Dual role for Fcγ receptors in host defense and disease in Borrelia burgdorferi-infected mice

Abstract

Arthritis in mice infected with the Lyme disease spirochete, Borrelia burgdorferi, results from the influx of innate immune cells responding to the pathogen in the joint and is influenced in part by mouse genetics. Production of inflammatory cytokines by innate immune cells in vitro is largely mediated by Toll-like receptor (TLR) interaction with Borrelia lipoproteins, yet surprisingly mice deficient in TLR2 or the TLR signaling molecule MyD88 still develop arthritis comparable to that seen in wild type mice after B. burgdorferi infection. These findings suggest that other, MyD88-independent inflammatory pathways can contribute to arthritis expression. Clearance of B. burgdorferi is dependent on the production of specific antibody and phagocytosis of the organism. As Fc receptors (FcγR) are important for IgG-mediated clearance of immune complexes and opsonized particles by phagocytes, we examined the role that FcγR play in host defense and disease in B. burgdorferi-infected mice. B. burgdorferi-infected mice deficient in the Fc receptor common gamma chain (FcεRγ(-/-) mice) harbored ~10 fold more spirochetes than similarly infected wild type mice, and this was associated with a transient increase in arthritis severity. While the elevated pathogen burdens seen in B. burgdorferi-infected MyD88(-/-) mice were not affected by concomitant deficiency in FcγR, arthritis was reduced in FcεRγ(-/-) MyD88(-/-) mice in comparison to wild type or single knockout mice. Gene expression analysis from infected joints demonstrated that absence of both MyD88 and FcγR lowers mRNA levels of proteins involved in inflammation, including Cxcl1 (KC), Xcr1 (Gpr5), IL-1beta, and C reactive protein. Taken together, our results demonstrate a role for FcγR-mediated immunity in limiting pathogen burden and arthritis in mice during the acute phase of B. burgdorferi infection, and further suggest that this pathway contributes to the arthritis that develops in B. burgdorferi-infected MyD88(-/-) mice.

Keywords: Borrelia burgdorferi; Fc receptor; Lyme disease; MyD88; arthritis; mice; toll-like receptors.

Figure 1

TNFα secretion by macrophages stimulated with OspC and OspC-immune complexes. Peritoneal macrophages from naïve WT and MyD88^−/− mice were stimulated in vitro with recombinant B. burgdorferi OspC protein or OspC-anti-OspC immune complexes for 24 h. The levels of secreted TNFα in the supernatants were measured by ELISA.

Figure 2

Quantitative PCR of B. burgdorferi in urinary bladders of infected mice. The copy number of the B. burgdorferi recA gene was normalized to the mouse β-actin gene by real-time PCR. Values are averages of 5–11 individual mice. Results are combined from 2 separate experiments. At day 14, the burdens in the FcεR_γ^−/− mice are statistically different from those in the WT mice (P = 0.0043) and those in the MyD88^−/− and FcεR_γ^−/− MyD88^−/− mice (P = 0.0029 and P = 0.0095, respectively). At day 21 the FcεR_γ^−/− mice maintain statistically higher burdens than the WT mice (P = 0.007) but lower burdens than the MyD88^−/− (P = 0.001) and the FcεR_γ^−/− MyD88^−/− (P ≤ 0.0001). At day 45 there is no difference between the WT and FcεR_γ^−/− mice but the MyD88^−/− and FcεR_γ^−/− MyD88^−/− burdens remain high. All P-values were calculated using the Mann-Whitney assay.

Figure 3

Arthritis severity is diminished in the combined absence of both FcγR and MyD88. Hind limb joints were analyzed, and each data point represents one joint. Arthritis was scored on a scale of 0 (negative) to 3 (severe). Four to 14 mice were used in each group and results were combined from 2 to 4 separate experiments. (A) shows arthritis severity at day 14, and (B) shows arthritis severity at day 21. Significant differences were observed in (A): WT vs. MyD88^−/− P = 0.007, FcεR_γ^−/− vs. MyD88^−/− P = 0.004, and MyD88^−/− vs. FcεR_γ^−/− MyD88^−/− P = 0.009. (B) FcεR_γ^−/− vs. WT P = 0.0026, FcεR_γ^−/− vs. MyD88^−/− P ≤ 0.0001, FcεR_γ^−/− vs. FcεR_γ^−/− MyD88^−/−P ≤ 0.0001, MyD88^−/− vs. FcεR_γ^−/− MyD88^−/−P = 0.0003, and WT vs. FcεR_γ^−/− MyD88^−/− P = 0.0013. P-values were determined using the unpaired Student t test.

Figure 4

Anti-B. burgdorferi IgG subclass titers are different in the single and double knockout mice. Anti-B. burgdorferi specific titers were determined by ELISA and are reported as the reciprocal of the endpoint positive titers. The FcεR_γ^−/− MyD88^−/− anti-B. burgdorferi IgG1 titers are significantly higher than the MyD88^−/− titers (P = 0.0113). The MyD88^−/− IgG 3 titers are significantly lower than the FcεR_γ^−/− MyD88^−/− titers (P = 0.0025) as well as the WT and FcεR_γ^−/− titers (P = 0.01 and.0025, respectively).