Multiple sclerosis
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2007 – 2009 Dr Hadwen Trust Research Technician: Functional analysis of the T cell immune response in multiple sclerosis by gene silencingDr Daniel Altmann and Dr Sabine Steinbach |
Professor Daniel M Altmann is head of the Human Disease Immunogenetics Group at the Hammersmith Hospital Campus of Imperial College London. His key research interests are the immunology of autoimmune disease and of bacterial infection. He is also Director of Research Strategy in the Department of Medicine.
The research technician supported by the Dr Hadwen Trust grant to work on this project was Dr Sabine Steinbach (pictured above).
Multiple sclerosis (MS) is the most common disease of central nervous system demyelination, and is associated with extreme erosion in quality of life and reduced life-expectancy; there is an urgent need for new treatments.
The disease involves stripping away of the myelin sheath that insulates the nerves, in time leading to death of those nerves and irreversible disability. The damage is caused by inflammatory attack by T lymphocytes. Since it is clear that healthy individuals also harbour autoimmune, anti-myelin T cells, much research has focused on elucidating those conditions that are associated with the breakage of functional ‘self-tolerance’ and resulting disease. Many aspects of immune control come into this, a key one being the expression of co-stimulatory molecules on antigen presenting cells.
The hypothesis is that differential expression of these co-stimulatory molecules can shift the immune response either towards or away from, autoimmunity. These co-stimulatory molecules, many of which belong to a family termed B7 have generally been studied by inducing disease in knockout mice. However, this Dr Hadwen Trust-funded study established an in vitro model system for investigating T-cells responses in multiple sclerosis, as an alternative to studies in knockout mice.
In a multi-system disease such as MS, much research emphasis has been placed on animal ‘models’ of the human illness. There have been more than 10,000 publications on the induction of the surrogate condition, experimental allergic encephalomyelitis (EAE), in rodents, guinea pigs, rabbits and monkeys. Animals suffer inflammation and damage to the nervous system that may result in paralysis, in experiments that can cause distress and suffering. However, it is becoming increasingly possible to generate functional data about the immunology of MS by studying patients’ T-cell responses.
This project started by selecting the optimal serum-free cell culture conditions (StemXVivo-based media) for the generation of monocyte-derived dendritic cells which could be used as antigen presenting cells.
Next, the first comprehensive comparison of co-stimulatory molecule expression by dendritic cells of MS patients relative to healthy control volunteers was conducted. This enabled to identification of suitable targets RNA interference technology and as a result B7-H1 was chosen as a candidate for knock down experiments.
To investigate the influence of B7-H1 on T-cell activation in vitro, methods for using small interfering RNA (siRNA) to knock down the expression of B7-H1 in immature dendritic cells were developed. B7-H1 silenced cells were then tested for their ability to induce T-cell responses in vitro. These assays showed that B7-H1 silencing increased proliferation of CD4+ T-cells from multiple sclerosis patients and healthy volunteers in vitro. Further experiments will be needed to verify these findings, but these results define B7-H1 as an inhibitory co-stimulatory molecule.
The finding that B7-H1 inhibits T-cell proliferation in MS patients is of medical significance. Drugs which could up-regulate the expression of B7-H1could inhibit auto-immune reactions. Another potential therapy might be to engineer in vitro B7-H1 silenced monocytes from patients using the siRNA approach developed by this project.
This project has demonstrated that, whereas the first-line method of choice for analysis of immune system co-stimulatory molecules has been the generation of a knockout mouse strain, it is feasible to conduct direct analysis in human cells. A further advantage is the opportunity of working directly with the cells from patients with the human disease and not at the distance of a partially related mouse ‘model’.
The established in vitro model using siRNA technology could be used in the future to study the effect of different co-stimulatory molecules on different cell populations from patients with different diseases. Such information would help in the development of new drugs. The established techniques of serum-free generation of MoDC and silencing co-stimulatory molecules from iMoDC could be used in the future to change monocytes of patients ex vivo into powerful MoDC for immunotherapy.