Oligodeoxynucleotide IMT504: Effects on Central Nervous System Repair Following Demyelination.

Multiple sclerosis (MS) is an immune-mediated central nervous system (CNS) disease characterized by demyelination resulting from oligodendrocyte loss and inflammation. Cuprizone (CPZ) administration experimentally replicates MS pattern-III lesions, generating an inflammatory response through microgliosis and astrogliosis. Potentially remyelinating agents include oligodeoxynucleotides (ODN) with a specific immunomodulatory sequence consisting of the active motif PyNTTTTGT. In this work, the remyelinating effects of ODN IMT504 were evaluated through immunohistochemistry and qPCR analyses in a rat CPZ-induced demyelination model. Subcutaneous IMT504 administration exacerbated the pro-inflammatory response to demyelination and accelerated the transition to an anti-inflammatory state. IMT504 reduced microgliosis in general and the number of phagocytic microglia in particular and expanded the population of oligodendroglial progenitor cells (OPCs), later reflected in an increase in mature oligodendrocytes. The intracranial injection of IMT504 and intravenous inoculation of IMT504-treated B lymphocytes rendered comparable results. Altogether, these findings unveil potentially beneficial properties of IMT504 in the regulation of neuroinflammation and oligodendrogenesis, which may aid the development of therapies for demyelinating diseases such as MS.

TGF-ß pro-oligodendrogenic effects on adult SVZ progenitor cultures and its interaction with the Notch signaling pathway.

Adult neural progenitor cells (NPCs) are capable of differentiating into neurons, astrocytes, and oligodendrocytes throughout life. Notch and transforming growth factor ß1 (TGF-ß) signaling pathways play critical roles in controlling these cell fate decisions. TGF-ß has been previously shown to exert pro-neurogenic effects on hippocampal and subventricular zone (SVZ) NPCs in vitro and to interact with Notch in different cellular types. Therefore, the aim of our work was to study the effect of TGF-ß on adult rat brain SVZ NPC glial commitment and its interaction with Notch signaling. Initial cell characterization revealed a large proportion of Olig2+, Nestin+, and glial fibrillary acidic protein (GFAP+) cells, a low percentage of platelet-derived growth factor receptor α (PDGFRα+) or NG2+ cells, and <1% Tuj1+ cells. Immunocytochemical analyses showed a significant increase in the percentage of PDGFRα+, NG2+, and GFAP+ cells upon four-day TGF-ß treatment, which demonstrates the pro-gliogenic effect of this growth factor on adult brain SVZ NPCs. Real-time polymerase chain reaction analyses showed that TGF-ß induced the expression of Notch ligand Jagged1 and downstream gene Hes1. Notch signaling inhibition in cultures treated with TGF-ß produced a decrease in the proportion of PDGFRα+ cells, while TGF-ß receptor II (TßRII) inhibition also rendered a decrease in the proportion of PDGFRα+ cells, concomitantly with a decrease in Jagged1 levels. These findings demonstrate the participation of Notch signaling in TGF-ß effects and illustrate the impact of TGF-ß on glial cell fate decisions of adult brain SVZ NPCs, as well as on oligodendroglial progenitor cell proliferation and maturation.

The peptide specificities of the autoantibodies elicited by mouse hepatitis virus A59.

Synthetic decapeptides (N=206) covering the entire sequence of mouse liver fumarylacetoacetate hydrolase (FAH) were used to analyze the specificities of the autoantibodies (autoAb) elicited towards this enzyme in mice infected with mouse hepatitis virus (MHV). These autoAb bound mainly to N- and C-terminal FAH peptides, the most reactive sequences being 1-50 and 390-420, respectively. Surprisingly, although FAH sequence 1-50 shares a high degree of homology with various MHV proteins, the C-terminal portion does not. Moreover, whereas the autoAb reacted with homologous peptides surrounding residues 70, 160 and 360, non-similar sequences around residues 130, 210, 240, 250, and 300 were also recognized, indicating that autoAb were not restricted to epitopes with sequence homologies. There was also a lack of correlation between the amount of anti-MHV or anti-FAH antibodies produced and the reactivity towards the peptides. Moreover, the spectrum of peptides recognized by the autoAb of a given mouse did not change significantly with time, which suggests that the MHV-elicited autoimmune response does not induce an epitope recognition spreading. Finally, anti-FAH Ab produced after immunization with rat liver FAH recognized essentially the same mouse FAH regions than autoAb from MHV-infected mice. Results indicated that the induction of the autoAb is not only related to molecular or structural mimicry, but rather supports the Danger model, in which any aggression, in this case the MHV infection, is susceptible to trigger the production of autoAb.

Sequence similarity and structural homologies are involved in the autoimmune response elicited by mouse hepatitis virus A59.

The features of autoantibodies (autoAb) to liver fumarylacetoacetate hydrolase (FAH) elicited in mice infected with mouse hepatitis virus (MHV) were studied by ELISA and western-blot competition assays. All sera tested contained Ab to cryptic FAH epitopes according with results from western-blot tests, whereas ELISA data indicated that some of these same sera did recognize native epitopes of the autoantigen (autoAg). Such differences were detected in individual sera from various mouse strains, and were ascribed to the fact that proteins insolubilized on solid supports expose a variety of conformational and cryptic antigenic determinants. On the other hand, whereas results from both experimental protocols showed that anti-MHV Ab did not cross-react with the soluble autoAg, the opposite situation did not show analogous results. Thus, binding of autoAb to insolubilized FAH could be inhibited by MHV depending on the mouse serum or the experimental protocol used. Additionally, a set of synthetic homologous peptides from mouse FAH and various viral proteins was employed to analyze the Ab repertoire of MHV-infected mice. Results indicated that two homologous peptides were recognized by most Ab: the N-terminal sequences (1-10) from FAH and the nucleocapside, both sharing 50% of identity, and sequence 2317-2326 of the RNA polymerase, a peptide showing 30% of identity with FAH 11-20. Results indicated that MHV-infection triggers at least three distinct Ab populations: anti-MHV, anti-FAH and cross-reacting Ab. This cross-reaction implies either sequential or conformational epitopes from both the viral proteins and the autoAg and may differ between individuals.

Detection of mouse hepatitis virus infection by assay of anti-liver autoantibodies.

The observation that mice infected with mouse hepatitis virus (MHV) develop autoantibodies directed mainly to liver fumarylacetoacetate hydrolase (FAH) enabled the development of an ELISA applicable to the detection of MHV-infection. The method, based on the titration of antibodies to semipurified FAH from rat liver, is easy, economical, and does not require the isolation of viral proteins from large MHV stocks. Furthermore, since sera from mice immunized with a purified fraction of the rat liver enzyme do react with its homologous protein, this antiserum can be used as a positive control avoiding the manipulation of samples from MHV-infected animals.