Cells responding to hedgehog signaling contribute to the theca of ovarian follicles. Cowan RG et al. (2021) Cell-fate mapping was used to identify cells that respond to the hedgehog (HH) signaling pathway and that are incorporated into the theca cell layer during ovarian follicle development. Expression of Gli1 is increased by HH signaling and can be used as a marker of cells responsive to HH in reporter mice. In transgenic Gli1ERcre/tdT mice, injection of tamoxifen (TAM) induces cre-mediated recombination and expression of td tomato (tdT) which leads to permanent fluorescent marking of cells expressing Gli1 and their progeny. The identity of tdT-positive cells was determined by co-staining ovaries for endothelial cells (CD31), pericytes (CSPG4), vascular smooth muscle cells (VSMC; smooth muscle actin) and steroidogenic cells (cytochrome P450 17A1). Gli1ERcre/tdT mice were injected with TAM on the day of birth. Cells positive for tdT in 2-day-old mice were identified as pericytes, located primarily in the medulla of the ovary in close proximity to endothelial cells. In both prepubertal mice and adult mice treated with equine chorionic gonadotropin to induce the formation of preovulatory follicles, tdT-positive cells were located within the theca cell layer and were identified as pericytes, VSMC and steroidogenic theca cells. Granulosa cells are known to express two HH ligands, Indian HH and desert HH (DHH). In DHHcre/tdT reporter mice, endothelial cells were marked as tdT-positive indicating that endothelial cells, in addition to granulosa cells, express Dhh in the ovary. These findings suggest that HH signaling may stimulate the development of the vasculature along with steroidogenic capacity of the theca layer during follicle development.//////////////////
NCBI Summary:
This gene encodes a member of the hedgehog family of proteins. The encoded preproprotein is proteolytically processed to generate multiple protein products, including an N-terminal fragment that is involved in signaling. Hedgehog family proteins are essential secreted signaling molecules that regulate a variety of developmental processes including growth, patterning and morphogenesis. The protein encoded by this gene specifically plays a role in bone growth and differentiation. Mutations in this gene are the cause of brachydactyly type A1, which is characterized by shortening or malformation of the fingers and toes. Mutations in this gene are also the cause of acrocapitofemoral dysplasia. [provided by RefSeq, Nov 2015]
General function
Ligand, Hormone, Growth factor
Comment
Cellular localization
Secreted
Comment
Ovarian function
Primary follicle growth, Steroid metabolism
Comment
The Hedgehog-Patched Signaling Pathway and Function in the Mammalian Ovary: A Novel Role for Hedgehog proteins in Stimulating Proliferation and Steroidogenesis of Theca cells. Spicer L et al. The expression of hedgehog (Hh) genes, their receptor, and the co-receptor in mice, rat and bovine ovaries were investigated. Reverse transcription-polymerase chain reaction (RT-PCR) of ovarian transcripts in mice showed amplification of transcripts for Indian (Ihh) and Desert (Dhh) hedgehog, Patched 1 (Ptch1), and Smoothened (Smo) genes. Semi-quantitative RT-PCR and Northern blot analyses showed that whole ovarian Ihh and Dhh transcripts were less 4-24 h after hCG versus 0-48 h after PMSG treatment in mice, whereas mouse Ptch1 and Smo transcripts were expressed throughout the gonadotropin treatments. Quantitative real-time RT-PCR (qRT-PCR) revealed the expression of the hedgehog-Patched signaling system with Ihh mRNA abundance in granulosa cells greater whereas Smo and Ptch1 mRNA abundance less in theca cells of small versus large follicles of cattle. In cultured rat and bovine theca-interstitial cells, qRT-PCR analyses revealed abundance of Gli1 and Ptch1 mRNAs were increased (P < 0.05) with SHH treatment. Additional studies using cultured bovine theca cells indicated that SHH induces proliferation and androstenedione production. IGF1 decreased Ihh mRNA abundance in bovine granulosa cells. The expression and regulation of Ihh transcripts in granulosa cells and Ptch1 mRNA in theca cells suggests a potential paracrine role of this system in bovine follicular development. These studies illustrate for the first time hedgehog activation of Gli1 transcriptional factor in theca cells and its stimulation of theca cell proliferation and androgen biosynthesis.
