Effect of Methotrexate on Collagen-Induced Arthritis Assessed by Micro-Computed Tomography and Histopathological Examination in Female Rats

  • Kim, Young Hee (Department of Biomedical Laboratory Science, Namseoul University) ;
  • Kang, Jin Seok (Department of Biomedical Laboratory Science, Namseoul University)
  • Received : 2014.11.12
  • Accepted : 2014.12.29
  • Published : 2015.03.01


We tested the hypothesis that micro-computed tomography (micro-CT) analysis provides a better quantitative readout of the therapeutic potential of methotrexate (MTX) for treating collagen-induced arthritis (CIA) in rats and compared to conventional histopathological examination. Rats were divided into three groups: Group 1 (G1) was treated with 0.9% saline, whereas groups 2 (G2) and 3 (G3) were boosted with type II collagen at days 0 and 7. Following the first collagen immunization, rats in G1 and G2 were treated with 0.9% saline and those in G3 were treated with 1.5 mg/kg MTX from day 14 to 28. All rats were sacrificed on day 28, at which point and all hind knee joints were analyzed by micro-CT and histopathological examination. Micro-CT analyses showed that bone volume and trabecular number were significantly decreased in G2 and G3 compared to G1 (p<0.01), as was percent bone volume (p<0.05 and p<0.01, respectively). However, bone surface/bone volume was significantly increased in G2 and G3 compared to G1 (p<0.05 and p<0.01, respectively). Trabecular separation was significantly increased in G3 compared to G1 (p<0.05). Histopathological examination showed that knee joints of rats in G2 and G3 showed severe joint destruction with inflammatory cell infiltration. However, cartilage destruction was slightly reduced in G3 compared to G2. Taken together, these results suggest that MTX treatment reduced cartilage destruction in rats with CIA, and micro-CT analyses made it possible to quantify arthritic bony lesion.


