Heat shock protein responses to aging and proteotoxicity in the olfactory bulb

Crum, Tyler S.; Gleixner, Amanda M.; Posimo, Jessica M.; Mason, Daniel M.; Broeren, Matthew T.; Heinemann, Scott D.; Wipf, Peter; Brodsky, Jeffrey L.; Leak, Rehana K.

doi:10.1111/jnc.13041

Cited by 15 publications

(14 citation statements)

References 114 publications

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“…Notably, aged allocortex exhibited lower levels of Hsp90 and CHIP than aged neocortex, an observation that supports further examination of these proteins in relation to diseases associated with aging. Finally, the age- and MG132-related increase in Hsp25 in allocortex is consistent with previous work that this Hsp is higher in the cortex, striatum, substantia nigra, and the olfactory bulb of aged animals relative to young controls (Crum et al, in press; Dickey et al, 2009; Gleixner et al, 2014; Gupte et al, 2010; Schultz et al, 2001). …”

Section: Discussionsupporting

confidence: 91%

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Heat shock protein defenses in the neocortex and allocortex of the telencephalon

Posimo

Weilnau

Gleixner

et al. 2015

Neurobiology of Aging

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The telencephalic allocortex develops protein inclusions before the neocortex in many age-related proteinopathies. One major defense mechanism against proteinopathic stress is the heat shock protein (Hsp) network. We therefore contrasted Hsp defenses in stressed primary neo- and allocortical cells. Neocortical neurons were more resistant to the proteasome inhibitor MG132 than neurons from three allocortical subregions: entorhinal cortex, piriform cortex, and hippocampus. However, allocortical neurons exhibited higher MG132-induced increases in Hsp70 and Hsc70. MG132-treated allocortical neurons also exhibited greater levels of protein ubiquitination. Inhibition of Hsp70/Hsc70 activity synergistically exacerbated MG132 toxicity in allocortical neurons more than neocortical neurons, suggesting that the allocortex is more reliant on these Hsp defenses. In contrast, astrocytes harvested from neo- or allocortex did not differ in their response to Hsp70/Hsc70 inhibition. Consistent with the idea that chaperones are maximally engaged in allocortical neurons, an increase in Hsp70/Hsc70 activity was protective only in neocortical neurons. Finally, the levels of select Hsps were altered in neocortex and allocortex in vivo with aging.

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Section: Discussionsupporting

confidence: 91%

“…Although Hsp70 induction is critical when protein homeostasis is compromised, Hsp70 is less likely to protect neurons against oxidative toxicity (Crum et al, in press). To test this hypothesis in the current model, we also treated neo/allocortical neuron cultures with paraquat in the presence or absence of VER115005 and MAL3-101.…”

Section: Resultsmentioning

confidence: 99%

Heat shock protein defenses in the neocortex and allocortex of the telencephalon

Posimo

Weilnau

Gleixner

et al. 2015

Neurobiology of Aging

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“…Thus, we tested the hypothesis that loss of Hsp70 activity would lead to loss of astrocytic neuroprotection and applied the Hsp70/Hsc70 inhibitor VER155008 to MG132-treated neuron/astrocyte co-cultures. As shown in our recent work in olfactory bulb and cortical neurons, we found strikingly synergistic effects of inhibitors of proteasomal activity and Hsp70/Hsc70 activity [86, 87]. Remarkably, severely stressed astrocytes protected neurons robustly against this synergistic, profound toxicity (Figure 7).…”

Section: Resultssupporting

confidence: 74%

Astrocytes Surviving Severe Stress Can Still Protect Neighboring Neurons from Proteotoxic Injury

