Multiple sclerosis (MS) is a disease that affects over 2.5 million people worldwide. It is typically first diagnosed in patients in their 20s and 30s and is most common among women (Trapp and Nave, 2008). MS is characterized by areas in which myelin, a collection of lipids and proteins that forms a protective and insulating layer around portions of neurons, is destroyed (also known as demyelination). This damage occurs most often in the brain and spinal cord, impairing the transmission of signals among nerve fibers. MS is considered to be an autoimmune disease because a person’s immune system attacks self tissues, in this case myelin and myelin-producing cells. A large amount of this damage it is thought to stem from the activation of autoreactive CD4+ T cells that recognize peptides from the myelin sheath. (In healthy patients, these CD4+ T cells - also known as “helper” T cells - are not autoreactive and help B cells produce antibody.) As the disease course of MS progresses, symptoms increase in severity and include dizziness, impaired thinking, tremors, and difficulty walking.
Tumor Necrosis Factor (TNF) is a cytokine (a subset of small cell-signalling proteins) that has been linked to MS and is expressed by many cell types (Grivennikov et al., 2005). This protein can trigger either cell death or cell survival and can it also mediate inflammation. Inflammation is a local response at a site of infection caused by the influx of leukocytes, which are cells that attempt to combat infection; infiltration of inflammatory cells is common in the brains and spinal cords of MS patients. High levels of TNF have also been observed in MS and other autoimmune disorders (Hohnoki et al., 1998); blockade of TNF is actually an approved treatment for some of these diseases, including rheumatoid arthritis (Feldmann and Maini, 2001). Nevertheless, the role that TNF plays in MS is still controversial.
A model called Experimental Autoimmune Encephalomyelitis (EAE) can be used among mice to study multiple sclerosis. EAE is induced by injecting mice with proteins that comprise myelin. In a recent study, published in the Journal of Immunology, Kruglov and colleagues (et al., 2011) used an EAE model to assess the roles of different cellular sources of TNF during EAE. The mice were scored daily on the basis of their clinical symptoms on a scale ranging from 0 to 6, where 0 = no disease and 6 = moribund.
The researchers used both TNF wild-type (WT) mice that expressed normal amounts of TNF and TNF knockout (KO) mice that had been genetically engineered so that they could not express TNF. The TNF deficient mice had a delayed disease onset due to a lower accumulation of inflammatory cells in their spinal cords, as revealed by staining these cells. After the onset of the disease, TNF deficient mice also had significantly more autoreactive CD4+T cells in their spleens than normal mice. These observations were made using a technique called flow cytometry, which is useful for categorizing cells. Therefore, TNF appears to play a dual role. TNF contributes to the initial infiltration of inflammatory cells in the central nervous system, but it also protects against EAE by curtailing the expansion of autoreactive T cell responses.
The researchers then wanted to know if TNF produced by T cells and myeloid cells (a subset of immune cells) was important for the development of EAE. Mice which had been genetically engineered to not produce TNF from their T cells or myeloid cells (MT-TNF KO mice) had a disease course and cell infiltration pattern very similar to that of the TNF KO mice. This indicates that myeloid cells and T cells are the main source of TNF during EAE.
To further distinguish between the effects of TNF from T cells or myeloid cells, the researchers genetically engineered mice so that either their T cells did not express TNF (T-TNF KO mice) or their myeloid cells did not express TNF (M-TNF KO mice). Disease symptoms were less severe in T-TNF KO mice than in normal mice and these T-TNK KO mice had less demyelination in the spinal cord, as revealed by Luxol staining. Surprisingly, T-TNF KO mice had higher numbers of total inflammatory infiltrating cells after the onset of the disease. Among these cells were increased levels of CD4+ T cell numbers (that were not all autoreactive) but decreased numbers of macrophages when compared to normal mice. Macrophages are cells in the immune system that can present peptides to CD4+ T cells, including myelin peptides. Additionally, the T-TNF KO mice had greater numbers of autoreactive Th1 and Th17 cells (which are subsets of CD4+ T cells) in their spleens after disease onset. Thus, TNF produced by T cells can be both beneficial and harmful. TNF can promote disease by causing accumulation of macrophages in the central nervous system, but it can also limit disease by restricting the development of autoreactive T cells.
Mice in which TNF production was eliminated from myeloid cells (M-TNF KO mice) had a later onset of disease when compared to normal mice. They also had lower numbers of infiltrating CD4+ cells shortly after this onset occurred but greater numbers of autoreactive Th1 and Th17 cells in their spleens. These findings show that TNF expressed by myeloid cells speeds up the onset of EAE but at the same time limits the development of autoreactive T cells.
The mechanism responsible for the increased numbers of autoreactive Th1 and Th17 CD4+T cells in the spleens of the T-TNF-KO and M-TNF KO mice was unclear. TNF could affect the production of other cytokines by cells such as macrophages. Cytokines affect the polarization of T cells and determine which subset of Th cells (such as Th1 or Th17) they differentiate into. IL-12 promotes the polarization of T cells into Th1 cells whereas IL-6 promotes the polarization of T cells into Th17 cells. Cells from mouse spleens were cultured shortly after the onset of the disease, as well as after clinical symptoms had reached a plateau, and cytokine levels were assessed. Significantly increased levels of IL-12 and IL-6 levels were seen among the cells from the T-TNF KO and M-TNF KO mice when compared to normal mice. Therefore, TNF produced by both T cells and myeloid cells regulates development of autoreactive T cells during EAE, though a mechanism involving the regulation of IL-12 and IL-6 production.
These findings reveal that TNF has both disease-causing and protective functions in an EAE model of multiple sclerosis. Additionally, TNF production by either T cells or myeloid cells can differently affect the course of this disease. Nevertheless, this elegant study could be refined further. The majority of human MS patients exhibit a relapsing and remitting disease course, but this model did exhibit any remissions due to the type of peptide chosen to induce EAE (Skundric 2005). Therefore, for future studies the authors could choose a different peptide for EAE induction that would more closely mimic human MS. An interesting future direction for these researchers would also be to investigate the mechanisms that control the production of TNF by T cells as well as myeloid cells.
This study is of significance because it helps individuals to understand the mechanisms underlying MS disease progression. It also explains why a non-specific TNF blockade in MS patients is not advisable and actually resulted in the exacerbation of disease in a 1999 clinical trial involving human patients. Research of this nature will help to expand our knowledge of the mechanisms underlying multiple sclerosis and allow scientists to develop effective therapeutic targets aimed to improve the lives of millions of people.
Kruglov, A. A., Lampropoulou, V., Fillatreau, S., & Nedospasov, S. A. (2011). Pathogenic and protective functions of TNF in neuroinflammation are defined by its expression in T lymphocytes and myeloid cells. Journal of Immunology (Baltimore, Md.: 1950), 187(11), 5660-5670. doi:10.4049/jimmunol.1100663
Trapp, B. D., & Nave, K. A. (2008). Multiple sclerosis: An immune or neurodegenerative disorder? Annual Review of Neuroscience, 31, 247-269. doi:10.1146/annurev.neuro.30.051606.094313
Hohnoki, K., Inoue, A., & Koh, C. S. (1998). Elevated serum levels of IFN-gamma, IL-4 and TNF-alpha/unelevated serum levels of IL-10 in patients with demyelinating diseases during the acute stage. Journal of Neuroimmunology, 87(1-2), 27-32.
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