Gelatinase B/matrix metalloproteinase-9 is a phase-specific effector molecule, independent from Fas, in experimental autoimmune encephalomyelitis.

Gelatinase B/matrix metalloproteinase-9 (MMP-9) triggers multiple sclerosis (MS) and the animal model of experimental autoimmune encephalomyelitis (EAE) by the breakdown of the blood-brain barrier. Interestingly, MMP-9 is beneficial in systemic autoimmunity caused by Fas-deficiency. Fas-deficient (faslpr) and Fas-ligand-deficient mice are protected against EAE. We here investigated the interaction between Fas and MMP-9 in the setting of induction of EAE and compared short- and long-term effects. We provoked EAE with myelin oligodendrocyte glycoprotein (MOG) peptide and compared EAE development in four genotypes (wild-type (WT), single knockout mmp-9-/-, faslpr, and mmp-9-/-/faslpr) and monitored leukocytes, cytokines and chemokines as immunological parameters. As expected, faslpr mice were resistant against EAE induction, whereas MMP-9 single knockout mice were not. In the double mmp-9-/-/ faslpr mice the effects on disease scores pointed to independent rather than interrelated disease mechanisms. On a short term, after EAE induction leukocytes infiltrated into the brain and cytokine and chemokine levels were significantly higher in all the four genotypes studied, even in the faslpr and mmp-9-/-/faslpr, which did not develop clinical disease. The levels of MMP-9 but not of MMP-2 were increased in the brain and in the peripheral organs after EAE induction. After 40 days all the animals recovered and did not show signs of EAE. However, the absence of MMP-9 in the remission phase suggested a protective role of MMP-9 in the late phase of the disease, because single mmp-9-/- mice presented a delayed remission in comparison with WT animals suggesting a phase-dependent role of MMP-9 in the disease. Nevertheless, the levels of some cytokines and chemokines remained higher than in control animals even 100 days after EAE induction, attesting to a prolonged state of immune activation. We thus yielded new insights and useful markers to monitor this activated immune status. Furthermore, MMP-9 but not MMP-2 levels remained increased in the brains and, to a higher extend, in the spleens of the WT mice even during the remission phase, which is in line with the role of MMP-9 as a useful marker and a protective factor for EAE in the remission phase.

Inhibition of gelatinase B/MMP-9 does not attenuate colitis in murine models of inflammatory bowel disease.

One third of patients with inflammatory bowel disease (IBD) inadequately respond to anti-TNF treatment and preclinical data suggest that matrix metalloproteinase-9 (MMP-9) is a novel therapeutic target. Here we show that IBD clinical and histopathological parameters found in wild type mice challenged with three different models of colitis, acute and chronic dextran sodium sulphate (DSS), and acute 2,4,6-trinitrobenzenesulfonic acid-induced colitis are not attenuated in MMP-9 knockout mice. We find similar colonic gene expression profiles in wild type and MMP-9 knockout mice in control and acute DSS conditions with the exception of eleven genes involved in antimicrobial response during colitis. Parameters of chronic colitis are similar in wild type and MMP-9 knockout mice. Pharmacological inhibition of MMP-9 with bio-active peptides does not improve DSS colitis. We suggest that MMP-9 upregulation is a consequence rather than a cause of intestinal inflammation and we question whether MMP-9 represents a disease target in IBD.

Insufficiently defined genetic background confounds phenotypes in transgenic studies as exemplified by malaria infection in Tlr9 knockout mice.

The use of genetically modified mice, i.e. transgenic as well as gene knockout (KO) and knock-in mice, has become an established tool to study gene function in many animal models for human diseases. However, a gene functions in a particular genomic context. This implies the importance of a well-defined homogenous genetic background for the analysis and interpretation of phenotypes associated with genetic mutations. By studying a Plasmodium chabaudi chabaudi AS (PcAS) malaria infection in mice bearing a TLR9 null mutation, we found an increased susceptibility to infection, i.e. higher parasitemia levels and increased mortality. However, this was not triggered by the deficient TLR9 gene itself. Instead, this disease phenotype was dependent on the heterogeneous genetic background of the mice, which appeared insufficiently defined as determined by single nucleotide polymorphism (SNP) analysis. Hence, it is of critical importance to study gene KO phenotypes on a homogenous genetic background identical to that of their wild type (WT) control counterparts. In particular, to avoid problems related to an insufficiently defined genetic background, we advocate that for each study involving genetically modified mice, at least a detailed description of the origin and genetic background of both the WT control and the altered strain of mice is essential.

Deficiency of gelatinase B/MMP-9 aggravates lpr-induced lymphoproliferation and lupus-like systemic autoimmune disease.

Gelatinase B/matrix metalloproteinase-9 (MMP-9) is a key enzyme involved in inflammatory, hematological, vascular and neoplastic diseases. In previous studies, we explored the intracellular substrate set or 'degradome' of MMP-9 and found many systemic autoantigens as novel intracellular gelatinase B substrates. Little is known, however, about the functional role of MMP-9 in the development of systemic autoimmunity in vivo. B6(lpr/lpr) mice with defective Fas-mediated apoptosis were used to investigate the functions of MMP-9 in lymphocyte proliferation and in the development of systemic autoimmunity. Combined Fas and gelatinase B deficiency resulted in extreme lymphoproliferative disease with enhanced lymphadenopathy and splenomegaly, and significantly reduced survival compared with single Fas deficiency. At the cellular level, this was corroborated by increased lymph node accumulation of 'double negative' T cells, B cells and myeloid cells. In addition, higher autoantibody titers and more pronounced autoimmune tissue injury were found in the absence of MMP-9, culminating in chronically enhanced systemic lupus erythematosus (SLE)-like autoimmunity. After cleavage by MMP-9 the SLE autoantigens U1snRNP A and ribosomal protein P0 were hardly recognized by plasma samples of both B6(lpr/lpr).MMP-9⁻/⁻ and B6(lpr/lpr).MMP-9+/+ mice, pointing to a destruction of B cell epitopes by MMP-9-mediated proteolysis. In addition, the same loss of immunodominant epitopes was observed with plasma samples from SLE patients, suggesting that MMP-9 suppresses systemic antibody-mediated autoimmunity by clearance of autoepitopes in immunogenic substrates. Thus, new protective functions for MMP-9 were revealed in the suppression of lymphoproliferation and dampening of systemic autoimmunity, cautioning against the long-term use of MMP inhibitors in autoimmune lymphoproliferative syndrome (ALPS) and SLE.