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Purpose: Oral squamous cell carcinoma (OSCC) is a significant public health challenge associated with high mortality rates primarily due to its invasive and metastatic behavior. This study aimed to evaluate the expression patterns of five critical biomarkers: β-catenin, E-cadherin, podoplanin (PDPN), CXCR4, and p53 in OSCC tissues and to investigate their correlations with clinicopathologic features and patient outcomes., Methods: We conducted an immunohistochemical analysis utilizing tissue microarrays (TMAs) from 95 patients diagnosed with primary OSCC. The expression levels of the five biomarkers were quantified using H-scores. Statistical analyses, including Kruskal-Wallis tests, Dunn's post-hoc tests, and correlation analyses, were performed to explore the associations between biomarker expression, clinicopathologic parameters, and overall patient survival., Results: The study found that loss of E-cadherin and β-catenin expression was significantly associated with increased tumor depth and lymphatic invasion, corroborating their role in the process of epithelial-mesenchymal transition (EMT). High levels of PDPN were noted in both early and late-stage OSCC, indicating its potential involvement in initiating invasive behaviors. Notably, CXCR4 expression exhibited positive correlations with E-cadherin and β-catenin, suggesting a hybrid invasion phenotype incorporating both EMT and collective invasion strategies. Although Cox regression analysis did not reveal significant associations between biomarker expression and overall survival (OS) or disease-specific survival (DSS), factors such as alcohol consumption, tumor size, lymph node involvement, and advanced clinical stage emerged as significant negative predictors of both OS and DSS., Conclusion: The expression profiles of β-catenin, E-cadherin, PDPN, CXCR4, and p53 in OSCC tissues provide valuable insights into a hybrid model of invasion that integrates mechanisms of EMT with an important rule in the tumor invasion. This nuanced understanding of OSCC progression highlights the potential of PDPN and CXCR4 as novel therapeutic targets, emphasizing the need for further investigation into their roles in OSCC biology and the development of targeted treatments that could improve patient outcomes and survival rates., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: We investigate the prognostic role of β-catenin and L1 neuronal cell-adhesion molecule (L1CAM) according to risk groups in endometrial carcinomas (EC)., Methods: A total of 335 EC patients were classified according to the Proactive Molecular Risk Classifier for Endometrial Cancer. We evaluated the expression of ß-catenin and L1CAM using immunohistochemistry, and their association with clinicopathological characteristics and survival., Results: The expressions of β-catenin and L1CAM were observed in 10.4% of all patients, respectively, and showed mutually exclusive pattern. While β-catenin expression was associated with endometrioid histology (p = 0.035) and low tumor grade (p = 0.045), L1CAM expression was associated with non-endometrioid histology (p < 0.001), high tumor grade (p < 0.001), lymphovascular space invasion (p = 0.006), and advanced International Federation of Gynecology and Obstetrics (FIGO) stage (p = 0.001). β-catenin expression was most frequent in the no specific molecular (NSMP) group (26/35, 74.3%), followed by the DNA polymerase-ε-mutated (POLE-mut) (6/35, 17.1%), and mismatch repair-deficiency (dMMR) (3/35, 8.6%). L1CAM expression was most frequent in the p53-abnormal group (22/35, 62.9%), followed by the NSMP (6/35, 17.1%), dMMR (4/35, 11.4%), and POLE-mut (3/35, 8.6%). Although both markers did not show statistical significance in multivariate analysis for both progression-free survival (PFS) and overall survival in entire cohort, β-catenin positivity was identified as the sole factor associated with worse PFS in the high-intermediate risk subgroup (p = 0.001)., Conclusion: The expression of nuclear β-catenin may serve as a potential biomarker for predicting recurrence and guiding therapeutic strategies in high-intermediate risk EC patients., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Acute exacerbation of chronic obstructive pulmonary disease (AECOPD) is the main factor that leads to the deterioration of the disease. Currently, the diagnosis of AECOPD mainly relies on clinical manifestations, good predictors or biomarkers are lacking. We aim to reveal specific biomarkers and potential pathogenesis of AECOPD and provide a research basis for the diagnosis and treatment., Methods: Four patients with AECOPD, four patients with stable COPD, and five control subjects were enrolled for RNA sequencing and KEGG analysis. The mRNA level of target genes was verified by quantitative real-time PCR (qPCR) with an expanded sample size (30 patients with AECOPD, 27 patients with stable COPD, and 35 control subjects). ELISA and immunofluorescence were used to identify the target proteins. Furthermore, the expression and function of WNT/β-catenin signaling pathway were assessed in animal models of COPD., Results: RNA sequencing showed that 54 genes were up-regulated and 111 genes were down-regulated in the AECOPD. Differentially expressed genes were mainly enriched in WNT signaling pathway, et al. QPCR revealed that multi-genes of the WNT/β-catenin signaling were significantly down-regulated in AECOPD (P < 0.05), and β-catenin protein was significantly decreased in plasma of AECOPD and stable COPD (P < 0.01), while phosphorylated β-catenin was significantly up-regulated in peripheral blood mononuclear cells of AECOPD (P < 0.05). Similarly, WNT ligands, WNT receptors, and downstream signaling molecules were down-regulated, with an increased phosphorylated β-catenin protein in animal models of COPD. Activation of WNT/β-catenin signaling pathway by lithium chloride reduced the expression of phosphorylated β-catenin and ameliorated the COPD-like airway inflammation in mice., Conclusion: WNT/β-catenin signaling pathway is down-regulated in AECOPD patients and in animal models of COPD. Increased expression of phosphorylated β-catenin in the blood might be a potential biomarker of AECOPD. Activation of WNT/β-catenin pathway may also represent a therapeutic target for AECOPD., (© 2022. The Author(s).)

