Quercetin effectively improves LPS-induced intestinal inflammation, pyroptosis, and disruption of the barrier function through the TLR4/NF-κB/NLRP3 signaling pathway in vivo and in vitro

  • Hui-Xin Zhang Department of Veterinary Clinic Medicine, College of Veterinary Medicine, China Agricultural University, Beijing, P. R. China
  • Ye-Ye Li Department of Veterinary Clinic Medicine, College of Veterinary Medicine, China Agricultural University, Beijing, P. R. China
  • Zhong-Jie Liu Department of Veterinary Clinic Medicine, College of Veterinary Medicine, China Agricultural University, Beijing, P. R. China
  • Jiu-Feng Wang Department of Veterinary Clinic Medicine, College of Veterinary Medicine, China Agricultural University, Beijing 100193, P. R. China
Keywords: quercetin; LPS; pyroptosis; TLR4; NLRP3; GSDMD; interleukins; ZO-1; claudins; IBD


Background: Inflammatory bowel diseases are characterized by the alterations of the mucosa and gastrointestinal physiology, and the core of these alterations is endothelial cells. Quercetin is a flavonoid presents in some traditional Chinese medicine, plants, and fruits. Its protective effects in several gastrointestinal tumors have been demonstrated, but its effects on bacterial enteritis and pyroptosis-related diseases have rarely been studied.

Objective: This study aimed to evaluate the effect of quercetin on bacterial enteritis and pyroptosis.

Design: In vitro experiments were performed using rat intestinal microvascular endothelial cells divided into seven groups: control group (no treatment), model group (10 μg/mL lipopolysaccharide (LPS)+1 mM adenosine triphosphate [ATP]), LPS group (10 μg/mL LPS), ATP group (1 mM ATP), and treatment groups (10 μg/mL LPS+1 mM ATP and 5, 10, and 20 μM quercetin). The expression of pyroptosis-associated proteins, inflammatory factors, tight junction proteins, and the percentage of late apoptotic and necrotic cells were measured. In vivo analysis was performed using specific pathogen-free Kunming mice pretreated with quercetin and the water extract of Cacumen Platycladi for 2 weeks followed by 6 mg/kg LPS on day 15. Inflammation in the blood and intestinal pathological changes were evaluated.

Results: Quercetin used in vitro significantly reduced the expression of Toll-like receptor 4 (TLR4), NOD-like receptor 3 (NLRP3), caspase-1, gasdermin D, interleukin (IL)-1β, IL-18, IL-6, and tumor necrosis factor-α. It also inhibited phosphorylation of nuclear factor-kappa B (NF-κB) p65 and increased cell migration and the expression of zonula occludens 1 and claudins, while reduced the number of late apoptotic cells. The in vivo results showed that Cacumen Platycladi and quercetin significantly reduced inflammation, protected the structure of the colon and cecum, and prevent fecal occult blood induced by LPS.

Conclusions: These findings suggested the ability of quercetin to reduce inflammation induced by LPS and pyroptosis through TLR4/NF-κB/NLRP3 pathway.


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Kaelin WG, Ratcliffe PJ. Oxygen sensing by metazoans: the central role of the HIF hydroxylase pathway. Mol Cell 2008; 30: 393–402. doi: 10.1016/j.molcel.2008.04.009

Tolstanova G, Deng X, French SW, Lungo W, Paunovic B, Khomenko T, et al. Early endothelial damage and increased colonic vascular permeability in the development of experimental ulcerative colitis in rats and mice. Lab Invest 2012; 92: 9–21. doi: 10.1038/labinvest.2011.122

Saijo H, Tatsumi N, Arihiro S, Kato T, Okabe M, Tajiri H, et al. Microangiopathy triggers, and inducible nitric oxide synthase exacerbates dextran sulfate sodium-induced colitis. Lab Invest 2015; 95: 728–48. doi: 10.1038/labinvest.2015.60

Meera CH, Munashe C. Diagnosis and treatment of infectious enteritis in neonatal and juvenile ruminants. Vet Clin North Am Food Anim Pract 2018; 34: 101–17. doi: 10.1016/j.cvfa.2017.08.001

