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To provide an update on the current understanding of the role of wingless/integrase-1 (Wnt) signaling in pediatric allergic asthma and other pediatric lung diseases.The Wnt signaling pathway is critical for normal lung development. Genetic and epigenetic human studies indicate a link between Wnt signaling and the development and severity of asthma in children. Mechanistic studies using animal models of allergic asthma demonstrate a key role for Wnt signaling in allergic airway inflammation and remodeling. More recently, data on bronchopulmonary dysplasia (BPD) pathogenesis points to the Wnt signaling pathway as an important regulator.Current data indicates that the Wnt signaling pathway is an important mediator in allergic asthma and BPD pathogenesis. Further studies are needed to characterize the roles of individual Wnt signals in childhood disease, and to identify potential novel therapeutic targets to slow or prevent disease processes.

Diabetic mellitus is a worldwide endocrine and metabolic disorder with insulin insensitivity or deficiency or both whose prevalence could rise up to 592 million by 2035. Consistent hyperglycemia leads to one of the most common comorbidities like Diabetic Peripheral Neuropathy (DPN). DPN is underlined with unpleasant sensory experience, such as tingling and burning sensation, hyperalgesia, numbness, etc. Globally, 50-60% of the diabetic population is suffering from such symptoms as microvascular complications. Consistent hyperglycemia during DM causes activation/inhibition of various pathways playing important role in the homeostasis of neurons and other cells. Disruption of these pathways results into apoptosis and mitochondrial dysfunctions, causing neuropathy. Among these, pathways like Polyol and PARP are some of the most intensively studied ones whereas those like Wnt pathway, Mitogen activated protein kinase (MAPK), mTOR pathway are comparatively newly discovered. Understanding of these pathways and their role in pathophysiology of DN underlines a few molecules of immense therapeutic value. The inhibitors or activators of these molecules can be of therapeutic importance in the management of DPN. This review, hence, focuses on these underlying molecular mechanisms intending to provide therapeutically effective molecular targets for the treatment of DPN.

OBJECTIVE: Since Wnt signaling plays an important role in both tooth agenesis and altered intestine homeostasis, the aim was to compare gastrointestinal symptoms in patients with isolated oligodontia caused by a Wnt pathway gene mutation and controls.METHODS: A case-control study was designed to compare self-reported gastrointestinal symptoms among patients with isolated oligodontia, caused by a Wnt signaling gene mutation, and fully dentate controls. The Gastrointestinal Symptom Rating Scale (GSRS) was used to assess gastrointestinal symptoms. Prevalence and severity of gastrointestinal symptoms among patients and age- and gender-matched controls was evaluated.RESULTS: Twenty patients with isolated oligodontia and a pathogenic variant in the wnt pathway genes WNT10A, LRP6 or PAX9 participated. The prevalence of gastrointestinal symptoms was higher in the oligodontia patients compared to their controls (Χ2 (1) = 87.33, p = .008). Mean GSRS total scores (p = .011) and domain scores for 'abdominal pain' (p = .022), 'reflux' (p = .003) and constipation (p = .030) were higher for these oligodontia patients compared to their controls.CONCLUSION: Gastrointestinal symptoms are more prevalent and more severe in patients with isolated oligodontia and a deficiency in a Wnt pathway related gene, when compared to controls without tooth agenesis.

Menopausal women often face long-term estrogen treatment. G protein-coupled estrogen receptor (GPER) expressed in intestinal crypt was activated by estrogen therapy, but it was unclear whether chronic GPER activation during menopause had an effect on intestinal stem cells (ISCs). We tested the effect of chronic GPER activation on ISCs of ovariectomized (OVX) mice by injection of the selective GPER agonist G-1 for 28 days, or G-1 stimulation of organoids derived from crypts of OVX mice. G-1 up-regulated crypt depth, the number of Ki67+, bromodeoxyuridine+ cells and Olfm4+ ISCs, and the expression of ISCs marker genes (Lgr5, Olfm4 and Axin2). G-1 administration promoted organoid growth, increased the number of EdU+ cells per organoid and protein expression of Cyclin D1 and cyclin B1 in organoids. After G-1 treatment in vivo or in vitro, Paneth cell–derived Wnt3, Wnt3 effector β-catenin and Wnt target genes c-Myc and Cyclin D1 increased in ileum or organoids. Once blocking the secretion of Wnt3 from Paneth cells, the effects of G-1 on organoids growth, ISCs marker genes and Wnt/β-catenin signaling were abolished. G-1 did not affect the number of Paneth cells in ex vivo organoids, while activated Mmp7/cryptdin program in Paneth cells, promoted their maturation, and increased the expression of lysozyme protein. G-1 pretreatment in OVX mice inhibited radiation-induced ISCs proliferation injury and enhanced the resistance of mice to intestinal injury. In conclusion, chronic GPER activation prompted the Wnt3 synthesis in Paneth cells, thus increased the proliferation of ISCs via activation of Wnt3/β-catenin signaling in OVX mice.

One of the central challenges for cancer therapy is the identification of factors in the tumor microenvironment that increase tumor progression and immune tolerance. In breast cancer, fibrosis is a histopathologic criterion for invasive cancer and poor survival that results from inflammatory factors and remodeling of the extracellular matrix to produce an immune tolerant microenvironment. To determine whether tolerance is associated with the immune checkpoint, Programmed Cell Death 1 (PD-1), NeuT/ATTAC mice, a conditional model of mammary fibrosis that we recently developed, were administered a murine-specific anti-PD-1 mAb related to pembrolizumab, and drug response was monitored by tumor development, imaging mass cytometry, immunohistochemistry and tumor gene expression by RNAseq. Tumor progression in NeuT/ATTAC mice was unaffected by weekly injection of anti-PD-1 over four months. Insensitivity to anti-PD-1 was associated with several processes, including increased tumor-associated macrophages (TAM), epithelial to mesenchymal transition (EMT), fibroblast proliferation, an enhanced extracellular matrix and the Wnt signaling pathway, including increased expression of Fzd5, Wnt5a, Vimentin, Mmp3, Col2a1 and Tgfβ1. These results suggest potential therapeutic avenues that may enhance PD-1 immune checkpoint sensitivity, including the use of tumor microenvironment targeted agents and Wnt pathway inhibitors.

Background. We aimed to evaluate the prognostic value of E2F4 expression in oral squamous cell carcinoma (OSCC) and clarify its influence on immune cell infiltration and biological functions. Methods. The Cancer Genome Atlas (TCGA) database, the STRING database, and related online tools as well as single-sample gene set enrichment analysis (ssGSEA) were used for the analyses in our study. Results. The E2F4 expression was elevated in OSCC tumor tissue compared with paracancerous tissues. The high expression of E2F4 was closely related to the poorer overall survival, disease-free survival, and progression-free interval of OSCC. In addition, pathway enrichment analyses revealed that the top 49 genes most closely related to E2F4 were strongly associated with the cell cycle. E2F4-related proteins were closely related to the following KEGG pathways: cell cycle, cellular senescence, PI3K-Akt signaling pathway, Wnt signaling pathway, and notch signaling pathway. It was also demonstrated that the E2F4 expression was negatively correlated with immune purity and strongly related to immune cell infiltration in OSCC. Conclusions. E2F4 can be used as a novel biomarker for the diagnosis and prognosis of OSCC.

