Age-Dependent Decline in Neuron Growth Potential and Mitochondria Functions in Cortical Neurons

Sutherland, Theresa C; Sefiani, Arthur; Horvat, Darijana; Huntington, Taylor E.; Lei, Yuanjiu; West, A. Phillip; Geoffroy, Cédric G.

doi:10.3390/cells10071625

Cited by 11 publications

(26 citation statements)

References 85 publications

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“…RT-qPCR assay was replicated as previously described 21 . Briefly, Directzol RNA micro-prep columns (Zymo, R2061) are used to extract RNA from neurons directly following neuron isolation.…”

Section: Methodsmentioning

confidence: 99%

“…Quantabio cDNA Synthesis kit (Quanta, 95047) was used to synthesize cDNA before conducting qPCR using the Quantabio PerfeCTa ® SYBR ® Green FastMix ® (Quanta, 95073) on the ViiA7 Real Time PCR system (Life Technologies). The neuron enrichment in the negative fraction was calculated as previously described 21 , −ΔCT of ΔNeuN against ΔGFAP and ΔGLAST using the formula: −ΔCT = −(ΔCT NeuN – (SQRT(ΔCT GFAP 2 + ΔCT GLAST 2 ))). CT was calculated for each group based on the absolute CT per primer subtracted by the respective CT of the negative control to reduce background noise.…”

Section: Methodsmentioning

confidence: 99%

“…Current therapeutic strategies in neurodegenerative diseases, such as glaucoma and Parkinson’s disease, aim at increasing axon regenerative capacity to mitigate pathology 19,20 . Aging itself reduces neurite regenerative capacity 21 and increases susceptibility of neurons to death and degeneration 22 which can produce worst outcomes after neurotrauma 23 and increases the prevalence of neurodegenerative diseases 24,25 . Therefore, it is imperative that novel therapeutic strategies can increase the survival and neurite regenerative capacity of neurons in older individuals as well.…”

Section: Introductionmentioning

confidence: 99%

“…Miltenyi recently developed a system to culture very young adult brain neurons (up to 4 weeks old) 46,48,49 . We modified this technique to culture middle- and advanced-age cortical neurons 21 . In the present report, we further adapted this protocol to screen for compounds that enhance survival and neurite outgrowth and determine the demographic the compounds have most efficacy in.…”

Section: Introductionmentioning

confidence: 99%

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Novel Adult Cortical Neuron Processing and Screening method illustrates Sex- and Age-dependent effects of pharmaceutical compounds

Geoffroy

Sefiani

Rusyn

2022

Preprint

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Neurodegenerative diseases and neurotraumatic injuries are typically age-associated disorders that can reduce neuron survival, neurite outgrowth, and synaptic plasticity leading to loss of cognitive capacity, executive function, and motor control. In pursuit of reducing the loss of said neurological functions, novel compounds are sought that promote neuron viability, neuritogenesis, and/or synaptic plasticity. Current high content in vitro screenings typically use cells that are iPSC-derived, embryonic, or originate from post-natal tissues; however, most patients suffering from neurodegenerative diseases and neurotrauma are of middle-age and older. The chasm in maturity between the neurons used in drug screens and those in a target population is a barrier for translational success of in vitro results. It has been historically challenging to culture adult neurons let alone conduct screenings; therefore, age-appropriate drug screenings have previously not been plausible. We have modified Miltenyi protocol to increase neuronal yield, neuron purity, and neural viability at a reduced cost to expand our capacity to screen compounds directly in primary adult neurons. Furthermore, we have developed the first morphology-based screening system using adult cortical neurons. By using primary adult cortical neurons from mice that were 4 to 48 weeks old for screening of a library of pharmaceuticals, we have demonstrated age- and sex-dependent effects on neuritogenesis and neuron survival in vitro. Utilizing age- and sex-appropriate in vitro models to find novel compounds increasing neuron survival and neurite outgrowth, made possible by our modified adult neuron processing method, will greatly increase the relevance of in vitro screening for finding neuroprotective compounds.

