Damaged mitochondria coincide with presynaptic vesicle loss and abnormalities in alzheimer's disease brain.

Loss of synapses is the most robust pathological correlate of Alzheimer's disease (AD)-associated cognitive deficits, although the underlying mechanism remains incompletely understood. Synaptic terminals have abundant mitochondria which play an indispensable role in synaptic function through ATP provision and calcium buffering. Mitochondrial dysfunction is an early and prominent feature in AD which could contribute to synaptic deficits. Here, using electron microscopy, we examined synapses with a focus on mitochondrial deficits in presynaptic axonal terminals and dendritic spines in cortical biopsy samples from clinically diagnosed AD and age-matched non-AD control patients. Synaptic vesicle density within the presynaptic axon terminals was significantly decreased in AD cases which appeared largely due to significantly decreased reserve pool, but there were significantly more presynaptic axons containing enlarged synaptic vesicles or dense core vesicles in AD. Importantly, there was reduced number of mitochondria along with significantly increased damaged mitochondria in the presynapse of AD which correlated with changes in SV density. Mitochondria in the post-synaptic dendritic spines were also enlarged and damaged in the AD biopsy samples. This study provided evidence of presynaptic vesicle loss as synaptic deficits in AD and suggested that mitochondrial dysfunction in both pre- and post-synaptic compartments contribute to synaptic deficits in AD.

C19orf12 ablation causes ferroptosis in mitochondrial membrane protein-associated with neurodegeneration.

Mitochondrial membrane protein-associated with neurodegeneration (MPAN) is a rare genetic disease characterized by aggressive neurodegeneration and massive iron accumulation in patients' brains. Genetics studies identified defects in C19orf12 locus being associated with MPAN which likely caused loss of function although underlying pathogenic mechanism(s) remain elusive. In the present study, we investigated C19orf12 knockout (KO) M17 neuronal cells and primary skin fibroblasts from MPAN patients with C19orf12 homozygous G58S or heterozygous C19orf12 p99fs*102 mutations as cellular models of MPAN. C19orf12 KO cells and MPAN fibroblast cells demonstrated mitochondrial fragmentation and dysfunction, iron overload and increased oxidative damage. Antioxidant NAC and iron chelator DFO rescued both oxidative stress and mitochondrial deficits. Moreover, C19orf12 KO cells and MPAN fibroblast cells were susceptible to erastin- or RSL3-induced ferroptosis which could be almost completely prevented by pretreatment of iron chelator DFO. Importantly, we also found mitochondrial fragmentation and increased ferroptosis related oxidative damage in neurons in the biopsied cortical tissues from an MPAN patient. Collectively, these results supported the notion that iron overload and ferroptosis likely play an important role in the pathogenesis of MPAN.

METTL3-dependent RNA m6A dysregulation contributes to neurodegeneration in Alzheimer's disease through aberrant cell cycle events.

BACKGROUND: N6-methyladenosine (m6A) modification of RNA influences fundamental aspects of RNA metabolism and m6A dysregulation is implicated in various human diseases. In this study, we explored the potential role of RNA m6A modification in the pathogenesis of Alzheimer disease (AD). METHODS: We investigated the m6A modification and the expression of m6A regulators in the brain tissues of AD patients and determined the impact and underlying mechanism of manipulated expression of m6A levels on AD-related deficits both in vitro and in vivo. RESULTS: We found decreased neuronal m6A levels along with significantly reduced expression of m6A methyltransferase like 3 (METTL3) in AD brains. Interestingly, reduced neuronal m6A modification in the hippocampus caused by METTL3 knockdown led to significant memory deficits, accompanied by extensive synaptic loss and neuronal death along with multiple AD-related cellular alterations including oxidative stress and aberrant cell cycle events in vivo. Inhibition of oxidative stress or cell cycle alleviated shMettl3-induced apoptotic activation and neuronal damage in primary neurons. Restored m6A modification by inhibiting its demethylation in vitro rescued abnormal cell cycle events, neuronal deficits and death induced by METTL3 knockdown. Soluble Aß oligomers caused reduced METTL3 expression and METTL3 knockdown exacerbated while METTL3 overexpression rescued Aß-induced synaptic PSD95 loss in vitro. Importantly, METTL3 overexpression rescued Aß-induced synaptic damage and cognitive impairment in vivo. CONCLUSIONS: Collectively, these data suggested that METTL3 reduction-mediated m6A dysregulation likely contributes to neurodegeneration in AD which may be a therapeutic target for AD.

