Maintenance and inactivation of mouse embryonic fibroblasts (MEFs) as feeder cells for human pluripotent stem cell culture
By onThis collection describes the maintenance and inactivation of mouse embryonic fibroblasts (MEFs) as feeder cells for human pluripotent stem cell (hPSC) culture.
Therapeutic Potential of PTB Inhibition Through Converting Glial Cells to Neurons in the Brain
By onCell replacement therapy represents a promising approach for treating neurodegenerative diseases. Contrary to the common addition strategy to generate new neurons from glia by overexpressing a lineage-specific transcription factor(s), a recent study introduced a subtraction strategy by depleting a single RNA-binding protein, Ptbp1, to convert astroglia to neurons not only in vitro but also in the brain. Given its simplicity, multiple groups have attempted to validate and extend this attractive approach but have met with difficulty in lineage tracing newly induced neurons from mature astrocytes, raising the possibility of neuronal leakage as an alternative explanation for apparent astrocyte-to-neuron conversion. This review focuses on the debate over this critical issue. Importantly, multiple lines of evidence suggest that Ptbp1 depletion can convert a selective subpopulation of glial cells into neurons and, via this and other mechanisms, reverse deficits in a Parkinson's disease model, emphasizing the importance of future efforts in exploring this therapeutic strategy.
Sectioning of Mouse Brain by Cryostat
By onThis protocol describes how to use the cryostat to prepare and slice mouse brain sections for Immunohistochemistry.
H9 ES AAVS1-NGN2 FAM134C/A/B-/-
By onES cells modified using CRISPR/Cas9 to lack ER-phagy receptor FAM134C, A & B. Introduced NEUROG2 gene in AAVS1 locus. Derived from human embryonic stem cells at blastocyst stage.
In vivo reduction of age-dependent neuromelanin accumulation mitigates features of Parkinson’s disease
By onHumanized rodents with neuromelanin can develop Parkinson's symptoms. Reducing neuromelanin accumulation mitigates PD features, showing a link between NM levels and disease pathology.
Assessing enrichment of proteins in the mitochondrial fraction in HEK cells
By onThis is a method for measuring protein enrichment on mitochondria in various conditions. In the resulting Western blot, one can assess the level of contamination of other organelles in the enrichment prep.
The impact of MG149, a KAT8 inhibitor, on mitophagy: cell-based in vitro assays
By onProtocol outlines in vitro assays (TMRM, mitoKeima) for evaluating MG149 and KAT8 inhibition effects on mitophagy.
The calcium sensor synaptotagmin-1 is critical for phasic axonal dopamine release in the striatum and mesencephalon, but is dispensable for basic motor behaviors in mice
By onIn this work, we conditionally deleted the calcium sensor synaptotagmin-1 (Syt1) in DA neurons (cKODA mice) to abrogate most activity-dependent axonal DA release in the striatum and mesencephalon. Syt1 cKODA mice showed intact performance in multiple unconditioned DA-dependent motor tasks, suggesting that activity-dependent DA release is dispensable for such basic motor functions. Basal extracellular levels of DA in the striatum were unchanged, suggesting that a basal tone of extracellular DA is sufficient to sustain basic movement. We also found multiple adaptations in the DA system of cKODA mice, similar to those happening at early stages of PD. Taken together, our findings reveal the striking resilience of DA-dependent motor functions in the context of a near-abolition of phasic DA release, shedding new light on why extensive loss of DA innervation is required to reveal motor dysfunctions in PD.
Non-ablative disease-modifying effects of magnetic resonance-guided focused ultrasound in neuromelanin-producing parkinsonian rodents
By onThe authors' findings indicate that tFUS treatment applied at prodromal/early disease stages provides extended structural and functional preservation of the nigrostriatal pathway in neuromelanin-producing parkinsonian rats.
AAV Purification Protocol with Iodixanol gradient
By onProtocol used in the Kaplitt and Marongiu labs to purify AAVs.
Fluorescent Immunolabelling for Alpha-Synuclein in neuronal primary culture (Testing PFF toxicity)
By onThis protocol is designed to perform fluorescent immunolabelling on neuronal primary culture after PFF incubation. The labelling of phosphorylated alpha-synuclein is considered as a marker of the PFF toxicity and should be done to validate the PFFs.
Nucleofection of hPSCs
By onThis protocol describes the standard procedure for the delivery of plasmids, mRNA, or ribonucleoprotein (RNP) into human pluripotent stem cells (hPSCs) using nucleofection.
Thawing of mouse embryonic fibroblasts (MEFs) for hPSC cultures
By onThis protocol describes the thawing of mouse embryonic fibroblasts (MEFs) as feeder cells for human pluripotent stem cell (hPSC) culture.
Vibrational Stabilization of Cluster Synchronization in Oscillator Networks
By onRestoring normal cluster synchronization is crucial for system functioning. Vibrational control can stabilize synchronization without state measurements, offering a solution for challenging scenarios.
Statistical analysis
By onThis protocol describes the statistical analysis applied to the quantifications in Chang et al. 2021 SA paper.
Inter-organellar Communication in Parkinson’s and Alzheimer’s Disease: Looking Beyond Endoplasmic Reticulum-Mitochondria Contact Sites
By onNeurodegenerative diseases (NDs) are generally considered proteinopathies but whereas this may initiate disease in familial cases, onset in sporadic diseases may originate from a gradually disrupted organellar homeostasis. Herein, endolysosomal abnormalities, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, and altered lipid metabolism are commonly observed in early preclinical stages of major NDs, including Parkinson's disease (PD) and Alzheimer's disease (AD). Among the multitude of underlying defective molecular mechanisms that have been suggested in the past decades, dysregulation of inter-organellar communication through the so-called membrane contact sites (MCSs) is becoming increasingly apparent. Although MCSs exist between almost every other type of subcellular organelle, to date, most focus has been put on defective communication between the ER and mitochondria in NDs, given these compartments are critical in neuronal survival. Contributions of other MCSs, notably those with endolysosomes and lipid droplets are emerging, supported as well by genetic studies, identifying genes functionally involved in lysosomal homeostasis. In this review, we summarize the molecular identity of the organelle interactome in yeast and mammalian cells, and critically evaluate the evidence supporting the contribution of disturbed MCSs to the general disrupted inter-organellar homeostasis in NDs, taking PD and AD as major examples.