Wnt signalling is an essential player in tissue formation, notably in the regulation of stem cell function. Wnt signalling is best known for its roles in G1/S progression. However, a complex Wnt programme that also mediates mitotic progression and asymmetric cell division (ACD) is emerging. Recent developments in this area have provided mechanistic insights as well as tools to engineer or target Wnt signalling for translational and therapeutic purposes. Here, we discuss the bidirectional relationship between Wnt activity and mitosis. We emphasise how various Wnt-dependent mechanisms control spindle dynamics, chromosome segregation, and ACD. Finally, we illustrate how knowledge about these mechanisms has been successfully employed in tissue engineering for regenerative medicine applications.

Background: Barbaloin, found in Aloe vera, exerts broad pharmacological activities, including antioxidant, anti-inflammatory, and anti-cancer. This study aims to investigate the effects of barbaloin on the osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs). Methods: Osteogenic induction of hBMSCs was performed in the presence or absence of barbaloin. Cell viability was determined by using the CCK-8 assay. The characteristic of hBMSCs was identified by using flow cytometry. Intracellular alkaline phosphatase (ALP) staining was performed to evaluate the ALP activity in hBMSCs. Alizarin Red S staining was performed to evaluate the matrix mineralization. The mRNA and protein levels of target genes were determined using qRT-PCR and western blotting, respectively. Results: Treatment with barbaloin (10 and 20 μg/mL) significantly increased cell viability of hBMSCs after 72 hours. In addition, treatment with barbaloin increased the mRNA expression levels of ALP and its activities. Treatment with barbaloin also increased matrix mineralization and the mRNA and protein levels of late-differentiated osteoblast marker genes BMP2, RUNX2, and SP7 in hBMSCs. The underlying mechanisms revealed that barbaloin increased the protein expressions of Wnt/β-catenin pathway-related biomarkers. Conclusion: Barbaloin promotes osteogenic differentiation of hBMSCs by the regulation of the Wnt/β-catenin signaling pathway.

Aberrant activation of Wnt/β-catenin induces renal dysfunction by initiating pro-apoptotic cascades, fibrosis, oxidative and inflammatory burden. This study tested the therapeutic effects of Wnt/β-catenin inhibitor pyrvinium against cisplatin-induced acute kidney injury (AKI) in rats. Cisplatin was administered at a single dose of 5 mg/kg (i.p.) and renal cisplatin accumulation and uptake in cortical slices were determined after the fifth day by atomic absorption spectroscopy. Levels of pro-inflammatory cytokines were checked by ELISA, and organic cation transporter-2 (OCT-2) transcription and expression in renal tissue were evaluated by RT-PCR and immunohistochemical technique. Cisplatin administration produced renal dysfunction manifested as increase in serum creatinine, blood urea nitrogen, proteinuria, reduced clearance and electrolyte imbalance. Oxidative stress indices, pro-inflammatory cytokines, fibronectin, and caspase-3 activity were elevated in cisplatin-challenged rats. Moreover, increased renal OCT-2 transcription and immunostaining were detected in cisplatin kidneys which resulted in platinum accumulation. Additional docking studies depicted strong interaction between the β-catenin and OCT-2 protein. These manifestations induced mitochondrial dysfunction, histological damage and fibrosis. Notably, Wnt/β-catenin inhibitor pyrvinium (60 µg/kg; p.o.) treatment reduced the renal OCT-2 gene transcription causing a decline in platinum levels. Thus, the present study concludes that Wnt/β-catenin inhibition attenuates cisplatin-induced AKI in rats, partly by down-regulating OCT-2 expression.

The microbiome plays a fundamental role in maintaining intestinal stem cell (ISC)-niche equilibrium. In this issue of Immunity, Kim and colleagues uncover a mechanism by which the microbiota drives macrophage WNT ligand-release to maintain ISC-niche homeostasis during early postnatal development.

Wnts are short-range signaling proteins, expressed in all metazoans from sponges to humans, critical for cell development and fate. There are 19 different Wnts in the human genome with varying expression levels and patterns, and post-translational modifications. Common to essentially all Wnts is the palmitoleation of a conserved serine by the O-acyltransferase PORCN in the endoplasmic reticulum (ER). All lipidated Wnts then bind a dedicated carrier Wntless (WLS), endowed with the task of transporting them from the ER to the plasma membrane, and ultimately facilitating their release to receptors on the Wnt-receiving cell to initiate signaling. Here, we will focus on the WLS-mediated transport step. There are currently two published structures, both obtained by single-particle cryo-electron microscopy of the Wnt/WLS complex: human Wnt8A-bound and human Wnt3A-bound WLS. We analyze the two Wnt/WLS structures — remarkably similar despite the sequence similarity between Wnt8A and Wnt3A being only ∼39% — to begin to understand the conserved nature of this binding mechanism, and ultimately how one carrier can accommodate a family of 19 different Wnts. By comparing how Wnt associates with WLS with how it binds to PORCN and FZD receptors, we can begin to speculate on mechanisms of Wnt transfer from PORCN to WLS, and from WLS to FZD, thus providing molecular-level insight into these essential steps of the Wnt signaling pathway.

Bladder cancer is a highly prevalent disease worldwide. We aimed to investigate the effect of miRNA/mRNA signaling on bladder urothelial carcinoma (BUC).MiRNA-139-3p wasselected from The Cancer Genome Atlas database, and its downstream target gene was predicted. The correlation between miRNA-139-3p and intersected mRNAs was analyzed. The mRNA expression levels of miRNA-139-3p and KIF18B in BUC were assayed via quantitative real-time polymerase chain reaction. Effects of miRNA-139-3p on cell proliferation, invasion, migration and cell cycle were detected via Cell Counting Kit-8, colony formation, transwell, wound healing and flow cytometry assays, respectively. Binding relationship between miRNA-139-3p and KIF18B was verified by dual-luciferase reporter gene detection. The protein expression levels of KIF18B, β-catenin and Cyclin D1 were detected by Western blot. Rescue assays were performed for verifying the interaction among miRNA-139-3p, KIF18B and Wnt/β-catenin signaling pathway, which revealed effects of miRNA-139-3p/KIF18B on BUC cells.MiRNA-139-3p was remarkably underexpressed, and KIF18B was dramatically overexpressed in BUC cells, respectively. It was also demonstrated that overexpressing miRNA-139-3p could prominently inhibit proliferation, invasion and migration of BUC, and block BUC cells at G0-G1 phase. Afterwards, we found that miRNA-139-3p could bind to KIF18B mRNA 3'UTR, and miRNA-139-3p had a negative regulatory effect with KIF18B. Subsequent experimental results presented that overexpressing KIF18B could reverse inhibitory effect of overexpressing miRNA-139-3p on BUC. Finally, this study also ascertained that miRNA-139-3p/KIF18B could repress oncogenic effects of BUC via modulating Wnt/β-catenin signaling pathway.MiRNA-139-3p/KIF18B/Wnt/β-catenin could significantly inhibit the malignant progression of BUC, and its targeting mechanism might provide an effective therapeutic target for BUC patients.

Proline/arginine-rich end leucine-rich repeat protein (PRELP) has been reported to contribute to the remodelling of cardiovascular tissues in the ischaemia-reperfusion injury model. However, research is lacking on the role of PRELP in myocardial fibrosis and ventricular remodelling, and the mechanism through which PRELP brings about these changes is not clear. This study aimed to evaluate the role of PRELP in ventricular remodelling and myocardial fibrosis following acute myocardial infarction (AMI) and to explore the underlying mechanism.In this study, we established AMI mouse and cellculture models in an oxygen-glucose deprivation environment.We found that over-expression of PRELP increased the infarct size and interstitial fibrotic area. Expression of the wnt/β-catenin pathway molecules, which are downstream of PRELP, increased more in the PRELP over-expression group than in the AMI group.Our results showed that PRELP, through the wnt/β-catenin signalling pathway, led to myocardial fibrosis and ventricular remodelling following AMI.

