DOI QR코드

DOI QR Code

Concentration-dependent in vitro Anti-osteoarthritis Effects of Mixed Formula - Pomegranate Concentrate Powder: Eucommiae Cortex: Achyranthis Radix 5:4:1 (g/g) on the Primary Cultured Rat Articular Chondrocytes

  • Choi, Beom Rak (Research Institute, HL Science Co., Ltd.) ;
  • Ku, Sae Kwang (Department of Histology and Anatomy, College of Korean Medicine, Daegu Haany University) ;
  • Kang, Su Jin (The Medical Research Center for Globalization of Herbal Medicine, Daegu Haany University) ;
  • Park, Hye Rim (Research Institute, HL Science Co., Ltd.) ;
  • Sung, Mi Sun (Research Institute, HL Science Co., Ltd.) ;
  • Lee, Young Joon (The Medical Research Center for Globalization of Herbal Medicine, Daegu Haany University) ;
  • Park, Ki Moon (Department of Foodscience and Biotechnology, Sungkyunkwan University)
  • Received : 2018.10.26
  • Accepted : 2019.02.25
  • Published : 2019.04.25

Abstract

The objective of present study is to evaluate concentration-dependent in vitro anti-osteoarthritic (OA) effects of synergic mixed formula consisted of dried pomegranate juice concentrate powder, Eucommiae Cortex aqueous extract and Achyranthis Radix aqueous extract 5:4:1 (g/g) mixture on the primary cultured rat articular chondrocytes. First, any cytotoxic effect of mixture was observed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium Bromide) assay. Next, cyto-protective effect of test substances was evaluated by using the recombinant human interleukin $(rhIL)-1{\alpha}$ induced chondrocytes. In addition, anti-inflammatory effects were also observed on the lipopolysaccaride (LPS) treated chondrocytes through prostaglandin $E_2(PGE_2)$ productions and 5-lipoxygenase (LPO) activities, and inhibitory effects on matrix metalloproteinase (MMP)-2 and MMP-9 activities were observed on $rhIL-1{\alpha}$ treated chondrocytes with their extracellular matrix (ECM) related mRNA expressions. No obvious cytotoxic effects of mixture were demonstrated. Inflammatory damages of chondrocytes and related ECM degradations induced by treatment of LPS or $rhIL-1{\alpha}$ were significantly and concentration-dependently inhibited by pretreatment of mixture from a concentration level of 0.001 mg/ml to 1 mg/ml. In addition, mixture showed $IC_{50}$ for $rhIL-1{\alpha}-induced$ MMP-2 and MMP-9 activities as 44.01 and $162.47{\mu}g/ml$, and also showed $EC_{50}$ for $rhIL-1{\alpha}-induced$ inhibition of collagen type II, SOX9 and aggrecan mRNA expression as 8.61, 10.79 and $4.47{\mu}g/ml$, respectively. It is observed that mixture showed concentration-dependent anti-inflammatory and cytoprotective ECM preserved effects on the primary cultured rat articular chondrocytes without cytotoxicity.

