Title: Optimisation of a flow-based human in vitro blood-brain barrier model to replace animal models for studying immunopathogenesis of viral brain infections
Funder: National Centre for the Replacement, Refinement and Reduction of Animals in Research

Abstract: Viral encephalitis is one of the most important types of neurological infection, because of the devastating impact on the quality of life of those affected, disproportionately large burden on the healthcare systems and community with large health economic and social costs. Encephalitis such as Japanese encephalitis (JE) is a massive burden on many developing countries. In the UK, encephalitis caused by herpes simplex virus and human immunodeficiency virus is a major problem. The blood-brain barrier (BBB) is the protective physiological barrier that prevents pathogens such as viruses entering the central nervous system (CNS). Studies have shown that BBB damage due to inflammation is a final common pathway in many brain infections including viral encephalitis, allowing pathogens and leukocyte entry into the brain. Currently studies that investigate the immunopathogenesis of viral brain infections are very dependent on animal models because of a lack of good in vitro human models. The most commonly used animal models for encephalitis are mouse and rat models. In mice, signs of infection include leg paralysis, a hunched back, ruffled fur, minimal activity and seizures. A human cell culture model of the BBB can give physiologically relevant data and will lead to a reduction of animals used for this kind of research.During my postdoctoral research over the last two years, I have developed a reliable two-dimensional cell culture model of the human BBB using immortalised human brain endothelial cells (iHBEC) and primary human astrocytes. I am investigating the mechanisms of Japanese encephalitis virus (JEV) entry through the BBB. However, there are certain limitations in this model; achieving a tight monolayer is difficult (compared with animal models) and the blood flow in vivo (important for leukocyte transmigration) cannot be simulated. I would now like to take this work forward with a David Sainsbury Fellowship by developing a multi-culture three-dimensional flow-based human BBB model for studying viral encephalitis in collaboration with researchers from the Cleveland Clinic Lerner Research Institute, USA. The model will mimic the blood flow in the brain capillaries. Primary HBEC or iHBEC will be seeded on the blood side and CNS cells such as astrocytes (main inducers of BBB phenotype on endothelial cells) and microglia (release cytokines/chemokines in response to CNS infection) in the brain side. Cell culture medium or serum from patients will be pumped through the lumen of the model to add shear stress on the endothelial cells. I will have access to JE serum and CSF samples through the Liverpool Group's extensive collaborations across Asia, funded by MRC and the Wellcome Trust and other samples from patients in the NIHR Encephalitis Programme Grant in Liverpool, headed by Prof. Tom Solomon. These modifications will allow an even better approximation to the real situation in humans. I will then use the model to understand the immunopathogenesis of JE. In particular, I will study the pathways of JEV-infected leukocyte trafficking through the BBB (not possible in the current model) and the effects of pro-inflammatory cytokines on this model. This model will be useful in the future for studying viral CNS infections including human immunodeficiency virus and herpes simplex virus, and for testing potential therapeutic drug candidates for viral encephalitis. Several UK and international biotech companies have expressed their strong interest on this as the model will be a useful tool for reducing/replacing the number of animals used in the first phase of drug discovery.


Research activity codes: 2.6 RESOURCES AND INFRASTRUCTURE (AETIOLOGY);2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION