Activation Of Wnt Signaling Research Articles

IGF1 drives Wnt-induced joint damage and is a potential therapeutic target for osteoarthritis

Osteoarthritis is the most common joint disease and a global leading cause of pain and disability. Current treatment is limited to symptom relief, yet there is no disease-modifying therapy. Its multifactorial etiology includes excessive activation of Wnt signaling, but how Wnt causes joint destruction remains poorly understood. Here, we identify that Wnt signaling promotes the transcription of insulin-like growth factor 1 (IGF1) in articular chondrocytes and that IGF1 is a major driver of Wnt-induced joint damage. Male mice with cartilage-specific Igf1 deficiency are protected from Wnt-triggered joint disease. Mechanistically, Wnt-induced IGF1 transcription depends on β-catenin and binding of Wnt transcription factor TCF4 to the IGF1 gene promoter. In a clinically relevant mouse model of post-traumatic osteoarthritis, cartilage-specific deletion of Igf1 protects against the disease in male mice. IGF1 silencing in chondrocytes from patients with osteoarthritis restores a healthy molecular profile. Our findings reveal that reducing Wnt-induced IGF1 is a potential therapeutic strategy for osteoarthritis.

Senescent lung-resident mesenchymal stem cells drive pulmonary fibrogenesis through FGF-4/FOXM1 axis

BackgroundIdiopathic pulmonary fibrosis (IPF) is an age-related disease featured with abnormal fibrotic response and compromised lung function. Cellular senescence is now considered as an essential driving mechanism for IPF. Given the poor knowledge of the mechanisms underpinning IPF progression, understanding the cellular processes and molecular pathways is critical for developing effective therapies of IPF.MethodsLung fibrosis was induced using bleomycin in C57BL/6 mice. Cellular senescence was measured by immunofluorescence. The effects of FGF-4 on fibroblast activation markers and signaling molecules were assessed with western blot and qPCR.ResultsWe demonstrated elevated abundance of senescent mesenchymal stem cells (MSCs) in IPF lung tissues, which was tightly correlated with the severity of pulmonary fibrosis in vivo. In addition, senescent MSCs could effectively induce the phenotype of pulmonary fibrosis both in vitro and in vivo. To further confirm how senescent MSCs regulate IPF progression, we demonstrate that FGF-4 is significantly elevated in senescent MSCs, which can induce the activation of pulmonary fibroblasts. In vitro, FGF-4 can activate Wnt signaling in a FOXM1-dependent manner. Inhibition of FOXM1 via thiostrepton effectively impairs FGF-4-induced activation of pulmonary fibroblast and dramatically suppresses the development of pulmonary fibrosis.ConclusionThese findings reveal that FGF-4 plays a crucial role in senescent MSCs-mediated pulmonary fibrogenesis, and suggests that strategies aimed at deletion of senescent MSCs or blocking the FGF-4/FOXM1 axis could be effective in the therapy of IPF.

The overexpression of R-spondin 3 affects hair morphogenesis and hair development along with the formation and maturation of the hair follicle stem cells.

Mice hair follicles (HFs) are a valuable model for studying various aspects of hair biology, including morphogenesis, development, and regeneration due to their easily observable phenotype and genetic manipulability. The initiation and progression of hair follicle morphogenesis, as well as the hair follicle cycle, are regulated by various signaling pathways, of which the main role is played by the Wingless-type MMTV integration site family (Wnt) and the Bone Morphogenic Protein (BMP). During the hair follicle cycle, the BMP pathway maintains hair follicle stem cells (HFSCs) in a dormant state while the Wnt pathway activates them for hair growth. Given the pivotal role of the Wnt pathway in hair biology and HFSCs regulation, we investigated the influence of the Wnt modulator - R-spondin 3 (Rspo3), in these processes. For this purpose, we developed a transgenic mice model with the overexpression of Rspo3 (Rspo3GOF) in the whole ectoderm and its derivatives, starting from early morphogenesis. Rspo3GOF mice exhibited a distinct phenotype with sparse hair and visible bald areas, caused by reduced proliferation and increased apoptosis of hair matrix progenitor cells, which resulted in a premature anagen-to-catagen transition with a shortened growth phase and decreased overall length of all hair types. In addition, Rspo3GOF promoted induction of auchene and awl, canonical Wnt-dependent hair type during morphogenesis, but the overall hair amount remained reduced. We also discovered a delay in the pre-bulge formation during morphogenesis and prolonged immaturity of the HFSC population in the bulge region postnatally, which further impaired proper hair regeneration throughout the mice's lifespan. Our data supported that Rspo3 function observed in our model works in HFSCs' formation of pre-bulge during morphogenesis via enhancing activation of the canonical Wnt pathway, whereas in contrast, in the postnatal immature bulge, activation of canonical Wnt signaling was attenuated. In vitro studies on keratinocytes revealed changes in proliferation, migration, and colony formation, highlighting the inhibitory effect of constitutive overexpression of Rspo3 on these cellular processes. Our research provides novel insights into the role of Rspo3 in the regulation of hair morphogenesis and development, along with the formation and maturation of the HFSCs, which affect hair regeneration.