Oxidative Stress Signaling in Blast TBI-Induced Tau Phosphorylation.

Traumatic brain injury caused by blast is associated with long-term neuropathological changes including tau phosphorylation and pathology. In this study, we aimed to determine changes in initial tau phosphorylation after exposure to a single mild blast and the potential contribution of oxidative stress response pathways. C57BL/6 mice were exposed to a single blast overpressure (BOP) generated by a compressed gas-driven shock tube that recapitulates battlefield-relevant open-field BOP, and cortical tissues were harvested at different time points up to 24 h after blast for Western blot analysis. We found that BOP caused elevated tau phosphorylation at Ser202/Thr205 detected by the AT8 antibody at 1 h post-blast followed by tau phosphorylation at additional sites (Ser262 and Ser396/Ser404 detected by PHF1 antibody) and conformational changes detected by Alz50 antibody. BOP also induced acute oxidative damage at 1 h post-blast and gradually declined overtime. Interestingly, Extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) were acutely activated in a similar temporal pattern as the rise and fall in oxidative stress after blast, with p38 showing a similar trend. However, glycogen synthase kinase-3 ß (GSK3ß) was inhibited at 1 h and remained inhibited for 24 h post blast. These results suggested that mitogen-activated protein kinases (MAPKs) but not GSK3ß are likely involved in mediating the effects of oxidative stress on the initial increase of tau phosphorylation following a single mild blast.

VPS35 D620N knockin mice recapitulate cardinal features of Parkinson's disease.

D620N mutation in the vacuolar protein sorting 35 ortholog (VPS35) gene causes late-onset, autosomal dominant familial Parkinson's disease (PD) and contributes to idiopathic PD. However, how D620N mutation leads to PD-related deficits in vivo remains unclear. In the present study, we thoroughly characterized the biochemical, pathological, and behavioral changes of a VPS35 D620N knockin (KI) mouse model with chronic aging. We reported that this VPS35 D620N KI model recapitulated a spectrum of cardinal features of PD at 14 months of age which included age-dependent progressive motor deficits, significant changes in the levels of dopamine (DA) and DA metabolites in the striatum, and robust neurodegeneration of the DA neurons in the SNpc and DA terminals in the striatum, accompanied by increased neuroinflammation, and accumulation and aggregation of α-synuclein in DA neurons. Mechanistically, D620N mutation induced mitochondrial fragmentation and dysfunction in aged mice likely through enhanced VPS35-DLP1 interaction and increased turnover of mitochondrial DLP1 complexes in vivo. Finally, the VPS35 D620N KI mice displayed greater susceptibility to MPTP-mediated degeneration of nigrostriatal pathway, indicating that VPS35 D620N mutation increased vulnerability of DA neurons to environmental toxins. Overall, this VPS35 D620N KI mouse model provides a powerful tool for future disease modeling and pharmacological studies of PD. Our data support the involvement of VPS35 in the development of α-synuclein pathology in vivo and revealed the important role of mitochondrial fragmentation/dysfunction in the pathogenesis of VPS35 D620N mutation-associated PD in vivo.

Mfn2 Ablation in the Adult Mouse Hippocampus and Cortex Causes Neuronal Death.

