Depending on the method of immunization, a single administration of CFA may result in the development of a local inflammatory process or chronic polyadjuvant-induced arthritis (AA). We administered naked DNA vaccines encoding MIP-1α, MCP-1, MIP-1β, and RANTES to Lewis rats and confirmed that each of these vaccines induced immunological memory to the corresponding gene product. Upon induction of disease, this memory effectively inhibited the development of the autoimmune condition. Self-specific Ab's developed in DNA-vaccinated animals were neutralizing in vitro and could adoptively transfer the beneficial effect of each vaccine. Repeated administration of the constructs encoding MCP-1, MIP-1α, or RANTES inhibited the development and progression of AA, even when each vaccine was administered only after the onset of disease. This suggests a highly effective way by which the immune system could be re-educated to generate protective immunity against its own harmful activities.

Cathepsins are involved in a variety of physiological processes including antigen processing and presentation and extracellular matrix degradation. In the present study, we evaluated whether expression levels of cathepsins S and B and their inhibitors cystatins B and C are affected by multiple sclerosis (MS) disease state (relapse and remission) and therapies (interferon-β[IFN-β] and the glucocorticoid [GC] methylprednisolone), and whether they are associated with the IFN-β response phenotype. Real-time PCR was employed to compare RNA expression levels in peripheral blood leucocytes (PBLs) and ELISA to determine serum protein levels of MS patients and matched healthy individuals. Cathepsin S RNA was higher in MS patients in the relapse state compared to controls (by 74%, P= 3 × 10−5, n= 30 versus n= 18) with a similar increase observed in serum (66%, P= 0.002, n= 18 versus n= 20). GC treatment reduced cathepsin S levels in PBL RNA (by 44%, P= 6 × 10−6, n= 27) and serum proteins (by 27%, P= 1 × 10−5, n= 26), reduced the serum protein levels of pro-cathepsin B (by 8%, P= 0.0007, n= 23), and in parallel increased the serum levels of their inhibitor cystatin C (by 82%, P= 8 × 10−6, n= 26). IFN-β therapy significantly elevated the RNA levels (n= 16) of cathepsin B (by 16%, P= 0.03), cystatin B (44%, P= 0.004) and cystatin C (48%, P= 0.011). In the serum, only cathepsin S levels were reduced by IFN-β (16%, P= 0.006, n= 25). Interestingly, pre-treatment serum cathepsin S/cystatin C ratio was higher in ‘good responders’ to IFN-β therapy compared to patients without a good response (by 94%, P= 0.003). These results suggest that cathepsin S and cystatin C may contribute to disease activity in MS, specifically in a subgroup of patients that are responsive to IFN-β therapy, and that these proteins should be further evaluated as biomarkers in MS.

DNA vaccination represents a novel means of expressing Ag in vivo for the generation of both humoral and cellular immune responses. The current study uses this technology to elicit protective immunity against experimental autoimmune encephalomyelitis (EAE), a T cell-mediated autoimmune disease of the central nervous system that serves as an experimental model for multiple sclerosis. RT-PCR verified by Southern blotting and sequencing of PCR products of four different C-C chemokines, macrophage-inflammatory protein-1α (MIP-1α), monocyte-chemotactic protein-1 (MCP-1), MIP-1β, and RANTES, were performed on brain samples from EAE rats to evaluate mRNA transcription at different stages of disease. Each PCR product was then used as a construct for naked DNA vaccination. The subsequent in vivo immune response to MIP-1α or MCP-1 DNA vaccines prevented EAE, even if disease was induced 2 mo after administration of naked DNA vaccines. In contrast, administration of the MIP-1β naked DNA significantly aggravated the disease. Generation of in vivo immune response to RANTES naked DNA had no notable effect on EAE. MIP-1α, MCP-1, and MIP-1β mRNA transcription in EAE brains peaked at the onset of disease and declined during its remission, whereas RANTES transcription increased in EAE brains only following recovery. Immunization of CFA without the encephalitogenic epitope did not elicit the anti-C-C chemokine regulatory response in DNA-vaccinated rats. Thus, modulation of EAE with C-C chemokine DNA vaccines is dependent on targeting chemokines that are highly transcribed at the site of inflammation at the onset of disease.