CD11B + CD36 + cells are bone anabolic macrophages that limit age-associated bone loss.

Disruptions in the bone remodeling cycle that occur with increasing age lead to degeneration of the skeleton and increased risk of fragility fractures. Our understanding of how the bone remodeling process within cortical bone is controlled and altered with age in males and females is limited. Here, we generated bone marrow chimeric mice to understand the impacts of age and sex on the bone remodeling process. We demonstrate that transplantation of aged male or female bone marrow into young lethally irradiated male hosts unexpectedly enhances cortical bone mass without an impacting cancellous bone. Our single cell RNA-sequencing data show that mice reconstituted with aged bone marrow exhibited subsets of cells marked by CD11B/CD36 expression that demonstrate enhanced production of anabolic cytokines as compared to young counterparts, and that these myeloid subsets exist under conditions of normal physiology in aged mice. Importantly, CD11B + CD36 + cells do not differentiate into osteoclasts in vitro, and CD36 does not mark TRAP+ cells in vivo. Instead, CD36 + cells localize to resorption sites, including within cortical bone defects, suggesting their involvement in cortical bone remodeling and healing. CD11B + CD36 + cells also express elevated levels of bone anabolic WNT ligands, especially Wnt6. In functional assays, we demonstrate that soluble factors produced by CD11B + CD36 + cells enhance osteoblast progenitor commitment, mineralization, and activation of WNT signaling in vitro. Moreover, CD11B/CD36 exquisitely mark a subset of anabolic myeloid cells within human bone marrow. In conclusion, our studies identified a novel population of aged macrophages that limit cortical bone loss.

The crucial role of beta-catenin in the osteoprotective effect of semaglutide in an ovariectomized rat model of osteoporosis.

Postmenopausal osteoporosis is a common chronic medical illness resulting from an imbalance between bone resorption and bone formation along with microarchitecture degeneration attributed to estrogen deficiency and often accompanied by other medical conditions such as weight gain, depression, and insomnia. Semaglutide (SEM) is a recently introduced GLP-1 receptor agonist (GLP-1RA) for the treatment of obesity and type 2 diabetes mellitus by mitigating insulin resistance. It has been discovered that the beneficial effects of GLP-1 are associated with alterations in lipolysis, adipogenesis, and anti-inflammatory processes. GLP-1 analogs transmit signals directly to adipose tissue. Mesenchymal stem cells (MSCs) are multidisciplinary cells that originate from bone marrow, migrate to injury sites, and promote bone regeneration. MSCs can differentiate into osteoblasts, adipose cells, and cartilage cells. Our aim is to investigate the role of semaglutide on bone formation and the Wnt signaling pathway. Osteoporosis was induced in female rats by ovariectomy, and the ovariectomized rats were treated with alendronate as standard treatment with a dose of 3mg/kg orally and semaglutide with two doses (150 mcg/kg and 300 mcg/kg) S.C. for 10 successive weeks. Semaglutide ameliorates bone detrimental changes induced by ovariectomy. It improves bone microarchitecture and preserves bone mineral content. Semaglutide ameliorates ovariectomy-induced osteoporosis and increases the expression of β-catenin, leading to increased bone formation and halted receptor activator of nuclear factor kappa-Β ligand (RANKL's) activation. Semaglutide can be used as a potential prophylactic and therapeutic drug against osteoporosis, possibly by activating Wnt signaling and decreasing bone resorption.

