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In vitro effects of triamcinolone acetonide and in combination with hyaluronan on canine normal and spontaneous osteoarthritis articular cartilage

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Abstract

The purposes of this study were to examine the cartilage degradation effects of triamcinolone acetonide (TA) on normal and osteoarthritic (OA) primary canine chondrocytes and cartilage explants and to examine the cartilage degradation effects of TA in combination with low-molecular-weight hyaluronan (LMWHA). To assess the effects of these drugs on cell culture, 3,[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and real-time PCR were used to measure chondrotoxicity and determine gene expression, respectively. Uronic acid and hydroxyproline remaining in cartilage and histopathology were used to estimate the effects of these drugs on cartilage explants. In chondrocyte cultures, TA reduced chondrocyte viability in a concentration-dependent manner. LMWHA 2.5 mg/ml combined with TA at IC20 (0.09 mg/ml) could increase the viability of normal chondrocytes when compared with TA-treated alone. TA at IC20 induced down-regulation of ACAN and induced up-regulation of ADAMTS5 in canine normal chondrocytes. TA at IC20 (0.11 mg/ml) up-regulated ADAMTS5, MMP2, MMP3, MMP13, and ACAN expression in canine OA chondrocytes. In explant culture, TA at 1.25, 2.5, and 5 mg/ml increased the severity of structural damage, chondrocyte loss and cluster formation, and proteoglycan loss in OA cartilage. LMWHA could decrease the chondrotoxicity of TA at IC20 only in normal chondrocytes, as observed by chondrocyte viability. The combination of LMWHA and TA did not show clearly beneficial effects in all other normal and OA samples. Consequently, using TA alone or in combination with LMWHA in OA cartilage should be of concern because it may lead to cartilage destruction.

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References

  • Adams ME, Atkinson MH, Lussier AJ, Schulz JI, Siminovitch KA, Wade JP, Zummer M (1995) The role of viscosupplementation with hylan G-F 20 (Synvisc®) in the treatment of osteoarthritis of the knee: a Canadian multicenter trial comparing hylan G-F 20 alone, hylan G-F 20 with non-steroidal anti-inflammatory drugs (NSAIDs) and NSAIDs alone. Osteoarthr Cartil 3:213–225

    Article  CAS  PubMed  Google Scholar 

  • Amiel D, Toyoguchi T, Kobayashi K, Bowden K, Amiel ME, Healey RM (2003) Long-term effect of sodium hyaluronate (Hyalgan®) on osteoarthritis progression in a rabbit model. Osteoarthr Cartil 11:636–643

    Article  CAS  PubMed  Google Scholar 

  • Archer CW, Francis-West P (2003) The chondrocyte. Int J Biochem Cell Biol 35:401–404

    Article  CAS  PubMed  Google Scholar 

  • Barreto RB, Sadigursky D, de Rezende MU, Hernandez AJ (2015) Effect of hyaluronic acid on chondrocyte apoptosis. Acta Ortop Bras 23:90–93

    Article  PubMed  PubMed Central  Google Scholar 

  • Bau B, Gebhard PM, Haag J, Knorr T, Bartnik E, Aigner T (2002) Relative messenger RNA expression profiling of collagenases and aggrecanases in human articular chondrocytes in vivo and in vitro. Arthritis Rheum 46:2648–2657

    Article  CAS  PubMed  Google Scholar 

  • Bland JH, Cooper SM (1984) Osteoarthritis: a review of the cell biology involved and evidence for reversibility. Management rationally related to known genesis and pathophysiology. Semin Arthritis Rheum 14:106–133

    Article  CAS  PubMed  Google Scholar 

  • Blumenkrantz N, Asboe-Hansen G (1973) New method for quantitative determination of uronic acids. Anal Biochem 54:484–489

    Article  CAS  PubMed  Google Scholar 

  • Bondeson J, Wainwright S, Hughes C, Caterson B (2008) The regulation of the ADAMTS4 and ADAMTS5 aggrecanases in osteoarthritis: a review. Clin Exp Rheumatol 26:139–145

    CAS  PubMed  Google Scholar 

  • Cook JL, Kuroki K, Visco D, Pelletier JP, Schulz L, Lafeber FP (2010) The OARSI histopathology initiative—recommendations for histological assessments of osteoarthritis in the dog. Osteoarthr Cartil 18:S66–S79

    Article  PubMed  Google Scholar 

  • Corrado EM, Peluso GF, Gigliotti S, de Durante C, Palmieri D, Savoia N, Oriani GO, Tajana GF (1995) The effects of intra-articular administration of hyaluronic acid on osteoarthritis of the knee: a clinical study with immunological and biochemical evaluations. Eur J Rheumatol Inflamm 15:47–56

