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Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by thrombosis and/or pregnancy complications. β2-glycoprotein I (β2GPI) complexed with phospholipid is recognized as a major target for autoantibodies in APS; however, less than half the patients with clinical manifestations of APS possess autoantibodies against the complexes. Therefore, the range of autoantigens involved in APS remains unclear. Recently, we found that human leukocyte antigen (HLA) class II molecules transport misfolded cellular proteins to the cell surface via association with their peptide-binding grooves. Furthermore, immunoglobulin G heavy chain/HLA class II complexes were specific targets for autoantibodies in rheumatoid arthritis. Here, we demonstrate that intact β2GPI, not peptide, forms a complex with HLA class II molecules. Strikingly, 100 (83.3%) of the 120 APS patients analyzed, including those whose antiphospholipid antibody titers were within normal range, possessed autoantibodies that recognize β2GPI/HLA class II complexes in the absence of phospholipids. In situ association between β2GPI and HLA class II was observed in placental tissues of APS patients but not in healthy controls. Furthermore, autoantibodies against β2GPI/HLA class II complexes mediated complement-dependent cytotoxicity against cells expressing the complexes. These data suggest that β2GPI/HLA class II complexes are a target in APS that might be involved in the pathogenesis.

Specific HLA class II alleles are strongly associated with susceptibility to rheumatoid arthritis (RA); however, how HLA class II regulates susceptibility to RA has remained unclear. Recently, we found a unique function of HLA class II molecules: their ability to aberrantly transport cellular misfolded proteins to the cell surface without processing to peptides. Rheumatoid factor (RF) is an autoantibody that binds to denatured IgG or Fc fragments of IgG and is detected in 70-80% of RA patients but also in patients with other diseases. Here, we report that intact IgG heavy chain (IgGH) is transported to the cell surface by HLA class II via association with the peptide-binding groove and that IgGH/HLA class II complexes are specifically recognized by autoantibodies in RF-positive sera from RA patients. In contrast, autoantibodies in RF-positive sera from non-RA individuals did not bind to IgGH/HLA class II complexes. Of note, a strong correlation between autoantibody binding to IgG complexed with certain HLA-DR alleles and the odds ratio for that allele's association with RA was observed (r = 0.81; P = 4.6 × 10(-5)). Our findings suggest that IgGH complexed with certain HLA class II alleles is a target for autoantibodies in RA, which might explain why these HLA class II alleles confer susceptibility to RA.

Nascent MHC class II molecules are associated with the invariant chain and are transported to the endolysosomal pathway, where MHC class II molecules acquire peptide antigens. On the other hand, misfolded endoplasmic reticulum (ER) proteins are generally degraded in the cells and are neither expressed on the cell surface nor secreted. Here, we found that MHC class II molecules associate with some misfolded ER proteins via the peptide-binding groove in competition with invariant chain. The misfolded proteins associated with MHC class II molecules are transported intact to the cell surface without processing to peptides. Furthermore, these complexes efficiently stimulate antigen-specific B cells. These findings reveal that MHC class II molecules function as a chaperone for the cell surface expression of misfolded ER proteins. In addition, we suggest that MHC class II molecules present not only peptides but also intact host-cell-derived proteins on the cell surface. These findings provide new insights into the function of MHC class II molecules.

Many patients with severe COVID-19 suffer from pneumonia and the elucidation of the mechanisms underlying the development of this severe condition is important. The in vivo function of the ORF8 protein secreted by SARS-CoV-2 is not well understood. Here, we analyzed the function of ORF8 protein by generating ORF8-knockout SARS-CoV-2 and found that the lung inflammation observed in wild-type SARS-CoV-2-infected hamsters was decreased in ORF8-knockout SARS-CoV-2-infected hamsters. Administration of recombinant ORF8 protein to hamsters also induced lymphocyte infiltration into the lungs. Similar pro-inflammatory cytokine production was observed in primary human monocytes treated with recombinant ORF8 protein. Furthermore, we demonstrated that the serum ORF8 protein levels are well-correlated with clinical markers of inflammation. These results demonstrated that the ORF8 protein is a SARS-CoV-2 viral cytokine involved in the immune dysregulation observed in COVID-19 patients, and that the ORF8 protein could be a novel therapeutic target in severe COVID-19 patients. [ABSTRACT FROM AUTHOR]

Author(s): Fumiji Saito; Kouyuki Hirayasu; Takeshi Satoh; Christian W. Wang; John Lusingu; Takao Arimori; Kyoko Shida; Nirianne Marie Q. Palacpac; Sawako Itagaki; Shiroh Iwanaga; Eizo Takashima; Takafumi Tsuboi; Masako Kohyama; [...]