It is believed that mitochondrial fragmentation cause mitochondrial dysfunction and neuronal deficits in Alzheimer's disease. We recently reported that constitutive knockout of the mitochondria fusion protein mitofusin2 (Mfn2) in the mouse brain causes mitochondrial fragmentation and neurodegeneration in the hippocampus and cortex. Here, we utilize an inducible mouse model to knock out Mfn2 (Mfn2 iKO) in adult mouse hippocampal and cortical neurons to avoid complications due to developmental changes. Electron microscopy shows the mitochondria become swollen with disorganized and degenerated cristae, accompanied by increased oxidative damage 8 weeks after induction, yet the neurons appear normal at the light level. At later timepoints, increased astrocyte and microglia activation appear and nuclei become shrunken and pyknotic. Apoptosis (Terminal deoxynucleotidyl transferase dUTP nick end labeling, TUNEL) begins to occur at 9 weeks, and by 12 weeks, most hippocampal neurons are degenerated, confirmed by loss of NeuN. Prior to the loss of NeuN, aberrant cell-cycle events as marked by proliferating cell nuclear antigen (PCNA) and pHistone3 were evident in some Mfn2 iKO neurons but do not colocalize with TUNEL signals. Thus, this study demonstrated that Mfn2 ablation and mitochondrial fragmentation in adult neurons cause neurodegeneration through oxidative stress and neuroinflammation in vivo via both apoptosis and aberrant cell-cycle-event-dependent cell death pathways.

Conditional Haploinsufficiency of ß-Catenin Aggravates Neuronal Damage in a Paraquat-Based Mouse Model of Parkinson Disease.

The canonical Wnt pathway is critical for both the development and adulthood survival and homeostatic maintenance of the midbrain dopaminergic (DA) neurons. Expanding evidence has demonstrated that genetic factors associated with familial Parkinson disease (PD) deregulate this important pathway, suggesting that a disturbed canonical Wnt pathway is likely involved in PD pathogenesis; yet, the specific role of this pathway in sporadic PD remains unclear. In this study, we aimed to determine the effects of specific inhibition of the canonical pathway by hemizygous knockout of ß-catenin, the obligatory component of the canonical Wnt pathway, on paraquat (PQ)-induced DA neuronal degeneration in the substantia nigra in vivo. We found that while hemizygous conditional knockout of ß-catenin in DA neurons did not cause any significant TH+ neuronal loss in the substantia nigra at basal level, it triggered elevated oxidative stress at basal level and further enhanced PQ-induced oxidative damage and loss of TH+ neurons in the substantia nigra and axonal termini in the striatum that manifested as exacerbated motor deficits. These data support the notion that reduced Wnt/ß-catenin signaling in sporadic PD likely contributes to DA neuronal loss through an enhanced oxidative stress-response pathway.

The sterol regulatory element-binding protein 2 is dysregulated by tau alterations in Alzheimer disease.

Disturbed neuronal cholesterol homeostasis has been observed in Alzheimer disease (AD) and contributes to the pathogenesis of AD. As the master switch of cholesterol biosynthesis, the sterol regulatory element-binding protein 2 (SREBP-2) translocates to the nucleus after cleavage/activation, but its expression and activation have not been studied in AD which is the focus of the current study. We found both a significant decrease in the nuclear translocation of N-terminal SREBP-2 accompanied by a significant accumulation of C-terminal SREBP-2 in NFT-containing pyramidal neurons in AD. N-terminal- SREBP-2 is also found in dystrophic neurites around plaques in AD brain. Western blot confirmed a significantly reduced nuclear translocation of mature SREBP-2 (mSREBP-2) in AD brain. Interestingly, reduced nuclear mSREBP-2 was only found in animal models of tauopathies such as 3XTg AD mice and P301L Tau Tg mice but not in CRND8 APP transgenic mice, suggesting that tau alterations likely are involved in the changes of mSREBP-2 distribution and activation in AD. Altogether, our study demonstrated disturbed SREBP-2 signaling in AD and related models, and proved for the first time that tau alterations contribute to disturbed cholesterol homeostasis in AD likely through modulation of nuclear mSREBP-2 translocation.

Mitofusin 2 Regulates Axonal Transport of Calpastatin to Prevent Neuromuscular Synaptic Elimination in Skeletal Muscles.