Failure to regenerate injured alveoli functionally and promptly causes a high incidence of fatality in coronavirus disease 2019 (COVID-19). How elevated plasminogen activator inhibitor-1 (PAI-1) regulates the lineage of alveolar type 2 (AT2) cells for re-alveolarization has not been studied. This study aimed to examine the role of PAI-1-Wnt5a-β catenin cascades in AT2 fate. Dramatic reduction in AT2 yield was observed in Serpine1Tg mice. Elevated PAI-1 level suppressed organoid number, development efficiency, and total surface area in vitro. Anti-PAI-1 neutralizing antibody restored organoid number, proliferation and differentiation of AT2 cells, and β-catenin level in organoids. Both Wnt family member 5A (Wnt5a) and Wnt5a-derived N-butyloxycarbonyl hexapeptide (Box5) altered the lineage of AT2 cells. This study demonstrates that elevated PAI-1 regulates AT2 proliferation and differentiation via the Wnt5a/β catenin cascades. PAI-1 could serve as autocrine signaling for lung injury repair.

Introduction: It has been shown that mechanical forces can induce or promote osteogenic differentiation as well as remodeling of the new created bone tissues. To apply this characteristic in bone tissue engineering, it is important to know which mechanical stimuli through which signaling pathway has a more significant impact on osteogenesis. Methods: In this systematic study, an electronic search was conducted using PubMed and Google Scholar databases. This study has been prepared and organized according to the preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines. Included studies were first categorized according to the in vivo and in vitro studies. Results: Six types of mechanical stresses were used in these articles and the most commonly used mechanical force and cell source were tension and bone marrow-derived mesenchymal stem cells (BMMSCs), respectively. These forces were able to trigger twelve signaling pathways in which Wnt pathway was so prominent. Conclusion: 1) Although specific signaling pathways are induced through specific mechanical forces, Wnt signaling pathways are predominantly activated by almost all types of force/stimulation, 2) All signaling pathways regulate expression of RUNX2, which is known as a master regulator of osteogenesis, 3) In Tension force, the mode of force administration, i.e, continuous or noncontinuous tension is more important than the percentage of elongation.

Bone is a rigid, mineralized connective tissue that constitutes part of the skeleton in most vertebrate animals. Bone remodeling is a complex process that involves the coordination of ossification and bone resorption activities by osteoblasts and osteoclasts, respectively, resulting in maintaining bone mass. This process involves several growth factors/cytokines and hormones regulating the various signaling pathways. Wnt is one of the major molecular signaling pathways that positively regulate the osteogenic differentiation of mesenchymal stem cells. Dysregulation in the Wnt signaling leads to serious bone-related disorders like osteoporosis and osteosclerosis. Recently, several studies reported the critical role of non-coding RNAs (ncRNAs), including microRNAs, long ncRNAs, and circular RNAs, in the regulation of bone homeostasis via modulating the Wnt signaling cascade. This review summarizes the importance of such ncRNAs in mediating the Wnt cascade and its effect on osteoblast differentiation. Understanding the regulatory role of these ncRNAs would serve as a novel therapeutic strategy for treating bone-related disorders.

We have synthesized an oxetane derivative of the benzimidazole compound mebendazole (OBD9) with enhanced solubility and strong anticancer activity in multiple types of cancer cells, especially colorectal cancer. In this report, we provide evidence that OBD9 suppresses colorectal cancer growth by interfering with the Wnt signaling pathway, a main driver of cell growth in colorectal cancer. Specifically, we find that OBD9 induces autophagic degradation of TNIK (traf2 and Nck-interacting kinase), which promotes T-cell factor-4 (TCF4)/beta-catenin–mediated gene expression. Thus, OBD9 as a TNIK inhibitor blocks Wnt/beta-catenin signaling at the final step of transcriptional activation. We suggest that OBD9 provides a potential novel autophagy-mediated, Wnt-damping therapeutic strategy for the treatment of colorectal cancer.

Dysfunction of cardiomyocytes contributes to the development of acute myocardial infarction (AMI). Nonetheless, the regulatory mechanism of lncRNA myocardial infarction-associated transcript (MIAT) in cardiomyocyte injury remains largely unclear. The cardiomyocyte injury was assessed via cell viability and apoptosis using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) and flow cytometry, respectively. The levels of MIAT, microRNA (miR)-488-3p, and Wnt5a were detected via quantitative real-time polymerase chain reaction and Western blot. After bioinformatical analysis, the binding between miR-488-3p and MIAT or Wnt5a was confirmed via dual-luciferase reporter, RNA immunoprecipitation, and RNA pull-down assays. Our results showed that MIAT expression was increased in AC16 cells after hypoxia treatment. Silencing of MIAT alleviated hypoxia-induced viability reduction, apoptosis increase, and Wnt/β-catenin pathway activation. MIAT directly targeted miR-488-3p. MiR-488-3p might repress hypoxia-induced cardiomyocyte injury, and its knockdown reversed the effect of MIAT depletion on cardiomyocyte injury. Wnt5a was validated as a target of miR-488-3p. Wnt5a expression restoration attenuated the influence of MIAT knockdown on hypoxia-triggered cardiomyocyte injury. Our findings demonstrated that downregulation of MIAT might mitigate hypoxia-induced cardiomyocyte injury partly through miR-488-3p mediated Wnt/β-catenin pathway.

This study investigated the preventive effects of α-ketoglutarate (α-KG, in the form of sodium salt) on a

Colorectal cancer (CRC) is one of the most frequent cancers leading to death worldwide. Different signaling pathways such as the canonical Wnt signaling pathway have many effects on the development of CRC. MicroRNAs are small non-coding RNAs and different evidence represent their importance in the development of cancer via regulating the expression of their target genes. miRNAs can affect CRC progression as oncogenes or tumor suppressors. Dysregulation in miRNA expression can occur for various reasons, causes different abnormalities in the Wnt signaling pathway, and contributes to CRC development. Identifying the exact interactions between microRNAs and mRNAs or other non-coding RNAs assists in designing effective therapeutic, diagnostic, and prognostic approaches. In this review, we aim to focus on microRNAs that regulate CRC through modulating the Wnt pathway and then present the perspective outlook on the implication of miRNA in liquid biopsy for the management of patients with CRC.

The interplay between apoptotic cancer cells and the tumor microenvironment modulates cancer progression and metastasis. Cancer-associated fibroblasts (CAFs) play a crucial role in promoting these events through paracrine communication. Here, we demonstrate that conditioned medium (CM) from lung CAFs exposed to apoptotic cancer cells suppresses TGF-β1-induced migration and invasion of cancer cells and CAFs. Direct exposure of CAFs to apoptotic 344SQ cells (ApoSQ) inhibited CAF migration and invasion and the expression of CAF activation markers. Enhanced secretion of Wnt‐induced signaling protein 1 (WISP-1) by CAFs exposed to ApoSQ was required for these antimigratory and anti-invasive effects. Pharmacological inhibition of Notch1 activation or siRNA-mediated Notch1 silencing prevented WISP-1 production by CAFs and reversed the antimigratory and anti-invasive effects. Enhanced expression of the Notch ligand delta-like protein 1 on the surface of ultraviolet-irradiated apoptotic lung cancer cells triggered Notch1-WISP-1 signaling. Phosphatidylserine receptor brain-specific angiogenesis inhibitor 1 (BAI1)-Rac1 signaling, which facilitated efferocytosis by CAFs, participated in crosstalk with Notch1 signaling for optimal production of WISP-1. In addition, a single injection of ApoSQ enhanced WISP-1 production, suppressed the expression of CAF activation markers in isolated Thy1+ CAFs, and inhibited lung metastasis in syngeneic immunocompetent mice via Notch1 signaling. Treatment with CM from CAFs exposed to ApoSQ suppressed tumor growth and lung metastasis, whereas treatment with WISP-1-immunodepleted CM from CAFs exposed to ApoSQ reversed the antitumorigenic and antimetastatic effects. Therefore, treatment with CM from CAFs exposed to apoptotic lung cancer cells could be therapeutically applied to suppress CAF activation, thereby preventing cancer progression and metastasis.

