No abstract

Glial fibrillary acidic protein (GFAP) is the main astrocytic intermediate filament (IF). GFAP splice isoforms show differential expression patterns in the human brain. GFAPδ is preferentially expressed by neurogenic astrocytes in the subventricular zone (SVZ), whereas GFAP+1 is found in a subset of astrocytes throughout the brain. In addition, the expression of these isoforms in human brain material of epilepsy, Alzheimer and glioma patients has been reported. Here, for the first time, we present a comprehensive study of GFAP isoform expression in both wild-type and Alzheimer Disease (AD) mouse models. In cortex, cerebellum, and striatum of wild-type mice, transcripts for Gfap-α, Gfap-β, Gfap-γ, Gfap-δ, Gfap-κ, and a newly identified isoform Gfap-ζ, were detected. Their relative expression levels were similar in all regions studied. GFAPα showed a widespread expression whilst GFAPδ distribution was prominent in the SVZ, rostral migratory stream (RMS), neurogenic astrocytes of the subgranular zone (SGZ), and subpial astrocytes. In contrast to the human SVZ, we could not establish an unambiguous GFAPδ localization in proliferating cells of the mouse SVZ. In APPswePS1dE9 and 3xTgAD mice, plaque-associated reactive astrocytes had increased transcript levels of all detectable GFAP isoforms and low levels of a new GFAP isoform, Gfap-ΔEx7. Reactive astrocytes in AD mice showed enhanced GFAPα and GFAPδ immunolabeling, less frequently increased vimentin and nestin, but no GFAPκ or GFAP+1 staining. In conclusion, GFAPδ protein is present in SVZ, RMS, and neurogenic astrocytes of the SGZ, but also outside neurogenic niches. Furthermore, differential GFAP isoform expression is not linked with aging or reactive gliosis. This evidence points to the conclusion that differential regulation of GFAP isoforms is not involved in the reorganization of the IF network in reactive gliosis or in neurogenesis in the mouse brain.

No abstract

In Alzheimer's disease (AD), amyloid plaques are surrounded by reactive astrocytes with an increased expression of intermediate filaments including glial fibrillary acidic protein (GFAP). Different GFAP isoforms have been identified that are differentially expressed by specific subpopulations of astrocytes and that impose different properties to the intermediate filament network. We studied transcript levels and protein expression patterns of all known GFAP isoforms in human hippocampal AD tissue at different stages of the disease. Ten different transcripts for GFAP isoforms were detected at different abundancies. Transcript levels of most isoforms increased with AD progression. GFAPδ-immunopositive astrocytes were observed in subgranular zone, hilus, and stratum-lacunosum-moleculare. GFAPδ-positive cells also stained for GFAPα. In AD donors, astrocytes near plaques displayed increased staining of both GFAPα and GFAPδ. The reading-frame-shifted isoform, GFAP(+1), staining was confined to a subset of astrocytes with long processes, and their number increased in the course of AD. In conclusion, the various GFAP isoforms show differential transcript levels and are upregulated in a concerted manner in AD. The GFAP(+1) isoform defines a unique subset of astrocytes, with numbers increasing with AD progression. These data indicate the need for future exploration of underlying mechanisms concerning the functions of GFAPδ and GFAP(+1) isoforms in astrocytes and their possible role in AD pathology.

Amyloid plaques in Alzheimer's disease (AD) mice are surrounded by activated microglia. The functional role of microglia activation in AD is not well understood; both detrimental and beneficial effects on AD progression have been reported. Here we show that the population of activated microglia in the cortex of the APPswe/PS1dE9 mouse AD model is divided into a CD11c-positive and a CD11c-negative subpopulation. Cd11c transcript levels and number of CD11c-positive microglia increase sharply when plaques start to occur and both parameters continue to rise in parallel with the age-related increasing plaque load. CD11c cells are localized near plaques at all stages of the disease development and constitute 23% of all activated microglia. No differences between these two populations were found in terms of proliferation, immunostaining intensity of Iba1, MHC class II, CD45, or immunoproteasome subunit LMP7/β5i. Comparison of the transcriptome of isolated CD11c-positive and CD11c-negative microglia from the cortex of aged APPswe/PS1dE9 with WT microglia showed that gene expression changes had a similar general pattern. However, a differential expression was found for genes involved in immune signaling (Il6, S100a8/Mrp8, S100a9/Mrp14, Spp1, Igf1), lysosome activation, and carbohydrate- and cholesterol/lipid-metabolism (Apoe). In addition, the increased expression of Gpnmb/DC-HIL, Tm7sf4/DC-STAMP, and Gp49a/Lilrb4, suggests a suppressive/tolerizing influence of CD11c cells. We show that amyloid plaques in the APP/PS1 model are associated with two distinct populations of activated microglia: CD11c-positive and CD11c-negative cells. Our findings imply that CD11c-positive microglia can potentially counteract amyloid deposition via increased Aβ-uptake and degradation, and by containing the inflammatory response.

