UMR 7221CNRS/MNHN Evolution des régulations endocriniennes Muséum National d Histoire Naturelle Paris - France

Transcription

1 Role of Foxl2 and Dlx5/6 on uterine development and function: implications for BPES UMR 7221CNRS/MNHN Evolution des régulations endocriniennes Muséum National d Histoire Naturelle Paris - France

2 TAKE HOME MESSAGES Foxl2 and Dlx5/6 are expressed in the ovary and in the uterus and can affect the development and function of both organs. In the uterus, Foxl2 controls stroma and myometrial formation while Dlx5 and Dlx6 control adenogenesis; uterine-specific inactivation of either of these genes results in infertility.

3 Dlx5 and Dlx6 : Roles in development The dll/dlx homeodomain gene family Dlx1 Dlx2 Dll Dll-A Dll-B Drosophilia Tunicates Dlx3 Dlx5 Dlx7 Dlx6 Mouse / Human Dlx5 LacZ/+ 9.5dpc ISH 11.5dpc Dlx5 LacZ/ dpc Early roles in the development of appendages, sensory organs, CNS and facial skeleton. Later roles in osteogenesis in the whole embryo in the limb Acampora et al. (1999), Samee et al. (2009)

4 Similar patterns of expression of dll and Dlx terminalia/genitalia Dlx5-lacZ Insect (adapted from Gorfinkiel et al. 1999) Mouse

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7 Perinatal phenotype

8 Low levels of testosterone and feminization (reduced AGD) in Dlx5/6 double mutant embryos. Nishida et al. 2008

9 Dlx5-lacZ testis

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12 StAR mrna expression pattern of control and Dlx5/6 double mutant embryos. Nishida et al. 2008

13 Dlx5 enhances the GATA-4-mediated promoter activation of the StAR gene Nishida et al. 2008

14 CONCLUSIONS The double inactivation of Dlx5 and Dlx6 in the mouse results in severe defects of genital organ development reminiscent of human hypospadias. Dlx5 and 6 are expressed in Leyding cells and participate to their differentiation. Dlx5 is an activator of steroidogenesis in the testis Dlx5 interacts with GATA4 to activate the STAR promotor Dlx5 and 6 might have a much more general role in the regulation of steroidogenesis

15 Dlx5/6 allelic reduction results in female subfertility ns WT females *** ns Dlx5/6 +/- females Males Dlx5 +/- Dlx5/6+/- Dlx6+/- Msx1+/- ; Dlx5/6+/- End1+/- Reduced litter size Reduced number of litters per female Shortened reproductive life (about 6 months) Average follicle number per section Bouhali et al. (2011) Ovaries of Dlx5/6 +/- heterozygous mice are smaller Reduced overall follicles number Reduced number of early follicles Higher percentage of mature follicles Premature follicular maturation reminiscent of certain forms of human POF

16 Defects of Dlx5/6 +/- ovaries WT Dlx5/6 +/- Ovaries of Dlx5/6 +/- heterozygous mice are smaller and have fewer follicles than those of normal littermates. Bouhali et al. (2011)

17 Genes suppressed by Dlx5/6 From Jeong et al., 2009

18 Bouhali et al. (2011)

19 Dlx5/6 and Foxl2 : a complementary expression in the ovary I II III mature Foxl2 Dlx5 Foxl2 is present in primordial and primary follicles and its expression decreases during follicular maturation : at the same time the expression of Dlx5 increases in granulosa cells. Bouhali et al. (2011)

20 DLX5/6 et FOXL2 in woman endometrium Microarray FOXL2 DLX5 Expression in different phases of the menstrual cycle As in the ovary? Dlx5/6 Foxl2 DLX6

21 Dlx5 and Foxl2 expression in the developing female reproductive tract uterus cervix oviduct ovary cervix

22 Dlx5 and Foxl2 during post-natal uterine maturation LE GE STROMA MYO

23 Foxl2-positive cells in the uterine stroma contribute to the inner myometrial layer S Foxl2 is expressed by mesenchymal progenitors of : - Stromal cells M - SMA+ cells of the inner myometrial layer -SMA+ cells surrounding blood vessels ROSA promoter stop LacZ x Foxl2 promoter Cre

