İn Vitro Lipopolisakkarit Uyarımlı Karaciğer Yangı Modelinde Alfa-Terpineol’ün Etkinliğinin Araştırılması

Year 2022, Volume: 33 Issue: 1, 71 - 78, 15.06.2022

Ayşe Pınar Tunçer Altuğ Küçükgül Mehmet Mustafa İşgör

https://doi.org/10.35864/evmd.1053710

Abstract

Terpinoller monoterpen bileşikler olup, birçok çalışmada antioksidan özelliklerinden dolayı antikanser, antikonvülsant ve antiülser gibi biyolojik etkileri ortaya konulmuştur. Araştırmanın amacını, LPS’in karaciğer hücrelerine uygulanarak oluşturulan yangı modelinde, α-terpineol’ün antiinflamatuar ve antiapoptotik biyofonksiyonlarının araştırılması oluşturmuştur. Denemede insan orijinli HepG2 (ATCC® HB-8065) hücreleri materyal olarak seçilmiştir. LPS ve α-terpineol, hücrelere farklı konsantrasyonlarda 24 saat uygulanmıştır ve etkin konsantrasyonları, hücre canlılık testleriyle (MTT) gerçekleştirilmiştir. Sonrasında, TNF-α, IL-1β, IL-10, Kaspaz 3, Bax ve Bcl-2 gen ekspresyon düzeyleri tam zamanlı kantatif polimeraz zincir reaksiyonuyla (qRT-PCR) araştırılmıştır. LPS’in 50 ng/ml konsantrasyonu %11,5 oranında hücre kayıplarına neden olduğu belirlenmiş ve modelleme için bu konsantrasyon seçilmiştir. Ancak, 10 μM konsantrasyonda α-terpineolün hücre kayıplarını % 2.12 oranında önlediği bulunmuştur. Çalışmada, LPS’in TNF-α ve IL-1β gen ekspresyonlarını arttırdığı, α-terpineol uygulamasının ise bu durumu tersine çevirdiği tespit edilmiştir. Yine, IL-10 gen ekspresyonu yüksek LPS konsantrasyonu ile baskılanırken, α-terpineol tarafından anlamlı düzeyde uyarıldığı da ortaya konulmuştur. Bununla birlikte, LPS’in kaspaz 3 ve Bax gen ekspresyonlarını arttırdığı, ancak α-terpineol’ün bu stimülasyonu baskıladığı tespit edilmiştir. Bcl-2 gen ekspresyonları ise LPS tarafından baskılanırken, α-terpineol tarafından uyarıldığı bulunmuştur. Özetle, α-terpineol’ün kısa süreli ve düşük olan proliferatif konsantrasyonunun özellikle patojen kaynaklı karaciğer rahatsızlıklarında alternatif bir tedavi ajanı olabileceği görülmüştür.

Keywords

α- terpineol, apoptoz, inflamasyon, lipopolisakkarit

Investigation of The Efficiency Of Alpha-Terpineol in the In Vitro Lipopolisacharide Stimulated Liver Inflammation Model

Abstract

Terpinols are monoterpene compounds, and biological effects like anticancer, anticonvulsant and antiulcer have been demonstrated in many studies due to their antioxidant properties. The aim of the research was to investigate the anti-inflammatory and antiapoptotic biofunctions of α-terpineol in LPS induced inflammation model of liver cells. HepG2 (ATCC® HB-8065) cells of human origin were selected as material. LPS and α-terpineol were applied to cells at different concentrations for 24 hours, and their effective concentrations were determined by cell viability tests (MTT). Afterwards, gene expression levels of TNF-α, IL-1β, IL-10, Caspase 3, Bax and Bcl-2 were investigated by qRT-PCR method. LPS at 50 ng/ml, caused 11.5% cell loss and was chosen for modeling. However, α-terpineol at 10 μM concentration prevented 2.12% of cell loss. In the study, LPS increased TNF-α and IL-1β gene expressions, and α-terpineol application reversed this situation. Again, while IL-10 was suppressed by high LPS concentration, it was significantly stimulated by α-terpineol. Besides, LPS increased caspase 3 and Bax gene expressions, but α-terpineol suppressed this stimulation. Bcl-2 gene expressions were suppressed by LPS and stimulated by α-terpineol. In summary, short-term and low proliferative concentration of α-terpineol can be an alternative treatment agent, especially in pathogen-induced liver disorders.

