Point mutation of tyrosine 759 of the IL-6 family cytokine receptor, gp130, augments collagen-induced arthritis in DBA/1J mice

Abstract

Background: Knock-in mice (gp130F759) with a Y759F point mutation in gp130, a signal transducing receptor subunit shared by members of the IL-6 cytokine family, show sustained activation of STAT3, enhanced acute-phase or immune responses, and autoimmune arthritis. We conducted a detailed analysis of collagen-induced arthritis (CIA) in gp130F759 with a DBA/1J background (D/J.gp130F759).

Methods: We backcrossed gp130F759 to C57BL/6 and DBA/1J, and compared the pathologic changes, including occurrence of arthritis, in the two distinct genetic backgrounds. We analyzed CIA in D/J.gp130F759 and investigated the effects of methotrexate (MTX) on CIA.

Results: C57BL/6 background gp130F759 mice, but not D/J.gp130F759, spontaneously developed polyarthritis and glomerulonephritis. On the other hand, keratitis of the eyes only developed in D/J.gp130F759, indicating the influence of genetic background on disease development in gp130F759 mice. Resistance of the DBA/1J background against spontaneous arthritis urged us to examine CIA in D/J.gp130F759. CIA in D/J.gp130F759 was more severe, with greater bone destruction, than the control mice. After collagen immunization, splenomegaly and serum levels of rheumatoid factor and anti-DNA antibody were augmented in D/J.gp130F759. Bio-Plex analysis of serum cytokines revealed increased IL-12p40 and PDGF-BB before immunization, and increased levels of IFN-gamma, IL-17, TNF-alpha, IL-9, and MIP-1beta 8 days after the booster dose. IL-6 and PDGF-BB in D/J.gp130F759 showed distinct kinetics from the other cytokines; higher levels were observed after arthritis development. MTX partially attenuated the development of arthritis and inhibited bone destruction in D/J.gp130F759, with reduction of anti-type II collagen antibody levels, suggesting that MTX mainly affects antigen-specific immune responses in CIA.

Conclusion: The Tyr-759 point mutation of the IL-6 family cytokine receptor subunit, gp130, caused autoimmune disease, and this was also influenced by the genetic background. CIA in D/J.gp130F759 is useful for evaluating drugs in a relatively short period because sustained activation of STAT3 may enhance the disease symptoms.

Figure 1

Severe arthritis developed in gp130F759 mice by collagen immunization. D/J.gp130F759 and wild-type mice were injected intradermally with collagen emulsified in FCA on day 0 and day 14. Values are expressed as mean ± standard error of 7 to 10 animals. *P < 0.05, **P < 0.01 versus wild-type mice by Student's t-test.

Figure 2

Marked splenomegaly in CIA of D/J.gp130F759 mice. Values are expressed as mean ± standard error of 7 to 10 animals of collagen-immunized mice on day 56 (a) and 3 to 4 animals of non-immunized mice (b). *P < 0.05, **P < 0.01 versus wild-type mice by Student's t-test.

Figure 3

Increased serum levels of rheumatoid factor in CIA of D/J.gp130F759 mice. Values are expressed as mean ± standard error of 7 to 10 animals of collagen-immunized mice on day 56 (a) and 3 to 4 animals of non-immunized mice (b). *P < 0.05, **P < 0.01 versus wild-type mice by Student's t-test.

Figure 4

Increased serum levels of anti-ssDNA antibody in CIA of D/J.gp130F759 mice. Values are expressed as mean ± standard error of 7 to 10 animals of collagen-immunized mice on day 56 (a) and 3 to 4 animals of non-immunized mice (b). *P < 0.05, **P < 0.01 versus wild-type mice by Student's t-test.

Figure 5

Serum levels of anti-collagen antibody in CIA of D/J.gp130F759 mice on day 56. Values are expressed as mean ± standard error of 7 to 10 animals on day 56. *P < 0.05 versus wild-type mice by Student's t-test.

Figure 6

Serum cytokines that showed different responses in D/J.gp130F759 mice (female). Serum cytokine levels were measured by Bio-Plex system. Values are expressed as mean ± standard error of 7 to 10 animals. *P < 0.05, **P < 0.01 versus wild-type mice at same day by Student's t-test.

Figure 7

IL-2 production in lymph node cells of D/J.gp130F759 mice (female) by collagen and concanavalin A stimulation. Values are expressed as mean ± standard error of 12 samples. **P < 0.01 versus non-treated group (Spon) by Student's t-test. #P < 0.05, ## P < 0.01 versus same condition group of wild type mice-derived lymph node cells by Student's t-test.

Figure 8

Methotrexate attenuated CIA (a), and delayed the onset of arthritis (b) in gp130F759 mice (male). Values are expressed as mean ± standard error of 11 to 12 animals. *P < 0.05, **P < 0.01 versus vehicle-treated group by Student's t-test. ^aP < 0.05, ^aaP < 0.01 versus Non-immunized group by Student's t-test.

Figure 9

Methotrexate attenuated the increase of COMP levels (a) and anti-collagen antibody levels (b) in the serum on day 42. Values are expressed as mean ± standard error of 11 to 12 animals. *P < 0.05, **P < 0.01 versus vehicle-treated group by Student's t-test. ^aaP < 0.01 versus Non-immunized group (NI) by Student's t-test.