Spadoni I, Zagato E, Bertocchi A, Paolinelli R, Hot E, Sabatino AD, et al. A gut-vascular barrier controls the systemic dissemination of bacteria. Science 2015; 350: 830–4. doi: 10.1126/science.aad0135

Jorgensen I, Miao EA. Pyroptotic cell death defends against intracellular pathogens. Immunol Rev 2015; 265 (1): 130–42. doi: 10.1111/imr.12287

Nirmala J.C, Lopus M. Cell death mechanisms in eukaryotes. Cell Biol Toxicol 2020; 36(2): 145–64. doi: 10.1007/s10565-019-09496-2

Chen XL, Liu GL, Yuan YY, Wu GT, Wang SL, Yuan LW. NEK7 interacts with NLRP3 to modulate the pyroptosis in inflammatory bowel disease via NF-κB signaling. Cell Death Dis 2019; 10: 906. doi: 10.1038/s41419-019-2157-1

Liu BH, Hou L, Luo WJ, Wang CC, Wen YJ, Liu YL, et al. LPS and ATP co-induce the activation of NLRP3 inflammasome in macrophages. J BJ Univ Agric 2018; 33: 74–8. doi: 0.13473/j.cnki.issn.1002-3186.2018.0119

He Y, Hara H, Núñez G. Mechanism and regulation of NLRP3 inflammasome activation. Trends Biochem Sci 2016; 41: 1012–21. doi: 10.1016/j.tibs.2016.09.002

Swanson KV, Deng M, Ting JPY. The NLRP3 inflammasome: molecular activation and regulation to therapeutics. Nat Rev Immunol 2019; 19: 477–89. doi: 10.1038/s41577-019-0165-0

He CL, Zhao Y, Jiang XL, Liang XX, Yin LZ, Yin ZQ, et al. Protective effect of Ketone musk on LPS/ATP-induced pyroptosis in J774A.1 cells through suppressing NLRP3/GSDMD pathway. Int Immunopharmacol 2019; 71: 328–35. doi: 10.1016/j.intimp.2019.03.054

Mezzasoma L, Antognelli C, Talesa VN. Atrial natriuretic peptide down-regulates LPS/ATP-mediated IL-1 beta release by inhibiting NF-kB, NLRP3 inflammasome and caspase-1 activation in THP-1 cells. Immunol Res 2016; 64: 303–12. doi: 10.1007/s12026-015-8751-0

Groß CJ, Mishra R, Schneider KS, Medard G, Wettmarshausen J, Dittlein DC, et al. K+ efflux-independent NLRP3 inflammasome activation by small molecules targeting mitochondria. Immunity 2016; 45: 761–73. doi: 10.1016/j.immuni.2016.08.010

Zhang J, Wu GH, Shan AS, Han Y, Jin YC, Fang HT, et al. Dietary glutamine supplementation enhances expression of ZO-1 and occludin and promotes intestinal development in Min piglets. Acta Agric Scand A Anim Sci 2017; 67: 15–21. doi: 10.1080/09064702.2017.1333133

Xu J, Liu ZH, Zhan W, Jiang R, Yang CH, Zhan H, et al. Recombinant TsP53 modulates intestinal epithelial barrier integrity via upregulation of ZO-1 in LPS-induced septic mice. Mol Med Rep 2018; 17: 1212–8. doi: 10.3892/mmr.2017.7946

Tian SY, Guo RX, Wei SC, Kong Y, Wei XL, Wang WW, et al. Curcumin protects against the intestinal ischemia-reperfusion injury: involvement of the tight junction protein ZO-1 and TNF-α related mechanism. Korean J Physiol Pharmacol 2016; 20(2): 147–52. doi: 10.4196/kjpp.2016.20.2.147

Hu RZ, He ZY, Liu M, Tan JJ, Zhang HF, Hou DX, et al. Dietary protocatechuic acid ameliorates inflammation and upregulates intestinal tight junction proteins by modulating gut microbiota in LPS-challenged piglets. J Anim Husbandry Biotechnol 2021; 12: 328–39. doi: 10.1186/s40104-020-00492-9

