Biology subjects

Cousin, L.

Publications and source records attributed to Cousin, L..

4 recordsLinked to original sources

bioRxiv2024

Morphological profiling in human dopaminergic neurons identifies mitochondrial uncoupling as a neuroprotective effect

Parkinsons disease (PD) involves multiple pathological processes in midbrain dopaminergic (mDA) neurons, including protein degradation defects, vesicular trafficking disruption, endolysosomal dysfunction, mitochondrial issues, and oxidative stress. Current PD models often lack complexity and focus on single phenotypes. We used patient-derived SNCA triplication (SNCA-4x) and isogenic control (SNCA-corr) mDA neurons, applying high-content imaging-based morphological profiling to identify and rescue multiple phenotypes. Screening 1,020 compounds, we identified top-scoring compounds that restored healthy profiles in SNCA-4x neurons, increasing Tyrosine hydroxylase (TH) and decreasing -synuclein (Syn) levels. Several hits were linked to mitochondrial biology. Tyrphostin A9, a mitochondrial uncoupler, and several of its structural analogues decreased ROS levels, normalized mitochondrial membrane potential, and increased respiration. Western blotting confirmed that Tyrphostin A9 reduces Syn levels. Our study highlights the neuroprotective potential of mild mitochondrial uncoupling in mDA neurons.

neuroscience↗

bioRxiv2024

Tunable bicontinuous macroporous cell culture scaffolds via kinetically controlled phase separation

Three-dimensional (3D) scaffolds enable biological investigations with a more natural cell conformation. However, the porosity of synthetic hydrogels is often limited to the nanometer scale, which confines the movement of 3D encapsulated cells and restricts dynamic cell processes. Precise control of hydrogel porosity across length scales remains a challenge and the development of porous materials that allow cell infiltration, spreading, and migration in a manner more similar to natural ECM environments is desirable. Here, we present a straightforward and reliable method for generating kinetically-controlled macroporous systems using liquid-liquid phase separation between poly(ethylene glycol) (PEG) and dextran. Photopolymerization-induced phase separation resulted in macroporous hydrogels with tunable pore size. Varying light intensity and hydrogel composition controlled polymerization kinetics, time to percolation, and complete gelation, which defined the average pore diameter (O = 1- 300 m) and final gel stiffness of the formed hydrogels. Critically, for biological applications, macroporous hydrogels were prepared from aqueous polymer solutions at physiological pH and temperature using visible light, allowing for direct cell encapsulation. We encapsulated human dermal fibroblasts in a range of macroporous gels with different pore sizes. Porosity improved cell spreading with respect to bulk gels and allowed migration in the porous systems.

bioengineering↗

bioRxiv2023

Pharmacological modulation of developmental and synaptic phenotypes in human SHANK3 deficient stem cell-derived neuronal models

One sentence summaryThis study describes the use of SHANK3 deficient stem cell-derived neuronal models to screen and characterize small molecules that partially rescued developmental and synaptic defects related to Phelan-McDermid syndrome (PMDS). Phelan-McDermid syndrome (PMDS) arises from mutations in the terminal region of chromosome 22q13, impacting the SHANK3 gene. The resulting deficiency of the postsynaptic density scaffolding protein SHANK3 is associated with autism spectrum disorder (ASD). We examined 12 different PMDS patient and CRISPR-engineered stem cell-derived neuronal models and controls and found that reduced expression of SHANK3 leads to neuronal hyperdifferentiation, increased synapse formation, and decreased neuronal activity. We performed automated imaging-based screening of 7,120 target-annotated small molecules and identified three compounds that rescued SHANK3-dependent neuronal hyperdifferentiation. One compound, Benproperine, rescued the decreased colocalization of Actin Related Protein 2/3 Complex Subunit 2 (ARPC2) with {beta}-actin and rescued increased synapse formation in SHANK3 deficient neurons when administered early during differentiation. Neuronal activity was only mildly affected, highlighting Benproperines effects as a neurodevelopmental modulator. This study demonstrates that small molecular compounds that reverse developmental phenotypes can be identified in human neuronal PMDS models.

neuroscience↗

bioRxiv2022

Machine learning-aided multidimensional phenotyping of Parkinson's disease patient stem cell-derived midbrain dopaminergic neurons

Combining multiple Parkinsons disease (PD) relevant cellular phenotypes might increase the accuracy of midbrain dopaminergic (mDA) in vitro models. We differentiated patient-derived induced pluripotent stem cells (iPSCs) with a LRRK2 G2019S mutation, isogenic control and genetically unrelated iPSCs into mDA neurons. Using automated fluorescence microscopy in 384-well plate format, we identified elevated levels of -synuclein and Serine 129 phosphorylation (pS129), reduced dendritic complexity, and mitochondrial dysfunction. Next, we measured additional image-based phenotypes and used machine learning (ML) to accurately classify mDA neurons according to their genotype. Additionally, we show that chemical compound treatments, targeting LRRK2 kinase activity or -synuclein levels, are detectable when using ML classification based on multiple image-based phenotypes. We validated our approach using a second isogenic patient derived SNCA gene triplication mDA neuronal model. This phenotyping and classification strategy improves the exploitability of mDA neurons for disease modelling and the identification of novel PD drug targets.

neuroscience↗