Background: Human aortic valve interstitial cells (hAVICs) are a key factor in the pathogenesis of calcific aortic valve disease (CAVD). This research examines the role and mechanism of microRNA miR-138-5p in osteogenic differentiation of hAVICs., Methods: RT-qPCR analysis was applied for detecting miR-138-5p and RUNX2 expression in valve tissues of CAVD patients and controls. On completion of induction of osteogenic differentiation of hAVICs, and after overexpression or interference of miR-138-5p expression, the condition of osteogenic differentiation and calcification of hAVICs was confirmed using alkaline phosphatase staining and alizarin red staining. Subsequently, western blot was utilized to detect the expression of osteogenesis-related proteins OPN and ALP, and Wnt/β-catenin signaling pathway-related proteins. Finally, the relationship between miR-138-5p and RUNX2 was validated by dual-luciferase reporter assay and Pearson's correlation test., Results: Down-regulation of miR-138-5p was found in CAVD patients and during osteogenic differentiation of hAVICs. Overexpression of miR-138-5p contribute to the inhibition of osteoblast differentiation and calcium deposition in hAVICs, and of ALP and OPN protein expression. RUNX2 was a target gene of miR-138-5p, and it was negatively correlated with miR-138-5p in CAVD. Additionally, overexpression of RUNX2 could reverse the inhibitory effect of miR-138-5p on osteogenic differentiation of hAVICs., Conclusion: miR-138-5p can act as a positive regulator of osteogenic differentiation in CAVD patients to involve in inhibiting valve calcification, which is achieved through RUNX2 and Wnt/β-catenin signaling pathway., (© 2022. The Author(s).)

DNA topoisomerase II alpha (TOP2A) reportedly plays a crucial role in several cancers, however, the precise regulatory role of TOP2A in metastatic characteristics of glioma is still poorly understood. Herein, we sought to elucidate the mechanisms by which TOP2A affects the metastatic phenotypes of glioma. We observed that a high level of TOP2A expression was dramatically linked with inferior survival in glioma patients while silencing of TOP2A impaired glioma cell proliferation and aggressiveness. TOP2A was found to directly interact with β-catenin and facilitated its translocation into the nucleus. Mechanistically, TOP2A effectively induced glioma cell growth and invasion in a β-catenin-dependent manner. Overall, we pinpoint TOP2A as a critical activator of the Wnt/β-catenin pathway in glioma, promoting cell growth, migration, and invasion.

Increased abnormal spindle-like microcephaly (ASPM) expression has been linked to clinical stage and poor prognosis in cancers, but the molecular mechanisms by which ASPM promotes cell metastasis in colorectal cancer (CRC) has not been identified. This study showed that the abilities of cell migration, invasion, and epithelial-mesenchymal transition (EMT) were attenuated in ASPM-deficient CRC cell lines. Furthermore, we reported that attenuation of ASPM expression inhibited CRC cell metastasis in vivo. Additionally, the expression of ASPM was positively correlated with β-catenin level in CRC tissues. Mechanistically, ASPM can upregulate β-catenin transcription by stimulating the β-catenin promoter and enhancing the nuclear translocation of β-catenin in CRC cells, which leads to the activation of the Wnt/β-catenin pathway. Finally, we showed that ASPM effectively induced CRC cell migration and invasion in a β-catenin-dependent manner., (© 2021 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia, Ltd on behalf of Kaohsiung Medical University.)

