Progression of Gene Expression Changes following a Mechanical Injury to Articular Cartilage as a Model of Early Stage Osteoarthritis

R S McCulloch, M S Ashwell, C Maltecca, A T O'Nan, P L Mente, R S McCulloch, M S Ashwell, C Maltecca, A T O'Nan, P L Mente

Abstract

An impact injury model of early stage osteoarthritis (OA) progression was developed using a mechanical insult to an articular cartilage surface to evaluate differential gene expression changes over time and treatment. Porcine patellae with intact cartilage surfaces were randomized to one of three treatments: nonimpacted control, axial impaction (2000 N), or a shear impaction (500 N axial, with tangential displacement to induce shear forces). After impact, the patellae were returned to culture for 0, 3, 7, or 14 days. At the appropriate time point, RNA was extracted from full-thickness cartilage slices at the impact site. Quantitative real-time PCR was used to evaluate differential gene expression for 18 OA related genes from four categories: cartilage matrix, degradative enzymes and inhibitors, inflammatory response and signaling, and cell apoptosis. The shear impacted specimens were compared to the axial impacted specimens and showed that shear specimens more highly expressed type I collagen (Col1a1) at the early time points. In addition, there was generally elevated expression of degradative enzymes, inflammatory response genes, and apoptosis markers at the early time points. These changes suggest that the more physiologically relevant shear loading may initially be more damaging to the cartilage and induces more repair efforts after loading.

Figures

Figure 1
Figure 1
Fold changes for shear versus axial specimens within each time point. Fold change is shown on the vertical axis as a log scale, and days in culture are shown on the horizontal axis. Error bars for fold changes are depicted and significant differences for the respective comparison are indicated with an asterisk (∗). A graph is shown for each gene category: cartilage matrix (a), degradative enzymes (b), inflammatory response (c), and apoptosis (d).