Escudero-Esparza A, Jiang WG, Martin TA. The Claudin family and its role in cancer and metastasis. Front Biosci 2011; 16: 1069–83. doi: 10.2741/3736

Ya ZC, Ozcan CU, Hismioguliari AA, Sunay FB, Ozcan T, Berksoy EA, et al. Protective effects of quercetin on necrotizing enterocolitis in a neonatal rat model. Am J Perinatol 2018; 35: 434–40. doi: 10.1055/s-0037-1608660

Chen WS, Zhou LL, Qiao Y. Quality evaluation of ilex asprella based on simultaneous determination of five bioactive components, chlorogenic acid, luteoloside, quercitrin, quercetin, and kaempferol, using UPLC-Q-TOF MS study. J AOAC Int 2019; 102: 1414–22. doi: 10.5740/jaoacint.18-0391

Truong, VL, Ko SY, Jun M, Jeong WS. Quercitrin from toona sinensis (Juss.) M.Roem. attenuates acetaminophen-induced acute liver toxicity in HepG2 cells and mice through induction of antioxidant machinery and inhibition of inflammation. Nutrients 2016; 8: 431. doi: 10.3390/nu8070431

Zhi KK, Li MQ, Bai J, Wu YF, Zhou SL, Zhang XP, et al. Quercitrin treatment protects endothelial progenitor cells from oxidative damage via inducing autophagy through extracellular signal-regulated kinase. Angiogenesis 2016; 19: 311–24. doi: 10.1007/s10456-016-9504-y

Zhou XH, Xu FL, Chen J. Effect of gemcitabine combined with Quercetin on apoptosis of pancreatic cancer cell Panc-1. Med Health Technol 2013; 35: 542–6.

Suzuki T, Hara H. Role of flavonoids in intestinal tight junction regulation. J Nutr Biochem 2011; 22: 401–8. doi: 10.1016/j.jnutbio.2010.08.001

Tamura M, Matsuo Y, Nakagawa H, Hoshi C, Hori S. Isolation of a quercetin-metabolizing bacterium 19- 20 from human feces. Food Sci Technol Res 2017; 23: 145–50. doi: 10.3136/fstr.23.145

Li R, Lin C, Yao GM, Yan HL, Wang L. Effects of Quercetin on diabetic retinopathy and its association with NLRP3 inflammasome and autophagy. Int J Ophthalmol 2021; 14: 42–9. doi: 10.18240/ijo.2021.01.06

Su KY, Yu CY, Chen YP, Hua KF, Chen YLS. 3,4-Dihydroxytoluene, a metabolite of rutin, inhibits inflammatory responses in lipopolysaccharide-activated macrophages by reducing the activation of NF-κB signaling. BMC Complement Med Ther 2014; 14: 21. doi: 10.1186/1472-6882-14-21

Liu P, Bian YF, Zhong J, Yang Y, Mu X, Liu ZJ. Establishment and characterization of a rat intestinal microvascular endothelial cell line. Tissue Cell 2021; 72: 101573. doi: 10.1016/j.tice.2021.101573

Coll RC, Robertson A, Chae JJ, Higgins SC, Muñoz-Planillo R, Inserra MC, et al. A small-molecule inhibitor of the NLRP3 inflammasome for the treatment of inflammatory diseases. Nat Med 2015; 21: 248–55. doi: 10.1038/nm.3806

Zhang X, Ke X, Chen JT, Gao YL, Zhou F. Dynamic expression and significance of IL-1β and IL-4 in ulcerative colitis rats with dampness and heat syndrome. J xi’an jiaotong Univ (med Sci) 2015; 36(5): 697–701. doi: 10.7652/jdxb201505024

Kong Q, Li Z, Zhao B, Duan WN, Qiu Z, Huang Q, et al. The role of TIPE2 in pyrophosis of mouse macrophages: evaluation by siRNA technique. Chin J Anesthesiol 2019; 39: 482–5. doi: 10.3760/cma.j.issn.0254-1416.2019.04.026

