This study aimed to examine the effect of dietary flavonoid isoquercitrin on ovarian granulosa cells using the immortalized human cell line HGL5. Cell viability, survival, apoptosis, release of steroid hormones 17beta-estradiol and progesterone, and human transforming growth factor-beta2 (TGF-beta2) and TGF-beta2 receptor as well as intracellular reactive oxygen species (ROS) generation were investigated after isoquercitrin treatment at the concentration range of 5-100 microg.ml-1. It did not cause any significant change (p>0.05) in cell viability as studied by AlamarBlue assay in comparison to control. No significant change was observed (p>0.05) in the proportion of live, dead and apoptotic cells as revealed by apoptotic assay using flow cytometry. Similarly, the release of 17beta-estradiol, progesterone, TGF-beta2 and its receptor were not affected significantly (p>0.05) by isoquercitrin as detected by ELISA, in comparison to control. Except for the highest concentration of 100 microg.ml-1, which led to oxidative stress, isoquercitrin exhibited antioxidative activity at lower concentration used in the study (5, 10, 25, and 50 microg.ml-1) by hampering the production of intracellular ROS, in comparison to control, as detected by chemiluminescence assay (p<0.05). Findings of the present study indicate an existence of the antioxidative pathway that involves inhibition of intracellular ROS generation by isoquercitrin in human ovarian granulosa cells.

Department of Animal Physiology Faculty of Biotechnology and Food Sciences Slovak University of Agriculture in Nitra Nitra Slovak Republic

Zobrazit více v PubMed

AI A, TANG Z, LIU Y, YU S, LI B, HUANG H, WANG X, CAO Y, ZHANG W. Characterization and identification of human immortalized granulosa cells derived from ovarian follicular fluid. Exp Ther Med. 2019;18:2167–2177. doi: 10.3892/etm.2019.7802. PubMed DOI PMC

AMADO NG, CERQUEIRA DM, MENEZES FS, Da SILVA JF, NETO VM, ABREU JG. Isoquercitrin isolated from hyptis fasciculata reduces glioblastoma cell proliferation and changes beta-catenin cellular localization. Anticancer Drugs. 2009;20:543–552. doi: 10.1097/CAD.0b013e32832d1149. PubMed DOI

AMADO NG, PREDES D, FONSECA BF, CERQUEIRA DM, REIS AH, DUDENHOEFFER AC, BORGES HL, MENDES FA, ABREU JG. Isoquercitrin suppresses colon cancer cell growth in vitro by targeting the wnt/β-catenin signaling pathway. J Biol Chem. 2014;289:35456–35467. doi: 10.1074/jbc.M114.621599. PubMed DOI PMC

APPLETON J. Evaluating the bioavailability of isoquercetin. Nat Med. 2010;2:1–6.

ARUMUGAM S, BANDIL K, PROKSCH P, MURUGIYAN K, BHARADWAJ M. Effects of A. marina-derived isoquercitrin on TNF-related apoptosis-inducing ligand receptor (TRAIL-R) expression and apoptosis induction in cervical cancer cells. Appl Biochem Biotechnol. 2017;182:697–707. doi: 10.1007/s12010-016-2355-6. PubMed DOI

BALDOVSKÁ S, MARUNIAKOVÁ N, SLÁMA P, PAVLÍK A, KOHÚT L, KOLESÁROVÁ A. Efficacy of phytonutrients from pomegranate peel on human ovarian cells in vitro. J Microbiol Biotechnol Food Sci. 2020;10:511–516. doi: 10.15414/jmbfs.2020.10.3.511-516. DOI

BALDOVSKA S, MICHALCOVA K, HALENAR M, CARBONELL-BARRACHINA AA, KOLESAROVA A. Polyphenol-rich pomoegranate extract as a potential modulator of steroidogenesis in human ovarian cells. J Microbiol Biotechnol Food Sci. 2019;8:1343–1346. doi: 10.15414/jmbfs.2019.8.6.1343-1346. DOI

BOURAKI G, METALLINOU C, SIMOPOULOU M, CHARALABOPOULOS K, ASIMAKOPOULOS B. Comparison of nine media in the culture of human ovarian granulosa lutein cells. In Vivo. 2012;26:823–825. PubMed

