Journal: bioRxiv

Article Title: Senescent cell heterogeneity and responses to senolytic treatment are related to cell cycle status during cell growth arrest

doi: 10.1101/2024.06.22.600200

Figure Lengend Snippet: High-content imaging workflow . a ) Sample preparation. Human lung primary microvascular endothelial cells (HMVEC-L) and fibroblasts (IMR-90) were induced to senescence using ionizing radiation (IR). IR and mock-irradiated cells (CTL) were either cultured in full-serum medium the entire time (FS) or switched to low-serum medium for the last 3 days of culture to induce quiescence in CTL cells (SS). b ) Staining for senescence markers. Prepared samples were either co-stained for senescence-associated beta- galactosidase activity (SA-β-Gal) and proliferation via EdU incorporation (EdU); or for other senescence markers (γH2AX, LaminB1, HMGB1, p21) using immunocytochemistry (ICC). c ) High-content image analysis was performed to identify senescent subpopulations.

Article Snippet: Primary human lung microvascular endothelial cells (HMVEC-L) were purchased from Lonza (CC-2527).

Techniques: Imaging, Sample Prep, Irradiation, Cell Culture, Staining, Activity Assay, Immunocytochemistry

Journal: bioRxiv

Article Title: Senescent cell heterogeneity and responses to senolytic treatment are related to cell cycle status during cell growth arrest

doi: 10.1101/2024.06.22.600200

Figure Lengend Snippet: Validation of senescence induction and population-level heterogeneity . a ) Representative images of senescence marker staining from full-serum (FS) samples. Top: mock-irradiated cells (CTL); bottom: ionizing-radiation-induced senescent cells (IR). For each co-staining, endothelial cells (HMVEC-L) are shown on the left; fibroblasts (IMR-90) are shown on the right. b-d ) Image quantification of HMVEC-L and IMR-90 cells for both FS and serum- starved (SS) conditions. CTL samples are in green, while IR samples are in purple. Data shown are from 2 independent experiments. b ) SA-β-Gal (left) and EdU (right) staining quantification. Each data point corresponds to one well (n = 18); bars indicate mean values. c ) Immunocytochemistry staining quantification. Top-left: γH2AX; top-right: p21; bottom-left: LaminB1; bottom-right: HMGB1. Each data point corresponds to one well (γH2AX n = 27; p21, LaminB1, and HMGB1 n = 9); bars indicate mean values. d ) Nuclear morphology feature quantification. Left: nuclear area; right: shape factor. Each data point corresponds to one well (n = 27); bars indicate mean values. ***: p-value < 10 -3 ; ****: p-value < 10 -4 ; non-significant values (p-value > 0.05) are shown.

Article Snippet: Primary human lung microvascular endothelial cells (HMVEC-L) were purchased from Lonza (CC-2527).

Techniques: Marker, Staining, Irradiation, Immunocytochemistry