References

    1. NIAMS . Osteoarthritis. In: NIAMS, editor. NIH. 06-4617 2006.
    1. Buckwalter J. A., Lane N. E. Athletics and osteoarthritis. American Journal of Sports Medicine. 1997;25(6):873–881. doi: 10.1177/036354659702500624.
    1. Donohue J. M., Buss D., Oegema T. R., Jr., Thompson R. C., Jr. The effects of indirect blunt trauma on adult canine articular cartilage. Journal of Bone and Joint Surgery. 1983;65(7):948–957.
    1. Jeffrey J. E., Brodie J. P., Aspden R. M. P131 the effects of a single fast impact load compared with slow severe loading on articular cartilage in vitro. Osteoarthritis and Cartilage. 2006;14:S81–S82. doi: 10.1016/S1063-4584(07)60583-4.
    1. Torzilli P. A., Grigiene R., Borrelli J., Jr., Helfet D. L. Effect of impact load on articular cartilage: cell metabolism and viability, and matrix water content. Journal of Biomechanical Engineering. 1999;121(5):433–441. doi: 10.1115/1.2835070.
    1. Ficklin T., Thomas G., Barthel J. C., Asanbaeva A., Thonar E. J., Masuda K., Chen A. C., Sah R. L., Davol A., Klisch S. M. Articular cartilage mechanical and biochemical property relations before and after in vitro growth. Journal of Biomechanics. 2007;40(16):3607–3614. doi: 10.1016/j.jbiomech.2007.06.005.
    1. Wilson W., van Burken C., van Donkelaar C. C., Buma P., van Rietbergen B., Huiskes R. Causes of mechanically induced collagen damage in articular cartilage. Journal of Orthopaedic Research. 2006;24(2):220–228. doi: 10.1002/jor.20027.
    1. Ashwell M. S., O'Nan A. T., Gonda M. G., Mente P. L. Gene expression profiling of chondrocytes from a porcine impact injury model. Osteoarthritis and Cartilage. 2008;16(8):936–946. doi: 10.1016/j.joca.2007.12.012.
    1. Aigner T., Zien A., Hanisch D., Zimmer R. Gene expression in chondrocytes assessed with use of microarrays. Journal of Bone and Joint Surgery. 2003;85(1, supplement 2):117–123. doi: 10.1302/0301-620X.85B1.12665.
    1. Salzmann G. M., Nuernberger B., Schmitz P., Anton M., Stoddart M. J., Grad S., Milz S., Tischer T., Vogt S., Gansbacher B., Imhoff A. B., Alini M. Physicobiochemical synergism through gene therapy and functional tissue engineering for in vitro chondrogenesis. Tissue engineering. Part A. 2009;15(9):2513–2524. doi: 10.1089/ten.tea.2008.0479.
    1. Aigner T., Dudhia J. Genomics of osteoarthritis. Current Opinion in Rheumatology. 2003;15(5):634–640. doi: 10.1097/00002281-200309000-00019.
    1. Fehrenbacher A., Steck E., Rickert M., Roth W., Richter W. Rapid regulation of collagen but not metalloproteinase 1, 3, 13, 14 and tissue inhibitor of metalloproteinase 1, 2, 3 expression in response to mechanical loading of cartilage explants in vitro. Archives of Biochemistry and Biophysics. 2003;410(1):39–47. doi: 10.1016/S0003-9861(02)00658-6.
    1. Fitzgerald J. B., Jin M., Dean D., Wood D. J., Zheng M. H., Grodzinsky A. J. Mechanical compression of cartilage explants induces multiple time-dependent gene expression patterns and involves intracellular calcium and cyclic AMP. The Journal of Biological Chemistry. 2004;279(19):19502–19511. doi: 10.1074/jbc.M400437200.
    1. Fitzgerald J. B., Jin M., Grodzinsky A. J. Shear and compression differentially regulate clusters of functionally related temporal transcription patterns in cartilage tissue. Journal of Biological Chemistry. 2006;281(34):24095–24103. doi: 10.1074/jbc.M510858200.
    1. Natoli R. M., Scott C. C., Athanasiou K. A. Temporal effects of impact on articular cartilage cell death, gene expression, matrix biochemistry, and biomechanics. Annals of Biomedical Engineering. 2008;36(5):780–792. doi: 10.1007/s10439-008-9472-5.
    1. Newberry W. N., Mackenzie C. D., Haut R. C. Blunt impact causes changes in bone and cartilage in a regularly exercised animal model. Journal of Orthopaedic Research. 1998;16(3):348–354. doi: 10.1002/jor.1100160311.
