The Clu‐rs2279590_h2kb variant increases axonal and neuritic damage in 5xFAD mice

Abstract

AbstractBackgroundGenome‐Wide Association Studies (GWAS) implicate clusterin (CLU, also known as apolipoprotein J) as a risk factor for late‐onset Alzheimer's disease (LOAD), with elevated expression of CLU being associated with increased risk of LOAD. The rs2279590 SNP risk allele is located within a putative enhancer element in intron 7 of CLU. Indeed, deletion of a 115bp region including this SNP reduces expression of CLU and neighboring genes (EPHX2 and PTK2B) on Chromosome 8. The region surrounding rs2279590 is poorly conserved within the mouse genome. Consequently, to model the effect of the rs2279590 risk allele on development of AD‐related pathology in mice we developed mice with a partially humanized allele of Clu.MethodCRISPR was used to substitute a 2kb region of human CLU (intron 7 to exon 9) containing the enhancer region and the GWAS AD‐risk rs2279590 SNP variant for the orthologous region of Clu within the C57BL/6J mouse genome. Clu‐rs2279590_h2kb mice were crossed with 5xFAD mice and aged to 4 months to study the effects of the partially humanized Clu allele on development of AD‐related pathologies in 5xFAD mice. Coronal brain sections were immunolabeled to visualize dense‐core plaques (Thioflavin‐S), microglia (IBA1), reactive astrocytes (GFAP), axonal and neuritic damage (neurofilament light chain (NfL) and LAMP1, respectively). Confocal images of hippocampal and cortical regions were analyzed using Imaris software. In addition, Aβ and NfL levels were quantified in the brain tissue and plasma NfL measured using meso scale discovery (MSD) technology.ResultFour‐month‐old 5xFAD hemizygous, Clu‐rs2279590_h2kb homozygous mice had reduced microglial density and volume compared to 5xFAD hemizygous controls. No change was found in astrocyte density or reactivity. Despite comparable plaque load between 5xFAD and 5xFAD x Clu‐rs2279590_h2kb mice, neuritic and axonal damage was elevated in the brain of 5xFAD x Clu‐rs2279590_h2kb mice compared to 5xFAD, while plasma NfL level did not differ. Insoluble Aβ42 levels mirrored plaque density.ConclusionDespite the lack of impact on Aβ plaque deposition, and moderate influence on gliosis, four‐month‐old 5xFAD hemizygous, Clu‐rs2279590_h2kb homozygous mice display elevated axonal and neuritic damage compared to 5xFAD hemizygous controls.