Skeletal muscles undergo atrophy in response to diseases and aging. Here we report that mitofusin 2 (Mfn2) acts as a dominant suppressor of neuromuscular synaptic loss to preserve skeletal muscles. Mfn2 is reduced in spinal cords of transgenic SOD1G93A and aged mice. Through preserving neuromuscular synapses, increasing neuronal Mfn2 prevents skeletal muscle wasting in both SOD1G93A and aged mice, whereas deletion of neuronal Mfn2 produces neuromuscular synaptic dysfunction and skeletal muscle atrophy. Neuromuscular synaptic loss after sciatic nerve transection can also be alleviated by Mfn2. Mfn2 coexists with calpastatin largely in mitochondria-associated membranes (MAMs) to regulate its axonal transport. Genetic inactivation of calpastatin abolishes Mfn2-mediated protection of neuromuscular synapses. Our results suggest that, as a potential key component of a novel and heretofore unrecognized mechanism of cytoplasmic protein transport, Mfn2 may play a general role in preserving neuromuscular synapses and serve as a common therapeutic target for skeletal muscle atrophy.

Rab10 Phosphorylation is a Prominent Pathological Feature in Alzheimer's Disease.

Alzheimer's disease (AD) is the leading cause of dementia in the elderly, characterized by neurofibrillary tangles (NFTs), senile plaques (SPs), and a progressive loss of neuronal cells in selective brain regions. Rab10, a small Rab GTPase involved in vesicular trafficking, has recently been identified as a novel protein associated with AD. Interestingly, Rab10 is a key substrate of leucine-rich repeat kinase 2 (LRRK2), a serine/threonine protein kinase genetically associated with the second most common neurodegenerative disease Parkinson's disease. However, the phosphorylation state of Rab10 has not yet been investigated in AD. Here, using a specific antibody recognizing LRRK2-mediated Rab10 phosphorylation at the amino acid residue threonine 73 (pRab10-T73), we performed immunocytochemical analysis of pRab10-T73 in hippocampal tissues of patients with AD. pRab10-T73 was prominent in NFTs in neurons within the hippocampus in all cases of AD examined, whereas immunoreactivity was very faint in control cases. Other characteristic AD pathological structures including granulovacuolar degeneration, dystrophic neurites and neuropil threads also contained pRab10-T73. The pRab10-T73 immunoreactivity was diminished greatly following dephosphorylation with alkaline phosphatase. pRab10-T73 was further found to be highly co-localized with hyperphosphorylated tau (pTau) in AD, and demonstrated similar pathological patterns as pTau in Down syndrome and progressive supranuclear palsy. Although pRab10-T73 immunoreactivity could be noted in dystrophic neurites surrounding SPs, SPs were largely negative for pRab10-T73. These findings indicate that Rab10 phosphorylation could be responsible for aberrations in the vesicle trafficking observed in AD leading to neurodegeneration.

Mfn2 ablation causes an oxidative stress response and eventual neuronal death in the hippocampus and cortex.

BACKGROUND: Mitochondria are the organelles responsible for energy metabolism and have a direct impact on neuronal function and survival. Mitochondrial abnormalities have been well characterized in Alzheimer Disease (AD). It is believed that mitochondrial fragmentation, due to impaired fission and fusion balance, likely causes mitochondrial dysfunction that underlies many aspects of neurodegenerative changes in AD. Mitochondrial fission and fusion proteins play a major role in maintaining the health and function of these important organelles. Mitofusion 2 (Mfn2) is one such protein that regulates mitochondrial fusion in which mutations lead to the neurological disease. METHODS: To examine whether and how impaired mitochondrial fission/fusion balance causes neurodegeneration in AD, we developed a transgenic mouse model using the CAMKII promoter to knockout neuronal Mfn2 in the hippocampus and cortex, areas significantly affected in AD. RESULTS: Electron micrographs of neurons from these mice show swollen mitochondria with cristae damage and mitochondria membrane abnormalities. Over time the Mfn2 cKO model demonstrates a progression of neurodegeneration via mitochondrial morphological changes, oxidative stress response, inflammatory changes, and loss of MAP2 in dendrites, leading to severe and selective neuronal death. In this model, hippocampal CA1 neurons were affected earlier and resulted in nearly total loss, while in the cortex, progressive neuronal death was associated with decreased cortical size. CONCLUSIONS: Overall, our findings indicate that impaired mitochondrial fission and fusion balance can cause many of the neurodegenerative changes and eventual neuron loss that characterize AD in the hippocampus and cortex which makes it a potential target for treatment strategies for AD.

