LRRK2-G2019S Synergizes with Ageing and Low-Grade Inflammation to Promote Gut and Peripheral Immune Cell Activation that Precede Nigrostriatal Degeneration
By onBackground Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most frequent cause of familial Parkinson’s disease (PD). The incomplete penetrance of LRRK2 mutations suggest that additional hits are required for disease onset. We hypothesized that chronic low-grade inflammation interacts with LRRK2 G2019S, the most frequent PD-associated mutation, to activate peripheral and central immune reactions and drive age-dependent neurodegeneration. Methods and Results We exposed wild-type and LRRK2 G2019S mice to a low chronic dose of lipopolysaccharide, and we performed a longitudinal analysis of central and peripheral immune reactions and neurodegeneration. Low-dose inflammation triggered nigrostriatal degeneration, macrophage/monocyte brain infiltration, and astro-/microgliosis. LRRK2 G2019S mice showed an early dysregulation of peripheral cytokines, increased CD4+ T-cell infiltration and α-synuclein aggregation in the colon. Interestingly, peripheral immune activation and colonic α-synuclein aggregation precede astro-/microgliosis and neurodegeneration. Conclusions Our study suggests an early role of the peripheral immune system and the gut in LRRK2 PD and provides a novel model to study early therapeutic immune targets and biomarkers.
Preparing MEF-cultured hPSCs for nucleofection
By onThis protocol describes the procedure or preparing MEF-cultured human pluripotent stem cells (hPSCs) for the delivery of plasmids, mRNA, or ribonucleoprotein (RNP) using nucleofection.
H9 ES AAVS1-NGN2 CCPG1-/-; PiggyBac-Keima-REEP5
By onES cells were modified using CRISPR/Cas9 to lack CCPG1 and express Keima-REEP5 ER-phagy flux reporter. NEUROG2 construct was introduced in AAVS1 locus. Cells are embryonic stem cells derived from a human female blastocyst stage.
Molecular cloning of SHIP164 plasmids for expression in mammalian cells
By onThis protocol is to help with the molecular cloning of SHIP164 and the sequences of other proteins.
Primary data associated with the manuscript “Whole proteome copy number dataset in primary mouse cortical neurons” (doi: 10.1016/j.dib.2023.109336)
By onRaw data reported in Supplementary table in "Whole Proteome Copy Number Dataset in Primary Mouse Cortical Neurons."
Lentivirus production
By onThis protocol is associated with the following preprint, published on February 19, 2022: “The AAA+ chaperone VCP disaggregates Tau fibrils and generates aggregate seeds.”
Striatal dopamine measurement through HPLC
By onProtocol for striatal dopamine measurement in mouse brain
Gene editing of YIPF4 in hESCs V3
By onThis protocol describes the creation of YIPF4 knockout cell lines in H9 hESC cells using CRISPR-Cas9. Associated with preprint: https://doi.org/10.1101/2022.12.06.519342
Live imaging to investigate mitophagy kinetics and NEMO recruitment in HeLa-M cells
By onProtocol for live imaging to investigate mitophagy kinetics and NEMO recruitment in HeLa-M cells.
Microscopy-based bead protein-protein interaction assay
By onAn assay to study protein-protein interaction
Dopaminergic axons track somatic signaling in behaving mice
By onHere, authors used genetic strategies to isolate key dopaminergic neuron subtypes and monitor their axonal and somatic signaling patterns in behaving mice.
Reconstitution of cargo-induced LC3 lipidation in mammalian selective autophagy
By onSelective autophagy is essential for maintaining cellular homeostasis. Using in vitro reconstitution, the authors explored the details of mitophagy initiation from autophagy receptor engagement through LC3 lipidation.
Primary data associated with the manuscript “CURTAIN – A Unique Web-based tool for exploration and sharing of MS-based proteomics data” (doi: https://doi.org/10.1101/2023.07.25.550405)
By onRaw immunoblotting data and their quantitation and graphs obtained using GraphPad Prism; PCR and sequencing data confirming that PPM1H-BromoTag clone E5 (used for all experiments in the manuscript) is a homozygous clone.
Raw tomogram of a TauRD-YFP aggregate in a TauRD-YFP expressing HEK293T cells and primary mouse neurons.
By onRaw tomogram of a TauRD-YFP aggregate in a TauRD-YFP expressing Hek293T cells. Accession numbers: HEK293T: https://www.ebi.ac.uk/emdb/EMD-13739 Primary mouse neurons: https://www.ebi.ac.uk/emdb/EMD-13740
CRISPResso analysis
By onThis protocol describes the process of using CRISPResso to analyze Next Generation Sequencing (NGS) data from CRISPR-targeted, genome-edited samples.
Stereotaxic Injections
By onStereotaxic injections for Conserved and cell type-specific transcriptional responses to IFN-γ in the ventral midbrain.