    Google Scholar 

  • Dean DD, Martel-Pelletier J, Pelletier JP, Howell DS, Woessner JF Jr (1989) Evidence for metalloproteinase and metalloproteinase inhibitor imbalance in human osteoarthritic cartilage. J Clin Invest 84:678–685

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dechant JE, Baxter GM, Frisbie DD, Trotter GW, McIlwraith CW (2003) Effects of dosage titration of methylprednisolone acetate and triamcinolone acetonide on interleukin-1-conditioned equine articular cartilage explants in vitro. Equine Vet J 35:444–450

    Article  CAS  PubMed  Google Scholar 

  • Denizot F, Lang R (1986) Rapid colorimetric assay for cell growth and survival. Modifications to the tetrazolium dye procedure giving improved sensitivity and reliability. J Immunol Methods 89:271–277

    Article  CAS  PubMed  Google Scholar 

  • Doyle AJ, Stewart AA, Constable PD, Eurell JAC, Freeman DE, Griffon DJ (2005) Effects of sodium hyaluronate and methylprednisolone acetate on proteoglycan synthesis in equine articular cartilage explants. Am J Vet Res 66:48–53

    Article  CAS  PubMed  Google Scholar 

  • Dragoo JL, Danial CM, Braun HJ, Pouliot MA, Kim HJ (2012) The chondrotoxicity of single-dose corticosteroids. Knee Surg Sports Traumatol Arthrosc 20:1809–1814

    Article  PubMed  Google Scholar 

  • Euppayo T, Siengdee P, Buddhachat K, Pradit W, Viriyakhasem N, Chomdej S, Ongchai S, Harada Y, Nganvongpanit K (2015) Effects of low molecular weight hyaluronan combined with carprofen on canine osteoarthritis articular chondrocytes and cartilage explants in vitro. In Vitro Cell Dev Biol Anim 51:857–865

    Article  CAS  PubMed  Google Scholar 

  • Evans CH, Kraus VB, Setton LA (2014) Progress in intra-articular therapy. Nat Rev Rheumatol 10:11–22

    Article  CAS  PubMed  Google Scholar 

  • Fan Z, Bau B, Yang H, Soeder S, Aigner T (2005) Freshly isolated osteoarthritic chondrocytes are catabolically more active than normal chondrocytes, but less responsive to catabolic stimulation with interleukin-1β. Arthritis Rheum 52:136–143

    Article  CAS  PubMed  Google Scholar 

  • Fox AJS, Bedi A, Rodeo SA (2009) The basic science of articular cartilage. Sports Health 1:461–468

    Article  Google Scholar 

  • Frizziero A, Maffulli N, Masiero S, Frizziero L (2014) Six-months pain relief and functional recovery after intra-articular injections with hyaluronic acid (mw 500–730 kDa) in trapeziometacarpal osteoarthritis. Muscles Ligaments Tendons J 4:256–261

    PubMed  PubMed Central  Google Scholar 

  • Ghosh P, Guidolin D (2002) Potential mechanism of action of intra-articular hyaluronan therapy in osteoarthritis: are the effects molecular weight dependent? Semin Arthritis Rheum 32:10–37

    Article  CAS  PubMed  Google Scholar 

  • Goldberg VM, Buckwalter JA (2005) Hyaluronans in the treatment of osteoarthritis of the knee: evidence for disease-modifying activity. Osteoarthr Cartil 13:216–224

    Article  CAS  PubMed  Google Scholar 

  • Goldring MB (2000) The role of the chondrocyte in osteoarthritis. Arthritis Rheum 43:1916–1926

    Article  CAS  PubMed  Google Scholar 

  • Grecomoro G, Piccione F, Letizia G (1992) Therapeutic synergism between hyaluronic acid and dexamethasone in the intra-articular treatment of osteoarthritis of the knee: a preliminary open study. Curr Med Res Opin 13:49–55

    Article  CAS  PubMed  Google Scholar 

  • Grishko V, Xu M, Ho R, Mates A, Watson S, Kim JT, Wilson GL, Pearsall AW 4th (2009) Effects of hyaluronic acid on mitochondrial function and mitochondria-driven apoptosis following oxidative stress in human chondrocytes. J Biol Chem 284:9132–9139

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Guidolin DD, Ronchetti IP, Lini E, Guerra D, Frizziero L (2001) Morphological analysis of articular cartilage biopsies from a randomized, clinical study comparing the effects of 500–730 kDa sodium hyaluronate (Hyalgan®) and methylprednisolone acetate on primary osteoarthritis of the knee. Osteoarthr Cartil 9:371–381

    Article  CAS  PubMed  Google Scholar 

  • Güngör İ, Yılmaz A, Ergün MA, Öztürk AM, Kaya K (2014) Does hyaluronic acid decrease the apoptotic effect of bupivacaine? Eklem Hastalık Cerrahisi 25:102–106