Liposomal honokiol isolated from the genus Magnolia has been found to have antiangiogenic, anti-inflammatory and antitumor properties. However, there has no report on its role in hair growth. Hair follicles are life-long cycled organelles that go through from anagen, catagen and telogen stages and are regulated by diverse signaling pathways, including Wnt/β-catenin, Notch, Epidermal growth factor (EGF) and Sonic hegehog (SHH). Wnt signals are essential for the initiation of hair follicle placode development and a new potential target of hair loss treatment. This study was designed to investigate the effect of liposomal honokiol (Lip-honokiol) on inducing hair anagen. We identified the hair grew out in advance in the shaving area of C57BL/6N mice after the treatment of liposomal honokiol (Lip-honokiol) by daily abdominal injection. We first demonstrated that Lip-Honokiol activated the Wnt3a/β-catenin pathway and downregulated the transforming growth factor-β1 (TGF-β1) to promote hair growth in mice via immunohistochemistry and immunofluorescence staining. These findings suggest that Lip-honokiol activated the Wnt/β-catenin pathway and accelerated the transfer from the telogen to anagen stage and finally promoted the hair growth., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Genome-wide association studies have shown that a gene variant in the Family with sequence similarity 13, member A (FAM13A) is strongly associated with reduced lung function and the appearance of respiratory symptoms in patients with chronic obstructive pulmonary disease (COPD). A key player in smoking-induced tissue injury and airway remodeling is the transforming growth factor-β1 (TGF-β1). To determine the role of FAM13A in TGF-β1 signaling, FAM13A -/- airway epithelial cells were generated using CRISPR-Cas9, whereas overexpression of FAM13A was achieved using lipid nanoparticles. Wild-type (WT) and FAM13A -/- cells were treated with TGF-β1, followed by gene and/or protein expression analyses. FAM13A -/- cells augmented TGF-β1-induced increase in collagen type 1 (COL1A1), matrix metalloproteinase 2 (MMP2), expression compared with WT cells. This effect was mediated by an increase in β-catenin (CTNNB1) expression in FAM13A -/- cells compared with WT cells after TGF-β1 treatment. FAM13A overexpression was partially protective from TGF-β1-induced COL1A1 expression. Finally, we showed that airway epithelial-specific FAM13A protein expression is significantly increased in patients with severe COPD compared with control nonsmokers, and negatively correlated with lung function. In contrast, β-catenin (CTNNB1), which has previously been linked to be regulated by FAM13A, is decreased in the airway epithelium of smokers with COPD compared with non-COPD subjects. Together, our data showed that FAM13A may be protective from TGF-β1-induced fibrotic response in the airway epithelium via sequestering CTNNB1 from its regulation on downstream targets. Therapeutic increase in FAM13A expression in the airway epithelium of smokers at risk for COPD, and those with mild COPD, may reduce the extent of airway tissue remodeling.

Pulmonary fibrosis (PF) is a chronic, progressive, and lethal disease with little response to available therapies. One of the major mechanisms of PF is the repeated injury and inadequate regeneration of the alveolar epithelium. In this study, we induced human umbilical cord mesenchymal stem cells (hUC-MSCs) to differentiate into type 2 alveolar epithelial cells (AEC2s), and we provided evidence that intratracheal transplantation of hUC-MSC-derived AEC2s (MSC-AEC2s) could improve mortality and alleviate fibrosis in bleomycin-induced PF mice. Transplantation of MSC-AEC2s could increase the AEC2 cell count in these mice, and the results of the cell tracing experiment exhibited that the increased AEC2s originated from the self-renewal of mouse alveolar epithelium. The AEC2 survival was controlled by the apoptosis of AEC2s via the expression of β-catenin in PF mice. In in vitro experiments, MSC-AEC2s could alleviate the apoptosis of MLE-12 cells induced by transforming growth factor beta (TGF-β1), which could be eliminated by using PRI-724, a β-catenin inhibitor, suggesting β-catenin signaling involved in the protection against apoptosis provided by MSC-AEC2s. Our study demonstrated that MSC-AEC2s could protect PF mice through regulating apoptosis mediated by β-catenin, which provided a viable strategy for the treatment of PF.

Osteoporosis (OP) behaves in different manners in different parts of the skeleton. This study aims to investigate the effects of curcumin on bone mass of the mandibular and femur from ovariectomized OP rats and to validate whether enhancer of zeste homolog 2 (EZH2)/Wnt/β-Catenin pathway is involved in this process. Curcumin was administered intragastrically into ovariectomized rats for 12 weeks. The bone parameters and the morphology of the trabecular bone of the left mandible and left femur were assessed by micro-computed tomography assay. Morphological changes of the left mandible and left femur were evaluated by hematoxylin and eosin staining. The mRNA levels of EZH2, β-Catenin, and Runx2 in the right mandible and right femur were examined by quantitative real-time polymerase chain reaction. Immunohistochemistry was performed to assess EZH2 expression. Both the mandible and femur exhibited OP-like changes in ovariectomized rats, while the mandible bone resorption was less than the femur bone resorption. Curcumin intragastric administration improved bone microstructure and promoted bone formation in the mandible and femur. Curcumin inhibited EZH2 mRNA level and induced that of β-Catenin and Runx2 in the mandible and femur. Collectively, curcumin exerts protective effects against OP, possibly by regulating the EZH2/Wnt/β-Catenin pathway., (© 2021 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia on behalf of Kaohsiung Medical University.)

Purpose: In a previous study we have shown in a mouse model that administration of nuclear factor-kappa B (NF-κB) inhibitor thalidomide has promising therapeutic effects on early radiation cystitis (ERC) and late radiation sequelae (LRS) of the urinary bladder. The aim of this study was to evaluate in the same mice the effect of thalidomide on adherens junction (AJ) proteins in ERC and LRS., Methods: Urothelial expressions of E‑cadherin and β‑catenin were assessed by immunohistochemistry in formalin-fixed paraffin-embedded (FFPE) bladder specimens over 360 days post single-dose irradiation on day 0. First, the effect of irradiation on AJ expression and then effects of thalidomide on irradiation-induced AJ alterations were assessed using three different treatment times., Results: Irradiation provoked a biphasic upregulation of E‑cadherin and β‑catenin in the early phase. After a mild decrease of E‑cadherin and a pronounced decrease of β‑catenin at the end of the early phase, both increased again in the late phase. Early administration of thalidomide (day 1-15) resulted in a steeper rise in the first days, an extended and increased expression at the end of the early phase and a higher expression of β‑catenin alone at the beginning of the late phase., Conclusion: Upregulation of AJ proteins is an attempt to compensate irradiation-induced impairment of urothelial barrier function. Early administration of thalidomide improves these compensatory mechanisms by inhibiting NF-κB signaling and its interfering effects.

