24 research outputs found
WNT4 is expressed in human fetal and adult ovaries and its signaling contributes to ovarian cell survival
A Secreted BMP Antagonist, Cer1, Fine Tunes the Spatial Organization of the Ureteric Bud Tree during Mouse Kidney Development
The epithelial ureteric bud is critical for mammalian kidney development as it generates the ureter and the collecting duct system that induces nephrogenesis in dicrete locations in the kidney mesenchyme during its emergence. We show that a secreted Bmp antagonist Cerberus homologue (Cer1) fine tunes the organization of the ureteric tree during organogenesis in the mouse embryo. Both enhanced ureteric expression of Cer1 and Cer1 knock out enlarge kidney size, and these changes are associated with an altered three-dimensional structure of the ureteric tree as revealed by optical projection tomography. Enhanced Cer1 expression changes the ureteric bud branching programme so that more trifid and lateral branches rather than bifid ones develop, as seen in time-lapse organ culture. These changes may be the reasons for the modified spatial arrangement of the ureteric tree in the kidneys of Cer1+ embryos. Cer1 gain of function is associated with moderately elevated expression of Gdnf and Wnt11, which is also induced in the case of Cer1 deficiency, where Bmp4 expression is reduced, indicating the dependence of Bmp expression on Cer1. Cer1 binds at least Bmp2/4 and antagonizes Bmp signalling in cell culture. In line with this, supplementation of Bmp4 restored the ureteric bud tip number, which was reduced by Cer1+ to bring it closer to the normal, consistent with models suggesting that Bmp signalling inhibits ureteric bud development. Genetic reduction of Wnt11 inhibited the Cer1-stimulated kidney development, but Cer1 did not influence Wnt11 signalling in cell culture, although it did inhibit the Wnt3a-induced canonical Top Flash reporter to some extent. We conclude that Cer1 fine tunes the spatial organization of the ureteric tree by coordinating the activities of the growth-promoting ureteric bud signals Gndf and Wnt11 via Bmp-mediated antagonism and to some degree via the canonical Wnt signalling involved in branching
Endometrial receptivity and implantation require uterine BMP signaling through an ACVR2A-SMAD1/SMAD5 axis.
During early pregnancy in the mouse, nidatory estrogen (E2) stimulates endometrial receptivity by activating a network of signaling pathways that is not yet fully characterized. Here, we report that bone morphogenetic proteins (BMPs) control endometrial receptivity via a conserved activin receptor type 2âA (ACVR2A) and SMAD1/5 signaling pathway. Mice were generated to contain single or double conditional deletion of SMAD1/5 and ACVR2A/ACVR2B receptors using progesterone receptor (PR)-cre. Female mice with SMAD1/5 deletion display endometrial defects that result in the development of cystic endometrial glands, a hyperproliferative endometrial epithelium during the window of implantation, and impaired apicobasal transformation that prevents embryo implantation and leads to infertility. Analysis of Acvr2a-PRcre and Acvr2b-PRcre pregnant mice determined that BMP signaling occurs via ACVR2A and that ACVR2B is dispensable during embryo implantation. Therefore, BMPs signal through a conserved endometrial ACVR2A/SMAD1/5 pathway that promotes endometrial receptivity during embryo implantation
Role of Wnt4 signaling in mammalian sex determination, ovariogenesis and female sex duct differentiation
Abstract
Mammalian female sex development was considered a default developmental pathway. However, the deletion of the Wnt4 gene, a member of the Wnt family of secreted signals, was shown to reverse the sex of XX female mouse embryo and caused exhibition of certain male characteristics. This indicated that the female sexual development cannot be default but depends on active signaling and cell-cell interaction.
The aim of the current study was to reveal the functional role of the Wnt4 gene in the control of sex determination, ovariogenesis and female sex duct formation.
This study demonstrates that testosterone is produced by the ovary of Wnt4-deficient female embryos. The inhibition of androgen action by an antiandrogen, flutamide, during gestation leads to complete degeneration of the Wolffian ducts in 80% of the Wnt4 mutant females. This suggests that testosterone is the possible mediator of the masculinization phenotype in Wnt4-deficient females. Wnt4 is expressed by ovarian somatic cells, which are vital for the control of female germline development. This work has shown that Wnt4 is the factor maintaining germ cell cysts, cell-cell interaction and early follicular gene expression. In addition, the findings indicate a critical role for Wnt4/5a signaling in meiosis. Our research has proven that Wnt4 has roles during postnatal ovary development as its defective signaling leads to premature ovarian failure associated with diminished Amh levels, defective basement membrane and cell polarization.
The Mullerian duct, the anlagen of oviduct, uterus and upper part of vagina, does not form in Wnt4-deficient females. This study indicates that Wnt4 is needed for migration initiation and maintenance during Mullerian duct formation prenatally. During the postnatal uterine differentiation Wnt4 is essential for endometrial gland formation.