Supported by : Ministry of Health and Welfare


  1. Afara, I. O., Prasadam, I., Crawford, R., Xiao, Y. and Oloyede, A. (2013) Near infrared (NIR) absorption spectra correlates with subchondral bone micro-CT parameters in osteoarthritic rat models. Bone 53, 350-357.
  2. Bevaart, L., Vervoordeldonk, M. J. and Tak, P. P. (2010) Evaluation of therapeutic targets in animal models of arthritis: how does it relate to rheumatoid arthritis? Arthritis Rheum. 62, 2192-2205.
  3. Brand, D. D., Latham, K. A. and Rosloniec, E. F. (2007) Collagen-induced arthritis. Nat. Protoc. 2, 1269-1275.
  4. Carvalheiro, H., da Silva, J. A. and Souto-Carneiro, M. M. (2013) Potential roles for CD8(+) T cells in rheumatoid arthritis. Autoimmun. Rev. 12, 401-409.
  5. Conway, J. R., Carragher, N. O. and Timpson, P. (2014) Developments in preclinical cancer imaging: innovating the discovery of therapeutics. Nat. Rev. Cancer 14, 314-328.
  6. Gerwin, N., Bendele, A. M., Glasson, S. and Carlson, C. S. (2010) The OARSI histopathology initiative - recommendations for histological assessments of osteoarthritis in the rat. Osteoarthritis Cartilage 18 Suppl 3, S24-34.
  7. Ishikawa, T., Nishigaki, F., Miyata, S., Hirayama, Y., Minoura, K., Imanishi, J., Neya, M., Mizutani, T., Imamura, Y., Naritomi, Y., Murai, H., Ohkubo, Y., Kagayama, A. and Mutoh, S. (2005) Prevention of progressive joint destruction in collagen-induced arthritis in rats by a novel matrix metalloproteinase inhibitor, FR255031. Br. J. Pharmacol. 144, 133-143.
  8. Koba, W., Jelicks, L. A. and Fine, E. J. (2013) MicroPET/SPECT/CT imaging of small animal models of disease. Am. J. Pathol. 182, 319-324.
  9. Maruotti, N., Cantatore, F. P., Crivellato, E., Vacca, A. and Ribatti, D. (2007) Macrophages in rheumatoid arthritis. Histol. Histopathol. 22, 581-586.
  10. Mello, S. B., Barros, D. M., Silva, A. S., Laurindo, I. M. and Novaes, G. S. (2000) Methotrexate as a preferential cyclooxygenase 2 inhibitor in whole blood of patients with rheumatoid arthritis. Rheumatology (Oxford) 39, 533-536.
  11. Myers, L. K., Rosloniec, E. F., Cremer, M. A. and Kang, A. H. (1997) Collagen-induced arthritis, an animal model of autoimmunity. Life Sci. 61, 1861-1878.
  12. Nagase, H. and Woessner, J. F., Jr. (1999) Matrix metalloproteinases. J. Biol. Chem. 274, 21491-21494.
  13. Otero, M. and Goldring, M. B. (2007) Cells of the synovium in rheumatoid arthritis. Chondrocytes. Arthritis Res. Ther. 9, 220.
  14. Perkins, D. J., St Clair, E. W., Misukonis, M. A. and Weinberg, J. B. (1998) Reduction of NOS2 overexpression in rheumatoid arthritis patients treated with anti-tumor necrosis factor alpha monoclonal antibody (cA2). Arthritis Rheum. 41, 2205-2210.<2205::AID-ART16>3.0.CO;2-Q
  15. Peterson, J. D., Labranche, T. P., Vasquez, K. O., Kossodo, S., Melton, M., Rader, R., Listello, J. T., Abrams, M. A. and Misko, T. P. (2010) Optical tomographic imaging discriminates between disease-modifying anti-rheumatic drug (DMARD) and non-DMARD efficacy in collagen antibody-induced arthritis. Arthritis Res. Ther. 12, R105.
  16. Ritsma, L., Vrisekoop, N. and van Rheenen, J. (2013) In vivo imaging and histochemistry are combined in the cryosection labelling and intravital microscopy technique. Nat. Commun. 4, 2366.
  17. Ryu, J. H., Lee, A., Chu, J. U., Koo, H., Ko, C. Y., Kim, H. S., Yoon, S. Y., Kim, B. S., Choi, K., Kwon, I. C., Kim, K. and Youn, I. (2011) Early diagnosis of arthritis in mice with collagen-induced arthritis, using a fluorogenic matrix metalloproteinase 3-specific polymeric probe. Arthritis Rheum. 63, 3824-3832.
  18. Schambach, S. J., Bag, S., Schilling, L., Groden, C. and Brockmann, M. A. (2010) Application of micro-CT in small animal imaging. Methods 50, 2-13.
  19. Scott, D. L. (2012) Biologics-based therapy for the treatment of rheumatoid arthritis. Clin. Pharmacol. Ther. 91, 30-43.
  20. Scott, D. L., Wolfe, F. and Huizinga, T. W. (2010) Rheumatoid arthritis. Lancet 376, 1094-1108.
  21. Segawa, Y., Yamaura, M., Aota, S., Omata, T., Tuzuike, N., Itokazu, Y., Oka, H., Tamaki, H. and Nakamura, T. (1997) Methotrexate maintains bone mass by preventing both a decrease in bone formation and an increase in bone resorption in adjuvant-induced arthritic rats. Bone 20, 457-464.
  22. Takaishi, H., Kimura, T., Dalal, S., Okada, Y. and D'Armiento, J. (2008) Joint diseases and matrix metalloproteinases: a role for MMP-13. Curr. Pharm. Biotechnol. 9, 47-54.
  23. Weinblatt, M. E., Kaplan, H., Germain, B. F., Merriman, R. C., Solomon, S. D., Wall, B., Anderson, L., Block, S., Small, R., Wolfe, F. and et al. (1991) Methotrexate in rheumatoid arthritis: effects on disease activity in a multicenter prospective study. J. Rheumatol. 18, 334-338.
  24. Weinblatt, M. E., Weissman, B. N., Holdsworth, D. E., Fraser, P. A., Maier, A. L., Falchuk, K. R. and Coblyn, J. S. (1992) Long-term prospective study of methotrexate in the treatment of rheumatoid arthritis. 84-month update. Arthritis Rheum. 35, 129-137.

Cited by

  1. Ramipril and haloperidol as promising approaches in managing rheumatoid arthritis in rats vol.765, 2015,
  2. Protective effects of fenofibrate and resveratrol in an aggressive model of rheumatoid arthritis in rats vol.54, pp.9, 2016,
  3. Old and new therapeutics for Rheumatoid Arthritis:in vivomodels and drug development vol.38, pp.1, 2016,
  4. Granisetron and carvedilol can protect experimental rats againstadjuvant-induced arthritis vol.39, pp.2, 2017,
  5. Possibilities for preventive treatment in rheumatoid arthritis? Lessons from experimental animal models of arthritis: a systematic literature review and meta-analysis vol.76, pp.2, 2017,
  6. Therapeutic effect of quercetin in collagen-induced arthritis vol.90, 2017,
  7. Identifying Reliable Diagnostic/Predictive Biomarkers for Rheumatoid Arthritis vol.13, pp.1177-2719, 2018,