Establishing the novel ex-vivo focal demyelination model of Multiple Sclerosis in other laboratories to reduce and replace live animal use.

Why did we fund this project?

This award aims to replace the use of mice in multiple sclerosis (MS) research using slices of brain or spinal cord tissue in a focal lesion ex vivo model.

MS is modelled in rodents by immunising against myelin, causing the immune system to target the myelin that insulates axons. This induces experimental autoimmune encephalomyelitis (EAE), which has similar myelin damage to that seen in patients with MS. EAE results in weakness, fore- and hindlimb paralysis and incontinence, although the degree of severity varies between animals. The studies are associated with a high level of suffering and the need for specialist husbandry and care. Ex vivo slice culture methods have previously been developed but these induce demyelination across the full tissue. Dr Ben Newland has developed a method for inducing focal lesions in ex vivo cultured slices, which then include both healthy and diseased tissue similar to the MS disease state in humans. Up to six brain slices and ten spinal cord slices can be isolated from each animal and demyelinating agent administered to the tissue using a cryogel scaffold. Ben and colleagues have previously validated the ex vivo cryogel method by comparing against post-mortem tissue from MS patients.

Ben will transfer this ex vivo method to the laboratories of Dr Yvonne Dombrowski and Professor Denise Fitzgerald at Queen’s University Belfast. He will also create detailed protocols and training videos to enable other researchers to take up the method. The cryogel scaffolds can be synthesised with a micro-stereolithography based 3D printer and will be made open source to enable easier manufacturing. Ben’s laboratory will also supply cryogels to research groups unable to produce the scaffolds themselves.

Until recently the only means of mimicking the patchy and dynamic nature of demyelination in multiple sclerosis (MS) was by using animal models such as the EAE model, or stereotactic injections of a myelin toxin into the brain or spinal cord, resulting in focal lesions. However, the labs of Williams and Newland recently created a focal demyelination model using brain and spinal cord slices in culture (Eigel et al., Acta Biomaterialia, 2019, 97, 216) which replicates both the patchy nature (areas of unaffected tissue directly adjacent to demyelinated tissue) and the dynamic process of spontaneous remyelination. Cryogel scaffolds, which are extremely soft (causing no tissue damage) yet robust (easy to pick up with tweezers etc) are used to deliver the demyelinating agent lysophosphatidylcholine to brain and spinal cord slice culture to cause these focal lesions.

This project seeks to transfer the skills and knowledge of this model firstly to the Dombrowski lab and the Fitzgerald lab, then to other groups around the world to replace live animal models and reduce the total number of animals required in this field of research.

A technician will learn the techniques in the Williams lab, create detailed protocols, record training videos and set up the model in the Dombrowski and Fitzgerald groups. In addition, standardisation of templates with a micro-stereolithography based 3D printer, from which the cryogel scaffolds can be reproducibly manufactured, will allow the Newland lab to roll out designs to other groups yet allow customisation for specific shape requirements

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Skills and Knowledge Transfer grant

Status

Active

Institution

Cardiff University

Grant reference number

NC/W000989/1

Award date

Sep 2021 - Aug 2023

Grant amount

£74,526