Fig. 6From: Chemoresistance is mediated by ovarian cancer leader cells in vitroLeader cells proliferate at the same rate in chemotherapy and do not arise due to cellular division and DNA replication, leader cell deficient lines are more susceptible to cisplatin treatment. A Flow cytometric analysis of Ki67 surface expression in LC+/− populations in ovarian cancer cell lines in the presence of cisplatin. B Cells were seeded at 300,000 cells/well in a 6-well plate, incubated for 18 h, and treated with 13 μg/ml cisplatin +/− aphidicolin (0.5 μg/ml) or tunicamycin (6 μg/ml) for 24 h. Following treatment, cells were collected and stained for Ki67 using a Ki67-BV786 antibody. GFP (indicative of KRT14 expression) and BV786 fluorescence was acquired using the BD LSRFortessa™ X-20 with data analysed using the FlowJo software (v10.5.0). C Analysis was performed using a non-parametric Mann-Whitney U test to determine statistical significance between groups; n = 2–4/cell line; ns = not significant. D Wild-type, LC-deficient (KRT14KO) and LC-enriched (KRT14OE) cells were seeded at 0.2 × 104 cells/0.1 ml/well in 96-well plates. Cells were synchronized in G0 by overnight incubation in serum-free media prior to treatment and treated with cisplatin concentrations ranging from 1.2–30 μg/ml for 48 h followed by the addition of cell viability dye alamarBlue™ (Invitrogen). Half maximal inhibitory (IC50) concentrations of cisplatin were calculated in GraphPad Prism (v9.0) and analysed by non-linear regression fit. E Long-term assessment of cellular proliferation in response to high doses of Cisplatin where 0.2 × 104 cells/0.1 ml/well wild-type, LC deficient (KRT14KO) and LC enriched (KRT14OE) cells were seeded onto RTCA xCELLigence E-plates and treated with 30 μg/ml cisplatin, cellular proliferation was monitored over 240-h and impedance readings taken every 15 minBack to article page