Journal: The EMBO Journal

Article Title: Cyclophilin D plays a critical role in the survival of senescent cells

doi: 10.1038/s44318-024-00259-2

Figure Lengend Snippet: ( A ) Immunodetection of CypD, p16, and p21 by western blots of total cell lysates separated by SDS–PAGE. The graphs show the densitometric quantification of the signals corresponding to each protein normalised to that of the β‐actin, from n = 3 independent biological replicates of IMR90 and A549 cells. ( B ) Relative cell viability (%) of proliferating (PRO, blue) or senescent (SEN, red) IMR90, A549, and SK-MEL cells after 7 days of treatment with anti-CypD siRNA, compared to the same cells treated with “scrambled” siRNA (siScrbl). Senescence was induced by treatment with X-ray irradiation (IMR90), or bleomycin (A549), or palbociclib (SK-MEL) for 7 days before the initiation of the treatments with siRNAs. Western blots show the residual levels of CypD in the three cellular cultures on the last day of the experiment. Lanes treated with siCypD are indicated as (+), lanes treated with siScrbl are indicated as (−). n = 3–5 independent experiments. ( C ) Relative cell viability (%) of proliferating (blue) or mTORi-quiescent (green) IMR90 and A549, after 7-days of treatment with anti-CypD siRNA, compared to the same cells treated with siRNA Scrbl. mTORi quiescence was induced with Spanisertib for 7 days before the initiation of the treatments with siRNAs. n = 4 independent experiments. ( D ) Survival curves obtained with increasing concentrations of Cyclosporin A (CSA) in IMR90, A549, SK-MEL, and LNCaP cells, proliferating (PRO) or senescent (SEN), compared to vehicle (DMSO). IC50 values (half maximal inhibitory concentration) and senolytic index (SI: ratio of IC50 values for senescent vs proliferative cells) are indicated for each cell type. Senescence was induced by treatment with ionizing irradiation (IMR90), bleomycin (A549), palbociclib (SK-MEL), or abemaciclib (LNCaP) for 7 days before the initiation of CSA treatment for 6 days. Viability of IMR90, A549, and SK-MEL cells was measured by CellTiter-Glo® Luminescent Cell Viability Assay. Viability of LNCaP cells was measured by Annexin V-BV421 and propidium iodide staining. n = 3-4 independent experiments. ( E ) Survival curves obtained with increasing concentrations of NIM811 in IMR90 and A549, proliferating or senescent, compared to vehicle (DMSO). Senescence was induced by treatment with doxorubicin (IMR90) or bleomycin (A549) for 7 days before the initiation of NIM811 treatment for 6 days. Viability was measured by CellTiter-Glo® Luminescent Cell Viability Assay. n = 3-4 independent experiments. ( F ) Tumor volume (mm 3 ) progression in nude mice over 10 days after oral gavage administration of Palbociclib (from day 1), intraperitoneal injection of CSA (from day 3, or the combination of both (Palbo+CSA), compared to controls. n = 8 per group. ( G ) Representative images of immunohistochemical analysis of p21 and quantification in tumors obtained from the nude mice used in ( E ). Scale bar = 200 μm. All the graphs show the mean ± SD. Statistical analyses were performed with Unpaired Student’s t test in ( A – C ), and two-way ANOVA multiple comparison with Tukey’s correction in ( E , F ). P values are indicated in the figure. P values in ( E ) indicate significance from day 7 to end of the experiment. .

Article Snippet: NIM811 , MedChemExpress , SDZ NIM811.

Techniques: Immunodetection, Western Blot, SDS Page, Irradiation, Concentration Assay, Cell Viability Assay, Staining, Injection, Immunohistochemical staining, Comparison

Journal: The EMBO Journal

Article Title: Cyclophilin D plays a critical role in the survival of senescent cells

doi: 10.1038/s44318-024-00259-2

Figure Lengend Snippet: Reagents and tools table

Article Snippet: NIM811 , MedChemExpress , SDZ NIM811.

Techniques: Recombinant, Sequencing, Diagnostic Assay, Staining, Cell Culture, Membrane, Western Blot, Transfection, Phospho-proteomics, DC Protein Assay, Software, Cell Viability Assay, cDNA Synthesis