    1. Thompson R. C., Jr., Oegema T. R., Jr., Lewis J. L., Wallace L. Osteoarthrotic changes after acute transarticular load. An animal model. Journal of Bone and Joint Surgery—Series A. 1991;73(7):990–1001.
    1. Flachsmann E. R., Broom N. D., Oloyede A. A biomechanical investigation of unconstrained sheer failure of the osteochondral region under impact loading. Clinical Biomechanics. 1995;10(3):156–165. doi: 10.1016/0268-0033(95)93706-Y.
    1. Tomatsu T., Imai N., Takeuchi N., Takahashi K., Kimura N. Experimentally produced fractures of articular cartilage and bone: the effects of shear forces on the pig knee. Journal of Bone and Joint Surgery B. 1992;74(3):457–462.
    1. Flannery C. R., Little C. B., Caterson B., Hughes C. E. Effects of culture conditions and exposure to catabolic stimulators (IL-1 and retinoic acid) on the expression of matrix metalloproteinases (MMPs) and disintegrin metalloproteinases (ADAMs) by articular cartilage chondrocytes. Matrix Biology. 1999;18(3):225–237. doi: 10.1016/S0945-053X(99)00024-4.
    1. Vincent T., Hermansson M., Bolton M., Wait R., Saklatvala J. Basic FGF mediates an immediate response of articular cartilage to mechanical injury. Proceedings of the National Academy of Sciences of the United States of America. 2002;99(12):8259–8264. doi: 10.1073/pnas.122033199.
    1. Fermor B., Weinberg J. B., Pisetsky D. S., Misukonis M. A., Fink C., Guilak F. Induction of cyclooxygenase-2 by mechanical stress through a nitric oxide-regulated pathway. Osteoarthritis and Cartilage. 2002;10(10):792–798. doi: 10.1053/joca.2002.0832.
    1. McCulloch R. S., Ashwell M. S., O'Nan A. T., Mente P. L. Identification of stable normalization genes for quantitative real-time PCR in porcine articular cartilage. Journal of Animal Science and Biotechnology. 2012;3(1, article 36) doi: 10.1186/2049-1891-3-36.
    1. Steibel J. P., Poletto R., Coussens P. M., Rosa G. J. M. A powerful and flexible linear mixed model framework for the analysis of relative quantification RT-PCR data. Genomics. 2009;94(2):146–152. doi: 10.1016/j.ygeno.2009.04.008.
    1. Benjamini Y., Hochberg Y. Controlling the false discovery rate: a practical and powerful approach to multiple testing. Journal of the Royal Statistical Society Series B: Methodological. 1995;57(1):289–300.
    1. Reiner A., Yekutieli D., Benjamini Y. Identifying differentially expressed genes using false discovery rate controlling procedures. Bioinformatics. 2003;19(3):368–375. doi: 10.1093/bioinformatics/btf877.
    1. Sandell L. J., Aigner T. Articular cartilage and changes in arthritis: cell biology of osteoarthritis. Arthritis Research. 2001;3(2):107–113. doi: 10.1186/ar148.
    1. Xu Y., Barter M. J., Swan D. C., et al. Identification of the pathogenic pathways in osteoarthritic hip cartilage: commonality and discord between hip and knee OA. Osteoarthritis and Cartilage. 2012;20(9):1029–1038. doi: 10.1016/j.joca.2012.05.006.
    1. Peffers M. J., Liu X., Clegg P. D. Transcriptomic signatures in cartilage ageing. Arthritis Research and Therapy. 2013;15(4, article R98) doi: 10.1186/ar4278.
    1. Zhang C., Wei X., Chen C., Cao K., Li Y., Jiao Q., Ding J., Zhou J., Fleming B. C., Chen Q., Shang X., Wei L. Indian hedgehog in synovial fluid is a novel marker for early cartilage lesions in human knee joint. International Journal of Molecular Sciences. 2014;15(5):7250–7265. doi: 10.3390/ijms15057250.
    1. Brew C., Clegg P., Boot-Handford R., Andrews G., Hardingham T. Gene expression in human chondrocytes in late osteoarthritis is changed in both fibrillated and intact cartilage without evidence of generalised chondrocyte hypertrophy. British Medical Journal. 2010;69(1):234–240.
    1. Lee C. M., Kisiday J. D., McIlwraith C. W., Grodzinsky A. J., Frisbie D. D. Development of an in vitro model of injuryinduced osteoarthritis in cartilage explants from adult horses through application of single-impact compressive overload. American Journal of Veterinary Research. 2013;74(1):40–47. doi: 10.2460/ajvr.74.1.40.
