Fig. 2

LASS2 inhibits the stemness and cisplatin resistance of BCSCs. A A diagram showing the establishment of cisplatin-resistant cells and BCSC sorting. B, C Flow cytometry analysis and quantitation of the CD44⁺ALDH1A1⁺ subpopulation in 5637 and 5637-CR cells. D Sphere formation assay of BCSCs and BCNSCs. Left: representative images of spheres. Right: comparison of the diameter of spheres. E Cell viability and IC₅₀ assay of BCSCs and BCNSCs. F Differentially expressed resistance-related genes in BCSCs were identified by an RT2 Profiler PCR Array (p < 0.05 and fold change > 5 genes were selected). G Immunoblotting determining the protein expression levels of LASS2, ABCC2, and CD44 in BCSCs and BCNSCs. H Cell viability and IC₅₀ assay for the indicated cells treated with 10 μg/mL cisplatin for 24 h. I In vitro extreme limiting dilution assay for the indicated cells treated with 10 μg/mL cisplatin for 24 h. J Sphere formation frequency of the indicated cells. ELDA software was used to calculate stem cell frequency (left panel) and perform the goodness-of-fit test (right panel). K–N Representative images (left panel) and quantification (right panel) of tumor sphere formation (K), EdU (L), soft agar colony formation (M), and apoptosis (N) assays for the indicated cells transfected with control or LASS2-overexpressing plasmid and treated with 10 μg/mL cisplatin for 24 h. Scale bars, 100 μm for the tumor sphere formation assay, and 50 μm for the EdU assay. Data are presented as the means ± SD of at least three technical replicates. **p < 0.01, ***p < 0.001, determined by one-way ANOVA