Inhibition of mitochondrial fragmentation protects against Alzheimer's disease in rodent model.

Mitochondrial dysfunction is an early prominent feature in susceptible neurons in the brain of patients with Alzheimer's disease, which likely plays a critical role in the pathogenesis of disease. Increasing evidence suggests abnormal mitochondrial dynamics as important underlying mechanisms. In this study, we characterized marked mitochondrial fragmentation and abnormal mitochondrial distribution in the pyramidal neurons along with mitochondrial dysfunction in the brain of Alzheimer's disease mouse model CRND8 as early as 3 months of age before the accumulation of amyloid pathology. To establish the pathogenic significance of these abnormalities, we inhibited mitochondrial fragmentation by the treatment of mitochondrial division inhibitor 1 (mdivi-1), a mitochondrial fission inhibitor. Mdivi-1 treatment could rescue both mitochondrial fragmentation and distribution deficits and improve mitochondrial function in the CRND8 neurons both in vitro and in vivo. More importantly, the amelioration of mitochondrial dynamic deficits by mdivi-1 treatment markedly decreased extracellular amyloid deposition and Aß1-42/Aß1-40 ratio, prevented the development of cognitive deficits in Y-maze test and improved synaptic parameters. Our findings support the notion that abnormal mitochondrial dynamics plays an early and causal role in mitochondrial dysfunction and Alzheimer's disease-related pathological and cognitive impairments in vivo and indicate the potential value of restoration of mitochondrial dynamics as an innovative therapeutic strategy for Alzheimer's disease.

TMEM230 Accumulation in Granulovacuolar Degeneration Bodies and Dystrophic Neurites of Alzheimer's Disease.

Transmembrane Protein 230 (TMEM230) is a newly identified protein associated with Parkinson's disease (PD) found in Lewy bodies and Lewy neurites of patients with PD or dementia with Lewy body disease. However, TMEM230 has not yet been investigated in the most common neurodegenerative disorder, Alzheimer's disease (AD). Here, we demonstrate that the expression of TMEM230 is specifically increased in neurons in AD patients. Importantly, both granulovacuolar degeneration (GVD) and dystrophic neurites (DNs), two prominent characteristic pathological structures associated with AD, contain TMEM230 aggregates. TMEM230 immunoreactivity can be detected in neurofibrillary tangles-containing neurons and hyperphosphorylated tau positive DNs. TMEM230 accumulation is also noted around senile plaques. These findings identifying TMEM230 as a component of GVD and DNs suggest TMEM230 dysregulation as a likely mechanism playing an important role in the pathogenesis of AD.

Mfn2 protects dopaminergic neurons exposed to paraquat both in vitro and in vivo: Implications for idiopathic Parkinson's disease.

Mitochondrial dynamics and quality control play a critical role in the maintenance of mitochondrial homeostasis and function. Pathogenic mutations of many genes associated with familial Parkinson's disease (PD) caused abnormal mitochondrial dynamics, suggesting a likely involvement of disturbed mitochondrial fission/fusion in the pathogenesis of PD. In this study, we focused on the potential role of mitochondrial fission/fusion in idiopathic PD patients and in neuronal cells and animals exposed to paraquat (PQ), a commonly used herbicide and PD-related neurotoxin, as models for idiopathic PD. Significantly increased expression of dynamin-like protein 1 (DLP1) and a trend towards reduced expression of Mfn1 and Mfn2 were noted in the substantia nigra tissues from idiopathic PD cases. Interestingly, PQ treatment led to similar changes in the expression of fission/fusion proteins both in vitro and in vivo which was accompanied by extensive mitochondrial fragmentation and mitochondrial dysfunction. Blockage of PQ-induced mitochondrial fragmentation by Mfn2 overexpression protected neurons against PQ-induced mitochondrial dysfunction in vitro. More importantly, PQ-induced oxidative damage and stress signaling as well as selective loss of dopaminergic (DA) neurons in the substantia nigra and axonal terminals in striatum was also inhibited in transgenic mice overexpressing hMfn2. Overall, our study demonstrated that disturbed mitochondrial dynamics mediates PQ-induced mitochondrial dysfunction and neurotoxicity both in vitro and in vivo and is also likely involved in the pathogenesis of idiopathic PD which make them a promising therapeutic target for PD treatment.