Colorectal cancer (CRC) is one of the leading cancers worldwide, accounting for high morbidity and mortality. The mechanisms governing tumor growth and metastasis in CRC require detailed investigation. The results of the present study indicated that the transcription factor (TF) myocyte enhancer factor 2A (MEF2A) plays a dual role in promoting proliferation and metastasis of CRC by inducing the epithelial-mesenchymal transition (EMT) and activation of WNT/β-catenin signaling. Aberrant expression of MEF2A in CRC clinical specimens was significantly associated with poor prognosis and metastasis. Functionally, MEF2A directly binds to the promoter region to initiate the transcription of ZEB2 and CTNNB1. Simultaneous activation of the expression of EMT-related TFs and Wnt/β-catenin signaling by MEF2A overexpression induced the EMT and increased the frequency of tumor formation and metastasis. The present study identified a new critical oncogene involved in the growth and metastasis of CRC, providing a potential novel therapeutic target for CRC intervention.

Purpose: To search the AKR1C3 and β-catenin expression in non-small cell lung cancer (NSCLC) and to explore the correlation between AKR1C3 and β-catenin and radiation resistance., Methods: Paraffin specimens from 61 patients with NSCLC were evaluated. These patients could not receive operation but received radical radiotherapy. The patients were divided into effective group and ineffective group with reference to RECIST evaluation criteria. The sites and intensity of AKR1C3 and β-catenin protein expression were detected by immunohistochemistry. The relationship between AKR1C3 and β-catenin and radiation resistance was analyzed by Mann-Whitney U test. The correlation between AKR1C3 and β-catenin was analyzed by Spearman's correlation test. Mann-Whitney U test was used to analyze the AKR1C3 overall expression in the effective group and the ineffective group after radiotherapy., Results: The nuclear expression in the two groups was statistically significant (p=0.033). The β-catenin protein was mainly expressed in the cytoplasm and the nucleus of tissues with NSCLC. The β-catenin nuclear expression was different between the two groups, with statistical significance (p=0.008). AKR1C3 nuclear expression was positively correlated with β-catenin nuclear expression (rs=0.382, p=0.002)., Conclusions: High AKR1C3 nuclear expression in NSCLC is related to radiation resistance. The higher the AKR1C3 nucleus expression, the worse short-term curative effects after radiotherapy. High β-catenin nuclear expression is related to radiation resistance, and the higher the β-catenin nuclear expression, the worse the short-term curative effects after radiotherapy. The nuclear aggregation of AKR1C3 during radiation resistance of non-small cell lung cancer (NSCLC) may have some synergistic relationship with nuclear aggregation of β-catenin.

Tumor-initiating cells or cancer stem cells are a subset of cancer cells that have tumorigenic potential in human cancer. Although several markers have been proposed to distinguish tumor-initiating cells from colorectal cancer cells, little is known about how this subpopulation contributes to tumorigenesis. Here, we characterized a tumor-initiating cell subpopulation from Caco-2 colorectal cancer cells. Based on the findings that Caco-2 cell subpopulations express different cell surface markers, we were able to discriminate three main fractions, CD44 - CD133 - , CD44 - CD133 + , and CD44 + CD133 + subsets, and characterized their biochemical and tumorigenic properties. Our results show that CD44 + CD133 + cells possessed an unusual capacity to proliferate and could form tumors when transplanted into NSG mice. Additionally, primary tumors grown from CD44 + CD133 + Caco-2 cells contained mixed populations of CD44 + CD133 + and non-CD44 + CD133 + Caco-2 cells, indicating that the full phenotypic heterogeneity of the parental Caco-2 cells was re-created. Notably, only the CD44 + CD133 + subset of Caco-2-derived primary tumors had tumorigenic potential in NSG mice, and the tumor growth of CD44 + CD133 + cells was faster in secondary xenografts than in primary transplants. Gene expression analysis revealed that the Wnt/β-catenin pathway was over-activated in CD44 + CD133 + cells, and the growth and tumorigenic potential of this subpopulation were significantly suppressed by small-molecule Wnt/β-catenin signaling inhibitors. Our findings suggest that the CD44 + CD133 + subpopulation from Caco-2 cells was highly enriched in tumorigenic cells and will be useful for investigating the mechanisms leading to human colorectal cancer development., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Purpose: Human amniotic epithelial cells (HAECs) have regenerative properties and low immunogenicity, which have enabled their use without immune rejection in regenerative medicine applications, such as wound repair, corneal surgery and burn repair. The aim of this study was to explore the potential role of HAECs in the proliferation of human corneal endothelial cells (HCEnCs) and the possible mechanism of regulation., Methods: HAECs and HCEnCs were isolated from donated tissue samples and were cultured; the collected HAEC culture medium (HAEC-Me) was added to the human corneal endothelium medium (CEM) to establish the HAEC-CM system. HCEnCs were cultured in CEM, 20%HAEC-Me, 20% HAEC-CM, 20% HAEC-CM supplemented with a GSK-3β inhibitor TWS119 or CEM supplemented with TWS119. Then, cell proliferation, apoptosis, cell cycle progression, telomerase activity, and Wnt/β-catenin pathway-related protein levels were assessed., Results: We found that the HCEnCs cultured in the 20% HAEC-CM had increased proliferative capacity, telomerase activity and β-catenin and Tcf4 expression levels, and they had a decrease in the rate of apoptosis and α-SMA expression when they were compared with the HCEnCs cultured in the 20% HAEC-Me. After GSK-3β was inhibited by TWS119, HCEnCs cultured in CEM or 20% HAEC-CM had an increased proliferative capacity, telomerase activity, β-catenin/Tcf4 expression and a decreased α-SMA expression, and they had a decreased apoptotic rate., Conclusions: These data indicate that the human amniotic epithelial cells microenvironment can promote the proliferation of human corneal endothelial cells, which may be related to regulating telomerase activity and epithelial-to-mesenchymal transition (EMT) via the Wnt/β-catenin pathway.

