Fig. 2. Time course and dose-dependence of retinoic acid effect on lumen formation. J3B1A cells were suspended in collagen gels at 3x10⁴ cells/ml and incubated with either 0.1% DMSO (control) or a wide range of RA concentrations. (A,B) Lumen formation was quantified after 3, 6 and 9 days as described in Materials and Methods. Results are expressed as a mean percentage of cysts (i.e. colonies containing a central cavity) in 25 randomly selected photographic fields (five fields from each of five separate experiments) per experimental condition. In A, values are plotted against time of treatment, whereas in B, the same values are plotted against RA concentration. P=0.01 for values of 100 pM RA at 3 days compared with the control; P<0.025 for values of 100 pM RA at 9 days compared with 3 days; P<0.01 for values of 1 nM and 10 nM RA at 9 days compared with 3 days; P<0.0025 for values of 10 µM RA at 6 days compared with 3 days and at 9 days compared with 6 days. (C) Lumen diameter was quantified on the micrographs taken at day 9 by measuring the minor axis of the central cavity in all cystic colonies. Data are expressed as mean±s.e.m. from five separate experiments. ^*P<0.01 compared with 100 pM RA; ^**P<0.0005 compared with 100 nM RA.