Wnt signaling activation confers a syncytiotrophoblast progenitor state on trophoblast stem cells of cynomolgus monkey.

Trophoblast stem cells (TSCs), derived from the trophectoderm of the blastocyst, are used as an in vitro model to reveal the mechanisms underlying placentation in mammals. In humans, suitable culture conditions for TSC derivation have recently been established. The established human TSCs (hTSCs) differentiate efficiently toward two trophoblast subtypes: syncytiotrophoblasts (STBs) and extravillous trophoblasts (EVTs). However, the efficiency of differentiation is lower in macaque TSCs than in hTSCs. Here, we demonstrate that the activation of Wnt signaling downregulated the expression of inhibitory G protein and induced trophoblastic lineage switching to the STB progenitor state. The treatment of macaque TSCs with a GSK-3 inhibitor, CHIR99021, upregulated STB progenitor markers and enhanced proliferation. Under the Wnt signaling-activated conditions, macaque TSCs effectively differentiated to STBs upon dbcAMP and forskolin treatment. RNA-seq analyses revealed the downregulation of inhibitory G protein, which may make macaque TSCs responsive to forskolin. Interestingly, this lineage switching appeared to be reversible as the macaque TSCs lost responsiveness to forskolin upon the removal of CHIR99021. The ability to regulate the direction of macaque TSC differentiation would be advantageous in elucidating the mechanisms underlying placentation in non-human primates.

Structural basis of Frizzled 4 in recognition of Dishevelled 2 unveils mechanism of WNT signaling activation

WNT signaling is fundamental in development and homeostasis, but how the Frizzled receptors (FZDs) propagate signaling remains enigmatic. Here, we present the cryo-EM structure of FZD4 engaged with the DEP domain of Dishevelled 2 (DVL2), a key WNT transducer. We uncover a distinct binding mode where the DEP finger-loop inserts into the FZD4 cavity to form a hydrophobic interface. FZD4 intracellular loop 2 (ICL2) additionally anchors the complex through polar contacts. Mutagenesis validates the structural observations. The DEP interface is highly conserved in FZDs, indicating a universal mechanism by which FZDs engage with DVLs. We further reveal that DEP mimics G-protein/β-arrestin/GRK to recognize an active conformation of receptor, expanding current GPCR engagement models. Finally, we identify a distinct FZD4 dimerization interface. Our findings delineate the molecular determinants governing FZD/DVL assembly and propagation of WNT signaling, providing long-sought answers underlying WNT signal transduction.

CasRx-based Wnt activation promotes alveolar regeneration while ameliorating pulmonary fibrosis in a mouse model of lung injury

Wnt/β-catenin signaling is an attractive target for regenerative medicine. A powerful driver of stem cell activity and hence tissue regeneration, Wnt signaling can promote fibroblast proliferation and activation, leading to fibrosis, while prolonged Wnt signaling is potentially carcinogenic. Thus, to harness its therapeutic potential, the activation of Wnt signaling must be transient, reversible and tissue-specific. In the lung, Wnt signaling is essential for alveolar stem cell activity and alveolar regeneration, which is impaired in lung fibrosis. Activation of Wnt/β-catenin signaling in lung epithelium may have anti-fibrotic effects. Here, we used intratracheal AAV6 injection to selectively deliver CasRx into lung epithelium, where it reversibly activates Wnt signaling by simultaneously degrading mRNAs encoding Axin1 and Axin2, negative regulators of Wnt/β-catenin signaling. Interestingly, CasRx mediated Wnt activation specifically in lung epithelium not only promotes alveolar type II cell (AT2) proliferation and alveolar regeneration, but also inhibits lung fibrosis resulted from bleomycin-induced injury, relevant in both preventive and therapeutic settings. Our study offers an attractive strategy for treating pulmonary fibrosis, with general implications for regenerative medicine.

CD11b/CD86 involved in the microenvironment of colorectal cancer by promoting Wnt signaling activation.