References

  1. Felson DT, Zhang Y. An update on the epidemiology of knee and hip osteoarthritis with a view to prevention. Arthritis Rheum. 1998;41(8):1343-13. https://doi.org/10.1002/1529-0131(199808)41:8<1343::AID-ART3>3.0.CO;2-9
  2. Goldring MB, Fukuo K, Birkhead JR, Dudek E, Sandell LJ. Transcriptional suppression by interleukin-1 and interferon-gamma of type II collagen gene expression in human chondrocytes. J Cell Biochem. 1994;54(1):85-99. https://doi.org/10.1002/jcb.240540110
  3. van de Loo FA, Joosten LA, van Lent PL, Arntz OJ, van den Berg WB. Role of interleukin-1, tumor necrosis factor alpha, and interleukin-6 in cartilage proteoglycan metabolism and destruction. Effect of in situ blocking in murine antigen- and zymosan-induced arthritis. Arthritis Rheum. 1995;38(2):164-72. https://doi.org/10.1002/art.1780380204
  4. Berenbaum F, Jacques C, Thomas G, Corvol MT, Bereziat G, Masliah J. Synergistic effect of interleukin-1 beta and tumor necrosis factor alpha on PGE2 production by articular chondrocytes does not involve PLA2 stimulation. Exp Cell Res. 1996;222(2):379-84. https://doi.org/10.1006/excr.1996.0047
  5. Grabowski PS, Macpherson H, Ralston SH. Nitric oxide production in cells derived from the human joint. Br J Rheumatol. 1996;35(3):207-12. https://doi.org/10.1093/rheumatology/35.3.207
  6. Stefanovic-Racic M, Morales TI, Taskiran D, McIntyre LA, Evans CH. The role of nitric oxide in proteoglycan turnover by bovine articular cartilage organ cultures. J Immunol. 1996;156(3):1213-20.
  7. Koolpe M, Pearson D, Benton HP. Expression of both P1 and P2 purine receptor genes by human articular chondrocytes and profile of ligand-mediated prostaglandin E2 release. Arthritis Rheum. 1999;42(2):258-67. https://doi.org/10.1002/1529-0131(199902)42:2<258::AID-ANR7>3.0.CO;2-O
  8. Appleton CT, McErlain DD, Pitelka V, Schwartz N, Bernier SM, Henry JL, et al. Forced mobilization accelerates pathogenesis: characterization of a preclinical surgical model of osteoarthritis. Arthritis Res Ther. 2007;9(1):R13. https://doi.org/10.1186/ar2120
  9. Tamura T, Ohmori K. Rhein, an active metabolite of diacerein, suppresses the interleukin-1alpha-induced proteoglycan degradation in cultured rabbit articular chondrocytes. Jpn J Pharmacol. 2001;85(1):101-4. https://doi.org/10.1254/jjp.85.101
  10. Qin J, Liu YS, Liu J, Li J, Tan Y, Li XJ, et al. Effect of Angelica sinensis Polysaccharides on Osteoarthritis In Vivo and In Vitro: A Possible Mechanism to Promote Proteoglycans Synthesis. Evid Based Complement Alternat Med. 2013;2013:794761.
  11. Na JY, Song KB, Kim S, Kwon YB, Kim DG, Lee JK, et al. Effects of HPL-04 on degenerative osteoarthritis. J Korean Soc Food Sci Nutr. 2014;43(1):30-9. https://doi.org/10.3746/jkfn.2014.43.1.030
  12. Nam DE, Kim OK, Shim TJ, Kim JH, Lee J. Effect of Boswellia Serrata Extracts on Degenerative Osteoarthritis in vitro and in vivo model. J Korean Soc Food Sci Nutr. 2014;43:631-40. https://doi.org/10.3746/jkfn.2014.43.5.631
  13. Langley P. Why a pomegranate? BMJ. 2000;321(7269): 1153-4. https://doi.org/10.1136/bmj.321.7269.1153
  14. Gil MI, Tomas-Barberan FA, Hess-Pierce B, Holcroft DM, Kader AA. Antioxidant activity of pomegranate juice and its relationship with phenolic composition and processing. J Agric Food Chem. 2000;48(10):4581-9. https://doi.org/10.1021/jf000404a
  15. Noda Y, Kaneyuki T, Mori A, Packer L. Antioxidant activities of pomegranate fruit extract and its anthocyanidins: delphinidin, cyanidin, and pelargonidin. J Agric Food Chem. 2002;50(1):166-71. https://doi.org/10.1021/jf0108765
  16. Singh RP, Chidambara Murthy KN, Jayaprakasha GK. Studies on the antioxidant activity of pomegranate (Punica granatum) peel and seed extracts using in vitro models. J Agric Food Chem. 2002;50(1):81-6. https://doi.org/10.1021/jf010865b
