Date Published: October 4, 2018
Publisher: Public Library of Science
Author(s): YuFeng Peng, Jose C. Crispin.
Defective clearance of apoptotic cells in MFG-E8 deficient mice results in lupus-like disease in the mixed B6x129, but not pure B6 background. The lack of overt autoimmunity in MFG-E8-/- B6 mice suggests that accumulation of apoptotic cells is not sufficient to break central tolerance. However, the delayed clearance of apoptotic cells in the follicles of MFG-E8-/- B6 mice provides an excellent opportunity to investigate how B cells respond to excessive apoptotic cells in the periphery under relatively non-inflammatory conditions. In MFG-E8-/- B6 mice, we found increased IgG2c production against apoptotic cells and oxidized LDL. Apoptotic cell induced antibody responses depended on MyD88 signal and T cell help. In addition, MFG-E8-/- B6 mice had enlarged MZ B cell compartments as well as an enhanced antibody response to NP-Ficoll. Moreover, a significant percentage of MZ B cells in aged MFG-E8-/- B6 mice migrated into follicles. Injecting apoptotic cells or oxidized LDL into wild type mice as well as physiological accumulation of LDL in ApoE-/- mice recapitulated the translocation of MZ B cells. To determine how MFG-E8 deficiency affects the functions of autoreactive B cells specific for nucleic acids in the periphery under non-inflammatory conditions, we utilized BCR transgenic mice to bypass central selection and compared the differentiation of TLR9 dependent anti-dsDNA 56R B cells and TLR7 dependent anti-ssRNA H564 B cells in MFG-E8-/- mice. In MFG-E8-/- 56R mice, anti-dsDNA specific 56R/Vκ38c B cells differentiated into MZ B cells but not AFCs. On the contrary, in MFG-E8-/-H564 mice, anti-ssRNA specific H564 B cells further differentiated into GC B cells and AFCs. Adoptive transfer of activated autoreactive B cells confirmed that H564 B cells were more sensitive to apoptotic cell antigens than 56R B cells. Our observations provide new insights about the MZ B cell translocation in lupus patients as well as the dichotomy of TLR9 and TLR7 signals in the pathogenesis of lupus.
Both central and peripheral tolerance play critical roles in controlling autoreactive B cells . Most antibodies encoded by the germline are autoreactive. In bone marrow, autoreactive immature B cells are either deleted, forced to undergo receptor editing, or become anergic. Once they arrive in the periphery, mature B cells can re-acquire auto-reactivity through somatic mutation during GC (germinal center) reaction. Antigens deposited on FDCs (follicular dendritic cells) in the GC play an important role in selecting mutated B cells: B cells with the highest affinities differentiate into memory cells, whereas those with low affinities, including potentially autoreactive clones are deleted. The selection of autoreactive B cells depends on the threshold of B cell activation. Numerous animal models have demonstrated that defects in both central and peripheral B cell tolerance are required to develop overt lupus-like disease.
In our study, we found MFG-E8 deficient mice had higher levels of IgG2b and 2c antibodies against apoptotic cells and oxidized LDL than w.t. mice. The class switching we observed is different than reported by Chikazawa et al., who described an increase of anti- IgM but not IgG against peroxidized lipids in MFG-E8-/- mice . Although the reason for this discrepancy is not clear, our results demonstrated excess apoptotic cells, without any adjuvant, can induce a T cell-dependent response. Since TCRα deficiency only partially reduced the anti-apoptotic cell IgG2c response, an innate B cell population such as MZ B cells may also contribute to this response. In fact, MZ B cells are capable of producing anti-viral particle IgG antibodies without T cell help . Surprisingly, we found both IgM and IgG2c responses against apoptotic cells were completely abrogated by MyD88 deficiency (Fig 4). Previous studies have firmly linked anti-dsDNA and anti-ssRNA antibody responses in murine lupus models to the TLR-MyD88 pathway [13, 14]. DsDNA and ssRNA together with their binding antibodies can engage TLR9 and TLR7 to provide essential co-stimulatory signals to perpetuate an autoreactive B cell reaction. However, antibodies against nucleic acids are absent in MFG-E8-/- mice in a B6 background. Therefore, apoptotic cells might provide other TLR ligands, such as HMGB-1 and oxidized LDL [37, 38], to co-stimulate the antibody response. As the B cell response to apoptotic cells is the precursor of autoantibody development, our results suggest the importance of the TLR-MyD88 pathway in autoantibody production is not limited to nucleic acid specific antibodies. The predominance of IgG2b and 2c subclasses in the autoantibody profile of 4 month old MFG-E8-/- mice differs from the IgM autoantibody induced by repeated injection of apoptotic cells [20, 39, 40]. Deposition of apoptotic cell fragments on FDCs in MFG-E8-/- mice may be required for class switching . It is interesting to note that the autoantibody responses triggered by apoptotic cells were not sustained in old MFG-E8-/- mice (Fig 3). The strength of costimulatory signal(s) provided by non-nucleic acid stimuli might be weaker than those provided by nucleic acids.