Colorectal cancer (CRC) is a malignancy that arises within the gastrointestinal tract. Despite ongoing research, the etiology and pathogenesis of CRC remain elusive; particularly, the distribution and characteristics of tumor-associated macrophages is currently an active area of investigation in understanding the pathological progression and prevention of CRC. This study utilized CRC patient surgical samples, mouse models of colitis-associated cancer, colonic organoid, and co-culture cell line to examine the changes in CD11b/CD86 at different pathological region and detect the Wnt signaling pathway activity. Our findings revealed a sensitive and increased expression of CD11b from the early to the advanced CRC tissues and correlated with poor prognosis, while CD86 expression was reduced in advanced CRC tissues. CD133 expression was also elevated in advanced CRC tissues and mainly co-localized with CD11b, suggesting a positive regulatory effect of CD11b and CD133 expression that may contribute to CRC progression. In AOM/DSS mouse models, activation of the Wnt signaling pathway was associated with increased CD133 and CD11b expression. Invitro, THP-1 cell was induced to high expression of CD11b, and the above conditional cultural medium enhanced HCT116 cell colony number and CD133 protein expression. Furthermore, colonic crypts from AOM/DSS mouse models were isolated to culture, and the colonic organoids exhibited dilation and significant increases expression of CD133 and β-Catenin/N-P-B-Catenin. CD11b might be an important factor to participate the progress of CRC. And the high CD11b of CRC microenviroment might potentially promote CD133 expression and associate with Wnt signal activation.

1465P Potential impact of APC mutation on survival via immune evasion through WNT signaling activation in HER2-positive gastric cancer treated with trastuzumab (tmab), nivolumab (nivo), and chemotherapy

1465P Potential impact of APC mutation on survival via immune evasion through WNT signaling activation in HER2-positive gastric cancer treated with trastuzumab (tmab), nivolumab (nivo), and chemotherapy

The Effect of Tideglusib Application on Type 1 and Type 3 Collagen Expressions by Human Dental-Pulp Derived Stem Cells: A Preliminary Study.

Although Tideglusib cytotoxicity studies and its effects on human dental pulp-derived stem cells (DPSCs) have been examined in previous studies, there is no study investigating the expression of type 1 collagen and type 3 collagen by Tideglusib. The purpose of this study is to examine the effect of Wnt signaling activation using Tideglusib execution on human DPSC to determine its potential efficacy in collagen expression. Stem cell isolation was performed from five human third molar wisdom tooth pulps. DPSCs identified in only one sample were treated with 50 nM Tideglusib for 24 h and 1 week. Axin-2, type 1 and type 3 collagen expressions were evaluated by Western blot analysis. DPSCs without treatment served as a negative control. The Mann-Whitney U test was used for statistical analysis. The levels of type 1 collagen and Axin-2 in the test group were significantly higher than those in the control group at 24 h (P = 0.000, P = 0.001, respectively). Compared to the control group, a slight increase in type 3 collagen expression was observed in the test group at 24 h (P value = 0.063). Application of 50 nM Tideglusib for 1 week revealed marked decreases in type 1 and type 3 collagen expressions (P = 0.029, P = 0.038, respectively). In contrast, there was a significant increase in the level of Axin-2 (P = 0.000) compared to the control group. The fact that Wnt signaling pathway activation obtained by Tideglusib application on DPSCs confirmed by the finding in the increase of Axin-2 at short and long-term evaluation periods which is resulted in the increase in the type 1 collagen expression at 24 h and decrease at 1 week together with the decrease in type 3 collagen expression at 1 week warrants further studies to evaluate the effect of Tideglusib on extracellular matrix expression.

Small molecule valproic acid enhances ventral patterning of human neural tube organoids by regulating Wnt and Shh signalling.

Valproic acid (VPA), a clinically approved small molecule, has been reported to activate Wnt signalling that is critical for dorsal-ventral (DV) patterning of neural tube. However, little is known about the impact of VPA on DV patterning process. Here, we show that even though VPA has a negative impact on the early formation of human neural tube organoids (hNTOs), it significantly enhances the efficiency of ventrally patterned hNTOs, when VPA is added during the entire differentiation process. RNA sequencing and RT-qPCR analysis demonstrates VPA activates endogenous Wnt signalling in hNTOs. Surprisingly, transcriptome analysis also identifies upregulation of genes for degradation of GLI2 and GLI3 proteins, whose truncated fragment are transcriptional repressors of Shh signalling. The Western-blot analysis confirms the increase of GLI3R proteins after VPA treatment. Thus, VPA might enhance ventral patterning of hNTOs through both activating Wnt, which can antagonise Shh signalling by inducing GLI3 expression, and/or inhibiting Shh signalling by inducing GLI protein degradation. We further obtain results to show that VPA still increases patterning efficiency of hNTOs with a weak influence on their early formation when the initiation time of VPA is delayed and its duration is reduced. Taken together, this study demonstrates that VPA enhances the generation of more reproducible hNTOs with ventral patterning, opening the avenues for the applications of hNTOs in developmental biology and regenerative medicine.