Tumor-associated macrophages (TAM) play an important role in supporting tumor growth and suppressing antitumor immune responses, and TAM infiltration has been associated with poor patient prognosis in various cancers. TAMs can be classified as pro-inflammatory, M1-like, or anti-inflammatory, M2-like. While multiple factors within the tumor microenvironment affect the recruitment, polarization, and functions of TAMs, accumulating evidence suggests that Wnt signaling represents an important, targetable driver of an immunosuppressive, M2-like TAM phenotype. TAM production of Wnt ligands mediates TAM-tumor cross-talk to support cancer cell proliferation, invasion, and metastasis. Targeting TAM polarization and the protumorigenic functions of TAMs through inhibitors of Wnt signaling may prove a beneficial treatment strategy in cancers where macrophages are prevalent in the microenvironment.

Intestinal fibrosis and stricture formation is an aggressive complication of Crohns disease (CD), linked to increased morbidity and costs. The present study investigates the contribution of Wingless-Int-1 (Wnt) signalling to intestinal fibrogenesis, considers potential cross-talk between Wnt and transforming growth factor β1 (TGFβ) signalling pathways, and assesses the therapeutic potential of small-molecule Wnt inhibitors. β-catenin expression was explored by immunohistochemistry (IHC) in formalin-fixed paraffin embedded (FFPE) tissue from patient-matched nonstrictured (NSCD) and strictured (SCD) intestine (n=6 pairs). Functional interactions between Wnt activation, TGFβ signalling, and type I collagen (Collagen-I) expression were explored in CCD-18Co cells and primary CD myofibroblast cultures established from surgical resection specimens (n=16) using small-molecule Wnt inhibitors and molecular techniques, including siRNA-mediated gene knockdown, immunofluorescence (IF), Wnt gene expression arrays, and western blotting. Fibrotic SCD tissue was marked by an increase in β-catenin-positive cells. In vitro, activation of Wnt-β-catenin signalling increased Collagen-I expression in CCD-18Co cells. Conversely, ICG-001, an inhibitor of β-catenin signalling, reduced Collagen-I expression in cell lines and primary CD myofibroblasts. TGFβ increased β-catenin protein levels but did not activate canonical Wnt signalling. Rather, TGFβ up-regulated WNT5B, a noncanonical Wnt ligand, and the Wnt receptor FZD8, which contributed directly to the up-regulation of Collagen-I through a β-catenin-independent mechanism. Treatment of CCD-18Co fibroblasts and patient-derived myofibroblasts with the FZD8 inhibitor 3235-0367 reduced extracellular matrix (ECM) expression. Our data highlight small-molecule Wnt inhibitors of both canonical and noncanonical Wnt signalling, as potential antifibrotic drugs to treat SCD intestinal fibrosis. They also highlight the importance of the cross-talk between Wnt and TGFβ signalling pathways in CD intestinal fibrosis.

Gastric cancer is a one of the most common malignant tumors with poor prognosis worldwide. Leucine-rich G-protein-coupled receptor 5 (LGR5) is determined as a modulator of Wnt signaling cascade and R-spondins are a family of secretory agonists in the Wnt signaling and act as ligands to interact with LGR5. However, the function of Rspondin-1 in GC remains obscure. Here, we identified the effect of Rspondin-1 on GC progression. Rspondin-1 and LGR5 were upregulated in clinical gastric cancer tissues. CCK-8 assay revealed that the viability of GC cells was reduced by Rspondin-1 depletion and enhanced by Rspondin-1 overexpression. The depletion of Rspondin-1 decreased while the overexpression of Rspondin-1 increased the numbers of colony formation and Edu-positive GC cells. The depletion of Rspondin-1 attenuated the invasion and migration ability of GC cells. Moreover, sphere formation assays revealed that the knockdown of Rspondin-1 reduced the stemness of GC cells. The expression of cancer stem cell markers, including Nanog, OCT3/4, and SOX2 were suppressed by Rspondin-1 depletion in GC cells. Rspondin-1 induced tumor growth of gastric cancer cells in vivo. Mechanically, the cell viability and invasion suppressed by the depletion of Rspondin-1 in GC cells were rescued by LGR5 overexpression. Besides, the overexpression of LGR5 reversed Rspondin-1 knockdown-inhibited Nanog and OCT3/4 expression. Consequently, we concluded that Rspondin-1 contributes to the progression and stemness of gastric cancer by LGR5.

The abnormal activation of the Wnt/β-catenin signaling pathway and epithelial-mesenchymal transition (EMT) in drug-resistant tumor cells and cancer stem cells (CSCs) stimulate tumor metastasis and recurrence. Here, a promising combined chemotherapeutic strategy of salinomycin (SL) and doxorubicin (DOX) with specific inhibition of tumor stemness by a targeted co-delivery nanosystem was developed to overcome this abnormal progression. This strategy could be benefit drugs to effectively penetrate and infiltrate into spheres of 3D-cultured breast cancer stem cells (BCSCs). The expression of the Wnt/β-catenin signaling pathway-related genes (β-catenin, LRP6, LEF1, and TCF12) and target genes (Cyclin D1, Cmyc, and Fibronectin) as well as CSC stemness-related genes (Oct4, Nanog, and Hes1) was downregulated by redox-sensitive co-delivery micelles decorated with oligohyaluronic acid as the active targeting moiety. The changes in EMT-associated gene expression (E-cadherin and Vimentin) in vitro showed that the EMT process was also effectively inverted. This strategy achieved a strong inhibitory effect on solid tumor growth and an effective reduction in the risk of tumor metastasis in 4T1 tumor-bearing mice in vivo and effectively alleviated splenomegaly caused by the malignant tumor. Immunohistochemical staining analysis of E-cadherin, Vimentin, and β-catenin confirmed that the inversion of the EMT was also achieved in solid tumors. These results highlight the potential of SL and DOX combined chemotherapeutic strategy for eliminating breast carcinoma. STATEMENT OF SIGNIFICANCE: Cancer stem cells (CSCs), as an important part of tumor heterogeneity, can survive against conventional chemotherapy and initiate tumorigenesis, recurrence, and metastasis. Moreover, non-CSCs can convert into the CSC state through the abnormal Wnt/β-catenin pathway, which is closely related to the epithelial-mesenchymal transition (EMT) process. Here, redox-degradable binary drug-loaded micelles (PPH/DOX+SL) were designed to target CSCs and overcome drug resistance of breast cancer cells. The combined chemotherapy of salinomycin (SL) and doxorubicin (DOX) reversed drug resistance, while the PPH/DOX+SL micelles enhanced the intracellular accumulation and drug penetration of BCSC spheres. The introduction of SL downregulated the expression of tumor stemness genes and the Wnt/β-catenin pathway-related genes and inverted the EMT process. PPH/DOX+SL continuously inhibited tumor growth and invasion in vivo.

