Figure 9. Co-culture of wild-type HSC on Tle4 null stromal cells impairs long term colony forming ability.

(a) Schematic of co-culture assay. Briefly, stromal cells were isolated from bones of 2 week old WT and KO animals and plated in aMEM with dexamethasone, ascorbic acid, and vitamin D₃ as described in Methods. WT LKS were plated and used in LTC-IC experiments (b) Bar graph showing a dramatic decreased frequency of Sca-1⁺c-Kit⁺ cells after two weeks of co-culture on the stroma from Tle4 null mice with representative flow plots (n = 2 biological replicates per genotype, in triplicate *: P<.01). (c) Long term culture-initiating cell assay (LTC-IC) after four weeks of co-culture with WT or KO stroma showed relatively few colonies obtained after culturing on Tle4 null bone marrow stroma (n = 2 biological replicates, each in 9-plicate, seeded in duplicates for CFU; *: P<.0001).