Motor-Coordinative and Cognitive Dysfunction Caused by Mutant TDP-43 Could Be Reversed by Inhibiting Its Mitochondrial Localization.

Dominant missense mutations in TAR DNA-binding protein 43 (TDP-43) cause amyotrophic lateral sclerosis (ALS), and the cytoplasmic accumulation of TDP-43 represents a pathological hallmark in ALS and frontotemporal lobar degeneration (FTD). Behavioral investigation of the transgenic mouse model expressing the disease-causing human TDP-43 M337V mutant (TDP-43M337V mice) is encumbered by premature death in homozygous transgenic mice and a reported lack of phenotype assessed by tail elevation and footprint in hemizygous transgenic mice. Here, using a battery of motor-coordinative and cognitive tests, we report robust motor-coordinative and cognitive deficits in hemizygous TDP-43M337V mice by 8 months of age. After 12 months of age, cortical neurons are significantly affected by the mild expression of mutant TDP-43, characterized by cytoplasmic TDP-43 mislocalization, mitochondrial dysfunction, and neuronal loss. Compared with age-matched non-transgenic mice, TDP-43M337V mice demonstrate a similar expression of total TDP-43 but higher levels of TDP-43 in mitochondria. Interestingly, a TDP-43 mitochondrial localization inhibitory peptide abolishes cytoplasmic TDP-43 accumulation, restores mitochondrial function, prevents neuronal loss, and alleviates motor-coordinative and cognitive deficits in adult hemizygous TDP-43M337V mice. Thus, this study suggests hemizygous TDP-43M337V mice as a useful animal model to study TDP-43 toxicity and further consolidates mitochondrial TDP-43 as a novel therapeutic target for TDP-43-linked neurodegenerative diseases.

Clinical and imaging characteristics of late onset mitochondrial membrane protein-associated neurodegeneration (MPAN).

Young onset dementias present significant diagnostic challenges. We present the case of a 35-year-old Kuwaiti man with social withdrawal, drowsiness, irritability, anxiety, aphasia, memory loss, hypereflexia, and Parkinsonism. Brain MRI showed bilateral symmetric gradient echo hypointensities in the globi pallidi and substantiae nigrae. Left cortical hypometabolism was seen on brain fluorodeoxyglucose positron emission tomography. A cortical brain biopsy revealed a high Lewy body burden. Genetic testing revealed a homozygous p.T11M mutation in the C19orf12 gene consistent with mitochondrial membrane protein-associated neurodegeneration. This is the oldest onset age of MPAN reported.

Correction: Individual Case Analysis of Postmortem Interval Time on Brain Tissue Preservation.

[This corrects the article DOI: 10.1371/journal.pone.0151615.].

The inhibition of TDP-43 mitochondrial localization blocks its neuronal toxicity.

