oxide and superoxide in inflammation and immune regulation. J. Physiol. Pharmacol.,  2003, 54(4), 469-487.
[PMID: 14726604] 
[56] 
Pacher, P.; Beckman, J.S.; Liaudet, L. Nitric oxide and peroxynitrite in health and disease. Physiol. Rev.,  2007, 87(1), 315-424.
[http://dx.doi.org/10.1152/physrev.00029.2006] [PMID:  17237348] 
[57] 
Radomski, M.W.; Palmer, R.M.; Moncada, S. Glucocorticoids inhibit the expression of an inducible, but not the constitutive, nitric oxide synthase in vascular endothelial cells. Proc. Natl. Acad. Sci. USA,  1990, 87(24), 10043-10047.
[http://dx.doi.org/10.1073/pnas.87.24.10043] [PMID:  1702214] 
[58] 
Kim, Y.W.; Zhao, R.J.; Park, S.J.; Lee, J.R.; Cho, I.J.; Yang, C.H.; Kim, S.G.; Kim, S.C. Anti-inflammatory effects of liquiritigenin as a conse-quence of the inhibition of NF-kappaB-dependent iNOS and proinflammatory cytokines production. Br. J. Pharmacol.,  2008, 154(1), 165-173.
[http://dx.doi.org/10.1038/bjp.2008.79] [PMID:  18332856] 
[59] 
Ye, Q. Cardiotoxicity evaluation and comparison of diterpene alkaloids on zebrafish. Drug Chem. Toxicol.,  2021, 44(3), 294-301.
[PMID: 30895830] 
[60] 
Zhu, H.; Liu, S.; Li, X.; Song, F.; Liu, Z.; Liu, S. Bioactivity fingerprint analysis of cyclooxygenase-2 ligands from radix Aconiti by ultrafil-tration-UPLC-MSn. Anal. Bioanal. Chem.,  2013, 405(23), 7437-7445.
[http://dx.doi.org/10.1007/s00216-013-7153-1] [PMID:  23831827] 
[61] 
Hikino, H.; Konno, C.; Takata, H.; Yamada, Y.; Yamada, C.; Ohizumi, Y.; Sugio, K.; Fujimura, H. Antiinflammatory principles of Aconitum roots. J. Pharmacobiodyn.,  1980, 3(10), 514-525.
[http://dx.doi.org/10.1248/bpb1978.3.514] [PMID:  7205533] 

Rights & Permissions Print Cite

Article Metrics

25

1

Journal Information

For Authors

For Editors

For Reviewers

Explore Articles

Open Access

Open Access Articles

For Visitors

DOI https://dx.doi.org/10.2174/1389200223666220411121614	Print ISSN 1389-2002
Publisher Name Bentham Science Publisher	Online ISSN 1875-5453

Current Drug Metabolism

Establishment of a High Content Image Platform to Measure NF-κB Nuclear Translocation in LPS-induced RAW264.7 Macrophages for Screening Anti-inflammatory Drug Candidates

Abstract Play Pause

Graphical Abstract

Related Journals

Related Books

Abstract