Molecular characterization of selectively vulnerable neurons in Alzheimer’s Disease

Abstract

Alzheimer's disease (AD) is characterized by the selective vulnerability of specific neuronal populations, the molecular signatures of which are largely unknown. To identify and characterize selectively vulnerable neuronal populations, we used single-nucleus RNA sequencing to profile the caudal entorhinal cortex and the superior frontal gyrus -brain regions where neurofibrillary inclusions and neuronal loss occur early and late in AD, respectively -from individuals spanning the neuropathological progression of AD. We identified RORB as a marker of selectively vulnerable excitatory neurons in the entorhinal cortex, and subsequently validated their depletion and selective susceptibility to neurofibrillary inclusions during disease progression using quantitative neuropathological methods. We also discovered an astrocyte subpopulation, likely representing reactive astrocytes, characterized by decreased expression of genes involved in homeostatic functions. Our characterization of selectively vulnerable neurons in AD paves the way for future mechanistic studies of selective vulnerability and potential therapeutic strategies for enhancing neuronal resilience. MAIN TEXTSelective vulnerability is a fundamental feature of neurodegenerative diseases, in which different neuronal populations show a gradient of susceptibility to degeneration 1, 2 . Selective vulnerability at the network level has been extensively explored in Alzheimer's disease (AD) 3-5 , currently the leading cause of dementia and lacking in effective therapies. However, little is known about the mechanisms underlying selective vulnerability at the cellular level in AD, which could provide insight into disease mechanisms and lead to therapeutic strategies.The entorhinal cortex (EC), an allocortex, is one of the first cortical brain regions to exhibit neuronal loss in AD 6 . Neurons in the external EC layers, especially in layer II (also known as alpha clusters of the lamina principalis externa, abbreviated "Pre-alpha") 7 , accumulate taupositive neurofibrillary changes and die early on in the course of AD 8-13 . However, these selectively vulnerable neurons have yet to be characterized extensively at the molecular level. Furthermore, it is unknown whether there are differences in vulnerability among subpopulations of these neurons. Although rodent models of AD have offered some insights [14][15][16] , the human brain has unique features with regard to cellular physiology, composition and aging [17][18][19] , limiting the extrapoloation of findings from animal models to address selective vulnerability.Previous studies have combined laser capture microdissection with microarray analysis of gene expression 20, 21 to characterize EC neurons in AD, but focused on disease-related changes in gene expression, rather than selective vulnerability. More recently, single-nucleus RNA-sequencing (snRNA-seq) has enabled large-scale characterization of transcriptomic profiles of individual cells from post-mortem human brain tissue 22, 23 . However, snRNA-seq studies of AD p...

Bayesian estimation of cell-type-specific gene expression per bulk sample with prior derived from single-cell data

“…Using bMIND, we estimated sample-level CTS expression and detected CTS-DEGs related to AD with FDR < 0.05 (Supplemental Table S1). Similar to the findings based on snRNA-seq of AD (Mathys et al, 2019), most identified CTS-DEGs were from excitatory neurons, a finding that comports with the observed selective vulnerability of excitatory neurons in the brain of AD samples (Leng et al, 2020). We compared the ExN-DEGs identified by the snRNA-seq study (Mathys et al, 2019) and bMIND from the three bulk datasets (Fig.…”

Astrocytes in Alzheimer´s Disease: Pathological Significance and Molecular Pathways

“…Subclustering of astrocyte nuclei revealed four subpopulations of cells with one subcluster called Ast1 enriched with AD cells that upregulated GLUL and the AD risk gene CLU (Mathys et al 2019) (Table 1), previously found upregulated in reactive astrocytes in response to neurodegeneration (Shin et al 2006). Recent singlenucleus sequencing of the entorhinal cortex and the superior frontal gyrus from human healthy brains (n = 3), early (n = 4) and advanced (n = 3) stages of AD also revealed an astrocyte subpopulation expressing higher levels of GFAP, called GFAP-high (Leng et al 2021). GFAP-high astrocytes upregulate CD44 and TNC, both involved in interactions with the extracellular matrix; as well as HSPB1 and HSP90AA1, chaperones involved in proteostasis.…”

“…GFAP-high astrocytes upregulate CD44 and TNC, both involved in interactions with the extracellular matrix; as well as HSPB1 and HSP90AA1, chaperones involved in proteostasis. Interestingly, GFAP-high astrocytes downregulated genes involved in glutamate and GABA homeostasis (SLC1A2, SLC1A3, GLUL and SLC6A11) and synaptic adhesion/maintenance (NRXN1, CADM2, PTN and GPC5), indicating they may have compromised homeostatic function (Leng et al 2021) (Table 1).…”

Molecular mechanisms of neurodegeneration in the entorhinal cortex that underlie its selective vulnerability during the pathogenesis of Alzheimer's disease

“…Importantly, it was found that although the AD risk gene apolipoprotein E (APOE) was repressed in astrocyte subpopulations and in AD-related oligodendrocyte progenitor cells, it was upregulated in an AD-specific microglial subpopulations within the EC, suggesting that these specialized cells contribute to disease susceptibility of the EC during AD. A selectively vulnerable subpopulation of excitatory neurons in the EC, which shows a corresponding selective depletion as the disease progresses, has also been identified using SnRNA-seq in post-mortem AD brains ( Leng et al, 2020 , preprint). Interestingly, this study identified nuclear receptor RAR-related orphan receptor beta (RORB), which is a developmental driver of neuronal subtype identity ( Jabaudon et al, 2012 ; Oishi et al, 2016 ), as a marker for the selective vulnerability of excitatory EC neurons.…”