BRISTOL SK, WOODRUFF TK. Follicle-restricted compartmentalization of transforming growth factor beta superfamily ligands in the feline ovary. Biol Reprod. 2004;70:846–859. doi: 10.1095/biolreprod.103.021857. PubMed DOI

BUONERBA C, De PLACIDO P, BRUZZESE D, PAGLIUCA M, UNGARO P, BOSSO D, RIBERA D, IACCARINO S, SCAFURI L, LIOTTI A. Isoquercetin as an adjunct therapy in patients with kidney cancer receiving first-line sunitinib (quasar): results of a phase I trial. Front Pharmacol. 2018;9:189. doi: 10.3389/fphar.2018.00189. PubMed DOI PMC

CHEN Q, LI P, LI P, XU Y, LI Y, TANG B. Isoquercitrin inhibits the progression of pancreatic cancer in vivo and in vitro by regulating opioid receptors and the mitogen-activated protein kinase signalling pathway. Oncol Rep. 2015;33:840–848. doi: 10.3892/or.2014.3626. PubMed DOI

CHEN F, CHEN X, YANG D, CHE X, WANG J, LI X, ZHANG Z, WANG Q, ZHENG W, WANG L, SONG X. Isoquercitrin inhibits bladder cancer progression in vivo and in vitro by regulating the PI3K/Akt and PKC signaling pathways. Oncol Rep. 2016;36:165–172. doi: 10.3892/or.2016.4794. PubMed DOI

CHEN M, DAI LH, FEI A, PAN SM, WANG HR. Isoquercetin activates the ERK1/2-NRF2 pathway and protects against cerebral ischemia-reperfusion injury in vivo and in vitro. Exp Ther Med. 2017;13:1353–1359. doi: 10.3892/etm.2017.4093. PubMed DOI PMC

DEVOS D, MOREAU C, DEVEDJIAN JC, KLUZA J, PETRAULT M, LALOUX C, JONNEAUX A, RYCKEWAERT G, GARÇON G, ROUAIX N. Targeting chelatable iron as a therapeutic modality in Parkinson’s disease. Antioxid Redox Signal. 2014;21:195–210. doi: 10.1089/ars.2013.5593. PubMed DOI PMC

DUONG TT, ANTAO S, ELLIS NA, MYERS SJ, WITTING PK. Supplementation with a synthetic polyphenol limits oxidative stress and enhances neuronal cell viability in response to hypoxia-re-oxygenation injury. Brain Res. 2008;1219:8–18. doi: 10.1016/j.brainres.2008.04.044. PubMed DOI

FANG YZ, ZHENG RL. Reactive oxygen species. In: FANG YZ, ZHENG RL, editors. Theory and Application of Free Radical Biology. 1st Edition. Scientific Press; Beijing, China: 2002. pp. 98–124.

FDA. Agency response letter gras notice no. Grn00220 [alpha-glycosyl isoquercitrin] U.S. Food and drug administration center for food safety and applied nutrition; 2007.

GERSTORFEROVÁ D, FLIEDROVÁ B, HALADA P, MARHOL P, KŘEN V, WEIGNEROVÁ L. Recombinant α-l-rhamnosidase from Aspergillus terreus in selective trimming of rutin. Process Biochem. 2012;47:828–835. doi: 10.1016/j.procbio.2012.02.014. DOI

HASUMURA M, YASUHARA K, TAMURA T, IMAI T, MITSUMORI K, HIROSE M. Evaluation of the toxicity of enzymatically decomposed rutin with 13-weeks dietary administration to Wistar rats. Food Chem Toxicol. 2004;42:439–444. doi: 10.1016/j.fct.2003.10.006. PubMed DOI

HAVELOCK JC, RAINEY WE, CARR BR. Ovarian granulosa cell lines. Mol Cell Endocrinol. 2004;228:67–78. doi: 10.1016/j.mce.2004.04.018. PubMed DOI

HUANG G, TANG B, TANG K, DONG X, DENG J, LIAO L, LIAO Z, YANG H, HE S. Isoquercitrin inhibits the progression of liver cancer in vivo and in vitro via the MAPK signalling pathway. Oncol Rep. 2014;31:2377–2384. doi: 10.3892/or.2014.3099. PubMed DOI