β-catenin is a multi-functional protein with a central role in regulating embryonic development and tissue homeostasis. The abnormal accumulation of β-catenin, due to disrupted β-catenin degradation or unregulated β-catenin synthesis, causes the development of cancer. A recent study showed that the overexpression of proto-oncogene serine/arginine-rich splicing factor 9 (SRSF9) promotes β-catenin accumulation via binding β-catenin mRNA and enhancing its translation in a manner that is dependent on the mechanistic target of rapamycin (mTOR). However, the regulation of the interaction between SRSF9 and mRNA of β-catenin remains unclear. Here, we show that AMP-activated protein kinase (AMPK) phosphorylates SRSF9 at the RNA-interacting SWQDLKD motif that plays a major role in determining substrate specificity. The phosphorylation by AMPK inhibits the binding of SRSF9 to β-catenin mRNA and suppresses β-catenin protein synthesis caused by SRSF9 overexpression without changing the β-catenin mRNA levels. Our findings suggest that AMPK activators are potential therapeutic targets for SRSF9-derived overproduction of β-catenin in cancer cells., Competing Interests: Declaration of competing interest We declare no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Hepatocellular carcinoma (HCC) is the most common primary hepatic malignancy in adults. Several studies have classified HCC into molecular subtypes aiming at detecting aggressive subtypes. The aim of the present study was to investigate the role of p53, β-catenin, CD133, and Ki-67 in subclassification of HCC into different aggressive subtypes and the correlation between those markers and the clinicopathologic characteristics of HCC patients. This retrospective study was conducted on paraffin-embedded blocks of 114 HCC specimens. Tissue microarray was constructed and immunostaining for p53, β-catenin, CD133, and Ki-67 was performed and HCC score was formulated. P53 expression was associated with old age (P=0.028), large tumor size (P=0.019), poorly differentiated HCC (P=0.012), hepatitis B virus (HBV) positivity (P=0.032), and hepatitis C virus (HCV) negativity (P =0.046). β-catenin expression was associated with small sized tumors (P=0.005), HBV negativity (P=0.027), early-staged tumors (P=0.029), and prolonged recurrence-free survival (P=0.045). High percentage of CD133 expression was associated with old patients (P=0.035) and HBV positivity (P= 0.045). Ki-67 expression was associated with large tumor size (P= 0.049), vascular invasion (P= 0.05), old age (P=0.035), and previous treatment of HCV by direct acting antiviral agents (P=0.005). Cases with high HCC score showed significant association with old patients (P=0.002), previous treatment of HCV by direct acting antiviral agents (P<0.001), large tumor size (P<0.001), and poorly differentiated tumors (P= 0.009). The proposed HCC score can divide HCC patients into subtypes necessitating tailoring of treatment strategy according to this proposed score to target and optimally treat the aggressive subtypes. This score needs to be further validated on large number of patients with longer follow-up period.

