cell culture experiments hela s3  (ATCC)

Journal: Scientific reports

Article Title: Highly specific Immunoproteasome inhibitor M3258 induces proteotoxic stress and apoptosis in KMT2A::AFF1 driven acute lymphoblastic leukemia.

doi: 10.1038/s41598-025-01657-0

Figure Lengend Snippet: Fig. 1. M3258, a highly specific inhibitor of β5i sites, induces apoptosis in ALL cells. (a) Inhibition of proteasome active sites was measured in extracts of RS4;11 and HeLa cells, and in the purified constitutive 26 S proteasomes; n = 2. (b, c) Cells were treated with Btz (b) or M3258 (c) for 48 h, and the viability was assessed by Alamar Blue; n = 2–4. (d) RS4;11 cells were treated with 75 nM and 300 nM M3258 for indicated times, and percentage of apoptotic cells was determined by flow cytometry; n = 2. (e) Cells were treated with M3258 for 12 h, and apoptosis was determined by flow cytometry. In a parallel experiment, RS4;11 cells were treated with M3258 for 4 h, and the chymotrypsin-like (β5) activity was measured by the Proteasome-Glo assay; n = 2. (f) The chymotrypsin-like (β5) activity of the proteasomes in cells treated with M3258 for times indicated was determined by the Proteasome-Glo assay; n = 2. (g) MM1.S cells were pre-treated with IFNγ for three days and then treated with M3258 for 48 h. Viability was measured with Alamar Blue; n = 2. Data on all panels are averages of n biological replicates, and the error bars indicate standard error of the mean.

Article Snippet: Cell lines and cell culture experiments HeLa S3 (RRID: CVCL_0058), RS4;11 (RRID: CVCL_0093), REH (RRID: CVCL_1650), and NALM-6 (RRID: CVCL_0092) cells were obtained from the American Tissue Culture collection; SEM (RRID: CVCL_0095) cells were obtained from DSMZ (Braunschweig, Germany).

Techniques: Inhibition, Purification, Flow Cytometry, Activity Assay, Glo Assay