  17. Ahmed S, Wang N, Hafeez BB, Cheruvu VK, Haqqi TM. Punica granatum L. extract inhibits IL-1beta-induced expression of matrix metalloproteinases by inhibiting the activation of MAP kinases and NF-kB in human chondrocytes in vitro. J Nutr. 2005;135(9):2096-102. https://doi.org/10.1093/jn/135.9.2096
  18. Shukla M, Gupta K, Rasheed Z, Khan KA, Haqqi TM. Bioavailable constituents/metabolites of pomegranate (Punica granatum L) preferentially inhibit COX2 activity ex vivo and IL-1beta-induced PGE2 production in human chondrocytes in vitro. J Inflamm (Lond). 2008;5:9. https://doi.org/10.1186/1476-9255-5-9
  19. Hadipour-Jahromy M, Mozaffari-Kermani R. Chondroprotective effects of pomegranate juice on monoiodoacetate-induced osteoarthritis of the knee joint of mice. Phytother Res. 2010;24(2):182-5. https://doi.org/10.1002/ptr.2880
  20. Rasheed Z, Akhtar N, Haqqi TM. Pomegranate extract inhibits the interleukin-1b-induced activation of MKK-3, p38a-MAPK and transcription factor RUNX-2 in human osteoarthritis chondrocytes. Arthritis Res Ther. 2010;12(5):R195. https://doi.org/10.1186/ar3166
  21. Hsieh CL, Yen GC. Antioxidant actions of du-zhong (Eucommia ulmoides Oliv.) toward oxidative damage in biomolecules. Life Sci. 2000;66(15):1387-1400. https://doi.org/10.1016/S0024-3205(00)00450-1
  22. Kwan CY, Chen CX, Deyama T, Nishibe S. Endothelium-dependent vasorelaxant effects of the aqueous extracts of the Eucommia ulmoides Oliv. leaf and bark: implications on their antihypertensive action. Vascul Pharmacol. 2003;40(5):229-35. https://doi.org/10.1016/j.vph.2003.09.001
  23. Zhao Y, Li Y, Wang X, Sun W. The experimental study of Cortex Eucommiae on meridian tropsim: the distribution study of aucubin in rat tissues. J Pharm Biomed Anal. 2008;46(2):368-73. https://doi.org/10.1016/j.jpba.2007.09.028
  24. Eum HA, Lee WY, Kim SH, Kim JY, Park SW, Lee JS, et al. Anti-inflammatory activity of CML-1: an herbal formulation. Am J Chin Med. 2005;33(1):29-40. https://doi.org/10.1142/S0192415X05002631
  25. Lee SY, Kwon HK, Lee SM. SHINBARO, a new herbal medicine with multifunctional mechanism for joint disease: first therapeutic application for the treatment of osteoarthritis. Arch Pharm Res. 2011;34(11):1773-7. https://doi.org/10.1007/s12272-011-1121-0
  26. Li J, Li HJ, Li P, Qi H. Simultaneous qualitation and quantification of four phytoecdysones in Radix Achyranthis Bidentatae by high-performance liquid chromatography with diode array detection. Biomed Chromatogr. 2007;21(8):823-8. https://doi.org/10.1002/bmc.824
  27. Yu F, Li X, Cai L, Li H, Chen J, Wong X, et al. Achyranthes bidentata polysaccharides induce chondrocyte proliferation via the promotion of the G1/S cell cycle transition. Mol Med Rep. 2013;7(3):935-40. https://doi.org/10.3892/mmr.2013.1286
  28. Weng X, Lin P, Liu F, Chen J, Li H, Huang L, et al. Achyranthes bidentata polysaccharides activate the Wnt/b-catenin signaling pathway to promote chondrocyte proliferation. Int J Mol Med. 2014;34(4):1045-50. https://doi.org/10.3892/ijmm.2014.1869
  29. Lee HS, Kim HH, Ku SK. Hepatoprotective effects of Artemisiae capillaris herba and Picrorrhiza rhizoma combinations on carbon tetrachloride-induced subacute liver damage in rats. Nutr Res. 2008;28(4):270-7. https://doi.org/10.1016/j.nutres.2008.02.001
  30. Choi JS, Kim JW, Kim KY, Cho HR, Choi IS, Ku SK. Antiosteoporotic effects of Polycan in combination with calcium lactate-gluconate in ovariectomized rats. Exp Ther Med. 2014;8(3):957-67. https://doi.org/10.3892/etm.2014.1793
  31. Choi JS, Kim JW, Kim KY, Lee JK, Sohn JH, Ku SK. Synergistic effect of fermented rice extracts on the probiotic and laxative properties of yoghurt in rats with loperamide-induced constipation. Evid Based Complement Alternat Med. 2014;2014:878503.