Genetic mutations in TAR DNA-binding protein 43 (TARDBP, also known as TDP-43) cause amyotrophic lateral sclerosis (ALS), and an increase in the presence of TDP-43 (encoded by TARDBP) in the cytoplasm is a prominent histopathological feature of degenerating neurons in various neurodegenerative diseases. However, the molecular mechanisms by which TDP-43 contributes to ALS pathophysiology remain elusive. Here we have found that TDP-43 accumulates in the mitochondria of neurons in subjects with ALS or frontotemporal dementia (FTD). Disease-associated mutations increase TDP-43 mitochondrial localization. In mitochondria, wild-type (WT) and mutant TDP-43 preferentially bind mitochondria-transcribed messenger RNAs (mRNAs) encoding respiratory complex I subunits ND3 and ND6, impair their expression and specifically cause complex I disassembly. The suppression of TDP-43 mitochondrial localization abolishes WT and mutant TDP-43-induced mitochondrial dysfunction and neuronal loss, and improves phenotypes of transgenic mutant TDP-43 mice. Thus, our studies link TDP-43 toxicity directly to mitochondrial bioenergetics and propose the targeting of TDP-43 mitochondrial localization as a promising therapeutic approach for neurodegeneration.

Upregulation of Glutaredoxin-1 Activates Microglia and Promotes Neurodegeneration: Implications for Parkinson's Disease.

AIMS: Neuroinflammation and redox dysfunction are recognized factors in Parkinson's disease (PD) pathogenesis, and diabetes is implicated as a potentially predisposing condition. Remarkably, upregulation of glutaredoxin-1 (Grx1) is implicated in regulation of inflammatory responses in various disease contexts, including diabetes. In this study, we investigated the potential impact of Grx1 upregulation in the central nervous system on dopaminergic (DA) viability. RESULTS: Increased GLRX copy number in PD patients was associated with earlier PD onset, and Grx1 levels correlated with levels of proinflammatory tumor necrosis factor-alpha (TNF-α) in mouse and human brain samples, prompting mechanistic in vitro studies. Grx1 content/activity in microglia was upregulated by lipopolysaccharide (LPS), or TNF-α, treatment. Adenoviral overexpression of Grx1, matching the extent of induction by LPS, increased microglial activation; Grx1 silencing diminished activation. Selective inhibitors/probes of nuclear factor κB (NF-κB) activation revealed glrx1 induction to be mediated by the Nurr1/NF-κB axis. Upregulation of Grx1 in microglia corresponded to increased death of neuronal cells in coculture. With a mouse diabetes model of diet-induced insulin resistance, we found upregulation of Grx1 in brain was associated with DA loss (decreased tyrosine hydroxylase [TH]; diminished TH-positive striatal axonal terminals); these effects were not seen with Grx1-knockout mice. INNOVATION: Our results indicate that Grx1 upregulation promotes neuroinflammation and consequent neuronal cell death in vitro, and synergizes with proinflammatory insults to promote DA loss in vivo. Our findings also suggest a genetic link between elevated Grx1 and PD development. CONCLUSION: In vitro and in vivo data suggest Grx1 upregulation promotes neurotoxic neuroinflammation, potentially contributing to PD. Antioxid. Redox Signal. 25, 967-982.

Individual Case Analysis of Postmortem Interval Time on Brain Tissue Preservation.

At autopsy, the time that has elapsed since the time of death is routinely documented and noted as the postmortem interval (PMI). The PMI of human tissue samples is a parameter often reported in research studies and comparable PMI is preferred when comparing different populations, i.e., disease versus control patients. In theory, a short PMI may alleviate non-experimental protein denaturation, enzyme activity, and other chemical changes such as the pH, which could affect protein and nucleic acid integrity. Previous studies have compared PMI en masse by looking at many different individual cases each with one unique PMI, which may be affected by individual variance. To overcome this obstacle, in this study human hippocampal segments from the same individuals were sampled at different time points after autopsy creating a series of PMIs for each case. Frozen and fixed tissue was then examined by Western blot, RT-PCR, and immunohistochemistry to evaluate the effect of extended PMI on proteins, nucleic acids, and tissue morphology. In our results, immunostaining profiles for most proteins remained unchanged even after PMI of over 50 h, yet by Western blot distinctive degradation patterns were observed in different protein species. Finally, RNA integrity was lower after extended PMI; however, RNA preservation was variable among cases suggesting antemortem factors may play a larger role than PMI in protein and nucleic acid integrity.