Keywords

α-terpineol, apoptosis, inflammation, lipopolysaccharide

References

• Armstrong MJ, Adams LA, Canbay A, Syn WK. (2014) Extrahepatic complications of nonalcoholic fatty liver disease. Hepatology. 59, 1174–1197.
• Ashtari S, Pourhoseingholi MA, Zali MR. (2015) Non-alcohol fatty liver disease in Asia: prevention and planning. World J Hepatol. 7, 1788–1796.
• Bauer K, Garbe D, Surburg H. (2001) Common Fragrance and Flavor materials: Preparations, properties, and uses. Fourth edition. New York: Wiley online boks.
• Brand C, Ferrante A, Prager RH, Riley TV, Carson CF, Finaly-Jones JJ, et al. (2001) The water-soluble components of the essential oil of Melaleuca alternifolia (tea tree oil) suppress the production of superoxide by human monocytes, but not neutrophils, activated in vitro. Inflamm. Res. 50(4), 213-219.
• Bubici C, Papa S, Pham CG, Zazzeroni F, Franzoso G. (2004) NF-kappaB and JNK: An intricate affair. Cell Cycle. 3, 1524-9.
• Buzzetti E, Pinzani M, Tsochatzis EA. (2016) The multiple-hit pathogenesis of non-alcoholic fatty liver disease (NAFLD). Metabolism. 65, 1038–1048.
• Capano M and Crompton M. (2002) Biphasic translocation of Bax to mitochondria. Biochem J. 367, 169-78.
• Chakraborti CK. (2015) New-found link between microbiota and obesity. World J Gastrointest Pathophysiol. 6, 110–119.
• Chalasani N, Younossi Z, Lavine JE, Charlton M, Cusi K, Rinella M, Harrison SA, Brunt EM, Sanyal AJ. (2018) The diagnosis and management of nonalcoholic fatty liver disease: practice guidance from the American Association for the Study of Liver Diseases. Hepatology. 67, 328–357.
• Choi Y, Sim WC, Choi HK, Lee S, Lee BH. (2013) α-Terpineol induces fatty liver in mice mediated by the AMP-activated kinase and sterol response element binding protein pathway. Food and Chemical Toxicology. 55, 129–136.
• Cintra DE, Pauli JR, Araujo EP, Moraes JC, de Souza CT, Milanski M, Morari J, Gambero A, Saad MJ and Velloso LA. (2008) Interleukin-10 is a protective factor against diet-induced insulin resistance in liver. Journal of Hepatology. 48, 628–637.
• De Oliveira MG, Marques RB, de Santana MF, Santos AB, Brito FA, Barreto EO, De Sousa DP, Almeida FR, Badauê-Passos D Jr, Antoniolli AR, Quintans-Júnior LJ. (2012) α-terpineol reduces mechanical hypernociception and inflammatory response. Basic Clin Pharmacol Toxicol. 111(2), 120-5.
• Dela Peña A, Leclercq I, Field J, George J, Jones B, Farrell G. (2005) NF-kappaB activation, rather than TNF, mediates hepatic inflammation in a murine dietary model of steatohepatitis. Gastroenterology. 129, 1663–1674.
• Duarte N, Coelho IC, Patarrão RS, Almeida JI, Penha-Gonçalves C, Macedo MP. (2015) How inflammation impinges on NAFLD: a role for Kupffer cells. BioMed Res Int. 2015, 984578.
• Hassan SB, Muhtasib HG, Goeransson H, Larsson R. (2010) Alpha-terpineol: a potential anticancer agent which acts through suppressing NF-κB signaling. Anticancer Res. 30(6), 1911-1920.
• Held S, Schieberle P, Somoza V. (2007) Characterization of alpha-terpineol as an anti-inflammatory component of orange juice by in vitro studies using oral buccal cells. J. Agric. Food Chem. 55(20), 8040-8046.
• Hirsova P and Gores GJ. (2015) Death receptor-mediated cell death and proinflammatory signaling in nonalcoholic steatohepatitis. Cell Mol Gastroenterol Hepatol. 1, 17–27. Khaleel C, Tabanca N, Buchbauer G. (2018) α-Terpineol, a natural monoterpene: A review of its biological properties. Open Chem. 16: 349–361.
• Kucukgul A and Erdogan S. (2014) Caffeic acid phenethyl ester (CAPE) protects lung epithelial cells against H2O2-induced inflammation and oxidative stress. HealthMED. 8(3), 329-338.
