Fig. 5. When added together with factors that promote branching morphogenesis, RA induces the formation of hollow tubular structures. (A) Serum-free conditioned medium from confluent monolayer cultures of J3B1A cells induces the formation of solid branching cords by J3B1A cells grown in collagen gels (10 day culture). (B) When incubated with conditioned medium for 3 days to allow initial cord formation and subsequently treated with 1 µM RA for a further 7 days, J3B1A cells form branching tubes enclosing a widely patent lumen. (C) Addition of 10 µM LPA stimulates colony branching without inducing lumen formation. (D) Concomitant treatment with 10 µM LPA and 10 nM RA results in the formation of arborized tubular structures that recapitulate the 3D organization of mammary gland ducts. Bar, 100 µM.