    Article  PubMed  Google Scholar 

  • Haddad IK (2000) Temporomandibular joint osteoarthrosis. Histopathological study of the effects of intra-articular injection of triamcinolone acetonide. Saudi Med J 21:675–679

    CAS  PubMed  Google Scholar 

  • Hepper CT, Halvorson JJ, Duncan ST, Gregory AJM, Dunn WR, Spindler KP (2009) The efficacy and duration of intra-articular corticosteroid injection for knee osteoarthritis: a systematic review of level I studies. J Am Acad Orthop Surg 17:638–646

    Article  PubMed  Google Scholar 

  • Héraud F, Héraud A, Harmand MF (2000) Apoptosis in normal and osteoarthritic human articular cartilage. Ann Rheum Dis 59:959–965

    Article  PubMed  PubMed Central  Google Scholar 

  • Hwang HS, Kim HA (2015) Chondrocyte apoptosis in the pathogenesis of osteoarthritis. Int J Mol Sci 16:26035–26054

    Article  PubMed  PubMed Central  Google Scholar 

  • Jubb RW, Piva S, Beinat L, Dacre J, Gishen P (2003) A one-year, randomised, placebo (saline) controlled clinical trial of 500–730 kDa sodium hyaluronate (Hyalgan) on the radiological change in osteoarthritis of the knee. Int J Clin Pract 57:467–474

    CAS  PubMed  Google Scholar 

  • Kawasaki K, Ochi M, Uchio Y, Adachi N, Matsusaki M (1999) Hyaluronic acid enhances proliferation and chondroitin sulfate synthesis in cultured chondrocytes embedded in collagen gels. J Cell Physiol 179:142–148

    Article  CAS  PubMed  Google Scholar 

  • Kim HA, Suh DI, Song YW (2001) Relationship between chondrocyte apoptosis and matrix depletion in human articular cartilage. J Rheumatol 28:2038–2045

    CAS  PubMed  Google Scholar 

  • Kim HJ, Braun HJ, Dragoo JL (2014) The effect of resveratrol on normal and osteoarthritic chondrocyte metabolism. Bone Joint Res 3:51–59

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kolar K (1990) Colorimetric determination of hydroxyproline as measure of collagen content in meat and meat products: NMKL collaborative study. J Assoc Off Anal Chem 73:54–57

    CAS  PubMed  Google Scholar 

  • Laurent TC, Laurent UB, Fraser JRE (1996) The structure and function of hyaluronan: an overview. Immunol Cell Biol 74:A1–A7

    Article  CAS  PubMed  Google Scholar 

  • Listrat V, Ayral X, Patarnello F, Bonvarlet JP, Simonnet J, Amor B, Dougados M (1997) Arthroscopic evaluation of potential structure modifying activity of hyaluronan (Hyalgan®) in osteoarthritis of the knee. Osteoarthr Cartil 5:153–160

    Article  CAS  PubMed  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔC T method. Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  • Nagase H, Woessner JF Jr (1999) Matrix metalloproteinases. J Biol Chem 274:21491–21494

    Article  CAS  PubMed  Google Scholar 

  • Nganvongpanit K, Boonsri B, Sripratak T, Markmee P (2013) Effects of one-time and two-time intra-articular injection of hyaluronic acid sodium salt after joint surgery in dogs. J Vet Sci 14:215

    Article  PubMed  PubMed Central  Google Scholar 

  • Onur TS, Sitron CS, Dang A (2013) Co-administration of hyaluronic acid with local anaesthetics shows lower cytotoxicity than local anaesthetic treatment alone in bovine articular chondrocytes. Bone Joint Res 2:270–275

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pelletier JP, Martel-Pelletier J, Abramson SB (2001) Osteoarthritis, an inflammatory disease: potential implication for the selection of new therapeutic targets. Arthritis Rheum 44:1237–1247

    Article  CAS  PubMed  Google Scholar 

  • Poole AR, Kojima T, Yasuda T, Mwale F, Kobayashi M, Laverty S (2001) Composition and structure of articular cartilage: a template for tissue repair. Clin Orthop Relat Res 391:S26–S33

    Article  PubMed  Google Scholar 

  • Rousseau JC, Delmas PD (2007) Biological markers in osteoarthritis. Nat Clin Pract Rheumatol 3:346–356

    Article  CAS  PubMed  Google Scholar 

  • Salter DM, Millward-Sadler SJ, Nuki G, Wright MO (2002) Differential responses of chondrocytes from normal and osteoarthritic human articular cartilage to mechanical stimulation. Biorheology 39:97–108

