Biology subjects

Itzhaki, E.

Publications and source records attributed to Itzhaki, E..

3 recordsLinked to original sources

bioRxiv2026

CytoLight: A Rapid and Versatile Fluorescent-Based Labeling Method for Extracellular Vesicle Characterization and Tracking

Efficient, aggregation-free extracellular vesicles (EVs) labeling is essential for studying their dynamics in-vitro and in-vivo. However, traditional dyes introduce limitations including aggregation, membrane intercalation, fluorescence transfer and inconsistent performance across EV sources thus distorting quantification, altering surface properties and confounding uptake and biodistribution analyses. Here, we systematically evaluated CytoLight, a luminal dye traditionally used for live-cell imaging, as an alternative for EV quantification, characterization, uptake analysis and in-vivo tracking, benchmarking it against PKH26, CFSE and ExoBrite across multiple platforms. CytoLight generated stable, intravesicular fluorescence without aggregation or membrane alteration, eliminating artifacts characteristic of conventional dyes. Using fluorescence-NTA and single-EV flow cytometry, CytoLight showed more consistent labeling across EV types than CFSE or ExoBrite, while avoiding PKH-related micelle-driven artifacts and exhibited compatibility with CD81 dual-detection. In uptake assays, CytoLight produced EV-specific endocytosis-dependent internalization signals exceeding labeled-BPS/protein controls. In-vivo, CytoLight-labeled EVs enabled fluorescent biodistribution mapping showing conventional EV tropism patterns distinguishable from labeled-PBS controls. These findings establish CytoLight as an effective, aggregation-free EV-labeling strategy. Its stability, specificity, compatibility with single-EV platforms and reliable performance in both cellular uptake and biodistribution studies position CytoLight as a practical, scalable alternative to current dyes, providing a stronger foundation for standardized and reproducible EV research.

cell biology↗

bioRxiv2023

Lowering mutant huntingtin by small molecules relieves Huntington's disease symptoms and progression

Huntingtons disease (HD) is an incurable inherited disorder caused by repeat expansion in the huntingtin gene (Htt). The mutant protein causes neuronal degeneration leading to severe motor and psychological abnormalities. Selective downregulation of the mutant Htt expression is considered the leading therapeutic approach for HD. We report the identification of novel small molecule inhibitors of Spt5-Pol II, SPI-24 and SPI-77, which selectively lower mutant Htt mRNA and protein levels in HD cells. In the BACHD mouse model, their direct delivery to the striatum diminished mutant Htt levels, ameliorated mitochondrial dysfunction, restored BDNF expression and improved motor and anxious-like phenotypes. Pharmacokinetic studies revealed that these SPIs pass the blood-brain-barrier and prolonged subcutaneous injection or oral administration to early-stage mice significantly delayed disease deterioration. SPI-24 long-term treatment had no side effects or global changes in gene expression. Thus, lowering mutant Htt levels by small molecules can be an effective therapeutic strategy for HD.

molecular biology↗

bioRxiv2019

Point mutations in topoisomerase I alter the mutation spectrum in E. coli and impact the emergence of drug resistance genotypes

Identifying the molecular mechanisms that give rise to genetic variation is essential for the understanding of evolutionary processes. Previously, we have used adaptive laboratory evolution to enable biomass synthesis from CO2 in E. coli. Genetic analysis of adapted clones from two independently evolving populations revealed distinct enrichment for insertion and deletion mutational events. Here, we follow these observations to show that mutations in the gene encoding for DNA Topoisomerase 1 (topA) give rise to mutator phenotypes with characteristic mutational spectra. Using genetic assays and mutation accumulation lines, we show that point mutations in topA increase the rate of sequence deletion and duplication events. Interestingly, we observe that a single residue substitution (R168C) results in a high rate of head-to-tail (tandem) short sequence duplications, which are independent of existing sequence repeats. Finally, we show that the unique mutation spectrum of topA mutants enhances the emergence of antibiotic resistance in comparison to mismatch-repair (mutS) mutators, and lead to new resistance genotypes. Our findings highlight a potential link between the catalytic activity of topoisomerases and the fundamental question regarding the emergence of de novo tandem repeats, which are known modulators of bacterial evolution.

microbiology↗