What is the proposed research strategy for the Sheffield Institute:
Current major research programmes and future direction

 

A major programme of research, integrating clinical and laboratory studies, aimed at identifying molecular mechanisms of neurodegeneration in MND was
established in Newcastle upon Tyne and transferred to the University of Sheffield in 2000. In Sheffield, supported by programme funding from the Wellcome
Trust and the MND Association, we have established a major national Care and Research Centre for Motor Neurone Disorders.
The Sheffield MND research team is in a unique position to make a major contribution tothe search for the causes and curative treatments for MND. Research resources underpinning the programme include:

• An extensive clinical database of 900 MND patients with high quality clinical details of all patients.

• The largest resource of human brain-bank material world- wide (CNS material donated by 140 MND patients), as well as DNA and cerebrospinal fluid banks.

• Robust cellular and other in vivoexperimental models of MND in which new treatment approaches can be verified.

• The research team assembled has multidisciplinary skills in neurology, pathology, gene therapy, molecular genetics, protein chemistry, cell biology and pharmacology to facilitate identification of cellular targets involved in motor neurone degeneration.

• MND is an excellent model for evaluating all forms of neuroprotective treatment which is why researchers at the Institute will work on finding treatments for Alzheimer, Parkinson’s and SMA which is a degenerative condition found in very young children. There will be several research programmes among which ‘Genetic susceptibility and disease modifying factors, Gene therapy, Brain reserve and stem-cell regeneration as factors determining cognitive state in Alzheimer patients’

Laboratory programmes

1 Using cellular models of motor neurone degeneration to define molecular mechanisms of neurodegeneration and to identify new therapeutic targets

We have established a robust cellular model to examine the molecular pathophysiology of motor neurone
injury in the first instance in a defined subtype of MND caused by mutations in the SOD1 gene. We have used a motor neuronal cell line NSC34 cells which are a hybrid mouse motor neurone/neuroblastoma cell line, which retain the ability to proliferate whilst exhibiting many motor neuronal characteristics. NSC34 cells have been transfected with 1 of several mutant forms of human SOD1(G93A, G37R, I113T), wild type SOD1 or vector only and single cell clones derived by limiting dilution. This cell line provides a good cellular model to investigate the effects of mutant SOD1 specifically in cells with motor neurone characteristics. Using this cellular model, we have demonstrated several important insights into the cell specific toxicity produced by the mutant SOD1 protein including:

1 Biochemical changes reflecting an increased tendency to programmed cell death or apoptosis, with increased expression of cleaved caspase 9 and annexin V staining on the cell surface under basal culture conditions ;

2 Increased cell death by apoptosis, with activation of caspases 9 and 3 when the cells are oxidatively stressed;

3 Altered intracellular handling and extracellular release of superoxide and nitric oxide (NO) free radical species;

4 Increased susceptibility to toxicity from exogenous nitric oxide;

5 Altered regulation in the expression at both gene and protein levels, of components of the motor neurone internal skeleton, neurofilament light and medium;

6 The development of abnormal swollen mitochondria (which are the energy generators
within cells), with impaired activity of complexes II and IV of the respiratory chain and impaired generation of energy in the form of ATP within the cells. The importance of these changes in the biochemical cascade of cellular injury produced by mutant SOD1 is reinforced by studies of apoptotic pathways, mitochondrial function and expression
of cytoskeletal components in the mouse SOD1 transgenic model and in human CNS tissue.