The present study provides new evidence for Wnt4 function during embryonic and adult female sexual differentiation.TiivistelmÀ
NisÀkkÀiden naaraspuolista kehitystÀ pidettiin aiemmin sukupuolisen erilaistumiskehityksen oletusarvona. Signaloivien proteiinien Wnt-perheeseen kuuluvan Wnt4-geenin puutteen todettiin kuitenkin johtavan XX naarasalkion sukupuolen kÀÀntymisen koiraaksi sekÀ aiheuttavan tiettyjÀ koiraille ominaisia piirteitÀ. TÀmÀ osoitti, ettei naaraspuolinen kehitys ole oletusarvo, vaan se riippuu aktiivisesta signaloinnista ja solujen vÀlisestÀ interaktiosta.
TÀmÀn vÀitöstutkimuksen tarkoitus oli selvittÀÀ Wnt4-geenin roolia sukupuolen mÀÀrÀytymisessÀ, munasarjojen kehittymisessÀ sekÀ naaraan sukupuolitiehyitten muodostumisessa.
Tutkimuksessa osoitettiin, ettĂ€ munasarjat tuottavat testosteronia niillĂ€ naaraspuolisilla alkioilla, joilta puuttuu Wnt4-geeni. 80 prosentilla naaraista, joilla on Wnt4-geenin puute, androgeenivaikutuksen esto raskauden aikana annettavalla antiandrogeenilla, flutamidilla, estÀÀ sukupuolen vaihtumisen fenotyypin. TĂ€mĂ€ viittaa siihen, ettĂ€ testosteroni toimii mahdollisena koiraan fenotyypin vĂ€littĂ€jĂ€nĂ€ naarailla, joilta puuttuu Wnt4-geeni. Wnt4 ilmentyy munasarjojen somaattisissa soluissa, jotka ovat tĂ€rkeitĂ€ naaraspuolisen ituradan kehityksen sÀÀtelyn kannalta. VĂ€itöstutkimus osoittaa, ettĂ€ Wnt4 on itusoluryppĂ€itĂ€, solujen vĂ€listĂ€ interaktiota sekĂ€ varhaista follikkeligeeni-ilmentymistĂ€ yllĂ€pitĂ€vĂ€ tekijĂ€. Tulokset osoittavat myös, ettĂ€ Wnt4/5a -signaloinnilla on tĂ€rkeĂ€ rooli meioosissa. Tutkimus osoittaa lisĂ€ksi, ettĂ€ Wnt4 vaikuttaa munasarjojen kehitykseen myös syntymĂ€n jĂ€lkeen. Puutteellinen signalointi alentaa Anti-MĂŒllerian hormonin tasoa, heikentÀÀ tyvikalvoa ja vĂ€hentÀÀ solujen polarisaatiota, joka johtaa ennenaikaiseen munasarjojen toiminnan hiipumiseen.
MĂŒllerin tiehyet, joista myöhemmin kehittyvĂ€t munanjohtimet, kohtu ja vaginan ylĂ€osa, jÀÀvĂ€t kokonaan muodostumatta naarailla, joilta puuttuu Wnt4-geeni. Tulokset viittaavat siihen, ettĂ€ Wnt4 on tarpeen alkioaikaiseen MĂŒllerin tiehyen muodostavien solujen liikkeellelĂ€htöön ja yllĂ€pitoon. Wnt4:llĂ€ on myös keskeinen rooli kohturauhasten muodostumisessa sukukypsyyden saavuttamisen aikana ja sen jĂ€lkeen
Wnt4 coordinates directional cell migration and extension of the MĂŒllerian duct essential for ontogenesis of the female reproductive tract
Wnt4 coordinates directional cell migration and extension of the MĂŒllerian duct essential for ontogenesis of the female reproductive tract
Abstract
The MĂŒllerian duct (MD) is the anlage of the oviduct, uterus and upper part of the vagina, the main parts of the female reproductive tract. Several wingless-type mouse mammary tumor virus (MMTV) integration site family member (Wnt) genes, including Wnt4, Wnt5a and Wnt7a, are involved in the development of MD and its derivatives, with Wnt4 particularly critical, since the MD fails to develop in its absence. We use, here, Wnt4EGFPCre-based fate mapping to demonstrate that the MD tip cells and the subsequent MD cells are derived from Wnt4+ lineage cells. Moreover, Wnt4 is required for the initiation of MD-forming cell migration. Application of anti-Wnt4 function-blocking antibodies after the initiation of MD elongation indicated that Wnt4 is necessary for the elongation as well, and consistent with this, cell culture wound-healing assays with NIH3T3 cells overexpressing Wnt4 promoted cell migration by comparison with controls. In contrast to the Wnt4 null embryos, some Wnt4monomeric cherry/monomeric cherry (Wnt4mCh/mCh) hypomorphic mice survived to adulthood and formed MD in âŒ45% of cases. Nevertheless, the MD of the Wnt4mCh/mCh females had altered cell polarization and basement membrane deposition relative to the controls. Examination of the reproductive tract of the Wnt4mCh/mCh females indicated a poorly coiled oviduct, absence of the endometrial glands and an undifferentiated myometrium, and these mice were prone to develop a hydro-uterus. In conclusion, the results suggest that the Wnt4 gene encodes signals that are important for various aspects of female reproductive tract development
HNF1B controls proximal-intermediate nephron segment identity in vertebrates by regulating Notch signalling components and Irx1/2
Abstract