Background: Histopathologically, scars can mimic superficial fibromatoses. Superficial fibromatoses are known to show nuclear β-catenin immunoexpression, although the tumor types do not harbor CTNNB1 or APC alterations. This study aimed to evaluate nuclear β-catenin immunoexpression in scars compared to that in superficial fibromatoses., Methods: Immunostaining with an anti-β-catenin antibody, clone 14, was performed on 8 superficial fibromatoses and 22 scars. The extent of β-catenin nuclear staining was classified as negative (<10%), focally positive (10-49%), or diffusely positive (50-100%). β-catenin staining intensity was semi-quantitatively graded as weak, moderate, or strong., Results: In 21 (95%) scars, nuclear β-catenin immunoexpression was detected in fibroblasts/myofibroblasts, with mainly diffuse (16/21) and moderate (14/21) to strong (5/21) staining. In contrast, seven (88%) of the eight superficial fibromatoses expressed β-catenin in the nuclei of the lesional spindle cells, at varying levels of staining intensity. Fibroblasts in normal papillary dermis always showed nuclear β-catenin expression to varying degrees but those in the reticular dermis did not., Conclusions: Scars typically exhibit nuclear β-catenin expression similar to that in superficial fibromatoses. Thus, β-catenin immunohistochemistry is not suitable for distinguishing superficial fibromatoses from scars., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Arrested alveolar development is the main pathological characteristic of neonatal bronchopulmonary dysplasia (BPD); however, a number of studies aiming to improve alveolarization have focused on alveolar epithelial cell damage and impairment. Previously, the authors reported that the Wnt signaling plays a key role in alveolar injury and repair by regulating alveolar epithelial type II cell (AECII) proliferation and differentiation. In the present study, the authors wished to investigate whether Yes‑associated protein (YAP), a transcriptional coactivator in the Hippo signaling pathway, affects AECII proliferation and differentiation via the Wnt/β‑catenin pathway in BPD. It was found that YAP regulated AECII proliferation and differentiation. A decreased expression of YAP, Wnt3a and nuclear β‑catenin was observed in lung tissues affected by BPD. In vitro, YAP and Wnt3a overexpression in BPD promoted AECII proliferation and differentiation into AECIs by increasing the nuclear transfer of β‑catenin and vice versa. The effects of a decreased Wnt3a expression in primary AECIIs in BPD were compensated by YAP overexpression, as were the effects of Wnt3a knockdown in primary AECIIs. On the whole, the findings of the present study demonstrate that YAP and Wnt3a independently promote AECII proliferation and differentiation in experimental BPD by increasing the nuclear β‑catenin levels. Therefore, Wnt3a or YAP may be candidate regulatory targets for improving the outcomes of BPD.

Wnt/beta-catenin signaling may play an essential role in endometrial decidualization, placentation, and the establishment of pregnancy. We investigate here the possible roles, immunolocalizations, and synthesis of the Wnt3, Wnt7a, and beta-catenin proteins in the rat endometrium during the estrous cycle and early postimplantation period. Wnt3 and Wnt7a had a similar localization and dynamic expression relative to the endometrial stages. Wnt7a immunostaining was not limited only to the luminal epithelial cells, but also to strong stainings in the stromal and endothelial cells. Wnt3, Wnt7a, and beta-catenin were highly synthesized and colocalized at the trophoblast-decidual interface; and were more obvious in the primary decidual zone, the GTCs, and the ectoplacental cone. Beta-catenin was strongly localized at the borders of the mature decidual cells; however, Wnt3 and Wnt7a immunolocalizations were decreased in those cells. As such, the immunolocalization of Wnt3, Wnt7a, and beta-catenin shifted with decidualization and placentation. The expression level of Wnt3, Wnt7a, and beta-catenin messenger RNAs increased in early pregnancy, and especially between Days 8.5 and 9.5. The dramatic changes in the expression of Wnt3, Wnt7a, and beta-catenin observed during the early days of pregnancy and the estrous cycle may indicate their roles in decidualization, stromal cell proliferation, and trophoblast invasion., (© 2020 Wiley Periodicals LLC.)

Accumulating evidence indicates the role of microglial activation and sustained neuroinflammation in the pathogenesis of cognitive dysfunction, a common feature associated with depressive disorders. It also indicates the role of Wnt/β-catenin pathway in regulation of microglia-mediated neuroinflammation. Amisulpride exhibits antidepressant and pro-cognitive activities in several clinical and experimental studies. Hitherto, the direct effects of amisulpride on Wnt/β-catenin signaling and microglial activity have not been thoroughly studied. This study aimed at investigating the effects of chronic amisulpride treatment on Wnt/β-catenin signaling and pro-inflammatory microglial activation and its role in alleviation of depressive-like behavior and cognitive deficits elicited by unpredictable chronic mild stress (UCMS). The effects of amisulpride (3 mg/kg/day) were investigated on behavioral/cognitive deficits, expression of Wnt/β-catenin pathway and microglial activation in the prefrontal cortex (PFC) of UCMS-exposed male Wistar rats. UCMS induced depressive-like behavior with impairment of performance in novel object recognition test and attentional set-shifting task. These behavioral deficits were associated with decreased total β-catenin and increased pro-inflammatory microglial activation. Amisulpride improved UCMS-induced behavioral/cognitive deficits, ameliorated Wnt/β-catenin signaling dysregulation and pro-inflammatory microglial activation. This work highlights the antidepressant and pro-cognitive effects of amisulpride in UCMS-exposed rats that could be mediated by modulation of Wnt/β-catenin pathway activity and amelioration of pro-inflammatory microglial activation in the prefrontal cortex. This could provide new insights into the putative mechanisms behind the antidepressant and pro-cognitive effects exerted by amisulpride., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Purpose: Canonical Wnt/ β-catenin pathway is one mechanism being activated in platinum-resistant epithelial ovarian cancer (EOC). Detecting potential targets for Wnt pathway modulation as a putative future therapeutic approach was the aim of this study., Methods: Biological effects of different Wnt modulators (SB216763, XAV939 and triptolide) on the EOC cell lines A2780 and its platinum-resistant clone A2780cis were investigated via multiple functional tests. Immunohistochemistry (IHC) was carried out to compare the expression levels of Wnt marker proteins (β-catenin, snail/ slug, E-cadherin) in patient specimens and to correlate them with lifetime data., Results: We could show that activated Wnt signaling of the platinum-resistant EOC cell line A2780cis can be reversed by Wnt manipulators through SB216763 or XAV939. All Wnt manipulators tested consecutively decreased cell proliferation and cell viability. Apoptosis of A2780 and A2780cis was enhanced by triptolide in a dose-dependent manner, whereas cell migration was inhibited by SB216763 and triptolide. IHC analyses elucidated significantly different expression patterns for Wnt markers in the serous subtype. Herein, higher plasmatic snail/ slug expression is associated with improved progression-free (PFS) and overall survival (OS)., Conclusion: According to the described effects on EOC biology, all three Wnt manipulators seem to have the potential to augment the impact of a platinum-based chemotherapy in EOC. This is promising as a dominance of this pathway was confirmed in serous histology.