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“…RT-qPCR assay was replicated as previously described 21 . Briefly, Directzol RNA micro-prep columns (Zymo, R2061) are used to extract RNA from neurons directly following neuron isolation.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Novel Adult Cortical Neuron Processing and Screening method illustrates Sex- and Age-dependent effects of pharmaceutical compounds

Geoffroy

Sefiani

Rusyn

2022

Preprint

Self Cite

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“…A significant increase in ROS production has been observed in aged neurons, and there also seems to be an increase in the markers associated with neuronal senescence, including higher expression levels of p53 and the increased activation of phosphor-JNK and phosphor-p38 MAPK [ 100 , 104 ]. A decrease in neurite outgrowth in older neurons has been detected in primary cortical neurons isolated and cultured from adult mice [ 105 ]. Aged neurons exhibit respiratory dysfunction, decreased mitochondrial membrane potential, changes in mitochondrial membrane transport proteins, and mitochondrial calcium accumulation [ 106 ].…”

Section: The Signaling Pathways Associated With Neuronal Senescence and Brain Agingmentioning

confidence: 99%

Human iPSC-Derived Neurons as A Platform for Deciphering the Mechanisms behind Brain Aging

et al. 2021

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With an increased life expectancy among humans, aging has recently emerged as a major focus in biomedical research. The lack of in vitro aging models—especially for neurological disorders, where access to human brain tissues is limited—has hampered the progress in studies on human brain aging and various age-associated neurodegenerative diseases at the cellular and molecular level. In this review, we provide an overview of age-related changes in the transcriptome, in signaling pathways, and in relation to epigenetic factors that occur in senescent neurons. Moreover, we explore the current cell models used to study neuronal aging in vitro, including immortalized cell lines, primary neuronal culture, neurons directly converted from fibroblasts (Fib-iNs), and iPSC-derived neurons (iPSC-iNs); we also discuss the advantages and limitations of these models. In addition, the key phenotypes associated with cellular senescence that have been observed by these models are compared. Finally, we focus on the potential of combining human iPSC-iNs with genome editing technology in order to further our understanding of brain aging and neurodegenerative diseases, and discuss the future directions and challenges in the field.

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Selective, Intrinsically Fluorescent Trk Modulating Probes

Pewklang,

Thompson,

Sefiani

et al. 2024

ACS Chem. Neurosci.

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Neurotrophins (NTs) elicit the growth, survival, and differentiation of neurons and other neuroectoderm tissues via activation of Trk receptors. Hot spots for NT•Trk interactions involve three neurotrophin loops. Mimicry of these using "cyclo-organopeptides" comprising loop sequences cyclized onto endocyclic organic fragments accounts for a few of the low molecular mass Trk agonists or modulators reported so far; the majority are nonpeptidic small molecules accessed without molecular design and identified in random screens. It has proven difficult to verify activities induced by low molecular mass substances are due to Trk activation (rather than via other receptors), enhanced Trk expression, enhanced NT expression, or other pathways. Consequently, identification of selective probes for the various Trk receptors (e.g., A, B, and C) has been very challenging. Further, a key feature of probes for early stage assays is that they should be easily detectable, and none of the compounds reported to date are. In this work, we designed novel cyclo-organopeptide derivatives where the organic fragment is a BODIPY fluor and found ones that selectively, though not specifically, activate TrkA, B, or C. One of the assays used to reach this conclusion (binding to live Trkexpressing cells) relied on intrinsic fluorescence in the tested materials. Consequently, this work established low molecular mass Trkselective probes exhibiting neuroprotective effects.

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Age-Dependent Decline in Neuron Growth Potential and Mitochondria Functions in Cortical Neurons

Cited by 11 publications

References 85 publications

Novel Adult Cortical Neuron Processing and Screening method illustrates Sex- and Age-dependent effects of pharmaceutical compounds

Novel Adult Cortical Neuron Processing and Screening method illustrates Sex- and Age-dependent effects of pharmaceutical compounds

Human iPSC-Derived Neurons as A Platform for Deciphering the Mechanisms behind Brain Aging

Selective, Intrinsically Fluorescent Trk Modulating Probes

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