Mounting evidence has demonstrated that a myriad of developmental signaling pathways, such as the Wnt, Notch, Hedgehog and Hippo, are frequently deregulated and play a critical role in regulating cancer stem cell (CSC) activity in human cancers, including gynecologic malignancies. In this review article, we describe an overview of various signaling pathways in human cancers. We further discuss the developmental roles how these pathways regulate CSCs from experimental evidences in gynecologic cancers. Moreover, we mention several compounds targeting CSCs in gynecologic cancers to enhance the treatment outcomes. Therefore, these signaling pathways might be the potential targets for developing targeted therapy in gynecologic cancers.

Imatinib, an inhibitor of tyrosine kinase, shows remarkable efficacy in chronic myeloid leukaemia (CML). Autophagy protects tumour cells against chemotherapeutic stimulation and contributes to imatinib resistance in CML. Kinesin family member 23 (KIF23) is involved in cytokinesis and associated with autophagy. The role of KIF23 in autophagy-induced imatinib resistance in CML was investigated. First, to induce drug resistance, CML cells were exposed to increasing concentrations of imatinib. The concentration of imatinib resistance in CML cells was screened through upregulation of 50% inhibitory concentration (IC

GNG5 is suggested to exert a critical effect on tumor development in human beings; however, its function and related mechanism within breast cancer (BC) are still unclear. In this regard, the present work focused on identifying and evaluating GNG5's function and revealing its possible molecular mechanism. Subcutaneous tumorigenesis model of nude mice and in-vitro cell model was established. The relationship between GNG5 expression and BC was studied through knockdown and overexpression experiments. The proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) of liver cancer cell lines overexpressing or silencing GNG5 were detected. Furthermore, the pathway mechanism of GNG5 was evaluated at the molecular level and was performed to further verify the possible targets and mechanisms of action. In comparison with that in normal tissue, GNG5 level within BC tissue was higher. In addition, GNG5 overexpression stimulated BC cell proliferation, invasion, migration and EMT. BC cells with reduced GNG5 expression exhibited significant decreases in glucose uptake, lactate levels, and ATP concentrations. In addition, GNG5 knockdown inhibited Wnt/β-catenin signaling. This study indicates that GNG5 may generate a vital function in BC. The results of the current work demonstrated GNG5's effect on BC pathological process, also providing a reference for developing new targeted therapies for BC.

Phospholipase D (PLD) is a potential therapeutic target against cancer. However, the contribution of PLD inhibition to the antitumor response remains unknown. We developed a potent and selective PLD1 inhibitor based on computer-aided drug design. The inhibitor enhanced apoptosis in colorectal cancer (CRC) cells but not in normal colonic cells, and in vitro cardiotoxicity was not observed. The inhibitor downregulated the Wnt/β-catenin signaling pathway and reduced the migration, invasion, and self-renewal capacity of CRC cells. In cancer, therapeutic engagement of immunogenic cell death (ICD) leads to more effective responses by eliciting the antitumor immunity of T cells. The CRC cells treated with the inhibitor showed hallmarks of ICD, including downregulation of “do not eat-me” signals (CD24, CD47, programmed cell death ligand 1 [PD-L1]), upregulation of “eat-me” signal (calreticulin), release of high-mobility group Box 1, and ATP. PLD1 inhibition subsequently enhanced the phagocytosis of cancer cells by macrophages through the surface expression of costimulatory molecules; as a result, the cancer cells were more susceptible to cytotoxic T-cell-mediated killing. Moreover, PLD1 inhibition attenuated colitis-associated CRC and orthotopically injected tumors, probably by controlling multiple pathways, including Wnt signaling, phagocytosis checkpoints, and immune signaling. Furthermore, combination therapy with a PLD1 inhibitor and an anti-PD-L1 antibody further enhanced tumor regression via immune activation in the tumor environment. Collectively, in this study, PLD1 was identified as a critical regulator of the tumor microenvironment in colorectal cancer, suggesting the potential of PLD1 inhibitors for cancer immunotherapy based on ICD and immune activation. PLD1 inhibitors may act as promising immune modulators in antitumor treatment via ICD.

Although many chemopreventive studies on colorectal tumors have been reported, no effective and safe preventive agent is currently available. We searched for candidate preventive compounds against colorectal tumor comprehensively from United States Food and Drug Administration (FDA)-approved compounds by using connectivity map (CMAP) analysis coupled with in vitro screening with colorectal adenoma (CRA) patient-derived organoids (PDOs). We generated CRA-specific gene signatures based on the DNA microarray analysis of CRA and normal epithelial specimens, applied them to CMAP analysis with 1309 FDA-approved compounds, and identified 121 candidate compounds that should cancel the gene signatures. We narrowed them down to 15 compounds, and evaluated their inhibitory effects on the growth of CRA-PDOs in vitro. We finally identified resveratrol, one of the polyphenolic phytochemicals, as a compound showing the strongest inhibitory effect on the growth of CRA-PDOs compared with normal epithelial PDOs. When resveratrol was administered to Apc

The small intestine is among the fastest self-renewing tissues in adult mammals. This rapid turnover is fueled by the intestinal stem cells residing in the intestinal crypt. Wnt signaling plays a pivotal role in regulating intestinal stem cell renewal and differentiation, and the dysregulation of this pathway leads to cancer formation. Several studies demonstrate that intestinal stem cells follow neutral drift dynamics, as they divide symmetrically to generate other equipotent stem cells. Competition for niche space and extrinsic signals in the intestinal crypt is the governing mechanism that regulates stemness versus cell differentiation, but the underlying molecular mechanisms are still poorly understood, and it is not yet clear how this process changes during disease. In this review, we highlight the mechanisms that regulate stem cell homeostasis in the small intestine, focusing on Wnt signaling and its regulation by RNF43 and ZNRF3, key inhibitors of the Wnt pathway. Furthermore, we summarize the evidence supporting the current model of intestinal stem cell regulation, highlighting the principles of neutral drift at the basis of intestinal stem cell homeostasis. Finally, we discuss recent studies showing how cancer cells bypass this mechanism to gain a competitive advantage against neighboring normal cells.

Non-alcoholic steatohepatitis (NASH) is a chronic liver disease that results from multiple metabolic disorders. Considering the complexity of the pathogenesis, the identification of a factor mediating the multiple pathogenic phenotypes of NASH will be important for treatment. In this study, we found that CXXC5, a negative feedback regulator of the Wnt/β-catenin pathway, was overexpressed with suppression of Wnt/β-catenin signaling and its target genes involved in hepatic metabolism in obese-NASH patients. Cxxc5−/− mice were found to be resistant to NASH pathogenesis with metabolic improvements. KY19334, a small molecule that activates the Wnt/β-catenin pathway via interference of the CXXC5-Dvl interaction, reversed the overall pathogenic features of NASH as Cxxc5−/− mice. The improvement in NASH by KY19334 is attributed to its regenerative effects through restorative activation of the suppressed Wnt/β-catenin signaling. Overall, the pronounced metabolic improvements with the stimulation of liver regeneration by interfering with the CXXC5-Dvl interaction provide a therapeutic approach for NASH.

Activation of the canonical Wingless-related integration site (Wnt) pathway has been shown to increase bone formation and therefore has therapeutic potential for use in orthopedic conditions. However, attempts at developing an effective strategy to achieve Wnt activation has been met with several challenges. The inherent hydrophobicity of Wnt ligands makes isolating and purifying the protein difficult. To circumvent these challenges, many have sought to target extracellular inhibitors of the Wnt pathway, such as Wnt signaling pathway inhibitors Sclerostin and Dickkopf-1, or to use small molecules, ions and proteins to increase target Wnt genes. Here, we review systemic and localized bioactive approaches to enhance bone formation or improve bone repair through antibody-based therapeutics, synthetic Wnt surrogates and scaffold doping to target canonical Wnt. We conclude with a brief review of emerging technologies, such as mRNA therapy and Clustered Regularly Interspaced Short Palindromic Repeats technology, which serve as promising approaches for future clinical translation.

