Fig. 4. MMP2 maturation correlates with Müllerian duct regression. (A) A schematic of MMP2 activation by MMP14 is shown. (B) Gelatin zymograms of conditioned media obtained from treated male and female genital ridges are shown for both the latent (pro-MMP2) and mature forms of MMP2 (mature MMP2). The first lane shows the control marker for purified recombinant (MMP2). The treatment is indicated above each lane and all zymograms are shown as negative images. (C) Female genital ridges treated with concanavalin A (Con A) and control analog undergo regression in the absence of MIS (Con A + control). GM6001 blocks con A-induced regression (Con A + GM6001). The Müllerian duct is not as clearly visible in con A + GM6001 ridges as in the untreated ridges, possibly because of incomplete inhibition of con A-stimulated MMP2 maturation, and mild toxicity of these compounds in organ culture. M, Müllerian duct; W, Wolffian duct. (D) Con A causes Müllerian duct regression independent of MIS receptor signaling. The relative luciferase activity of the Tlx2 promoter fused to luciferase reporter in P19 cells over a wide concentration range of con A (10-200 µg/ml) is shown. No enhancement of the MIS receptor signaling is observed in the absence of MIS, despite high doses of con A. In genital ridge organ cultures 50 µg/ml of con A is sufficient to cause full Müllerian duct regression in the absence of MIS (see 4C).