• Lampronti I, Saab AM, Gambari A. (2006) Antiproliferative activity of essential oils derived from plants belonging to the Magnoliophyta division. Int. J. Oncol. 29(4), 989-995. Lucas C, Lucas G, Lucas N, Krzowska-Firych J, Tomasiewicz K. (2018) A systematic review of the present and future of non-alcoholic fatty liver disease. Clin Exp Hepatol. 4, 165–174.
• Machado MV and Diehl AM. (2016) Pathogenesis of Nonalcoholic Steatohepatitis. Gastroenterology. 150, 1769–77.
• Mazzotti A, Caletti MT, Sasdelli AS, Brodosi L, Marchesini G. (2016) Pathophysiology of nonalcoholic fatty liver disease: lifestyle-gut-gene interaction. Dig Dis. Suppl 1, 3–10. Nishio K, Horie M, Yoko MA, Hitoshi S, Yoshihisa I, Yasukazu H, Niki YE. (2013) Attenuation of lipopolysaccharide (LPS)-induced cytotoxicityby tocopherols and tocotrienols. Redox Biology. 1(1):97-103.
• Nogueira MNM, Aquino SG, Rossa JC, Spolidorio DMP. (2014) Terpinen-4-ol and alpha-terpineol (tea tree oil components) inhibit the production of IL-1b, IL-6 and IL-10 on human macrophages. Inflamm. Res. 63, 769–778.
• Poeta M, Pierri L, Vajro P. (2017) Gut-liver axis derangement in non-alcoholic fatty liver disease. Children (Basel). 4(8), 66.
• Pourbakhsh H, Taghiabadi E, Abnous K, Timcheh Hariri A, Hosseini SM, Hosseinzadeh H. (2014) Effect of Nigella sativa fixed oil on ethanol toxicity in rats. Iran J Basic Med Sci. 17(12), 1020-1031.
• Rossol M, Heine H, Meusch U, Quandt D, Klein C, Sweet MJ, Hauschildt S. (2011) LPS-induced cytokine production in human monocytes and macrophages. Critical Reviews in Immunology. 31(5), 379-446.
• Sabino CK, Ferreria-Filho ES, Mendes MB, Da Silva-Filho JC, et al. (2013) Cardiovascular effects induced by α-terpineol in hypertensive rats. Flavour Frag. J. 28(5), 333-339.
• Sales A, Felipe LO, Bicas JL. (2020) Production, Properties, and Applications of α-Terpineol. Food and Bioprocess Technology. doi: 10.1007/s11947-020-02461-6.
• Sell C. (2003) A Fragrant Introduction to terpenoid chemistry, 1st ed., The Royal Society of Chemistry, Cambridge, UK.
• Shi H, Yang YG, Binlin L, Xiaoyu SG, Lu YJS. (2016) The in vitro effect of lipopolysaccharide on proliferation, inflammatoryfactors and antioxidant enzyme activity in bovine mammary epithelial cells. Animal Nutrition. 2, 99-104.
• Sousa GM, Cazarin CBB, Maróstica Junior MR, et al. (2020) The effect of α-terpineol enantiomers on biomarkers of rats fed a high-fat diet. Heliyon. 6(4), e03752.
• Stone WL, Qui M, Smith M. (2003) Lipopolysaccharide enhances the cytotoxicity of 2-chloroethyl ethyl sulfide. BMC Cell Biology. 4, 1-7.
• Ting Ge Y, Zhong AQ, Xu GF, Lu Ying. (2019) Resveratrol Protects BV2 Mouse Microglial Cells Against LPS-induced Inflammatory Injury by Altering the miR-146a-5p/TRAF6/NF-κB Axis. Immunopharmacol Immunotoxicol. 41(5), 549-557.
• Valentijn AJ, Metcalfe AD, Kott J, Streuli CH, Gilmore AP. (2003) Spatial and temporal changes in Bax subcellular localization during anoikis. J Cell Biol. 162, 599-612.
• Wolff A, 1904. Uber Grundgesetze der Immunitat. Zentralbl Bakteriol. 37, 390–397.
• Yang H, Young DW, Gusovsky F and Chow JC. (2000) Cellular events mediated by lipopolysaccharide-stimulated tolllike receptor 4. MD-2 is required for activation of mitogen-activated protein kinases and Elk-1. J Biol Chem. 275, 20861–20866.
• Younossi ZM, Koenig AB, Abdelatif D, et al. (2016) Global epidemiology of nonalcoholic fatty liver disease-meta analytic assessment of prevalence, incidence, and outcomes. Hepatology. 64, 73–84.
• Zhang L, Xing D, Liu L, Gao X, Chen M. (2007) TNF-α Induces Apoptosis Through JNK/BaxDependent Pathway in Differentiated, but not Naïve PC12 Cells. Cell Cycle. 6(12), 1479-1486.
• Zhu J, Liu M, Kennedy RH, Liu SJ. (2006) TNFalpha-induced impairment of mitochondrial integrity and apoptosis mediated by caspase-8 in adult ventricular myocytes. Cytokine. 34, 96-105.