DYRK1B inhibition exerts senolytic effects on endothelial cells and rescues endothelial dysfunctions.

Aging is the major risk factor for chronic disease development. Cellular senescence is a key mechanism that triggers or contributes to age-related phenotypes and pathologies. The endothelium, a single layer of cells lining the inner surface of a blood vessel, is a critical interface between blood and all tissues. Many studies report a link between endothelial cell senescence, inflammation, and diabetic vascular diseases. Here we identify, using combined advanced AI and machine learning, the Dual Specificity Tyrosine Phosphorylation Regulated Kinase 1B (DYRK1B) protein as a possible senolytic target for senescent endothelial cells. We demonstrate that upon induction of senescence in vitro DYRK1B expression is increased in endothelial cells and localized at adherens junctions where it impairs their proper organization and functions. DYRK1B knock-down or inhibition restores endothelial barrier properties and collective behavior. DYRK1B is therefore a possible target to counteract diabetes-associated vascular diseases linked to endothelial cell senescence.

Evaluation of Chemo- and Photo-toxicity of a Live Fluorescent Dye for Cell Analysis.

Live cell imaging is used to track the dynamic adaptation of cell size and motility to various external factors. Bright-field configuration can be used for these experiments; however, the analysis can be challenging and difficult to automate. In this direction, a superior alternative is represented by the use of live cell dyes, which provide intense fluorescence from subcellular structures of living cells. Yet, the potential chemo- and photo-toxicity of the fluorophores poses the necessity of an accurate protocol optimization to avoid artefacts. Toxicity studies generally focus on cell proliferation and apoptosis, neglecting the cellular activities under investigation. Here, we present the case of SYTO 13 in combination with primary endothelial cells. The optimization of the staining procedure is tested comparing cell proliferation and motility rate. In addition, the combined effect of staining and fluorescent illumination, reporting for photochemical toxicity, is evaluated. We demonstrate that while cell viability and proliferation are mainly unaffected by the staining and imagining protocols, a significant reduction of the motility rate is induced both by the chemical dye alone and in combination with fluorescent illumination. The general implications for this procedure are discussed.