Deep penetrating nevi (DPNs) are intermediate grade lesions which have the capacity to recur, metastasize, or progress to melanoma. Differentiating DPN from other melanocytic lesions including blue and cellular blue nevi can be diagnostically challenging, and markers to distinguish these entities can be useful. Mutations of the β-catenin and mitogen-activated protein kinase pathways have recently been elucidated as distinctive of DPN. This pathway can subsequently activate lymphoid enhancer-binding factor 1 (LEF1), a transcription factor shown to facilitate the epithelial-mesenchymal transition to propagate tumorigenesis. Seventy-two cases in total were examined on hematoxylin and eosin sections and with β-catenin and LEF1 immunohistochemistry. This included: DPN (14), cellular blue nevi (19), blue nevi (15), congenital melanocytic nevi (12), and melanoma (12). Nuclear expression of LEF1, present throughout the entire depth of the lesion, was noted in 13/14 (93%) of DPN, 0/19 (0%) of cellular blue nevi, 0/15 (0%) of blue nevi, 1/12 (8%) of congenital melanocytic nevi, and 9/12 (75%) of melanoma cases. Nuclear expression of β-catenin, present throughout the entire depth of the lesion, was noted in 14/14 (100%) of DPN, 0/18 (0%) of cellular blue nevi, 0/15 (0%) of blue nevi, 1/12 (8%) of congenital melanocytic nevi, and 1/12 (8%) of melanoma cases. A majority of congenital melanocytic nevi demonstrated a gradient of LEF1 and β-catenin expression with more intense staining superficially and loss of staining with increasing depth. Deep, uniform nuclear LEF1 combined with β-catenin immunohistochemical staining can be useful in distinguishing DPN from histologic mimics.

MicroRNAs (miRNAs) are important gene regulators whose dysregulations can be involved in tumorigenesis. β-catenin, the main agent in the Wnt/β-catenin pathway, controls various genes and its over-expression has been discovered in different kinds of cancers including Hepatocellular Carcinoma (HCC). Extensive research demonstrated that the Wnt signaling is one of the major affected pathways in HCC. This study aimed to find miRNA targeting β-catenin gene by bioinformatic approaches and confirm this correlation to propose new therapeutic targets for HCC. Prediction of miRNAs targeting 3'-Untranslated Regions (UTR) of β-catenin mRNA, were done using different types of credible bioinformatic databases. The luciferase assay was also recruited for further confirmation of the bioinformatic predictions. In the first step, the expression of β-catenin was assessed in the HepG2 cell line by real-time PCR technique. Next, transduction of HepG2 cells were done by lentiviral vectors containing the desired miRNA. Then, the expression level of miRNA and the β-catenin gene were evaluated. Based on the results obtained from different bioinformatic databases, miR-214 was selected as the potential miRNA with the highest probability in targeting β-catenin. Furthermore, Luciferase assay results confirmed the accuracy of our bioinformatic prediction. In line with our hypothesis, after the overexpression of miR-214 in HepG2 cells, β-catenin gene expression was reduced significantly. Gathered results indicate the miRNAs role in the down-regulation of their target genes. Hence, the results propose that miR-214 can prevent HCC development by suppressing β-catenin and may supply a newfound approach towards HCC therapy in humans.

