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The Mycoplasma arthritidis mitogen (MAM) superantigen (SAg) is a potent activator of human and murine cells and is produced by an organism that is a cause of acute and chronic arthritis of rodents. It is phylogenetically unrelated to other bacterial SAgs and exhibits a number of unique features. We recently demonstrated that MAM differentially regulates the cytokine responses of different mouse strains following in vivo administration. Here we show that the presence in inbred C3H/HeJ mice of the mutant Lps(d) gene, which is associated with a defect in Toll-like receptor 4 (TLR4), influences MAM regulation of cytokine profiles in vivo. Whereas the levels of type 1 cytokines (interleukin-2 [IL-2], gamma interferon, IL-12, and tumor necrosis factor alpha) were depressed in cells from MAM-injected wild-type C3H/HeSnJ mice, they were elevated in cells from C3H/HeJ mice. Furthermore, the levels of type 2 cytokines (IL-4, IL-6, and IL-10) were elevated in Lps(n) C3H/HeSnJ mice but depressed in Lps(d) C3H/HeJ mice. The transcript for IL-12 p40 was highly expressed in C3H/HeJ but not C3H/HeSnJ mice. F(1) mice exhibited the same cytokine profile as C3H/HeJ mice, indicating that the mutant gene exhibited dominant-negative inheritance. In addition, C3H/HeJ mice were highly susceptible to toxic death in comparison with C3H/HeSnJ mice after injection with live M. arthritidis organisms. Our results suggest that MAM interacts with the lipopolysaccharide signaling pathway, possibly involving TLR4 or a combinatorial Toll complex.

Mycoplasma arthritidis mitogen (MAM) is a potent superantigen secreted by M. arthritidis, an agent of murine arthritis. Here we compare the abilities of MAM to induce a panel of cytokines in vitro and in vivo in BALB/c and C3H/HeJ mouse strains that differ in susceptibility to mycoplasmal arthritis. Splenocytes from both mouse strains produced high levels of all cytokines by 24 h following in vitro exposure to MAM. No differences in cytokine profiles were seen irrespective of the MAM dose. However, there were striking differences in cytokine profiles present in supernatants of splenocytes that had been collected from mice after intravenous (i.v. ) injection of MAM and subsequently rechallenged with MAM in vitro. Splenocytes collected 24 and 72 h after i.v. injection of MAM and challenged in vitro with MAM showed the most marked divergence in the secreted cytokines. Type 1 cytokines were markedly elevated in C3H/HeJ cell supernatants, whereas they were depressed or remained low in BALB/c cell supernatants. In contrast, the levels of type 2 cytokines were all greatly increased in BALB/c cell cultures but were decreased or remained low in C3H/HeJ supernatants. Interleukin-12 mRNA and protein was also markedly elevated in C3H/HeJ mice, as were the levels of immunoglobulin G2a. The data indicate a major skewing in cytokine profiles to a type 1 inflammatory response in C3H/HeJ mice but to a protective type 2 response in BALB/c mice. These cytokine changes appear to be associated with the severe arthritis in C3H/HeJ mice following injection of M. arthritidis in comparison to the mild disease seen in injected BALB/c mice.

Mycoplasma arthritidis, an agent of rodent arthritis, produces a potent superantigen (SAg), MAM. Previous work established that MAM is presented to T cells by murine H-2E or the homologous human HLA-DR molecules and that lymphocytes lacking a functional H-2E molecule fail to respond to MAM. Recently, more potent and purified preparations of MAM of known protein content have become available. This enabled us to more effectively compare the response of MAM with that of other SAgs by using lymphocytes from mice whose cells express different H-2A and HLA-DQ molecules. Here we demonstrate that cells from some H-2E-negative mouse strains respond to higher concentrations of MAM. By use of inbred, congenic, and recombinant mice, we show that these differences are, in fact, exercised at the level of the major histocompatibility complex (MHC) and that allelic polymorphisms at H-2A influence reactivity to MAM. In addition, polymorphisms at HLA-DQ, the human homolog of H-2A, also influence responsiveness to MAM. Cells expressing DQw6 (HLA-DQA1*0103 and DQBI*0601 chains) gave much higher responses to MAM than did cells expressing DQw8 (DQA1*0301 and DQB1*0302 chains). In fact, responses of lymphocytes expressing DQB1*0601 chains homozygously were as high as those observed for cells expressing a functional H-2E molecule. Murine lymphocytes responded less well to staphylococcal enterotoxin B (SEB) and SEA, but mouse cells expressing human MHC molecules gave much higher responses. The patterns of reactivity observed with cells expressing the various murine and human alleles differed for MAM, SEB, and SEA, suggesting that each of these SAgs interacts with different regions or residues on MHC molecules. It has been hypothesized that SAgs might play a role in susceptibility to autoimmune disease. Allelic polymorphisms at MHC loci might therefore influence susceptibility to autoimmune disease by affecting immunoreactivity to specific superantigens.