    CAS  PubMed  Google Scholar 

  • Schaefer EC, Stewart AA, Durgam SS, Byron CR, Stewart MC (2009) Effects of sodium hyaluronate and triamcinolone acetonide on glucosaminoglycan metabolism in equine articular chondrocytes treated with interleukin-1. Am J Vet Res 70:1494–1501

    Article  CAS  PubMed  Google Scholar 

  • Siengdee P, Radeerom T, Kuanoon S, Euppayo T, Pradit W, Chomdej S, Ongchai S, Nganvongpanit K (2015) Effects of corticosteroids and their combinations with hyaluronan on the biochemical properties of porcine cartilage explants. BMC Vet Res 11:298

    Article  PubMed  PubMed Central  Google Scholar 

  • Soma LR, Uboh CE, You Y, Guan F, Boston RC (2011) Pharmacokinetics of intra-articular, intravenous, and intramuscular administration of triamcinolone acetonide and its effect on endogenous plasma hydrocortisone and cortisone concentrations in horses. Am J Vet Res 72:1234–1242

    Article  CAS  PubMed  Google Scholar 

  • Sternlicht MD, Werb Z (2001) How matrix metalloproteinases regulate cell behavior. Annu Rev Cell Dev Biol 17:463–516

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Stoker AM, Cook JL, Kuroki K, Fox DB (2006) Site-specific analysis of gene expression in early osteoarthritis using the Pond-Nuki model in dogs. J Orthop Surg Res 1, 8. http://doi.org/10.1186/1749-799X-1-8

  • Syed HM, Green L, Bianski B, Jobe CM, Wongworawat MD (2011) Bupivacaine and triamcinolone may be toxic to human chondrocytes: a pilot study. Clin Orthop Relat Res 469:2941–2947

    Article  PubMed  PubMed Central  Google Scholar 

  • Takahashi K, Hashimoto S, Kubo T, Hirasawa Y, Lotz M, Amiel D (2000) Effect of hyaluronan on chondrocyte apoptosis and nitric oxide production in experimentally induced osteoarthritis. J Rheumatol 27:1713–1720

    CAS  PubMed  Google Scholar 

  • Wenz W, Breusch SJ, Graf J, Stratmann U (2000) Ultrastructural findings after intraarticular application of hyaluronan in a canine model of arthropathy. J Orthop Res 18:604–612

    Article  CAS  PubMed  Google Scholar 

  • Wernecke C, Braun HJ, Dragoo JL (2015) The effect of intra-articular corticosteroids on articular cartilage: a systematic review. Orthop J Sports Med 3:2325967115581163

    Article  PubMed  PubMed Central  Google Scholar 

  • Yates AC, Stewart AA, Byron CR, Pondenis HC, Kaufmann KM, Constable PD (2006) Effects of sodium hyaluronate and methylprednisolone acetate on proteoglycan metabolism in equine articular chondrocytes treated with interleukin-1. Am J Vet Res 67:1980–1986

    Article  CAS  PubMed  Google Scholar 

  • Zhou PH, Liu SQ, Peng H (2008) The effect of hyaluronic acid on IL-1β-induced chondrocyte apoptosis in a rat model of osteoarthritis. J Orthop Res 26:1643–1648

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This study was kindly supported by the 50th CMU Anniversary Ph.D. program. The authors are also grateful for additional research funding via a CMU Research Group Grant (2015), Chiang Mai University, Thailand. TE designed and performed the experiments, analyzed primary data, and wrote the manuscript. PS oversaw laboratory work and writing of the manuscript. KP, SC, and SO gave advice on laboratory work and assisted in discussion of the manuscript. KN designed and conducted the overall study and writing of the manuscript. All authors have read and approved the final manuscript.

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Authors and Affiliations

  1. Animal Bone and Joint Research Laboratory, Department of Veterinary Biosciences and Public Health, Faculty of Veterinary Medicine, Chiang Mai University, Chiang Mai, 50100, Thailand

    Thippaporn Euppayo, Puntita Siengdee, Kittisak Buddhachat & Korakot Nganvongpanit

  2. Science and Technology Research Institute, Chiang Mai University, Chiang Mai, 50200, Thailand

    Waranee Pradit

  3. Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai, 50200, Thailand

    Siriwadee Chomdej

  4. Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Chiang Mai, 50200, Thailand

    Siriwan Ongchai

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  1. Thippaporn Euppayo
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  2. Puntita Siengdee
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  3. Kittisak Buddhachat
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Correspondence to Korakot Nganvongpanit.

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Euppayo, T., Siengdee, P., Buddhachat, K. et al. In vitro effects of triamcinolone acetonide and in combination with hyaluronan on canine normal and spontaneous osteoarthritis articular cartilage. In Vitro Cell.Dev.Biol.-Animal 52, 723–735 (2016). https://doi.org/10.1007/s11626-016-0022-4

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