Skip to main content
NCBI home page
Annals of the Rheumatic Diseases logoLink to Annals of the Rheumatic Diseases
. 2003 Dec;62(12):1139–1144. doi: 10.1136/ard.2003.007401

p53 in rheumatoid arthritis synovial fibroblasts at sites of invasion

C Seemayer 1, S Kuchen 1, M Neidhart 1, P Kuenzler 1, V Rihoskova 1, E Neumann 1, M Pruschy 1, W Aicher 1, U Muller-Ladner 1, R Gay 1, B Michel 1, G Firestein 1, S Gay 1
PMCID: PMC1754413  PMID: 14644850

Abstract

Objective: To analyse the functional response of p53 in rheumatoid arthritis synovial fibroblasts (RASF) in vitro and in vivo and to investigate whether activation of p53 modulates the destructive process of RASF.

Methods: RASF and controls grown on chamber slides were either directly examined with DO7 anti-p53 antibodies by immunofluorescence or irradiated with 10 Gy x rays and analysed time dependently for the expression of p53. The percentage of positive cells was evaluated by a quantitative scoring system. RASF and normal (N) SF cultured in vitro were co-implanted with human cartilage in SCID mice for 60 days. Consecutively, the invasion score was evaluated, and the number of p53 positive cells was determined at the sites of invasion by immunohistochemistry. In addition, synovial tissues from RA, osteoarthritis, and normal synovia were stained with DO7 antibodies.

Results: In vitro the rate of expression of p53 in RASF was low (<5%), but transiently inducible by ionising irradiation (50%). In vitro low p53 expressing RASF disclosed, when invading articular cartilage, a nuclear p53 signal in 20% of the cells, indicating the induction of p53 in a distinct population of RASF during the invasive process.

Conclusions: These data suggest an inductive p53 response at sites of cartilage invasion during the destructive process driven by activated RASF.

Full Text

The Full Text of this article is available as a PDF (435.0 KB).

Figure 1 .

Figure 1

Open in a new tab

Immunofluorescence for p53 in irradiated and non-irradiated RASF: (A) RASF 6 in passage 3 irradiated with 10 Gy x rays; (B) RASF 6 in passage 3 non-irradiated; (C) RASF 7 in passage 11 irradiated with 10 Gy x rays; (D) RASF 7 in passage 11 non-irradiated. Magnification x400.

Figure 2 .

Figure 2

Open in a new tab

Irradiation of RASF, NSF, and FSFB with 10 Gy x rays and fixation after distinct times. Expression of p53 was determined by immunofluorescence. p53 was transiently induced by ionising irradiation after 2.5 hours in all investigated fibroblast types. Twenty hours after irradiation almost p53 background expression levels were obtained.

Figure 3 .

Figure 3

Open in a new tab

Immunohistochemistry of p53 using DO7 antibodies in SCID mice sections of NSF, RASF, and SV40 transformed cells. In NSF, only limited invasion, but hardly any p53 signal was detectable. In RASF, expression of p53 was found in some cartilage invading cells. Strong p53 expression occurred in SV40 transformed RASF (positive control). Magnification x400.

Figure 4 .

Figure 4

Open in a new tab

Quantification of p53 positive cells at sites of invasion in the SCID mouse co-implantation model of RA. Data are expressed as means (SD). On average, p53 was expressed during the invasive process in 20% of cartilage invading RASF. In vitro, the same RASF were largely p53 negative (<5%).

Figure 5 .