IWASE AB, KIYONO T, TAKIKAWA S, GOTO M, NAKAMURA T, NAGATOMO Y, NAKAHARA T, KOTANI T, KOBAYASHI H. Establishment of a human nonluteinized granulosa cell line that transitions from the gonadotropin-independent to the gonadotropin-dependent status. Endocrinology. 2012;153:2851–2860. doi: 10.1210/en.2011-1810. PubMed DOI

JFA. Japanese Specifications and Standards for Food Additives. Tokyo: Japan: 2007. p. 8.

JUNG SH, KIM BJ, LEE EH, OSBORNE NN. Isoquercitrin is the most effective antioxidant in the plant Thuja orientalis and able to counteract oxidative-induced damage to a transformed cell line (RGC-5 cells) Neurochem Int. 2010;57:713–721. doi: 10.1016/j.neuint.2010.08.005. PubMed DOI

KIM JH, QUILANTANG NG, KIM HY, LEE S, CHO EJ. Attenuation of hydrogen peroxide-induced oxidative stress in SH-SY5Y cells by three flavonoids from Acer okamotoanum. Chem Papers. 2019;73:1135–1144. doi: 10.1007/s11696-018-0664-7. DOI

KNIGHT PG, GLISTER C. TGF-beta superfamily members and ovarian follicle development. Reproduction. 2006;132:191–206. doi: 10.1530/rep.1.01074. PubMed DOI

LI X, JIANG Q, WANG T, LIU J, CHEN D. Comparison of the antioxidant effects of quercitrin and isoquercitrin: Understanding the role of the 6″-oh group. Molecules. 2016;21:1246. doi: 10.3390/molecules21091246. PubMed DOI PMC

LU Z, WANG J, LIN S, ZHAN Y. Degradation of rutin into isoquercitrin by Bacillus litoralis strain C44. IOSR J Engin. 2013;2:1154–1161. doi: 10.9790/3021-020511541161. DOI

MAGALINGAM KB, RADHAKRISHNAN A, HALEAGRAHARA N. Protective effects of quercetin glycosides, rutin, and isoquercetrin against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in rat pheochromocytoma (PC-12) cells. Int J Immunopathol Pharmacol. 2016;29:30–39. doi: 10.1177/0394632015613039. PubMed DOI PMC

MICHALCOVA K, ROYCHOUDHURY S, HALENAR M, TVRDA E, KOVACIKOVA E, VASICEK J, CHRENEK P, BALDOVSKA S, SANISLO L, KREN V, KOLESAROVA A. In vitro response of human ovarian cancer cells to dietary bioflavonoid isoquercitrin. J Environ Sci Health B. 2019;54:752–757. doi: 10.1080/03601234.2019.1633214. PubMed DOI

NEVEU V, PEREZ-JIMÉNEZ J, VOS F, CRESPY V, Du CHAFFAUT L, MENNEN L, KNOX C, EISNER R, CRUZ J, WISHART D, SCALBERT A. Phenol-Explorer: an online comprehensive database on polyphenol contents in foods. Database (Oxford) 2010;2010:bap024. doi: 10.1093/database/bap024. PubMed DOI PMC

PAULKE A, SCHUBERT-ZSILAVECZ M, WURGLICS M. Determination of St. John’s wort flavonoid-metabolites in rat brain through high performance liquid chromatography coupled with fluorescence detection. J Chromatogr B Analyt Technol Biomed Life Sci. 2006;832:109–113. doi: 10.1016/j.jchromb.2005.12.043. PubMed DOI

PUTTABYATAPPA M, BROGAN RS, VANDEVOORT CA, CHAFFIN CL. EGF-like ligands mediate progesterone’s anti-apoptotic action on macaque granulosa cells. Biol Reprod. 2013;88:1–10. doi: 10.1095/biolreprod.112.103002. PubMed DOI PMC

RAINEY WH, SAWETAWAN C, SHAY JW, MICHAEL MD, MATHIS JM, KUTTEH W, BYRD W, CARR BR. Transformation of human granulosa cells with the e6 and e7 regions of human papillomavirus. J Clin Endocrinol Metab. 1994;78:705–710. doi: 10.1210/jcem.78.3.8126145. PubMed DOI