    1. Sanchez-Adams J., Leddy H. A., McNulty A. L., O'Conor C. J., Guilak F. The mechanobiology of articular cartilage: bearing the burden of osteoarthritis. Current Rheumatology Reports. 2014;16(10):451.
    1. van der Kraan P. M., van den Berg W. B. Chondrocyte hypertrophy and osteoarthritis: role in initiation and progression of cartilage degeneration? Osteoarthritis and Cartilage. 2012;20(3):223–232. doi: 10.1016/j.joca.2011.12.003.
    1. Dieppe P. A., Lohmander L. S. Pathogenesis and management of pain in osteoarthritis. The Lancet. 2005;365(9463):965–973. doi: 10.1016/S0140-6736(05)71086-2.
    1. Ding L., Guo D., Homandberg G. A., Buckwalter J. A., Martin J. A. A single blunt impact on cartilage promotes fibronectin fragmentation and upregulates cartilage degrading stromelysin-1/matrix metalloproteinase-3 in a bovine ex vivo model. Journal of Orthopaedic Research. 2014;32(6):811–818. doi: 10.1002/jor.22610.
    1. Lee J. H., Fitzgerald J. B., DiMicco M. A., Grodzinsky A. J. Mechanical injury of cartilage explants causes specific time-dependent changes in chondrocyte gene expression. Arthritis and Rheumatism. 2005;52(8):2386–2395. doi: 10.1002/art.21215.
    1. Tchetina E. V., Squires G., Poole A. R. Increased type II collagen degradation and very early focal cartilage degeneration is associated with upregulation of chondrocyte differentiation related genes in early human articular cartilage lesions. The Journal of Rheumatology. 2005;32(5):876–886.
    1. Swingler T. E., Waters J. G., Davidson R. K., et al. Degradome expression profiling in human articular cartilage. Arthritis Research & Therapy. 2009;11(3, article R96) doi: 10.1186/ar2741.
    1. Blaney Davidson E. N., van der Kraan P. M., van den Berg W. B. TGF-β and osteoarthritis. Osteoarthritis and Cartilage. 2007;15(6):597–604. doi: 10.1016/j.joca.2007.02.005.
    1. Allen R. T., Robertson C. M., Harwood F. L., Sasho T., Williams S. K., Pomerleau B. A., Amiel D. Characterization of mature vs aged rabbit articular cartilage: analysis of cell density, apoptosis-related gene expression and mechanisms controlling chondrocyte apoptosis. Osteoarthritis and Cartilage. 2004;12(11):917–923. doi: 10.1016/j.joca.2004.08.003.
    1. Pennock A. T., Robertson C. M., Emmerson B. C., Harwood F. L., Amiel D. Role of apoptotic and matrix-degrading genes in articular cartilage and meniscus of mature and aged rabbits during development of osteoarthritis. Arthritis and Rheumatism. 2007;56(5):1529–1536. doi: 10.1002/art.22523.
    1. Kim H. A., Lee Y. J., Seong S. C., Choe K. W., Song Y. W. Apoptotic chondrocyte death in human osteoarthritis. Journal of Rheumatology. 2000;27(2):455–462.
    1. Ashwell M. S., Gonda M. G., Gray K., Maltecca C., O'Nan A. T., Cassady J. P., Mente P. L. Changes in chondrocyte gene expression following in vitro impaction of porcine articular cartilage in an impact injury model. Journal of Orthopaedic Research. 2013;31(3):385–391. doi: 10.1002/jor.22239.
    1. Desjardin C., Vaiman A., Mata X., et al. Next-generation sequencing identifies equine cartilage and subchondral bone miRNAs and suggests their involvement in osteochondrosis physiopathology. BMC Genomics. 2014;15, article 798
    1. Gonzalez A. Osteoarthritis year 2013 in review: genetics and genomics. Osteoarthritis and Cartilage. 2013;21(10):1443–1451. doi: 10.1016/j.joca.2013.07.001.
    1. Peffers M. J., Liu X., Clegg P. D. Transcriptomic profiling of cartilage ageing. Genomics Data. 2014;2:27–28. doi: 10.1016/j.gdata.2014.03.001.
    1. Tew S. R., Mcdermott B. T., Fentem R. B., Peffers M. J., Clegg P. D. Transcriptome-wide analysis of mRNA decay in normal and osteoarthritic human articular chondrocytes. Arthritis & Rheumatology. 2014;66(11):3052–3061. doi: 10.1002/art.38849.

Source: PubMed

Sponsorer og samarbeidspartnere

Medisinsk tilstand

Legemiddelintervensjoner

3
Abonnere