Figure 5

Open in a new tab

Immunohistochemistry of p53 with DO7 antibodies of RA tissue specimens and controls. Limited expression of p53 was detected in RA synovium, but no expression in normal synovium. Strong nuclear p53 expression was found in a neurological tumour (positive control). Magnification RA2 x200, all others x400.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arnett F. C., Edworthy S. M., Bloch D. A., McShane D. J., Fries J. F., Cooper N. S., Healey L. A., Kaplan S. R., Liang M. H., Luthra H. S. The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis. Arthritis Rheum. 1988 Mar;31(3):315–324. doi: 10.1002/art.1780310302. [DOI] [PubMed] [Google Scholar]
  2. Firestein G. S., Echeverri F., Yeo M., Zvaifler N. J., Green D. R. Somatic mutations in the p53 tumor suppressor gene in rheumatoid arthritis synovium. Proc Natl Acad Sci U S A. 1997 Sep 30;94(20):10895–10900. doi: 10.1073/pnas.94.20.10895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Firestein G. S., Nguyen K., Aupperle K. R., Yeo M., Boyle D. L., Zvaifler N. J. Apoptosis in rheumatoid arthritis: p53 overexpression in rheumatoid arthritis synovium. Am J Pathol. 1996 Dec;149(6):2143–2151. [PMC free article] [PubMed] [Google Scholar]
  4. Gay S., Gay R. E., Koopman W. J. Molecular and cellular mechanisms of joint destruction in rheumatoid arthritis: two cellular mechanisms explain joint destruction? Ann Rheum Dis. 1993 Mar;52 (Suppl 1):S39–S47. doi: 10.1136/ard.52.suppl_1.s39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Inazuka M., Tahira T., Horiuchi T., Harashima S., Sawabe T., Kondo M., Miyahara H., Hayashi K. Analysis of p53 tumour suppressor gene somatic mutations in rheumatoid arthritis synovium. Rheumatology (Oxford) 2000 Mar;39(3):262–266. doi: 10.1093/rheumatology/39.3.262. [DOI] [PubMed] [Google Scholar]
  6. Kullmann F., Judex M., Neudecker I., Lechner S., Jüsten H. P., Green D. R., Wessinghage D., Firestein G. S., Gay S., Schölmerich J. Analysis of the p53 tumor suppressor gene in rheumatoid arthritis synovial fibroblasts. Arthritis Rheum. 1999 Aug;42(8):1594–1600. doi: 10.1002/1529-0131(199908)42:8<1594::AID-ANR5>3.0.CO;2-#. [DOI] [PubMed] [Google Scholar]
  7. Lee C. S., Portek I., Edmonds J., Kirkham B. Synovial membrane p53 protein immunoreactivity in rheumatoid arthritis patients. Ann Rheum Dis. 2000 Feb;59(2):143–145. doi: 10.1136/ard.59.2.143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Migita K., Tanaka F., Yamasaki S., Shibatomi K., Ida H., Kawakami A., Aoyagi T., Kawabe Y., Eguchi K. Regulation of rheumatoid synoviocyte proliferation by endogenous p53 induction. Clin Exp Immunol. 2001 Nov;126(2):334–338. doi: 10.1046/j.1365-2249.2001.01677.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Müller-Ladner U., Gay R. E., Gay S. Activation of synoviocytes. Curr Opin Rheumatol. 2000 May;12(3):186–194. doi: 10.1097/00002281-200005000-00005. [DOI] [PubMed] [Google Scholar]
  10. Müller-Ladner U., Kriegsmann J., Franklin B. N., Matsumoto S., Geiler T., Gay R. E., Gay S. Synovial fibroblasts of patients with rheumatoid arthritis attach to and invade normal human cartilage when engrafted into SCID mice. Am J Pathol. 1996 Nov;149(5):1607–1615. [PMC free article] [PubMed] [Google Scholar]
  11. Pap T., Aupperle K. R., Gay S., Firestein G. S., Gay R. E. Invasiveness of synovial fibroblasts is regulated by p53 in the SCID mouse in vivo model of cartilage invasion. Arthritis Rheum. 2001 Mar;44(3):676–681. doi: 10.1002/1529-0131(200103)44:3<676::AID-ANR117>3.0.CO;2-6. [DOI] [PubMed] [Google Scholar]
  12. Pap T., van der Laan W. H., Aupperle K. R., Gay R. E., Verheijen J. H., Firestein G. S., Gay S., Neidhart M. Modulation of fibroblast-mediated cartilage degradation by articular chondrocytes in rheumatoid arthritis. Arthritis Rheum. 2000 Nov;43(11):2531–2536. doi: 10.1002/1529-0131(200011)43:11<2531::AID-ANR21>3.0.CO;2-V. [DOI] [PubMed] [Google Scholar]
  13. Parak W. J., Dannöhl S., George M., Schuler M. K., Schaumburger J., Gaub H. E., Müller O., Aicher W. K. Metabolic activation stimulates acid production in synovial fibroblasts. J Rheumatol. 2000 Oct;27(10):2312–2322. [PubMed] [Google Scholar]
  14. Rème T., Travaglio A., Gueydon E., Adla L., Jorgensen C., Sany J. Mutations of the p53 tumour suppressor gene in erosive rheumatoid synovial tissue. Clin Exp Immunol. 1998 Feb;111(2):353–358. doi: 10.1046/j.1365-2249.1998.00508.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Sugiyama M., Tsukazaki T., Yonekura A., Matsuzaki S., Yamashita S., Iwasaki K. Localisation of apoptosis and expression of apoptosis related proteins in the synovium of patients with rheumatoid arthritis. Ann Rheum Dis. 1996 Jul;55(7):442–449. doi: 10.1136/ard.55.7.442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Tak P. P., Klapwijk M. S., Broersen S. F., van de Geest D. A., Overbeek M., Firestein G. S. Apoptosis and p53 expression in rat adjuvant arthritis. Arthritis Res. 2000 Mar 1;2(3):229–235. doi: 10.1186/ar92. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Tak P. P., Smeets T. J., Boyle D. L., Kraan M. C., Shi Y., Zhuang S., Zvaifler N. J., Breedveld F. C., Firestein G. S. p53 overexpression in synovial tissue from patients with early and longstanding rheumatoid arthritis compared with patients with reactive arthritis and osteoarthritis. Arthritis Rheum. 1999 May;42(5):948–953. doi: 10.1002/1529-0131(199905)42:5<948::AID-ANR13>3.0.CO;2-L. [DOI] [PubMed] [Google Scholar]
  18. Tak P. P., Zvaifler N. J., Green D. R., Firestein G. S. Rheumatoid arthritis and p53: how oxidative stress might alter the course of inflammatory diseases. Immunol Today. 2000 Feb;21(2):78–82. doi: 10.1016/s0167-5699(99)01552-2. [DOI] [PubMed] [Google Scholar]
  19. Yamanishi Yuji, Boyle David L., Pinkoski Michael J., Mahboubi Artin, Lin Tesu, Han Zuoning, Zvaifler Nathan J., Green Douglas R., Firestein Gary S. Regulation of joint destruction and inflammation by p53 in collagen-induced arthritis. Am J Pathol. 2002 Jan;160(1):123–130. doi: 10.1016/S0002-9440(10)64356-8. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Annals of the Rheumatic Diseases are provided here courtesy of BMJ Publishing Group

RESOURCES