<scp>TGF</scp>‐β2 and <scp>TGF</scp>‐β3 from cultured β‐amyloid‐treated or 3xTg‐<scp>AD</scp>‐derived astrocytes may mediate astrocyte–neuron communication

Tapella, Laura; Cerruti, M.; Biocotino, Isabella; Stevano, Alessio; Rocchio, Francesca; Canonico, Pier Luigi; Grilli, Mariagrazia; Genazzani, Armando A.; Lim, Dmitry

doi:10.1111/ejn.13819

Cited by 25 publications

(20 citation statements)

References 82 publications

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“…Our data suggest that these changes concern multiple aspects of astroglial biology, characteristic for AD as the deregulation of cellular bioenergetics and Ca 2+ signaling, mitochondria-ER interaction and proteostasis. The effects of these changes, in turn, may have multiple consequences, e.g., we recently have shown that conditioned medium from 3xTg-AD astrocytes reduces synaptic proteins in cultured neurons 50 , while 3Tg-iAstros fail to support the integrity of the blood-brain barrier in vitro 51 . In light of this, our work exemplifies the importance of multidisciplinary approach in studying such complex phenomena as AD and illustrates the use of iAstro cells as versatile and cost-effective astrocytic cellular model of AD.…”

Section: Discussionmentioning

confidence: 99%

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

et al. 2020

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The pathogenesis of Alzheimer's disease (AD), a slowly-developing age-related neurodegenerative disorder, is a result of the action of multiple factors including deregulation of Ca 2+ homeostasis, mitochondrial dysfunction, and dysproteostasis. Interaction of these factors in astrocytes, principal homeostatic cells in the central nervous system, is still poorly understood. Here we report that in immortalized hippocampal astrocytes from 3xTg-AD mice (3Tg-iAstro cells) bioenergetics is impaired, including reduced glycolysis and mitochondrial oxygen consumption, and increased production of reactive oxygen species. Shotgun proteomics analysis of mitochondria-ER-enriched fraction showed no alterations in the expression of mitochondrial and OxPhos proteins, while those related to the ER functions and protein synthesis were deregulated. Using ER-and mitochondria-targeted aequorin-based Ca 2+ probe we show that, in 3Tg-iAstro cells, ER was overloaded with Ca 2+ while Ca 2+ uptake by mitochondria upon ATP stimulation was reduced. This was accompanied by the increase in short distance (≈8-10 nm) contact area between mitochondria and ER, upregulation of ER-stress/unfolded protein response genes Atf4, Atf6 and Herp, and reduction of global protein synthesis rate. We suggest that familial AD mutations in 3Tg-iAstro cells induce mitochondria-ER interaction changes that deregulate astrocytic bioenergetics, Ca 2+ homeostasis and proteostasis. These factors may interact, creating a pathogenic loop compromising homeostatic and defensive functions of astroglial cells predisposing neurons to dysfunction.

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Section: Discussionmentioning

confidence: 99%

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

et al. 2020

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“…We, therefore, investigated the levels of mRNA of TGFβ2 and TGFβ3 in both non-stimulated and LPS-treated iAstro lines. As shown in Supplementary Figure 3, in line with cultured primary astrocytes 42 , TGFβ3 is the most expressed TGFβ isoform in iAstro lines. Moreover, TGFβ3 was significantly up-regulated in 3Tg-iAstro (0.267 ± 0.02 ΔC(t) TGFβ3/S18, n = 4) compared to WT-iAsro cells (0.094 ± 0.014 ΔC(t) TGFβ3/S18, n = 4; p = 0.0012) (Supplementary Figure 3B).…”

Section: Resultsmentioning

confidence: 64%

“…We recently reported that transforming growth factor-β2 (TGFβ2) and TGFβ3 were up-regulated at mRNA levels in 3Tg-primary astrocytes as compared to WT astroglial cultures 42 . We, therefore, investigated the levels of mRNA of TGFβ2 and TGFβ3 in both non-stimulated and LPS-treated iAstro lines.…”

Section: Resultsmentioning

confidence: 99%

Gene expression, proteome and calcium signaling alterations in immortalized hippocampal astrocytes from an Alzheimer’s disease mouse model

