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Alzheimer’s disease (AD) has been associated with cardiovascular and cerebrovascular risk factors (CVRFs) during middle age and later and is frequently accompanied by cerebrovascular pathology at death. An interaction between CVRFs and genetic variants might explain the pathogenesis. Genome-wide, gene by CVRF interaction analyses for AD, in 6568 patients and 8101 controls identified FMNL2 (p = 6.6 × 10–7). A significant increase in FMNL2 expression was observed in the brains of patients with brain infarcts and AD pathology and was associated with amyloid and phosphorylated tau deposition. FMNL2 was also prominent in astroglia in AD among those with cerebrovascular pathology. Amyloid toxicity in zebrafish increased fmnl2a expression in astroglia with detachment of astroglial end feet from blood vessels. Knockdown of fmnl2a prevented gliovascular remodeling, reduced microglial activity and enhanced amyloidosis. APP/PS1dE9 AD mice also displayed increased Fmnl2 expression and reduced the gliovascular contacts independent of the gliotic response. Based on this work, we propose that FMNL2 regulates pathology-dependent plasticity of the blood–brain-barrier by controlling gliovascular interactions and stimulating the clearance of extracellular aggregates. Therefore, in AD cerebrovascular risk factors promote cerebrovascular pathology which in turn, interacts with FMNL2 altering the normal astroglial-vascular mechanisms underlying the clearance of amyloid and tau increasing their deposition in brain.

In this chapter, we present the methodology currently used in our laboratory to generate a starPEG-MMP (starPEG)- and heparin maleimide HM06 (heparin)-based 3D cell culture system, in a hydrogel, that can be used to study human neuronal development and Alzheimer's disease (AD) pathology. A 3D cell culture system can mimic the in vivo cellular environment better than a 2D format, in which these cells exhibit neural network formation, electrophysiological activity, tissue-specific extracellular matrix (ECM) deposition, and neurotransmitter responsiveness. When treated with amyloid beta-42 (Aβ42) peptides, this system recapitulates many of the pathological effects of AD, including reduced neural stem cell proliferation, impaired neuronal network formation, dystrophic axonal ends, synaptic loss, failure to deposit ECM, elevated tau hyperphosphorylation, and formation of neurofibrillary tangles. Culturing human primary cortical astrocyte (pHA)- or induced pluripotent stem cell (iPSC)-derived human neural stem cells in this biohybrid hydrogel system has led to the discovery of novel regulatory pathways underlying neurodegenerative pathology in different phases of AD.

Neurogenesis relates to the brain resilience and is reduced in Alzheimer’s disease (AD). Restoring healthy levels of neurogenesis could have beneficial effects for coping with neurodegeneration. However, molecular mechanisms that could enhance neurogenesis from astroglial progenitors in AD pathology are largely unknown. We used lentiviruses to expressNgfrin the hippocampus of the APP/PS1dE9 mouse model of AD, histologically analyzed the changes in proliferation of neural stem cells and neurogenesis; performed single-cell transcriptomics, spatial proteomics, and functional knockdown studies. We found that expression ofNgfr, a neurogenic determinant in pathology-induced neuroregeneration in zebrafish, stimulated proliferative and neurogenic outcome in the APP/PS1dE9 AD mouse model. Ngfr suppressed reactive astrocyte marker Lipocalin-2 (Lcn2) in astroglia. Blockage of Lcn2 receptor, Slc22a17, recapitulated the neurogenic effects of NGFR, and long-term Ngfr expression reduced amyloid plaques and Tau phosphorylation. Furthermore, immunostaining on postmortem human hippocampi with AD or primary age-related Tauopathy and 3D human astroglial cultures showed that elevated LCN2 levels correlate with gliosis. By comparing transcriptional changes in mouse hippocampus, zebrafish brain, and human AD brains in terms of cell intrinsic differential gene expression analyses as well as weighted gene co-expression network analysis, we observed common potential downstream effectors of NGFR signaling,C4BandPFKP, that are relevant to AD. Our study links the regulatory role of an autocrine molecular mechanism in astroglia to the neurogenic ability and modulatory effects on amyloid and tau phosphorylation, opening new research avenues and suggesting that neurogenesis-oriented therapeutic approaches could be a potential clinical intervention for AD.