MTF2 facilitates the advancement of osteosarcoma through mediating EZH2/SFRP1/Wnt signaling

BackgroundOsteosarcoma is a soft tissue neoplasm with elevated recurrence risk and highly metastatic potential. Metal response element binding transcriptional factor 2 (MTF2) has been revealed to exert multiple activities in human tissues. The present research was conducted to explore the functions and related response mechanism of MTF2 in osteosarcoma which have not been introduced yet.MethodsBioinformatics tools identified the differential MTF2 expression in osteosarcoma tissues. MTF2 expression in osteosarcoma cells was examined with Western blot. Cell Counting Kit-8 (CCK-8) assay, 5-Ethynyl-2’-deoxyuridine (EDU) staining, wound healing as well as transwell assays measured cell proliferation, migration and invasion, respectively. Flow cytometry assay detected the cellular apoptotic level. Western blot also measured the expressions of proteins associated with epithelial mesenchymal transition (EMT), apoptosis and enhancer of zeste homolog 2 (EZH2)/secreted frizzled-related protein 1 (SFRP1)/Wnt signaling. Co-immunoprecipitation (Co-IP) assay confirmed MTF2-EZH2 interaction.ResultsMTF2 expression was increased in osteosarcoma tissues and cells. MTF2 interference effectively inhibited the proliferation, migration and invasion of osteosarcoma cells and promoted the cellular apoptotic rate. MTF2 directly bound to EZH2 and MTF2 silence reduced EZH2 expression, activated SFRP1 expression and blocked Wnt signaling in osteosarcoma cells. EZH2 upregulation or SFRP1 antagonist WAY-316606 partly counteracted the impacts of MTF2 down-regulation on the SFRP1/Wnt signaling and the biological phenotypes of osteosarcoma cells.ConclusionsMTF2 might down-regulate SFRP1 to activate Wnt signaling and drive the progression of osteosarcoma via interaction with EZH2 protein.

Three-dimensional culture in a bioengineered matrix and somatic cell complementation to improve growth and survival of bovine preantral follicles.

Here we explored poly(ethylene glycol) (PEG) bioengineered hydrogels for bovine preantral follicle culture with or without ovarian cell co-culture and examined the potential for differentiation of bovine embryonic stem cells (bESCs) towards gonadal somatic cells to develop a system more similar to the ovarian microenvironment. Bovine preantral follicles were first cultured in two-dimensional (2D) control or within PEG hydrogels (3D) and then co-cultured within PEG hydrogels with bovine ovarian cells (BOCs) to determine growth and viability. Finally, we tested conditions to drive differentiation of bESCs towards the intermediate mesoderm and bipotential gonad fate. Primary follicles grew over the 10-day culture period in PEG hydrogels compared to 2D control. Early secondary follicles maintained a similar diameter within the PEG while control follicles decreased in size. Follicles lost viability after co-encapsulation with BOCs; BOCs lost stromal cell signature over the culture period within hydrogels. Induction of bESCs towards gonadal somatic fate under WNT signaling was sufficient to upregulate intermediate mesoderm ( LHX1 ) and early coelomic epithelium/bipotential gonad markers ( OSR1 , GATA4 , WT1 ). Higher BMP4 concentrations upregulated the lateral plate mesoderm marker FOXF1 . PAX3 expression was not induced, indicating absence of the paraxial mesoderm lineage. Culture of primary stage preantral follicles in PEG hydrogels promoted growth compared to controls; BOCs did not maintain identity in the PEG hydrogels. Collectively, we demonstrate that PEG hydrogels can be a potential culture system for early preantral follicles pending refinements, which could include addition of ESC-derived ovarian somatic cells using the protocol described here. We demonstrate that three-dimensional bioengineered hydrogels could aid in the survival and growth of small bovine preantral follicles. Moreover, bovine embryonic stem cells have the potential to differentiate towards precursors of somatic gonadal cell types, presenting an alternative cell source for preantral follicle co-culture.