Shi DH, Dai YP, Wang LF, Zhou Q, Zhang XL, Zhang J. Analysis of chemical constituents of Platycladus orientalis leaves before and after charcoal frying based on UHPLC-QTOF-MS/MS identification. Chin J Exp Formulae 2021; 27: 107–16. doi: 10.13422/j.cnki.syfjx.20202354

Lei M, Dai JK, Cao D, Liu X, Wang LQ, Zhao QQ, et al. Research progress on chemical constituents and pharmacological effects of platycladus orientalis leaves and seeds. Chem Life 2018; 38: 281–9. doi: 10.13488/j.smhx.20180216

Zhao L, Li MY, Sun KC, Su S, Geng TT, Sun H. Hippophae rhamnoides polysaccharides protect IPEC-J2 cells from LPS-induced inflammation, apoptosis and barrier dysfunction in vitro via inhibiting TLR4/NF-kappa B signaling pathway. Int J Biol Macromol 2020; 155: 1202–15. doi: 10.1016/j.ijbiomac.2019.11.088

Zhang RY, Dong LN, Li FX, Liu P, Wang JP. Correlation between fecal and intestinal mucosal flora and tight junction protein ZO-1 in patients with functional gastrointestinal disease. Chin Med 2017; 12: 1193–6. doi: 10.3760/cma.j.issn.16734777.2017.08.019

Tornavaca O, Chia M, Dufton N, Almagro LO, Conway DE, Randi AM, et al. ZO-1 controls endothelial adherens junctions, cell-cell tension, angiogenesis, and barrier formation. J Cell Biol 2015; 208: 821–38. doi: 10.1083/jcb.201404140

Garcia-Hernandez V, Quiros M, Nusrat A. Intestinal epithelial claudins: expression and regulation in homeostasis and inflammation. HHS Public Access 2017; 1397(1): 66–7. doi: 10.1111/nyas.13360

Barrett KE. Claudin-2 pore causes leak that breaches the dam in intestinal inflammation. J Clin Invest 2020; 130(10): 5100–1. doi: 10.1172/JCI140528

Bu N, Wang Y, Wang R, Sun LT, Fan YQ. Experimental study of tripterygium polyglycosides through JAK2/STAT3 signaling pathway to reduce intestinal mucosal cell apoptosis and inflammation in rats with ulcerative colitis. Mod Digest Interv Diagn Treat 2019; 24: 466–70. doi: 10.3969/j.issn.1672-2159.2019.05.004

Lin CF, Kuo YT, Chen TY, Chien CT. Quercetin-rich guava (psidium guajava) juice in combination with trehalose reduces autophagy, apoptosis and pyroptosis formation in the kidney and pancreas of type II diabetic rats. Molecules 2016; 21: 334. doi: 10.3390/molecules21030334

Han XJ, Xu TS, Fang QJ, Zhang HJ, Yue LJ, Hu G, et al. Quercetin hinders microglial activation to alleviate neurotoxicity via the interplay between NLRP3 inflammasome and mitophagy. Redox Biol 2021; 44: 102010. doi: 10.1016/j.redox.2021.102010

Luo X, Bao XY, Weng XZ, Bai XX, Feng Y, Huang JX, et al. The protective effect of Quercetin on macrophage pyroptosis via TLR2/Myd88/NF-kappaB and ROS/AMPK pathway. Life Sci 2021; 291: 120064. doi: 10.1016/j.lfs.2021.120064

Huan F, Cheng J, Jin SX, Xu JH, Wang J, Xiao H. Study on the genotoxicity of Quercetin in mammalian cells in vitro. Chin J Prev Med 2010; 11: 797–800. doi: 10.16506/j.1009-6639.2010.08.011

Qiao Y. TLR-induced NF-KB activation regulates NLRP3 expression in murine macrophages. FEBS Lett 2012; 586(7): 1022–6. doi: 10.1016/j.febslet.2012.02.045