WNT signaling represents an attractive target for cancer therapy due to its widespread oncogenic role. However, the molecular players involved in WNT signaling and the impact of their perturbation remain unknown for numerous recalcitrant cancers. Here, we characterize WNT pathway activity in small cell lung cancer (SCLC) and determine the functional role of WNT signaling using genetically engineered mouse models. β-Catenin, a master mediator of canonical WNT signaling, was dispensable for SCLC development, and its transcriptional program was largely silenced during tumor development. Conversely, WNT5A, a ligand for β-catenin–independent noncanonical WNT pathways, promoted neoplastic transformation and SCLC cell proliferation, whereas WNT5A deficiency inhibited SCLC development. Loss of p130 in SCLC cells induced expression of WNT5A, which selectively increased Rhoa transcription and activated RHOA protein to drive SCLC. Rhoa knockout suppressed SCLC development in vivo, and chemical perturbation of RHOA selectively inhibited SCLC cell proliferation. These findings suggest a novel requirement for the WNT5A–RHOA axis in SCLC, providing critical insights for the development of novel therapeutic strategies for this recalcitrant cancer. This study also sheds light on the heterogeneity of WNT signaling in cancer and the molecular determinants of its cell-type specificity. Significance: The p130–WNT5A–RHOA pathway drives SCLC progression and is a potential target for the development of therapeutic interventions and biomarkers to improve patient treatment.

Background Long non-coding RNAs (lncRNAs) play an essential role in biological processes. However, the expression patterns of lncRNAs that regulate the non-Mendelian inheritance feather phenotypes remain unknown. Objective This study aimed to compare the expression profiles of lncRNAs in the follicles of the late-feathering cocks (LC) and late-feathering hens (LH) that followed genetic rules and the early-feathering hen (EH) and early-feathering cock (EC) that did not conform to the genetic laws. Methods We performed RNA sequencing and investigated the differentially expressed lncRNAs (DElncRNAs) between the early- and late-feathering chickens, which function by cis-acting or participate in the competing endogenous RNA (ceRNA) network. Results A total of 53 upregulated and 43 downregulated lncRNAs were identified in EC vs. LC, and 58 upregulated and 109 downregulated lncRNAs were identified in EH vs. LH. The target mRNAs regulated by lncRNAs in cis were enriched in the pentose phosphate pathway, TGF-β signaling pathway and Jak-STAT signaling pathway in EC vs. LC and were associated with the TGF-β signaling pathway, Wnt signaling pathway, p53 signaling pathway and Jak-STAT signaling pathway in EH vs. LH. In addition, the lncRNA-mediated ceRNA regulatory pathways of hair follicle formation were mainly enriched in the TGF-β signaling pathway, Wnt signaling pathway, melanogenesis, and calcium signaling pathways. The levels of ENSGALG00000047626 were significantly higher in the late-feathering chickens than in the early-feathering chickens, which regulated the expression of SSTR2 by gga-miR-1649-5p. Conclusion This study provides a novel molecular mechanism of lncRNA’s response to the feather rate that does not conform to the genetic laws in chickens.

MicroRNA (miRNA)-222-3p is overexpressed in numerous tumors, where it acts as an oncogene. Although miRNA-222 is highly expressed in acute myeloid leukemia (AML), its functions and the mechanisms underlying these functions have not yet been fully elucidated. This study aimed to investigate the regulatory roles of miRNA-222-3p in AML and the molecular mechanisms underlying these roles. In this study, we observed that miRNA-222-3p increased the viability and suppressed the apoptosis of AML cells. Axin2 was demonstrated to be a direct target of miRNA-222-3p, which when overexpressed, inhibited Axin2 expression and stimulated the Wnt/β-catenin pathway. In contrast, upregulation of Axin2 expression levels reduced the viability and enhanced the apoptosis of AML cells. Moreover, it partially reversed the effects of the miRNA-222-3p mimic on the proliferation and apoptosis of, and modulation of the Wnt/β-catenin pathway in, AML cells. Taken together, this study provides strong evidence that miRNA-222-3p can serve as a molecular target for AML treatment.

Wnt-1 inducible signaling pathway protein 2 (WISP-2/CCN5) is a recently identified adipokine that has been described as an important mediator of canonical Wnt activation in adipogenic precursor cells. In osteoarthritis (OA), the most common form of arthritis, chondrocytes exhibit aberrant and increased production of pro-inflammatory mediators and matrix degrading enzymes such as IL-1β and MMP-13. Although recent evidence suggests a role for Wnt signaling in OA physiopathology, little is known about the involvement of WISP-2 in cartilage degradation. In the present study, we determined the expression of WISP-2 in healthy and OA human chondrocytes. WISP-2 expression is modulated along chondrocyte differentiation and downregulated at the onset of hypertrophy by inflammatory mediators. We also investigated the effect of WISP-2 on cartilage catabolism and performed WISP-2 loss-of-function experiments using RNA interference technology in human T/C-28a2 immortalized chondrocytes. We demonstrated that recombinant human WISP-2 protein reduced IL-1β-mediated chondrocyte catabolism, that IL-1β and WNT/b-catenin signaling pathways are involved in rhWISP-2 protein and IL-1β effects in human chondrocytes, and that WISP-2 has a regulatory role in attenuating the catabolic effects of IL-1β in chondrocytes. Gene silencing of WISP-2 increased the induction of the catabolic markers MMP-13 and ADAMTS-5 and the inflammatory mediators IL-6 and IL-8 triggered by IL-1β in human primary OA chondrocytes in a Wnt/β-catenin dependent manner. In conclusion, here we have shown for the first time that WISP-2 may have relevant roles in modulating the turnover of extracellular matrix in the cartilage and that its downregulation may detrimentally alter the inflammatory environment in OA cartilage. We also proved the participation of Wnt/β-catenin signaling pathway in these processes. Thus, targeting WISP-2 might represent a potential therapeutical approach for degenerative and/or inflammatory diseases of musculoskeletal system, such as osteoarthritis.

Tankyrase inhibition stabilises AXINs and antagonises the Wnt/β-catenin pathway in adenomatous polyposis coli (APC)-mutated colorectal cancer (CRC), suggesting that tankyrase is a potential therapeutic target for APC-mutated CRC. However, clinical trials on reported tankyrase inhibitors have been severely limited by on-target toxicity in the gastrointestinal (GI) tract. Herein, we report a new tankyrase-selective inhibitor, STP1002, having preclinical antitumour efficacy without on-target toxicity in APC-mutated CRC models.STP1002 was developed and characterised using in vitro and in vivo functional studies; its pharmacokinetics, antitumour efficacy and toxicity were evaluated in vivo.STP1002 showed potent, selective inhibition of tankyrase 1/2 but not of members of the poly (ADP-ribose) polymerase 1/2 (PARP1/2). STP1002 exerted antitumour activity by stabilising AXINs and antagonising the Wnt/β-catenin pathway in a subset of APC-mutated CRC cell lines but not in inhibitor-resistant cells and APC-wild-type CRC cell lines. STP1002 showed favourable pharmacokinetic profiles for oral administration once daily. STP1002 inhibited tumour growth of APC-mutated CRC xenograft animal models but not of APC-wild type models in a dose-dependent manner. The antitumour efficacy of STP1002 was confirmed using APC-mutated CRC patient-derived tumour xenograft models. STP1002 showed no significant on-target toxicity in the GI tract compared to G007-LK, which shows severe ileum toxicity in preclinical animal models.These results demonstrate that STP1002, a novel, orally active tankyrase inhibitor, shows preclinical antitumour efficacy without on-target toxicity in the GI tract. Our data provide a rationale for a clinical trial on STP1002 as a potential tankyrase-targeted drug in patients with APC-mutated CRC.