Rocchio¹,

Tapella²,

Manfredi³

et al. 2019

Cell Death Dis

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Evidence is rapidly growing regarding a role of astroglial cells in the pathogenesis of Alzheimer’s disease (AD), and the hippocampus is one of the important brain regions affected in AD. While primary astroglial cultures, both from wild-type mice and from rodent models of AD, have been useful for studying astrocyte-specific alterations, the limited cell number and short primary culture lifetime have limited the use of primary hippocampal astrocytes. To overcome these limitations, we have now established immortalized astroglial cell lines from the hippocampus of 3xTg-AD and wild-type control mice (3Tg-iAstro and WT-iAstro, respectively). Both 3Tg-iAstro and WT-iAstro maintain an astroglial phenotype and markers (glutamine synthetase, aldehyde dehydrogenase 1 family member L1 and aquaporin-4) but display proliferative potential until at least passage 25. Furthermore, these cell lines maintain the potassium inward rectifying (Kir) current and present transcriptional and proteomic profiles compatible with primary astrocytes. Importantly, differences between the 3Tg-iAstro and WT-iAstro cell lines in terms of calcium signaling and in terms of transcriptional changes can be re-conducted to the changes previously reported in primary astroglial cells. To illustrate the versatility of this model we performed shotgun mass spectrometry proteomic analysis and found that proteins related to RNA binding and ribosome are differentially expressed in 3Tg-iAstro vs WT-iAstro. In summary, we present here immortalized hippocampal astrocytes from WT and 3xTg-AD mice that might be a useful model to speed up research on the role of astrocytes in AD.

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“…Primary cultures were prepared as described previously (Tapella et al, 2018) with slight modifications. In brief, for purified astroglial cultures, cerebella were dissected from 3-to 5-day-old pups in cold HBSS, digested in 0.25% trypsin (37 °C, 20 min), washed in complete culture medium and resuspended in cold HBSS supplemented with 10% FBS.…”

Section: Primary Astroglial and Mixed Culturesmentioning

confidence: 99%

Deletion of calcineurin from GFAP‐expressing astrocytes impairs excitability of cerebellar and hippocampal neurons through astroglial Na⁺/K⁺ ATPase

Tapella

Soda

Mapelli

et al. 2019

Glia

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Astrocytes perform important housekeeping functions in the nervous system including maintenance of adequate neuronal excitability, although the regulatory mechanisms are currently poorly understood. The astrocytic Ca 2+ /calmodulin-activated phosphatase calcineurin (CaN) is implicated in the development of reactive gliosis and neuroinflammation, but its roles, including the control of neuronal excitability, in healthy brain is unknown. We have generated a mouse line with conditional knockout (KO) of CaN B1 (CaNB1) in glial fibrillary acidic protein (GFAP)-expressing astrocytes (astroglial calcineurin knock-out, ACN-KO). Here we report that postnatal and astrocyte-specific ablation of CaNB1 did not alter normal growth and development as well as adult neurogenesis. Yet, we found that specific deletion of astrocytic CaN selectively impairs intrinsic neuronal excitability in hippocampal CA1 pyramidal neurons and cerebellar granule cells (CGCs).This impairment was associated with a decrease in after-hyperpolarization in CGC, while passive properties were unchanged, suggesting impairment of K + homeostasis. Indeed, blockade of Na + /K + -ATPase (NKA) with ouabain phenocopied the electrophysiological alterations observed in ACN-KO CGCs. In addition, NKA activity was significantly lower in cerebellar and hippocampal lysates and in pure astrocytic cultures from ACN-KO mice. While no changes were found in protein levels, NKA activity was inhibited by the specific CaN inhibitor FK506 in both cerebellar lysates and primary astroglia from control mice, suggesting that CaN directly modulates NKA activity and in this manner controls neuronal excitability. In summary, our data provide formal evidence for the notion that astroglia is fundamental for controlling basic neuronal functions and place CaN center-stage as an astrocytic Ca 2+ -sensitive switch.

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TGF‐β2 and TGF‐β3 from cultured β‐amyloid‐treated or 3xTg‐AD‐derived astrocytes may mediate astrocyte–neuron communication

Cited by 25 publications

References 82 publications

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

Gene expression, proteome and calcium signaling alterations in immortalized hippocampal astrocytes from an Alzheimer’s disease mouse model

Deletion of calcineurin from GFAP‐expressing astrocytes impairs excitability of cerebellar and hippocampal neurons through astroglial Na⁺/K⁺ ATPase

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TGF‐β2 and TGF‐β3 from cultured β‐amyloid‐treated or 3xTg‐AD‐derived astrocytes may mediate astrocyte–neuron communication

Cited by 25 publications

References 82 publications

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice

Gene expression, proteome and calcium signaling alterations in immortalized hippocampal astrocytes from an Alzheimer’s disease mouse model

Deletion of calcineurin from GFAP‐expressing astrocytes impairs excitability of cerebellar and hippocampal neurons through astroglial Na+/K+ ATPase

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Deletion of calcineurin from GFAP‐expressing astrocytes impairs excitability of cerebellar and hippocampal neurons through astroglial Na⁺/K⁺ ATPase