The Hedgehog Signaling Pathway in the Mouse Ovary. Russell MC et al. The hedgehog (HH) signaling pathway plays an essential role in the Drosophila ovary, regulating cell proliferation and differentiation, but a role in the mammalian ovary has not been defined. Expression of components of the HH pathway in the mouse ovary and effects of altering HH signaling in vitro were determined. RT-PCR analyses show developmentally regulated expression of sonic (Shh), indian (Ihh) and desert (Dhh) in the ovary. Expression is detected in whole ovary, granulosa cells and corpora lutea. The mRNAs for the two receptors, patched homolog 1 and 2 (Ptch1, Ptch2), and the signal transducer, smoothened (Smo), are also expressed. Immunohistochemistry using an antibody that detects all three HH ligands demonstrated HH protein primarily in granulosa cells of follicles from primary to antral stages of development. Follicles also stained for PTCH1 and SMO in both granulosa and theca cells. Treatment of cultured preantral follicles and granulosa cells with recombinant SHH increased growth and proliferation while treatment with the HH pathway inhibitor, cyclopamine, had no effect. Therefore, activation of HH signaling can increase cell proliferation and follicle growth but is not essential for these processes in vitro. Treatment of granulosa cells with SHH increased levels of mRNA for Gli1, a transcriptional target of HH signaling, while cyclopamine decreased expression. SHH had no effect on production of progesterone by cultured granulosa cells, while cyclopamine increased progesterone production. The results demonstrate a functional HH pathway in the follicle and identify granulosa cells as at least one of the potential targets of HH signaling.
Expression regulated by
Growth Factors/ cytokines
Comment
GDF9////////The hedgehog system in ovarian follicles of cattle selected for twin ovulations and births: evidence of a link between the IGF and hedgehog systems. Aad PY et al. (2012) Hedgehog signaling is involved in regulation of ovarian function in Drosophila, but its role in regulating mammalian ovarian folliculogenesis is less clear. Therefore, gene expression of Indian hedgehog (IHH) and its type 1 receptor, patched 1 (PTCH1), were quantified in bovine granulosa (GC) or theca (TC) cells of small (1-5 mm) antral follicles by in situ hybridization and of larger (5-17 mm) antral follicles by real-time RT-PCR from ovaries of cyclic cows genetically selected (Twinner) or not selected (control) for twin ovulations. Expression of IHH mRNA was localized to GC and cumulus cells, whereas PTCH1 mRNA was greater in TC than in GC. Estrogen-active (E-A; follicular fluid concentration of estradiol > progesterone) versus estrogen-inactive follicles had a greater abundance of mRNA for IHH in GC and PTCH1 in TC. Abundance of IHH mRNA in GC was not affected by cow genotype, whereas TC PTCH1 mRNA was less in large E-A follicles of Twinners than in controls. In vitro, estradiol and wingless-type (WNT) 3A increased IHH mRNA in IGF1-treated GC. IGF1 and BMP4 treatments decreased PTCH1 mRNA in small TC. Estradiol and LH increased PTCH1 mRNA in IGF1-treated TC from large and small follicles, respectively. In summary, functional status of ovarian follicles was associated with differences in hedgehog signaling in GC and TC. We hypothesize that as follicles grow and develop, increased free IGF1 may suppress expression of IHH mRNA by GC and PTCH1 mRNA by TC, and these effects are regulated in a paracrine way by estradiol and other intra- and extragonadal factors.//////////////////
Ovarian localization
Granulosa
Comment
Owens GE, et al reported ovulatory surges of human CG prevent hormone-induced granulosa cell tumor formation leading to the identification of
tumor-associated changes in the transcriptomes.
The authors used a
transgenic mouse model in which granulosa cell tumors occur with 100%
penetrance in CF-1 mice that harbor a novel transgene encoding a chimeric
LHbeta subunit. When this transgene is expressed in other strains of mice,
including (C57BL/6female x CF-1male,Tg) F-1 hybrids, luteomas develop even
though levels of LH remain high. This dichotomous response permits a
longitudinal comparison of global changes in transcriptomes uniquely
associated with either granulosa cell tumors or luteomas. The authors report
numerous changes in the transcriptome, including a decrease in LH receptor
mRNA and increases in several mRNAs that encode secreted proteins previously
associated with granulosa cell tumors. They identified a
constellation of mRNAs that encode proteins that may serve as new markers for
this tumor phenotype. Additional experiments indicated that periodic treatment
with human CG prevented formation of granulosa cell tumors in mice genetically
predisposed to tumor development and, instead, led to the appearance of
luteomas. More importantly, ovarian transcriptomes from the luteomas induced
by ovulatory doses of human CG permitted refined confirmation of gene
expression changes that were uniquely associated with either granulosa cell
tumors in the permissive CF-1 genetic background or in luteomas in the F-1
hybrids. Together, these dynamic changes in the ovarian transcriptome indict
various signaling pathways potentially involved in mediating the actions of LH
over time and, depending on genetic background, the formation of either a
luteoma or a granulosa cell tumor. Indian hedgehog is a gene that is up regulated in granulosa cell tumors.