Stem cell therapy has been extensively studied to improve heart function following myocardial infarction; however, its therapeutic potency is limited by low rates of engraftment, survival, and differentiation. Here, we aimed to determine the roles of the β-catenin/Oct4 signaling axis in the regulation of long-term survival and angiogenesis of peripheral blood mesenchymal stem cells (PBMSCs). These cells were obtained from rat abdominal aortic blood. We showed that β-catenin promotes the self-renewal, antiapoptotic effects, and long-term survival of PBMSCs by activating the Oct4 pathway through upregulation of the expression of the antiapoptotic factors Bcl2 and survivin and the proangiogenic cytokine bFGF and suppression of the levels of the proapoptotic factors Bax and cleaved caspase-3. β-Catenin overexpression increased Oct4 expression. β-Catenin knockdown suppressed Oct4 expression in PBMSCs. However, β-catenin levels were not affected by Oct4 overexpression or knockdown. Chromatin immunoprecipitation assays proved that β-catenin directly regulates Oct4 transcription in PBMSCs. In vivo, PBMSCs overexpressing β-catenin showed high survival in infarcted hearts and resulted in better myocardial repair. Further functional analysis identified Oct4 as the direct upstream regulator of Ang1, bFGF, HGF, VEGF, Bcl2, and survivin, which cooperatively drive antiapoptosis and angiogenesis of engrafted PBMSCs. These findings revealed the regulation of β-catenin in PBMSCs by the Oct4-mediated antiapoptotic/proangiogenic signaling axis and provide a breakthrough point for improving the long-term survival and therapeutic effects of PBMSCs.

Understanding the regulatory mechanisms of glioblastoma growth is crucial for developing novel therapies. In this study, the expression and function of zinc finger protein 501 (ZNF501) in human glioblastoma cells were characterized. ZNF501 was abundantly expressed in human glioblastoma cell lines U251, U118, U87, and U138. ZNF501 ablation in these cell lines caused inhibition of proliferation, sphere formation, and the expression of SOX2 and OCT4. Besides, ZNF501 ablation increased the sensitivity of these cell lines to Temozolomide but did not influence cell migration. Orthotopic implantation of U251 cells and U118 cells indicated that ZNF501 ablation suppressed glioblastoma cell proliferation and tumor formation in vivo. ZNF501 ablation induced down-regulation of Frizzled-6 (FZD6), a component of the Wnt signaling. Lentivirus-mediated FZD6 overexpression restored the proliferation, sphere formation, drug sensitivity, and SOX2 and OCT4 expression in these cell lines. ZNF501 ablation also incurred down-regulation of JNK phosphorylation which is an indicator of the non-canonical Wnt signaling, but did not change the expression of active β-catenin which is the hallmark of the canonical Wnt signaling. Inhibition of the non-canonical Wnt signaling abrogated the effects of ZNF501 and FZD6. Therefore, for the first time, we showed that ZNF501 is essential for the growth and stemness of glioblastoma cells possibly by maintaining FZD6 expression and the non-canonical Wnt signaling.

Despite advances in colorectal cancer (CRC) treatment, most advanced CRC patients who experience disease progression after chemotherapy, targeted therapy, and immunotherapy face a situation in which there is no available medicine. Thus, new therapeutic drugs for CRC are urgently needed. Studies have shown that cholesteryl ester transfer protein (CETP) has a vital role in tumor development and is a possible target for CRC therapy. We found that Evacetrapib, a CETP inhibitor, suppressed CRC cell growth by inhibiting the Wnt/β-catenin signaling pathway and activating the c-Jun NH2-terminal kinase (JNK) signaling pathway in CRC. Therefore, Evacetrapib displays an anti-cancer effect and is a possible option for treating CRC.

Tooth agenesis is a common structural birth defect in humans that results from failure of morphogenesis during early tooth development. The homeobox transcription factor Msx1 and the canonical Wnt signaling pathway are essential for “bud to cap” morphogenesis and are causal factors for tooth agenesis. Our recent study suggested that Msx1 regulates Wnt signaling during early tooth development by suppressing the expression of Dkk2 and Sfrp2 in the tooth bud mesenchyme, and it demonstrated partial rescue of Msx1-deficient molar teeth by a combination of DKK inhibition and genetic inactivation of SFRPs. In this study, we found that Sostdc1/Wise, another secreted Wnt antagonist, is involved in regulating the odontogenic pathway downstream of Msx1. Whereas Sostdc1 expression in the developing tooth germ was not increased in Msx1−/− embryos, genetic inactivation of Sostdc1 rescued maxillary molar, but not mandibular molar, morphogenesis in Msx1−/− mice with full penetrance. Since the Msx1−/−; Sostdc1−/− embryos exhibited ectopic Dkk2 expression in the developing dental mesenchyme, similar to Msx1−/− embryos, we generated and analyzed tooth development in Msx1−/−; Dkk2−/− double and Msx1−/−; Dkk2−/−; Sostdc1−/− triple mutant mice. The Msx1−/−; Dkk2−/− double mutants showed rescued maxillary molar morphogenesis at high penetrance, with a small percentage also exhibiting mandibular molars that transitioned to the cap stage. Furthermore, tooth development was rescued in the maxillary and mandibular molars, with full penetrance, in the Msx1−/−; Dkk2−/−; Sostdc1−/− mice. Together, these data reveal 1) that a key role of Msx1 in driving tooth development through the bud-to-cap transition is to control the expression of Dkk2 and 2) that modulation of Wnt signaling activity by Dkk2 and Sostdc1 plays a crucial role in the Msx1-dependent odontogenic pathway during early tooth morphogenesis.

Rho guanine nucleotide exchange factor 40 (ARHGEF40) is a member of the Dbl-family of guanine nucleotide factor proteins. However, its expression pattern and biological function in malignant tumors, notably in nonsmall cell lung cancer (NSCLC) are currently unknown. The present study demonstrated that ARHGEF40 was highly expressed in NSCLC specimens and that its expression was significantly associated with advanced TNM stage (p 0.001), lymph node metastasis (p = 0.002), and poor prognosis (p = 0.0056). In addition, ARHGEF40 accelerated nuclear translocation of the key component β-catenin and increased the expression levels of the Wnt signaling pathway targets c-myc, cyclin D1 and MMP7. Moreover, it promoted lung cancer cell proliferation and invasion in vitro and in vivo. To elucidate the underlying molecular mechanism, the current study demonstrated that ARHGEF40 could induce activation of the Wnt signaling pathway by increasing the phosphorylation levels of AKT and GSK3β via interaction with RhoA. Moreover, the Dbl homology (DH)-pleckstrin homology (PH) domain of ARHGEF40 was responsible for this interaction. Its deletion abolished the binding, which blocked the activation of the Wnt signaling. Taken together, the data indicated that ARHGEF40 promoted the malignant phenotype of lung cancer cells by activating the AKT-Wnt axis. This was achieved by its interaction with RhoA via the DH-PH domain. ARHGEF40 may serve as a novel target for NSCLC treatment.