  32. Choi JS, Shin HS, Kim KY, Ku SK, Choi IS, Kim JW. Effect of Polycalcium, a mixture of Polycan and calcium lactate-gluconate in a 1:9 weight ratio, on rats with surgery-induced osteoarthritis. Exp Ther Med. 2015;9(5):1780-90. https://doi.org/10.3892/etm.2015.2332
  33. Kang SJ, Choi BR, Kim SH, Yi HY, Park HR, Kim DC, et al. Dried pomegranate potentiates anti-osteoporotic and anti-obesity activities of red clover dry extracts in ovariectomized rats. Nutrients. 2015;7(4):2622-47. https://doi.org/10.3390/nu7042622
  34. Choi BR, Ku SK, Kang SJ, Park HR, Sung MS, Lee YJ, et al. Anti-osteoarthritis effects of Pomegranate, Eucommiae cortex and Achyranthis radix extracts on the primary cultured rat articular chondrocytes. J Soc Prev Korean Med. 2017;21(3):87-98. https://doi.org/10.25153/spkom.2017.21.3.009
  35. Choi BR, Ku SK, Kang SJ, Park HR, Sung MS, Lee YJ, et al. Selection of the optimal herbal composition of pomegranate concentrated powder from aqueous extracts of Eucommiae Cortex and Achyranthis Radix to treat osteoarthritis in rats. J Korean Med. 2017;38(4):11-40. https://doi.org/10.13048/jkm.17037
  36. Kim KH, Park SJ, Lee JE, Lee YJ, Song CH, Choi SH, et al. Anti-skin-aging benefits of exopolymers from Aureobasidium pullulans SM2001. J Cosmet Sci. 2014;65(5):285-98.
  37. Kang SJ, Lee JE, Lee EK, Jung DH, Song CH, Park SJ, et al. Fermentation with Aquilariae Lignum enhances the anti-diabetic activity of green tea in type II diabetic db/db mouse. Nutrients. 2014;6(9):3536-71. https://doi.org/10.3390/nu6093536
  38. Smit HF, Woerdenbag HJ, Singh RH, Meulenbeld GJ, Labadie RP, Zwaving JH. Ayurvedic herbal drugs with possible cytostatic activity. J Ethnopharmacol. 1995;47(2):75-84. https://doi.org/10.1016/0378-8741(95)01255-C
  39. Kratzke RA, Kramer BS. Evaluation of in vitro chemosensitivity using human lung cancer cell lines. J Cell Biochem Suppl. 1996;24:160-4.
  40. Ghosh J, Myers CE. Arachidonic acid stimulates prostate cancer cell growth: critical role of 5-lipoxygenase. Biochem Biophys Res Commun. 1997;235(2):418-23. https://doi.org/10.1006/bbrc.1997.6799
  41. Sailer ER, Schweizer S, Boden SE, Ammon HP, Safayhi H. Characterization of an acetyl-11-keto-b-boswellic acid and arachidonate-binding regulatory site of 5-lipoxygenase using photoaffinity labeling. Eur J Biochem. 1998;256(2):364-8. https://doi.org/10.1046/j.1432-1327.1998.2560364.x
  42. Vane JR, Bakhle YS, Botting RM. Cyclooxygenases 1 and 2. Annu Rev Pharmacol Toxicol. 1998;38:97-120. https://doi.org/10.1146/annurev.pharmtox.38.1.97
  43. Bensen WG, Fiechtner JJ, McMillen JI, Zhao WW, Yu SS, Woods EM, et al. Treatment of osteoarthritis with celecoxib, a cyclooxygenase-2 inhibitor: a randomized controlled trial. Mayo Clin Proc. 1999;74(11):1095-105. https://doi.org/10.4065/74.11.1095
  44. Hardy MM, Seibert K, Manning PT, Currie MG, Woerner BM, Edwards D, et al. Cyclooxygenase 2-dependent prostaglandin E2 modulates cartilage proteoglycan degradation in human osteoarthritis explants. Arthritis Rheum. 2002;46(7):1789-1803. https://doi.org/10.1002/art.10356
  45. Bresnihan B. Pathogenesis of joint damage in rheumatoid arthritis. J Rheumatol. 1999;26(3):717-19.
  46. Nagase H, Woessner JF, Jr. Matrix metalloproteinases. J Biol Chem. 1999;274(31):21491-4. https://doi.org/10.1074/jbc.274.31.21491
  47. Halliwell B. Oral inflammation and reactive species: a missed opportunity? Oral Dis. 2000;6(3):136-7. https://doi.org/10.1111/j.1601-0825.2000.tb00324.x
  48. Guzman RE, Evans MG, Bove S, Morenko B, Kilgore K. Mono-iodoacetate-induced histologic changes in subchondral bone and articular cartilage of rat femorotibial joints: an animal model of osteoarthritis. Toxicol Pathol. 2003;31(6):619-24. https://doi.org/10.1080/01926230390241800