Cheon PJ, Nam JC, Nan HH, Choo L, Kil B. The functional effects on anti-oxidant and anti-inflammation of veronica persica Poir. Extracts. Korea J Org Agric 2018; 26: 661–76. doi: 10.11625/KJOA.2018.26.4.661

Li HX, Zhou LL, Zhi ZK, Lv XR, Wei ZH, Zhang X, et al. Lipopolysaccharide upregulates miR-132/212 in Hirschsprung-associated enterocolitis, facilitating pyroptosis by activating NLRP3 inflammasome via targeting Sirtuin 1 (SIRT1). AGING-US 2020; 12: 18588–602. doi: 10.18632/aging.103852

Liu ZX, Chen YZ, Xu JM, An N, Zhang J, Gu SL, et al. Heme-activated pyrin domain-containing NOD-like receptor family protein 3 (NLRP3) inflammasome induces renal tubular epithelial cell pyrolysis. J Cell Mol Immunol 2020; 36: 809–14. doi: 10.13423/j.cnki.cjcmi.009067

Dong WJ, Zhu QS, Yang BW, Qin Q, Wang YW, Xia XM, et al. Polychlorinated biphenyl quinone induces caspase 1-mediated pyroptosis through induction of pro-inflammatory HMGB1-TLR4-NLRP3-GSDMD signal axis. Chem Res Toxicol 2019; 32: 1051–7. doi: 10.1021/acs.chemrestox.8b00376

Jiang WD, Wen HL, Liu Y, Jiang J, Kuang SY, Wu P, et al. The tight junction protein transcript abundance changes and oxidative damage by tryptophan deficiency or excess are related to the modulation of the signalling molecules, NF-kappa B p65, TOR, caspase-(3,8,9) and Nrf2 mRNA levels, in the gill of young grass carp (Ctenopharyngodon idellus). Fish Shellfish Immunol 2015; 46: 168–80. doi: 10.1016/j.fsi.2015.06.002

Feng L, Gan L, Jiang WD, Wu P, Liu Y, Jiang J, et al. Gill structural integrity changes in fish deficient or excessive in dietary isoleucine: towards the modulation of tight junction protein, inflammation, apoptosis and antioxidant defense via NF-kappa B, TOR and Nrf2 signaling pathways. Fish Shellfish Immunol 2017; 63: 127–38. doi: 10.1016/j.fsi.2017.02.010

Park SY, Jang H, Kim SY, Kim D, Park Y, Kee SH. Expression of E-cadherin in epithelial cancer cells increases cell motility and directionality through the localization of ZO-1 during collective cell migration. Bioengineering 2020; 8: 65. doi: 10.3390/bioengineering8050065

Bai YL, Zhu XD, Lan YZ, Xu KY, Song N, Fan LH. Clinical application and dosage of Arborvitae orientalis. J CH Univ Tradit Chin Med 2020; 36(06): 1123–6. doi: 10.13463/j.cnki.cczyy.2020.06.009

Yang HF, Li YJ, Lu GF, Jiang CM. Clinical expansion trial of rhubarb orientalis leaf mixture. Jiangsu Agric Sci 2012; 40(12): 235–6. doi: 10.15889/j.issn.1002-1302.2012.12.163

Kawabata K, Kato Y, Sakano T, Baba N, Hagiwara K, Tamura A, et al. Effects of phytochemicals on in vitro anti-inflammatory activity of Bifidobacterium adolescentis. Biosci Biotechnol Biochem 2015; 79: 799–807. doi: 10.1080/09168451.2015.1006566
How to Cite
Zhang H.-X., Li Y.-Y., Liu Z.-J., & Wang J.-F. (2022). Quercetin effectively improves LPS-induced intestinal inflammation, pyroptosis, and disruption of the barrier function through the TLR4/NF-κB/NLRP3 signaling pathway<em> in vivo</em> and <em>in vitro</em&gt;. Food & Nutrition Research, 66. https://doi.org/10.29219/fnr.v66.8948
Original Articles