R-spondin 1 (RSPO1) is a ligand for the intestinal stem cell (ISC) marker Lgr5 in the crypt, which functions to amplify canonical Wnt signaling to stimulate the division of ISCs. Despite the crucial role of recombinant human RSPO1 (rhRSPO1) in homeostasis and regeneration, little is known about RSPO1 among different species. Here, we cloned the porcine RSPO1 (pRSPO1) gene and obtained rpRSPO1 protein through the expression system of the recombinant

This study aimed to investigate the effects of scaffold matrix attachment region binding protein 1 (SMAR1) on the development of bladder cancer (BCa). SMAR1 expression in paired tumor and corresponding adjacent normal tissues from 55 BCa patients was detected by quantitative reverse transcription-polymerase chain reaction. BCa cells were transfected to regulate SMAR1 expression. BCa cells were treated with XAV-939, LiCl and 2-deoxyglucose. The effect of SMAR1 on the viability, proliferation, migration, invasion and Warburg effect of BCa cells was researched by counting kit-8, colony formation assay, Transwell and aerobic glycolysis assays. Western blot was performed to detect protein expression. BCa cell growth in vivo was recorded in nude mice. Immunohistochemical staining was performed for clinical and xenografted tumor tissue specimens. SMAR1 expression was down-regulated in BCa patients, associating with worse prognoses. SMAR1 knockdown enhanced the viability, proliferation, migration, invasion, EMT and Warburg effect of BCa cells. The opposite effect was found in the SMAR1 overexpression BCa cells. XAV-939 treatment reversed the elevation of β-catenin, c-Myc and Cyclin D1 proteins expression and Warburg effect in Bca cells post-SMAR1 knockdown. LiCl treatment abrogated the inhibition of β-catenin, c-Myc and Cyclin D1 proteins expression and Warburg effect proteins due to SMAR1 overexpression in BCa cells. SMAR1 overexpression inhibited the growth of BCa cells in vivo. SMAR1 might suppress the Wnt/β-catenin signaling pathway activity to inhibit the progression of BCa. It might be an effective treatment target for BCa.

The tumor microenvironment plays a central role in cancer initiation and progression. CD248 is expressed in tumor-associated stromal cells, particularly fibroblasts and pericytes. Exploring the function of CD248 has the potential to provide biological insights into tumor-supportive stroma and potential therapeutic targets. Here, we investigated the role of stromal CD248 in lung cancer. In orthotopic lung cancer transplantation models, tumor volume, density of vessels and pericytes, and functionality of tumor vessels were all lower in mice lacking Cd248 (Cd248LacZ/LacZ) compared with Cd248 wild-type or haploinsufficient mice. Two angiogenic factors, OPN and SERPINE1, were decreased in Cd248LacZ/LacZ pericytes, and supplementation with both factors rescued their proliferation and endothelial cell tube formation–promoting ability. Mechanistically, Wnt/β-catenin signaling induced Opn and Serpine1 expression and was suppressed in Cd248LacZ/LacZ pericytes. CD248 interacted with Wnt pathway repressors IGFBP4 and LGALS3BP, leading to increased Wnt/β-catenin signaling. Correspondingly, administration of a β-catenin inhibitor in Cd248+/LacZ mice mimicked the effect of Cd248 loss and blocked the growth of transplanted lung tumor cells that were resistant to this inhibitor in vitro. In addition, CD248+ pericytes coexpressed OPN and SERPINE1 and correlated with increased tumor size in human lung cancer. Additionally, high expression of CD248, OPN, and SERPINE1 was associated with poor survival in lung cancer patients. In summary, CD248 derepresses Wnt signaling and upregulates OPN and SERPINE1 in pericytes, resulting in enhanced angiogenesis and lung cancer growth. This novel axis of CD248–Wnt signaling–angiogenic factors in pericytes provides a potential target for lung cancer therapy. Significance: These findings demonstrate that CD248 maintains pericyte function in lung cancer through the Wnt signaling pathway and present CD248 as a potential therapeutic target.

To date, therapeutic approaches for cementum regeneration are limited and outcomes remain unpredictable. A significant barrier to improve therapies for cementum regeneration is that the cementocyte and its intracellular signal transduction mechanisms remain poorly understood. This study aims to elucidate the regulatory mechanism of Wnt pathway in cementogenesis.The effects of canonical Wnt signaling were compared in vitro using immortalized murine cementocyte cell line IDG-CM6 and osteocyte cell line IDG-SW3 by quantitative real-time polymerase chain reaction, Western blot, confocal microscopy, alkaline phosphatase (ALP) assay, and Alizarin red S staining. In vivo, histological changes of cementum and bone formation were examined in transgenic mice in which constitutive activation of β-catenin is driven by Dmp1 promoter.Expression of components of the Wnt/β-catenin pathway were much greater in the IDG-SW3 cells compared with the IDG-CM6 cells resulting in much lower expression of Sost/sclerostin in the IDG-SW3 cells. In the IDG-CM6 cells, low dose Wnt3a (20 ng/ml) had a modest effect while high dose (200 ng/ml) inhibited runt-related transcription factor 2, osterix, ALP, and osteopontin in contrast to the IDG-SW3 cells where high dose Wnt3a dramatically increased mRNA expression of these same markers. However, high Wnt3a significantly increased mRNA for components of Wnt/β-catenin signaling pathway in both IDG-CM6 and IDG-SW3 cells. In vivo, constitutive activation of β-catenin in the Dmp1-lineage cells in mice leads to bone hyperplasia and cementum hypoplasia.These findings indicate that Wnt signaling has distinct and different effects on the regulation of long bone as compared with cementum.

Osteosarcoma (OS) is the most prevalent malignant bone tumor with poor prognosis. Developing new drugs for the chemotherapy of OS has been a focal point and a major obstacle of OS treatment. Nitazoxanide (NTZ), a conventional anti-parasitic agent, has got increasingly noticed because of its favorable antitumor potential. Herein, we investigated the effect of NTZ on human OS cells in vitro and in vivo. The results obtained in vitro showed that NTZ inhibited the proliferation, migration and invasion, arrested cell cycle at G1 phase, while induced apoptosis of OS cells. Mechanistically, NTZ suppressed the activity of AKT/mTOR and Wnt/β-catenin signaling pathways of OS cells. Consistent with the results in vitro, orthotopic implantation model of 143B OS cells further confirmed that NTZ inhibited OS cells growth and lung metastasis in vivo. Notably, NTZ caused no apparent damage to normal cells/tissues. In conclusion, NTZ may inhibit tumor growth and metastasis of human OS cells through suppressing AKT/mTOR and Wnt/β-catenin signaling pathways.

The PI3K/Akt and Wnt/β-catenin pathways play an important role in the acquisition of the malignant phenotype in cancer. However, there are few data regarding the role of the interplay between both pathways in colorectal cancer (CRC) progression. The mutational status and the clinicopathological characteristics of PI3K/Akt and Wnt/β-catenin pathways were accessed by bioinformatic analysis whereas that the impact of the interplay between the activity of both pathways to explain tumorigenic potential was performed in vitro using IGF-1 and Wnt3a treatments in CRC cell models. The mutational status of these pathways did not influence the survival of CRC patients, but an association between clinicopathological characteristics in patients with mutations in one, but not in both pathways was observed. A potentiating effect on the activation of both pathways and enhanced cellular migration and proliferation was observed when both pathways were activated simultaneously with IGF-1 and Wnt3a. In addition, these effects were hindered after pretreatment with LY294002, a specific PI3K inhibitor, suggesting some dependence between these two signaling cascades. Our findings show that, regardless of mutational status, there is an interplay between the activity of PI3K/Akt and Wnt/β-catenin pathways that contributes to events related to CRC progression and that the reversal of such events using a PI3K inhibitor highlights the value of targeting these pathways for potential directed therapies in CRC patients.