SignificanceThe adult mammalian heart does not functionally repair itself after injury. Therefore, identification of cardiac stem cell (CSC) populations is of great interest for regenerative intervention. However, the significance of such CSC populations remains heavily debated. Using single-cell mRNA sequencing and genetic lineage tracing, we interrogate the existence of CSCs with unbiased mouse models of proliferation. Cycling cardiomyocytes were only robustly observed in the early postnatal growth phase, while cycling cells in homoeostatic and damaged adult myocardium consisted mainly of various noncardiomyocyte cell types. Injury-activated cardiac fibroblasts that acquire a gene expression profile similar to that of neonatal cardiac fibroblasts signal—in an autocrine fashion—to prevent cardiac rupture. We find no evidence for the existence of a quiescent CSC population.

The proteasome is the major protein degradation system within the cell, comprised of different proteolytic subunits; amyloid-β is thought to impair its activity in Alzheimer's disease. Neuroinflammation is a prominent hallmark of Alzheimer's disease, which may implicate an activation of the immunoproteasome, a specific proteasome variant induced by immune signalling that holds slightly different proteolytic properties than the constitutive proteasome. Using a novel cell-permeable proteasome activity probe, we found that amyloid-β enhances proteasome activity in glial and neuronal cultures. Additionally, using a subunit-specific proteasome activity assay we showed that in the cortex of the APPswePS1dE9 plaque pathology mouse model, immunoproteasome activities were strongly increased together with increased messenger RNA and protein expression in reactive glia surrounding plaques. Importantly, this elevated activity was confirmed in human post-mortem tissue from donors with Alzheimer's disease. These findings are in contrast with earlier studies, which reported impairment of proteasome activity in human Alzheimer's disease tissue and mouse models. Targeting the increased immunoproteasome activity with a specific inhibitor resulted in a decreased expression of inflammatory markers in ex vivo microglia. This may serve as a potential novel approach to modulate sustained neuroinflammation and glial dysfunction associated with Alzheimer's disease.

Reactive astrocytes with an increased expression of intermediate filament (IF) proteins Glial Fibrillary Acidic Protein (GFAP) and Vimentin (VIM) surround amyloid plaques in Alzheimer's disease (AD). The functional consequences of this upregulation are unclear. To identify molecular pathways coupled to IF regulation in reactive astrocytes, and to study the interaction with microglia, we examined WT and APPswe/PS1dE9 (AD) mice lacking either GFAP, or both VIM and GFAP, and determined the transcriptome of cortical astrocytes and microglia from 15- to 18-month-old mice. Genes involved in lysosomal degradation (including several cathepsins) and in inflammatory response (including Cxcl5, Tlr6, Tnf, Il1b) exhibited a higher AD-induced increase when GFAP, or VIM and GFAP, were absent. The expression of Aqp4 and Gja1 displayed the same pattern. The downregulation of neuronal support genes in astrocytes from AD mice was absent in GFAP/VIM null mice. In contrast, the absence of IFs did not affect the transcriptional alterations induced by AD in microglia, nor was the cortical plaque load altered. Visualizing astrocyte morphology in GFAP-eGFP mice showed no clear structural differences in GFAP/VIM null mice, but did show diminished interaction of astrocyte processes with plaques. Microglial proliferation increased similarly in all AD groups. In conclusion, absence of GFAP, or both GFAP and VIM, alters AD-induced changes in gene expression profile of astrocytes, showing a compensation of the decrease of neuronal support genes and a trend for a slightly higher inflammatory expression profile. However, this has no consequences for the development of plaque load, microglial proliferation, or microglial activation.