Follicle stages
Primary, Secondary, Antral
Comment
Wijgerde M, et al reported hedgehog signaling in mouse ovary: Ihh and Dhh from granulosa cells induce target gene expression in developing theca cells.
Follicle development in the mammalian ovary requires interactions between the oocyte, granulosa cells, and theca cells, co-ordinating gametogenesis and steroidogenesis. Here, we show that granulosa cells of growing follicles in mouse ovary act as a source of hedgehog signaling. Expression of indian hedgehog (Ihh) and desert hedgehog (Dhh) mRNAs initiates in granulosa cells at the primary follicle stage, and we find induced expression of the hedgehog target genes Ptch1 and Gli1, in the surrounding pre-theca cell compartment. Cyclopamine, a highly specific hedgehog signaling antagonist, inhibits this induced expression of target genes, in cultured neonatal mouse ovaries. The theca cell compartment remains a target of hedgehog signaling throughout follicle development, showing induced expression of the hedgehog target genes Ptch1, Ptch2, Hip1, and Gli1. In peri-ovulatory follicles, a dynamic synchrony between loss of hedgehog expression and loss of induced target gene expression is observed. Oocytes are unable to respond to hedgehog, since they lack expression of the essential signal transducer Smo (smoothened). The present results point to a prominent role of hedgehog signaling in the communication between granulosa cells and developing theca cells.
Phenotypes
Mutations
2 mutations
Species: mouse
Mutation name: None
type: targeted overexpression fertility: infertile - ovarian defect Comment: Dominant activation of the hedgehog signaling pathway in the ovary alters theca development and prevents ovulation. Ren Y et al. The role of the hedgehog (HH) signaling pathway in ovarian function was examined in transgenic mice in which expression of a dominant active allele of the signal transducer smoothened (SmoM2) was directed to the ovary and M?an duct by cre-mediated recombination (Amhr2(cre/+)SmoM2). Mutant mice were infertile and had ovarian and reproductive tract defects. Ovaries contained follicles of all sizes and corpora lutea (CL), but oocytes were rarely recovered from the oviducts of superovulated mice and remained trapped in preovulatory follicles. Measures of luteinization did not differ. Cumulus expansion appeared disorganized, and in vitro analyses confirmed a reduced expansion index. Microarray analysis indicated that expression levels of genes typical of smooth muscle were reduced in mutant mice and RT-PCR showed that levels of expression of muscle genes were reduced in the non-granulosa, theca-interstitial cell-enriched fraction. While a layer of cells in the outer theca was positively stained for smooth muscle actin in control ovaries, this staining was reduced or absent in mutant ovaries. Expression of a number of genes in granulosa cells that are known to be important for ovulation did not differ in mutants and controls. Expression of components of the HH pathway was observed in both granulosa cells and in the non-granulosa, residual ovarian tissue and changed in response to treatment with eCG/hCG. The results show that appropriate signaling through the HH pathway is required for development of muscle cells within the theca and that impaired muscle development is associated with failure to release the oocyte at ovulation.
Species: mouse
Mutation name: type: null mutation fertility: infertile - ovarian defect Comment: Lineage specification of ovarian theca cells requires multicellular interactions via oocyte and granulosa cells. Liu C et al. (2015) Organogenesis of the ovary is a highly orchestrated process involving multiple lineage determination of ovarian surface epithelium, granulosa cells and theca cells. Although the sources of ovarian surface epithelium and granulosa cells are known, the origin(s) of theca progenitor cells have not been definitively identified. Here we show that theca cells derive from two sources: Wt1(+) cells indigenous to the ovary and Gli1(+) mesenchymal cells that migrate from the mesonephros. These progenitors acquire theca lineage marker Gli1 in response to paracrine signals Desert hedgehog (Dhh) and Indian hedgehog (Ihh) from granulosa cells. Ovaries lacking Dhh/Ihh exhibit theca layer loss, blunted steroid production, arrested folliculogenesis and failure to form corpora lutea. Production of Dhh/Ihh in granulosa cells requires growth differentiation factor 9 (GDF9) from the oocyte. Our studies provide the first genetic evidence for the origins of theca cells and reveal a multicellular interaction critical for the formation of a functional theca.//////////////////