REUTER S, GUPTA SC, CHATURVEDI MM, AGGARWAL BB. Oxidative stress, inflammation, and cancer: How are they linked? Free Radic Biol Med. 2010;49:1603–1616. doi: 10.1016/j.freeradbiomed.2010.09.006. PubMed DOI PMC

ROY SK, HUGHES J. Ontogeny of granulosa cells in the ovary: Lineage-specific expression of transforming growth factor beta 2 and transforming growth factor beta 1. Biol Reprod. 1994;51:821–830. doi: 10.1095/biolreprod51.5.821. PubMed DOI

SHARMA R, ROYCHOUDHURY S, SINGH N, SARDA Y. Methods to measure reactive oxygen species (ROS) and total antioxidant capacity (TAC) in the reproductive system. In: AGARWAL A, SHARMA R, GUPTA S, HARLEV A, AHMAD G, Du PLESSIS SS, ESTEVES SC, WANG SW, DURAIRAJANAYAGAM D, editors. Oxidative Stress in Human Reproduction. Springer; Cham: 2017. pp. 17–46. DOI

TROMBLY DJ, WOODRUFF TK, MAYO KE. Roles for transforming growth factor beta superfamily proteins in early folliculogenesis. Semin Reprod Med. 2009;27:14–23. doi: 10.1055/s-0028-1108006. PubMed DOI PMC

TVRDÁ E, TUŠIMOVÁ E, KOVÁČIK A, PAÁL D, GREIFOVÁ H, ABDRAMANOV A, LUKÁČ N. Curcumin has protective and antioxidant properties on bull spermatozoa subjected to induced oxidative stress. Anim Reprod Sci. 2016;172:10–20. doi: 10.1016/j.anireprosci.2016.06.008. PubMed DOI

VALENTOVÁ K, VRBA J, BANCÍŘOVÁ M, ULRICHOVÁ J, KŘEN V. Isoquercitrin: Pharmacology, toxicology, and metabolism. Food Chem Toxicol. 2014;68:267–682. doi: 10.1016/j.fct.2014.03.018. PubMed DOI

VANDER A, SHERMAN J, LUCIANO D, TSAKOPOULOS M. The mechanisms of body function. In: PASCHALIDIS PH, editor. Human Physiology. 8th edition. Medical Editions; Athens: 2001. pp. 860–872.

WEIGNEROVÁ L, MARHOL P, GERSTORFEROVÁ D, KŘEN V. Preparatory production of quercetin-3-β-d-glucopyranoside using alkali-tolerant thermostable α-l-rhamnosidase from aspergillus terreus. Bioresour Technol. 2012;115:222–227. doi: 10.1016/j.biortech.2011.08.029. PubMed DOI

WU P, LIU S, SU J, CHEN J, LI L, ZHANG R, CHEN T. Apoptosis triggered by isoquercitrin in bladder cancer cells by activating the AMPK-activated protein kinase pathway. Food Funct. 2017;8:3707–3722. doi: 10.1039/C7FO00778G. PubMed DOI

XIE W, WANG M, CHEN C, ZHANG X, MELZIG MF. Hepatoprotective effect of isoquercitrin against acetaminophen-induced liver injury. Life Sci. 2016;152:180–189. doi: 10.1016/j.lfs.2016.04.002. PubMed DOI

ZHOU J, YOSHITOMI H, LIU T, ZHOU B, SUN W, QIN L, GUO X, HUANG L, WU L, GAO M. Isoquercitrin activates the amp-activated protein kinase (AMPK) signal pathway in rat h4iie cells. BMC Complement Altern Med. 2014;14:42. doi: 10.1186/1472-6882-14-42. PubMed DOI PMC

ZHU M, LI J, WANG K, HAO X, GE R, LI Q. Isoquercitrin inhibits hydrogen peroxide-induced apoptosis of ea. y926 cells via the PI3K/AKT/GSK3β signaling pathway. Molecules. 2016;21:356. doi: 10.3390/molecules21030356. PubMed DOI PMC