Genetic regulation of autoimmune neuroinflammation

Abstract

Multiple sclerosis (MS) is a complex disease characterized by inflammation and demyelination in the central nervous system (CNS), causing neurological deficits. Although there is an increased familial aggregation rate and evidence for a genetic influence on disease susceptibility, environmental factors are also of importance. MS is a complex disease, meaning that no single mode of inheritance is evident. Disease regulatory genes have been difficult to identify with linkage analysis and association studies in humans due to genetic heterogeneity, polygeneity, incomplete penetrance, gene-gene interactions and environmental variations. To reduce the effects of these obstacles we have used an animal model, Experimental Autoimmune Encephalomyelitis (EAE) in the rat, in which genetic heterogeneity is reduced, the environment is controlled, the disease phenotypes are well defined and the sample size is not limited. EAE is induced by immunization with myelin oligodendrocyte glycoprotein (MOG) and results in a CNS-specific chronic relapsing inflammatory disease that closely resembles human MS.

Only the MHC region has proven to be of significance for disease development in both MS and EAE. However, there is evidence for great impact of non-MHC genes on disease susceptibility. The aim of this thesis work has thus been to identify non-MHC loci/genes regulating MOG-EAE. Firstly, linkage analysis has performed in a (LEW.1AV1 x PVG.1AV1) F2 intercross, where the LEW background is susceptible and PVG background is protective in MOG-EAE. We identified Eae16 and Eae17 on chromosomes 8 and 13, respectively. These regions differed from previously identified EAE loci in other strain combinations.

Secondly, there are speculations about autoimmune diseases sharing common pathways and common susceptibility genes. We therefore characterized congenic strains for arthritis-linked loci on chromosome 4 in MOG-EAE. We evaluated three recombinant strains: C4R1, C4R2 and C4R3 and the full-length congenic DA.(C4)PVG. PVG compared to DA alleles in C4R1 did not have any effect in arthritis, but displayed a down-regulatory effect in MOG-EAE. In contrast to the down-regulatory effect in arthritis, PVG alleles in C4R3 resulted in tendencies of a worsened MOG-EAE compared to DA. The full-length congenic, C4, displayed an intermediary EAE disease, suggesting that the C4R1 and the C4R3 fragments counteract eachother.

Previously, Eae18 on rat chromosome 10 was identified in a (DA x ACI) F2 intercross and the biological effect of this locus was demonstrated in a congenic strain. This congenic has been narrowed down to a 15cM large fragment. Eae18 was further mapped in an advanced intercross line (AIL) between the susceptible DA and protective PVG.1AV1 strains and linkage analysis performed on animals in the F7 generation. The AIL has the advantage of more recombinational events, resulting in ~3.5 fold reduction of the confidence interval in a F7 compared to a F2 population in a sample of the same size. Eae18 was resolved into three separate loci between 2.5-4Mb in size, all of which are syntenic to regions with linkage in human MS materials.

An EAE regulatory effect of the previously identified Eae5 locus was demonstrated using the congenic strain BN.DA-Eae5. The AIL was utilized to resolve this locus on rat chromosome 12. Linkage analysis performed resulted in a confidence interval of ~1.4Mb, comprising 20 genes in total, of which Ncf-1 is one of eight confirmed genes in the rat. Ncf-1 has been identified as a regulatory gene in pristane-induced arthritis and is also a prime candidate in MOG-EAE. The identification of EAE regulatory genes will hopefully improve the understanding of MS pathogenesis and unravel new, more efficient therapeutic targets for MS patients.