Tissue macrophages comprise a heterogeneous group of cell types differing in location, surface markers and function (1). Red pulp macrophages are a distinct splenic subset involved in removing senescent red blood cells (2). Transcription factors such as PU.1 (also known as Sfpil) and C/EBPa (Cebpa) have general roles in myelomonocytic development (3,4), but the transcriptional basis for producing tissue macrophage subsets remains unknown. Here we show that Spi-C (encoded by Spic), a PU.1-related transcription factor, selectively controls the development of red pulp macrophages. Spi-C is highly expressed in red pulp macrophages, but not monocytes, dendritic cells or other tissue macrophages. [Spic.sup.-/-] mice have a cell-autonomous defect in the development of red pulp macrophages that is corrected by retroviral Spi-C expression in bone marrow cells, but have normal monocyte and other macrophage subsets. Red pulp macrophages highly express genes involved in capturing circulating haemoglobin and in iron regulation. [Spic.sup.-/-] mice show normal trapping of red blood cells in the spleen, but fail to phagocytose these red blood cells efficiently, and develop an iron overload localized selectively to splenic red pulp. Thus, Spi-C controls development of red pulp macrophages required for red blood cell recycling and iron homeostasis., Spi-C belongs to the Spi subfamily of Ets transcription factors, which includes PU.1 and Spi-B (5,6), and was initially reported to be expressed in B cells (5,7). We compared expression [...]

We analyzed the cytolytic activity of intra-epithelial T cells (IEL) isolated from the small intestines of 2- to 3-month-old mutant mice rendered deficient in different gene(s) in which the number of IEL expressing either T cell receptor (TCR)-[Alpha][Beta] ([Alpha][Beta]-IEL) or TCR-[Gamma][Delta] ([Gamma][Delta]-IEL) were absent or markedly diminished. When compared with wild-type littermates, cytolytic activity of [Gamma][Delta]-IEL was sharply attenuated in TCR-[Beta] mutant mice but remained unaltered in TCR-[Alpha] mutant mice in which a minor population of dull TCR-[[Beta].sup.+] ([[Beta].sup.dim])-IEL was also present. Cytolytic activity of [Gamma][Delta]-IEL was maintained in mice doubly homozygous for [[Beta].sub.2]-microglobulin and transporter associated with antigen processing 1 gene mutations in which a conspicuous decrease was noted in absolute numbers of [Alpha][Beta]-IEL. In contrast, both TCR-[Delta] and IL-7 receptor-[Alpha] gene mutations that lead to lack of [Gamma][Delta]-IEL generation did not affect the development or cytolytic activity of the remaining [Alpha][Beta]-IEL. The anti-CD3 and anti-TCR-[Gamma][Delta] mAb-induced IFN-[Gamma] production of [Gamma][Delta]-IEL showed the same TCR-[Alpha] and TCR-[Beta] mutation-dependent variability. These results indicate that cytolytic and IFN-[Gamma]-producing activities of [Gamma][Delta] T cells in mouse intestinal epithelium are TCR-[Beta]-chain-dependent.

Objective: Specific HLA class II alleles are associated with susceptibility to systemic lupus erythematosus (SLE). The role of HLA class II molecules in SLE pathogenesis remains unclear, although anti‐DNA antibodies are specific to SLE and correlate with disease activity. We previously demonstrated that misfolded proteins bound to HLA class II molecules are specific targets for the autoantibodies produced in autoimmune diseases. This study was undertaken to validate our hypothesis that DNA binds to HLA class II molecules in a manner similar to that of misfolded proteins and that DNA bound to HLA class II molecules is involved in SLE pathogenesis. Methods: We analyzed the binding of DNA to HLA class II molecules, as well as the response of cells expressing anti‐DNA B cell receptors (BCRs) to cells expressing the DNA/HLA class II complex. Results: Efficient binding of DNA to HLA class II molecules was observed in risk alleles of SLE, such as HLA–DRB1*15:01. The efficiency of DNA binding to each HLA–DR allele was positively associated with the risk of SLE conferred by the HLA–DR allele. In addition, reporter cells carrying anti‐DNA BCRs were activated by cells expressing DNA/HLA class II complexes. Conclusion: These results provide evidence that DNA bound to HLA class II molecules is involved in SLE pathogenesis. [ABSTRACT FROM AUTHOR]