Gut-derived memory γδ T17 cells exacerbate sepsis-induced acute lung injury in mice

Sepsis is a critical global health concern linked to high mortality rates, often due to acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). While the gut-lung axis involvement in ALI is recognized, direct migration of gut immune cells to the lung remains unclear. Our study reveals sepsis-induced migration of γδ T17 cells from the small intestine to the lung, triggering an IL-17A-dominated inflammatory response in mice. Wnt signaling activation in alveolar macrophages drives CCL1 upregulation, facilitating γδ T17 cell migration. CD44+ Ly6C– IL-7Rhigh CD8low cells are the primary migratory subtype exacerbating ALI. Esketamine attenuates ALI by inhibiting pulmonary Wnt/β-catenin signaling-mediated migration. This work underscores the pivotal role of direct gut-to-lung memory γδ T17 cell migration in septic ALI and clarifies the importance of localized IL-17A elevation in the lung.

Epigenetic Regulation and Molecular Mechanisms of Burn Injury-Induced Nociception in the Spinal Cord of Mice.

Epigenetic mechanisms, including histone post-translational modifications (PTMs), play a critical role in regulating pain perception and the pathophysiology of burn injury. However, the epigenetic regulation and molecular mechanisms underlying burn injury-induced pain remain insufficiently explored. Spinal dynorphinergic (Pdyn) neurons contribute to heat hyperalgesia induced by severe scalding-type burn injury through p-S10H3-dependent signaling. Beyond p-S10H3, burn injury may impact various other histone H3 PTMs. Double immunofluorescent staining and histone H3 protein analyses demonstrated significant hypermethylation at H3K4me1 and H3K4me3 sites and hyperphosphorylation at S10H3 within the spinal cord. By analyzing Pdyn neurons in the spinal dorsal horn, we found evidence of chromatin activation with a significant elevation in p-S10H3 immunoreactivity. We used RNA-seq analysis to compare the effects of burn injury and formalin-induced inflammatory pain on spinal cord transcriptomic profiles. We identified 98 DEGs for burn injury and 86 DEGs for formalin-induced inflammatory pain. A limited number of shared differentially expressed genes (DEGs) suggest distinct central pain processing mechanisms between burn injury and formalin models. KEGG pathway analysis supported this divergence, with burn injury activating Wnt signaling. This study enhances our understanding of burn injury mechanisms and uncovers converging and diverging pathways in pain models with different origins.

Abstract We091: Maternal hyperglycemia disrupts human cardiac cell lineage differentiation

Maternal diabetes is associated with a 4-fold increased risk of offspring developing congenital heart defects (CHDs). It remains unknown how maternal diabetes interferes with cardiac cell lineage determination and leads to malformations in the heart. In this study, we aim to elucidate the cellular mechanisms by which maternal hyperglycemia causes high risk of CHDs in newborns. Here we leverage an in vitro hyperglycemic model using human induced pluripotent stem cells (iPSCs), which could recapture cardiac differentiation and cell lineage commitment during embryonic heart development. We collected differentiating cells at D5 (cardiac mesoderm), D10 (cardiac progenitors), and D14 (early cardiomyocytes) during cardiac differentiation under normal and hyperglycemic conditions, and performed single-cell transcriptomic analysis. We found that hyperglycemia significantly impedes cardiac differentiation of human iPSCs as robust cardiac differentiation is rarely observed in multiple iPSC lines (n=10) under hyperglycemia. At the cellular level, hyperglycemia interferes with cardiac differentiation of human iPSCs in response to WNT signaling activation, which is manifested by reduced number of cardiac mesoderm (MESP1+ PDGFRA+) cells at D5 of differentiation. In contrast, neural differentiation is enhanced under hyperglycemia, with a high proportion of neural cell lineage (SOX2+ PAX6+). At D10, differentiated neural cell lineage dominates the cell population under hyperglycemia at the expense of cardiac progenitors and early cardiomyocytes. At D14, the prevalence of neural lineage persist whereas early cardiomyocytes only accounts for a small portion of cell population under hyperglycemia. Moreover, we treated D30 iPSC-derived cardiomyocytes (iPSC-CMs) with high glucose concentration (25 mM) for 7 days and found that iPSC-CMs show reduced mitochondria respiration and ATP production, but elevated apoptosis and reactive oxygen species (ROS) generation. Together, our data suggest that maternal hyperglycemia could interrupt human embryonic heart morphogenesis through overriding WNT-medicated cardiac differentiation and promoting neural cell lineage determination by default.