et al. 2015

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Astrocytes are one of the major cell types to combat cellular stress and protect neighboring neurons from injury. In order to fulfill this important role, astrocytes must sense and respond to toxic stimuli, perhaps including stimuli that are severely stressful and kill some of the astrocytes. The present study demonstrates that primary astrocytes that managed to survive severe proteotoxic stress were protected against subsequent challenges. These findings suggest that the phenomenon of preconditioning or tolerance can be extended from mild to severe stress for this cell type. Astrocytic stress adaptation lasted at least 96 hours, the longest interval tested. Heat shock protein 70 (Hsp70) was raised in stressed astrocytes, but inhibition of neither Hsp70 nor Hsp32 activity abolished their resistance against a second proteotoxic challenge. Only inhibition of glutathione synthesis abolished astrocytic stress adaptation, consistent with our previous report. Primary neurons were plated upon previously stressed astrocytes and the co-cultures were then exposed to another proteotoxic challenge. Severely stressed astrocytes were still able to protect neighboring neurons against this injury and the protection was unexpectedly independent of glutathione synthesis. Stressed astrocytes were even able to protect neurons after simultaneous application of proteasome and Hsp70 inhibitors, which otherwise elicited synergistic, severe loss of neurons when applied together. Astrocyte-induced neuroprotection against proteotoxicity was not elicited with astrocyte-conditioned media, suggesting that physical cell-to-cell contacts may be essential. These findings suggest that astrocytes may adapt to severe stress so that they can continue to protect neighboring cell types from profound injury.

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“…Tissue from the hippocampus was harvested from Sprague Dawley rat brains on postnatal day 1 or 2 (Charles River, Wilmington, MA, USA). Tissue chunks were dissociated and plated as previously described in Neurobasal‐A media with the supplement B27 (Crum et al, ; Posimo et al, ). Cultures were treated with MG132 (EMD Millipore, Billerica, MA, USA) or paraquat (Sigma‐Aldrich, St. Louis, MO, USA) on day in vitro 5 (DIV5) for 24 h. This was referred to as the first hit and was added to the existing media as a 10× solution.…”

Section: Methodsmentioning

confidence: 99%

“…In order to determine the neuronal purity of the cultures, cells were immunocytochemically labeled for the neuronal marker MAP2 and the astrocyte marker glial fibrillary acidic protein (GFAP) using visible‐range secondary antibodies for higher resolution microscopy, as previously described (Crum et al, ; Posimo et al, ). For the latter experiments, nuclei were stained with Hoechst 33258 (10 μg/mL, bisBenzimide) in phosphate‐buffered saline with 0.3% Triton‐X for 15 min.…”

Section: Methodsmentioning

confidence: 99%

Synergistic stress exacerbation in hippocampal neurons: Evidence favoring the dual‐hit hypothesis of neurodegeneration

et al. 2016

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The dual hit hypothesis of neurodegeneration states that severe stress sensitizes vulnerable cells to subsequent challenges so that the two hits are synergistic in their toxic effects. Although the hippocampus is vulnerable to a number of neurodegenerative disorders, there are no models of synergistic cell death in hippocampal neurons in response to combined proteotoxic and oxidative stressors, the two major characteristics of these diseases. Therefore, we developed a relatively high-throughput dual hit model of stress synergy in primary hippocampal neurons. In order to increase the rigor of our study and strengthen our interpretations, we employed three independent, unbiased viability assays at multiple timepoints. Stress synergy was elicited when hippocampal neurons were treated with the proteasome inhibitor MG132 followed by exposure to the oxidative toxicant paraquat, but only after 48h. MG132 and paraquat only elicited additive effects 24h after the final hit and even loss of heat shock protein 70 activity and glutathione did not promote stress synergy at this early timepoint. Dual hits of MG132 elicited modest glutathione loss and slightly synergistic toxic effects 48h after the second hit, but only at some concentrations and only according to two viability assays (metabolic fitness and cytoskeletal integrity). The thiol N-acetyl cysteine protected hippocampal neurons against dual MG132/MG132 hits but not dual MG132/paraquat hits. Our findings support the view that proteotoxic and oxidative stress propel and propagate each other in hippocampal neurons, leading to synergistically toxic effects, but not as the default response and only after a delay. The neuronal stress synergy observed here lies in contrast to astrocytic responses to dual hits, because astrocytes that survive severe proteotoxic stress resist additional cell loss following second hits. In conclusion, we present a new model of hippocampal vulnerability in which to test therapies, because neuroprotective treatments that are effective against severe, synergistic stress are more likely to succeed in the clinic.

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Heat shock protein responses to aging and proteotoxicity in the olfactory bulb

Cited by 15 publications

References 114 publications

Heat shock protein defenses in the neocortex and allocortex of the telencephalon

Heat shock protein defenses in the neocortex and allocortex of the telencephalon

Astrocytes Surviving Severe Stress Can Still Protect Neighboring Neurons from Proteotoxic Injury

Synergistic stress exacerbation in hippocampal neurons: Evidence favoring the dual‐hit hypothesis of neurodegeneration

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