Dendritic cells (DCs) are critical for immune homeostasis. To target DCs, we generated a mouse line with Flip deficiency in cells that express cre under the CD11c-promoter (CD11c-Flip-KO). CD11c-Flip-KO mice spontaneously develop erosive, inflammatory arthritis, resembling rheumatoid arthritis, which is dramatically reduced when these mice are crossed with Rag−/− mice. The CD8α+ DC subset is significantly reduced, along with alterations in NK cells and macrophages. Autoreactive CD4+ T cells and autoantibodies specific for joint tissue are present and arthritis severity correlates with the number of autoreactive CD4+ T cells and plasmablasts in the joint draining lymph nodes. Reduced T regulatory cells (Tregs) inversely correlate with arthritis severity, and the transfer of Tregs ameliorates arthritis. This KO line identifies a model that will permit in depth interrogation of the pathogenesis of rheumatoid arthritis, including the role of CD8α+ DCs and other cells of the immune system.

Little is known about the mechanisms regulating the transition of circulating monocytes into pro- or anti-inflammatory macrophages in chronic inflammation. Here, we took advantage of our novel mouse model of rheumatoid arthritis, in which Flip is deleted under the control of a CD11c promoter (HUPO mice). During synovial tissue homeostasis, both monocyte-derived F4/80int and self-renewing F4/80hi tissue–resident, macrophage populations were identified. However, in HUPO mice, decreased synovial tissue–resident macrophages preceded chronic arthritis, opened a niche permitting the influx of activated monocytes, with impaired ability to differentiate into F4/80hi tissue–resident macrophages. In contrast, Flip-replete monocytes entered the vacated niche and differentiated into tissue-resident macrophages, which suppressed arthritis. Genes important in macrophage tissue residency were reduced in HUPO F4/80hi macrophages and in leukocyte-rich rheumatoid arthritis synovial tissue monocytes. Our observations demonstrate that the macrophage tissue–resident niche is necessary for suppression of chronic inflammation and may contribute to the pathogenesis of rheumatoid arthritis.

We previously observed that SNAPIN, which is an adaptor protein in the SNARE core complex, was highly expressed in rheumatoid arthritis synovial tissue macrophages, but its role in macrophages and autoimmunity is unknown. To identify SNAPIN's role in these cells, we employed siRNA to silence the expression of SNAPIN in primary human macrophages. Silencing SNAPIN resulted in swollen lysosomes with impaired CTSD (cathepsin D) activation, although total CTSD was not reduced. Neither endosome cargo delivery nor lysosomal fusion with endosomes or autophagosomes was inhibited following the forced silencing of SNAPIN. The acidification of lysosomes and accumulation of autolysosomes in SNAPINsilenced cells was inhibited, resulting in incomplete lysosomal hydrolysis and impaired macroautophagy/ autophagy flux. Mechanistic studies employing ratiometric color fluorescence on living cells demonstrated that the reduction of SNAPIN resulted in a modest reduction of H C pump activity; however, the more critical mechanism was a lysosomal proton leak. Overall, our results demonstrate that SNAPIN is critical in the maintenance of healthy lysosomes and autophagy through its role in lysosome acidification and autophagosome maturation in macrophages largely through preventing proton leak. These observations suggest an important role for SNAPIN and autophagy in the homeostasis of macrophages, particularly long-lived tissue resident macrophages.

The reduction of synovial tissue macrophages is a reliable biomarker for clinical improvement in patients with rheumatoid arthritis (RA), and macrophages are reduced in synovial tissue shortly after initiation of TNF inhibitors. The mechanism for this initial response is unclear. These studies were performed to identify the mechanisms responsible for the initial reduction of macrophages following TNF inhibition, positing that efflux to draining lymph nodes was involved. RA synovial tissue and synovial fluid macrophages expressed CCR7, which was increased in control macrophages following incubation with TNF-α. Human TNF transgenic (hTNF-Tg) mice were treated with infliximab after development of arthritis. Ankles were harvested and examined by histology, immunohistochemistry, quantitative RT-PCR, ELISA, and flow cytometry. hTNF-Tg mice treated with infliximab demonstrated significant clinical and histologic improvement 3 d after the initiation of therapy, at which time Ly6C macrophages were significantly reduced in the ankles. However, no evidence was identified to support a role of macrophage efflux to draining lymph nodes following treatment with infliximab. In contrast, apoptosis of Ly6C macrophages in the ankles and popliteal lymph nodes, decreased migration of monocytes into the ankles, and a reduction of CCL2 were identified following the initiation of infliximab. These observations demonstrate that Ly6C macrophage apoptosis and decreased ingress of circulating monocytes into the joint are responsible for the initial reduction of macrophages following infliximab treatment in hTNF-Tg mice.

Objective Macrophages are critical in the pathogenesis of rheumatoid arthritis (RA). We recently demonstrated that FLIP (FLICE-like inhibitory protein) is necessary for the differentiation and/or survival of macrophages. We also identified that FLIP is highly expressed in RA synovial macrophages. This study was performed to determine if the reduction of Flip in macrophages would reduce synovial tissue macrophages and ameliorate serum transfer induced arthritis (STIA). Methods Mice with Flip deleted in myeloid cells (Flipf/fLysMc/+ mice) and littermate controls were employed. STIA was induced by intraperitoneal injection of K/BxN serum. Arthritis was evaluated by clinical score and change in thickness of the ankles. Joints were examined by histology and immunohistochemistry. Cells were isolated from the ankles and bone marrow and examined by flow cytometry, qRT-PCR or Western blot. Results In contrast to expectations, Flipf/fLysMc/+ mice developed more severe arthritis early in the clinical course, however the peak arthritis was attenuated and the resolution phase more complete. Prior to the induction of STIA, tissue resident macrophages were reduced. Further at day 9 post arthritis induction the number of F4/80hi macrophages in the joints of the Flipf/fLysMc/+ mice was not decreased, but increased. Flip was reduced in the F4/80hi macrophages in the ankles of the Flipf/fLysMc/+ mice, while F4/80hi population expressed an anti-inflammatory phenotype in both the Flipf/fLysMc/+ and control mice. Conclusions These observations suggest that reduction of FLIP in macrophages, by increasing the number of anti-inflammatory macrophages, may be an effective therapeutic approach to suppress inflammation, depending upon the disease stage.