GSK3β Inhibitors Inhibit TGFβ Signaling in the Human Trabecular Meshwork.

Primary open-angle glaucoma (POAG) is a leading cause of blindness, and its primary risk factor is elevated intraocular pressure (IOP) due to pathologic changes in the trabecular meshwork (TM). We previously showed that there is a cross-inhibition between TGFβ and Wnt signaling pathways in the TM. In this study, we determined if activation of the Wnt signaling pathway using small-molecule Wnt activators can inhibit TGFβ2-induced TM changes and ocular hypertension (OHT). Primary human TM (pHTM) cells and transduced SBE-GTM3 cells were treated with or without Wnt and/or TGFβ signaling activators and used for luciferase assays; for the extraction of whole-cell lysate, conditioned medium, cytosolic proteins, and nuclear proteins for Western immunoblotting (WB); or for immunofluorescent staining. Human donor eyes were perfusion cultured to study the effect of Wnt activators on IOP. We found that the small-molecule Wnt activators (GSK3β inhibitors) (BIO, SB216763, and CHIR99021) activated canonical Wnt signaling in pHTM cells without toxicity at tested concentrations. This activation inhibited TGFβ signaling as well as TGFβ2-induced extracellular matrix deposition and formation of cross-linked actin networks in pHTM cells or SBE-GTM3 cells. We also observed nuclear translocation of both Smad4 and β-catenin in pHTM cells, which suggested that the cross-inhibition between the TGFβ and Wnt signaling pathways may occur in the nucleus. Using our ex vivo model, we found that CHIR99021 inhibited TGFβ2-induced OHT in perfusion-cultured human eyes. Our results showed that small-molecule Wnt activators have the potential for treating TGFβ signaling-induced OHT in patients with POAG.

Wnt-5a/Ca2+ pathway modulates endogenous current and oocyte structure of Xenopus laevis

Wnt signaling plays an essential role in cellular processes like development, maturation, and function maintenance. Xenopus laevis oocytes are a suitable model to study not only the development but also the function of different receptors expressed in their membranes, like those receptors expressed in the central nervous system (CNS) including Frizzled 7. Here, using frog oocytes and recordings of endogenous membrane currents in a two-electrode path configuration along with morphological observations, we evaluated the role of the non-canonical Wnt-5a ligand in oocytes. We found that acute application of Wnt-5a generated changes in endogenous calcium-dependent currents, entry oscillatory current, the membrane's outward current, and induced membrane depolarization. The incubation of oocytes with Wnt-5a caused a reduction of the membrane potential, potassium outward current, and protected the ATP current in the epithelium/theca removed (ETR) model. The oocytes exposed to Wnt-5a showed increased viability and an increase in the percentage of the germinal vesicle breakdown (GVBD), at a higher level than the control with progesterone. Altogether, our results suggest that Wnt-5a modulates different aspects of oocyte structure and generates calcium-dependent endogenous current alteration and GVDB process with a change in membrane potential at different concentrations and times of the exposition. These results help to understand the cellular effect of Wnt-5a and present the use of Xenopus oocytes to explore the mechanism that could impact the activation of Wnt signaling.

Unraveling Cancer's Wnt Signaling: Dynamic Control through Protein Kinase Regulation.

Since the initial identification of oncogenic Wnt in mice and Drosophila, the Wnt signaling pathway has been subjected to thorough and extensive investigation. Persistent activation of Wnt signaling exerts diverse cancer characteristics, encompassing tumor initiation, tumor growth, cell senescence, cell death, differentiation, and metastasis. Here we review the principal signaling mechanisms and the regulatory influence of pathway-intrinsic and extrinsic kinases on cancer progression. Additionally, we underscore the divergences and intricate interplays of the canonical and non-canonical Wnt signaling pathways and their critical influence in cancer pathophysiology, exhibiting both growth-promoting and growth-suppressing roles across diverse cancer types.