Figure 5. A short - and long-term evaluation of the clearance effect of ABT263 on OA chondrocytes (P1) in monolayer culture. (A) Schematic of the monolayer experiments in b–f. (B) Gross observation (left; scale bar: 5 mm) and microscope images (right; scale bar: 500 μm) of OA chondrocytes after treatment with 2.5μM ABT263 with SA-β-gal staining. (C) Representative flow cytometric plots measuring cell apoptosis from two independent experiments. (D) Immunofluorescence of HMGB1 in cultured OA chondrocytes at day 3 and day 10. Scale bar: 100 μm. (E) Statistical analysis of the percentage of apoptotic cells and fluorescence intensity of HMGB1 from three independent experiments are shown. (F) Western blot analysis of proteins level for c-caspase3 and p21 at Day 3 and Day 10. (G) qPCR analysis of mRNA levels for CDKN1A and CDKN2A (N = 3 for each group).