24 Foxl2 is absent from stromal cells during decidualisation 5,5dpc

25 Foxl2 conditional inactivation in the mouse uterus E11,5 E13,5 E15 P0 P5 P6 P15 P25 P30 Histological maturation PGR2::Cre Reproductive Function : Cyclicity Implantation

26 Foxl2 conditional inactivation in the uterus

27 Foxl2 conditional inactivation in the uterus results in infertility

28 Post-natal deletion of Foxl2 in the uterus results in a stromal defect with increased myometrial differentiation Decrease of stromal layer Disorganized and hypertrophic inner myometrial layer

29 Decrease of stromal layer Disorganized and hypertrophic inner myometrial layer Supplemetary bundles of SMA+ cells inthe adult No SMA+ blood vessels in the adult stroma

30 Post-natal deletion of Foxl2 in the uterus results in smaller uterine glands

31 No blood vessel SMA+ in the stroma in P25 and adult mutants

32 Consequences for BPES BPES is an autosomal dominant genetic disorder characterized by narrow palpebral fissures and eyelid levator muscle defects. In female BPES patients, eyelid malformations are associated (BPES type I) or not (BPES type II) to premature ovarian failure. FOXL2 is the only gene known to be mutated in BPES. What is the uterine status of BPES patients?

33 Foxl2 is expressed both by Cranial Neural Crest Cells and by Cranial Mesodermal Cells, which give rise to skeletal and muscular components of the head.

34 Expression of Foxl2 in Cranial Neural Crest Cells, but not in mesodermal cells, is essential for the development of extraocular muscles including the levator palpebrae

35 Dlx5 expression in the adult uterus

36 Conditional inactivation of Dlx5/6 in the uterus inhibits adenogenesis No uterine glands in the mutant

37 Uterus-specific inactivation of Dlx5 and Dlx6 results in loss of adenogenesis No uterine glands in the mutant

38 Uterus-specific inactivation of Dlx5 and Dlx6 results in loss of adenogenesis No uterine glands in the mutant

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40 DLX5 and FOXL2 in endometriosis Endometriosis Endometrial tissues external to the uterine cavity 10-15% of women of reproductive age Painful menstrual period, chronic pelvic pain, subfertility associated 30% of infertility case Under hormonal influence Unkown aetiology Most accepted theory, endometrial debris transplanted after retrograde menstrual flow Endometriotic tissue vs. Endometrium Strong (p < 0,00001) decrease in DLX5 expression Strong increase (p < 0,001) in Foxl2 expression

41 In endometrioid adenocarcinoma, the expression of DLX5 is profoundly modified depending of epithelial morphological changes and stages of tumor differentiation At variance from normal endometrial epithelia which present a uniform DLX5 staining, in endometriosis, DLX5 is present in few sparse cells of the epithelium of endometriotic lesions.

42 Museum National d Histoire Naturelle/CNRS Paris, France Brice BELLESORT Kamal BOUHALI Anne BACHELOT Gladys ALFAMA Anastasia FONTAINE Nicolas NARBOUX-NÊME Yorick GITTON Eglantine HEUDE UE Grants CRESCENDO, IDEAL Institut Jacques Monod Paris, France Prof. Marc FELLOUS Dr. Daniel VAIMANS Sandrine CABURET Berlin, Germany Mathias TREIER Kumamoto University, Japan Gen YAMADA University of Genova, Italy Ottavia BARBIERI

43 Transgenic testis with Dlx5 reporter

44 Increased expression of FoxL2 in mouse pharingeal arch 1 following Dlx5 and Dlx6 inactivation. From Jeong et al., 2009

45 Opposite regulation of Dlx5/6 and Foxl2 in steroidogenesis