Oxidative regulation of NCF1 in B cells and mouse models of arthritis

Abstract

Autoimmune diseases (ADs) are chronic pathologies that result from a dysregulation of autoreactive B and T cells leading to autoantibody production and ultimately destruction of self.

In paper I, we studied inflammation associated with autoimmunity. The glycolipid alpha- galactosylceramide (αGalCer) has been used as an adjuvant for vaccines, cancer treatment, and autoimmunity regulation, and shown to be effective. We show that when combined with a sterile mouse model of inflammation by injecting the pro-inflammatory cytokine IL-18, invariant natural killer T cells (iNKT) promote the expansion of autoreactive B cells instead of regulating them. This study offers insight into the usage of αGalCer in the context of chronic inflammation and implications in autoimmunity.

Identifying genetic polymorphisms in autoimmune diseases is crucial for understanding how the immune system operates to find innovative therapies to treat human diseases. Genetic mapping of quantitative trait loci (QTL) and differentially expressed genes (DEGs) are helpful tools, however, the identification of single nucleotide polymorphisms (SNPs) is a difficult task. One of the major SNP associated with autoimmune arthritis was identified in rat models, using pristane-induced arthritis (PIA). An amino acid replacement in the NCF1 protein at position 153 from threonine to methionine (T153M) decreased the oxidative burst capacity of the cells, which led to increased arthritis severity. The NOX2 complex and its subunits are expressed in many cell types, predominantly in phagocytes (neutrophils and macrophages) but also in antigen-presenting cells (APCs). This complex is essential to produce reactive oxygen species (ROS). Several studies have pinpointed an association of SNPs in different subunits of the NOX2 complex with different autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), confirming the findings in animal models. The fact that a lower ROS response was associated with arthritis severity contradicts the prevailing dogma that ROS are detrimental, making it important to explain how ROS can shape and protect against autoimmunity.

In paper II, we found that a point mutation in the NCF4 subunit of the NOX2 complex, has an effect in murine models of arthritis, shaping B cell responses and consequently plasma cells that are attracted to the sites of inflammation such as the synovium, secreting pathogenic antibodies that contribute to disease progression.

In papers III and IV, we focused on the role of NCF1 in B cells and germinal center B cells (GC-B), respectively. Using conditional mice flipping Ncf1 alleles, we show that ROS regulates the activation and differentiation of B cells, with profound effects on T cells. Generally, mice expressing the low ROS allele (NCF1153M) exclusively in B or GC-B cells have reduced ROS levels and increased arthritis severity as compared to littermate controls harboring the high ROS allele (NCF1T153). In parallel, sera levels of anti-collagen type II (COL2) antibodies were increased in the NCF1153M mice. T cell responses were affected in both models, with increased pro-inflammatory cytokines release, immune shaping of regulatory T cells, and T follicular helper (Tfh) cells. These findings provide evidence on the pleiotropic activity of NCF1-restricted ROS and present a previously undisclosed role of NCF1 in regulating B and GC-B cells in autoimmune diseases (ADs).

It is well known that B or T cells that react against self-antigens are deleted or inactivated by central tolerance mechanisms in the primary lymphoid organs bone marrow (BM) or thymus, however, some autoreactive clones can escape to the periphery and instigate ADs. In paper V, we studied antigen-specific B cells that opposite to what has been postulated are positively selected in the BM and found in the periphery of mice, rats, and even humans. These cells instead of causing autoimmunity, prevent it. COL2-transgenic mouse models do not develop spontaneous arthritis but are instead protected, mainly by the induction of regulatory T cells.