The nephron is a highly specialised segmented structure that provides essential filtration and resorption renal functions. It arises by formation of a polarised renal vesicle that differentiates into a comma-shaped body and then a regionalised S-shaped body (SSB), with the main prospective segments mapped to discrete domains. The regulatory circuits involved in initial nephron patterning are poorly understood. We report here that HNF1B, a transcription factor known to be involved in ureteric bud branching and initiation of nephrogenesis, has an additional role in segment fate acquisition. Hnf1b conditional inactivation in murine nephron progenitors results in rudimentary nephrons comprising a glomerulus connected to the collecting system by a short tubule displaying distal fates. Renal vesicles develop and polarise normally but fail to progress to correctly patterned SSBs. Major defects are evident at late SSBs, with altered morphology, reduction of a proximo-medial subdomain and increased apoptosis. This is preceded by strong downregulation of the Notch pathway components Lfng, Dll1 and Jag1 and the Irx1/2 factors, which are potential regulators of proximal and Henleâs loop segment fates. Moreover, HNF1B is recruited to the regulatory sequences of most of these genes. Overexpression of a HNF1B dominant-negative construct in Xenopus embryos causes downregulation specifically of proximal and intermediate pronephric segment markers. These results show that HNF1B is required for the acquisition of a proximo-intermediate segment fate in vertebrates, thus uncovering a previously unappreciated function of a novel SSB subcompartment in global nephron segmentation and further differentiation
Improving signal detection in emission optical projection tomography via single source multi-exposure image fusion
Abstract
We demonstrate a technique to improve structural data obtained from Optical Projection Tomography (OPT) using Image Fusion (IF) and contrast normalization. This enables the visualization of molecular expression patterns in biological specimens with highly variable contrast values. In the approach, termed IF-OPT, different exposures are fused by assigning weighted contrasts to each. When applied to projection images from mouse organs and digital phantoms our results demonstrate the capability of IF-OPT to reveal high and low signal intensity details in challenging specimens. We further provide measurements to highlight the benefits of the new algorithm in comparison to other similar methods
Optical studies of nanodiamond-tissue interaction:skin penetration and localization
Abstract
In this work, several optical-spectroscopic methods have been used to visualize and investigate the penetration of diamond nanoparticles (NPs) of various sizes (3â150 nm), surface structures and fluorescence properties into the animal skin in vitro. Murine skin samples have been treated with nanodiamond (ND) water suspensions and studied using optical coherence tomography (OCT), confocal and two-photon fluorescence microscopy and fluorescence lifetime imaging (FLIM). An analysis of the optical properties of the used nanodiamonds (NDs) enables the selection of optimal optical methods or their combination for the study of nanodiamondâskin interaction. Among studied NDs, particles of 100 nm in nominal size were shown to be appropriate for multimodal imaging using all three methods. All the applied NDs were able to cross the skin barrier and penetrate the different layers of the epidermis to finally arrive in the hair follicle niches. The results suggest that NDs have the potential for multifunctional applications utilizing multimodal imaging
A FRET-based high-throughput screening platform for the discovery of chemical probes targeting the scaffolding functions of human tankyrases
Abstract
Tankyrases catalyse poly-ADP-ribosylation of their binding partners and the modification serves as a signal for the subsequent proteasomal degradation of these proteins. Tankyrases thereby regulate the turnover of many proteins involved in multiple and diverse cellular processes, such as mitotic spindle formation, telomere homeostasis and Wnt/ÎČ-catenin signalling. In recent years, tankyrases have become attractive targets for the development of inhibitors as potential therapeutics against cancer and fibrosis. Further, it has become clear that tankyrases are not only enzymes, but also act as scaffolding proteins forming large cellular signalling complexes. While many potent and selective tankyrase inhibitors of the poly-ADP-ribosylation function exist, the inhibition of tankyrase scaffolding functions remains scarcely explored. In this work we present a robust, simple and cost-effective high-throughput screening platform based on FRET for the discovery of small molecule probes targeting the proteinâprotein interactions of tankyrases. Validatory screening with the platform led to the identification of two compounds with modest binding affinity to the tankyrase 2 ARC4 domain, demonstrating the applicability of this approach. The platform will facilitate identification of small molecules binding to tankyrase ARC or SAM domains and help to advance a structure-guided development of improved chemical probes targeting tankyrase oligomerization and substrate protein interactions