Figure 4

TRAIL-expressing MSCs reduce the clonogenic potential of H357 SP and non-SP cells and produce additional SP cancer cell killing to mitoxantrone treatment. (A, B) In all, 200 DiI-labelled, SP (A) or non-SP (B) H357 cells were plated for colony forming assays before the addition of 5 × 10⁴ TRAIL-expressing MSCs (MSCFLT). (C) Quantification of large colony numbers from (A, B) demonstrates a reduction in large colonies with doxycyline (dox)-induced TRAIL expression in both SP and non-SP cells compared with the co-cultures without dox (ND). Furthermore, SP cells produced more colonies than non-SP cells. (D) Phase-contrast and (E) fluorescent microscopy demonstrate the green fluorescent protein (green) from the doxycycline (dox)-activated MSCFLTs surrounding the DiI-labelled (red) H357 colonies. (F) SP cells were exposed to mitoxantrone (Mitox) and then co-cultured with MSCFLT cells. Bar chart represents triplicate flow cytometry experiments and demonstrates a further increase in death and apoptosis of the SP cells with the addition of doxycycline and activation of MSCFLT TRAIL expression. ^***P<0.001, ^**P<0.01, ns, non-significant. Scale bars represent 25 μm.