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doi: 10.1242/10.1242/dev.00591

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Endothelial and steroidogenic cell migration are regulated by WNT4 in the developing mammalian gonad

¹ Section of Gene Function and Regulation, Institute of Cancer Research, 237 Fulham Road, London SW3 6JB, UK
² Duke University Medical Center, Durham, NC 27710, USA

View larger version (58K): [in a new window]   Fig. 2. Genes normally expressed in the coelomic vessel in the testis are found associated with the ectopic coelomic vessel in the Wnt4-/- XX gonad. Whole-mount in situ hybridisation for Jag1 and Pdgfra expression was performed on gonads from Wnt4-/- or Wnt4+/+ XX and XY 12.5 dpc embryos as indicated.

View larger version (65K): [in a new window]   Fig. 1. The coelomic blood vessel forms in the Wnt4-/- ovary. (A) Light microscope images and Pecam stained images of kidney (kid), adrenal (ad) and ovary (ov) from 15.5 dpc embryos that were wild-type (+/+) and homozygous (-/-) for the mutant Wnt4 allele as indicated. (B) Gonads from 12.5 dpc embryos that were wild-type (+/+) or homozygous (-/-) for the mutant Wnt4 allele stained with a Pecam antibody, which labels the endothelium and germ cells at 12.5 dpc (speckled pattern in XX +/+ sample). In all images anterior is towards the left. Coelomic blood vessels are indicated by arrows and the ectopic vessel in Wnt4-/- adrenals is indicated by an arrowhead.

View larger version (92K): [in a new window]   Fig. 3. Mesonephric endothelial cells migrate into Wnt4-/- XX gonads. (A) Mesonephroi from 12.5 dpc embryos ubiquitously expressing lacZ were incubated with gonads from 12.5 dpc embryos that were Wnt4+/+ XX and XY or Wnt4-/- XX and stained for ß-galactosidase activity. (B) Mesonephroi from 11.5 dpc embryos ubiquitously expressing green fluorescent protein (GFP) (green) were incubated with gonads from 11.5 dpc embryos that were Wnt4+/+ XX and XY or Wnt4-/- XX. After incubation the sample was stained with an antibody against Pecam (red). Pecam marks endothelial cells and germ cells (red arrow indicates Pecam-stained germ cells; yellow arrow indicates Pecam-stained endothelial cells).

View larger version (89K): [in a new window]   Fig. 4. Ectopic steroidogenic cells are clustered at the anterior region of the Wnt4-/- gonad. Whole-mount in situ hybridisation for 3ß-hydroxysteroid dehydrogenase (3ß-HSD) expression was performed on gonads and mesonephric region from Wnt4+/+ or Wnt4-/- XX 12.5 dpc embryos as indicated. The top panels show the area within the urogenital region where the adrenal forms, which is indicated by the arrows. The asterisks indicate the anterior region of the gonad where the ectopic steroidogenic cells are found in the Wnt4-/- embryos. The position of the gonad is marked `g'. In all images, anterior is leftwards.

View larger version (69K): [in a new window]   Fig. 5. Steroidogenic cells migrate from the anterior region of the mesonephros into the Wnt4-/- XX gonad. Gonads from Wnt4-/- XX 11.5-12.5 dpc embryos were incubated with mesonephroi from Cyp11a1:lacZ 11.5-12.5 dpc embryos and stained for ß-galactosidase activity. (Top) The anterior region of the mesonephros was included in the co-culture, and (bottom) the anterior region of the mesonephros was removed. The lines indicate the boundary between gonad and mesonephros in the co-cultures.

View larger version (60K): [in a new window]   Fig. 6. Wnt4 is expressed in testis of Sf1:Wnt4 transgenic animals. Whole-mount in situ hybridisation for Wnt4 expression was performed on 13.5 dpc testis from transgenic and non-transgenic (NT) embryos and for Sf1 expression on 13.5 dpc testis from a non-transgenic.

View larger version (95K): [in a new window]   Fig. 7. Vascular and steroidogenic phenotype observed in the Sf1:Wnt4 transgenic XY gonads. (A) Coelomic blood vessel pattern is affected in Sf1:Wnt4 transgenic testis. Testis from transgenic (a-c) and non-transgenic (NT) (d) 13.5 dpc embryos were stained with an antibody to Pecam. (B) Leydig cell differentiation occurs in Sf1:Wnt4 transgenic testis. Whole-mount in situ hybridisation for Cyp11a1 expression on 13.5 dpc XY gonads from a Sf1:Wnt4 transgenic embryo (left) and a non-transgenic embryo (NT) (right). The transgenic gonads were derived from different integration events (1-4).