Studies have shown that cancer stem cells (CSCs) are involved in resistance and metastasis of cancer; thus, therapies targeting CSCs have been proposed. Here, we report that heat shock 70-kDa protein 1-like (HSPA1L) is partly involved in enhancing epithelial-mesenchymal transition (EMT) and CSC-like properties in non-small cell lung cancer (NSCLC) cells. Aldehyde dehydrogenase 1 (ALDH1) is considered a CSC marker in some lung cancers. Here, we analyzed transcriptional changes in genes between ALDH1high and ALDH1low cells sorted from A549 NSCLC cells and found that HSPA1L was highly expressed in ALDH1high cells. HSPA1L played two important roles in enhancing CSC-like properties. First, HSPA1L interacts directly with IGF1Rβ and integrin αV to form a triple complex that is involved in IGF1Rβ activation. HSPA1L/integrin αV complex-associated IGF1Rβ activation intensified the EMT-associated cancer stemness and γ-radiation resistance through its downstream AKT/NF-κB or AKT/GSK3β/β-catenin activation pathway. Secondly, HSPA1L was also present in the nucleus and could bind directly to the promoter region of β-catenin to function as a transcription activator of β-catenin, an important signaling protein characterizing CSCs by regulating ALDH1 expression. HSPA1L may be a novel potential target for cancer treatment because it both enhances IGF1Rβ activation and regulates γβ-catenin transcription, accumulating CSC-like properties.

Oropharyngeal squamous cell carcinoma (OPSCC) is subclassified by the World Health Organization into two different entities: human papillomavirus (HPV)-positive and HPV-negative tumors. HPV infection promotes the epithelial-to-mesenchymal transition (EMT) and transformation of keratinocyte stem cells into cancer stem cells. EMT is a crucial process in the carcinogenesis of epithelial-derived malignancies, and we aimed to study the role of its markers in OPSCC. This study consists of 202 consecutive OPSCC patients diagnosed and treated with curative intent. We examined E-cadherin, β-catenin, and vimentin expression using immunohistochemistry and compared these with tumor and patient characteristics and treatment outcome. We found that the cell-membranous expression of β-catenin was stronger in HPV-positive than in HPV-negative tumors, and it was stronger in the presence of regional metastasis. The stromal vimentin expression was stronger among HPV-positive tumors. A high E-cadherin expression was associated with tumor grade. No relationship between these markers and survival emerged. In conclusion, β-catenin and vimentin seem to play different roles in OPSCC: the former in the tumor tissue itself, and the latter in the tumor stroma. HPV infection may exploit the β-catenin and vimentin pathways in carcinogenic process. More, β-catenin may serve as a marker for the occurrence of regional metastasis.

Gastric cancer (GC) is one of the most prevalent cancers and a major cause of cancer related mortality worldwide. Incidence of GC is affected by various factors, including genetic and environmental factors. Despite extensive research has been done for molecular characterization of GC, it remains largely unknown. Therefore, further studies specially conducted among various ethnicities in different geographic locations, are required to know the precise molecular mechanisms leading to tumorigenesis and progression of GC. The expression patterns of seven candidate genes, including β-catenin, Notch1, GATA6, CDX2, miR-34a, miR-181a, and miR-93 were determined in 24 paired GC tissues and corresponding non-cancerous tissues by quantitative Real-Time PCR. The association between the expression of these genes and clinicopathologic factors were also investigated. Our results demonstrated that overall mRNA levels of GATA6 were significantly decreased in the tumor samples in comparison with the non-cancerous tissues (median fold change (FC) = 0.3143; P = 0.0003). Overall miR-93 levels were significantly increased in the tumor samples relative to the non-cancerous gastric tissues (FC = 2.441; P = 0.0002). β-catenin mRNA expression showed a strong positive correlation with miR-34a (r = 0.5784; P = 0.0031), and miR-181a (r = 0.5652; P = 0.004) expression. miR-34a and miR-181a expression showed a significant positive correlation (r = 0.4862; P = 0.016). Moreover, lower expression of Notch1 was related to distant metastasis in GC patients with a borderline statistical significance (p = 0.0549). These data may advance our understanding of the molecular biology that drives GC as well as provide potential targets for defining novel therapeutic strategies for GC treatment.

Anorectal malformations (ARMs) is one of the most common gastrointestinal anomalies. Previous research revealed that miR-92a-2-5p was upregulated in ARMs. However, the underlying roles remains unknown. The current study was to further investigate the spatiotemporal expression patterns of miR-92a-2-5p and its target gene protein kinase C alpha (PRKCA) predicted by bioinformatic method, and to explore their potential functions in anorectal malformations (ARMs). Rat models with ethylenethiourea-induced ARMs were made for subsequent experiments. Direct target relationship between miR-92a-2-5p and PRKCA was validated using a luciferase reporter assay. The spatiotemporal expression pattern of miR-92a-2-5p was evaluated using fluorescence in situ hybridization (FISH), while the expression of PRKCA was revealed by immunohistochemical staining and western blotting. IEC-6 cells were transfected with mimics/mimics NC (Negative control)/inhibitor/inhibitor NC of miR-92a-2-5p or si-PRKCA/si-PRKCA NC, respectively. Then the downstream molecules of miR-92a-2-5p, PRKCA and β-catenin, were subsequently detected. Meanwhile, apoptosis and viability assays were measured. Dual luciferase assay confirmed the direct regulatory relationship between miR-92a-2-5p and PRKCA. FISH revealed that miR-92a-2-5p was expressed with a higher level in ARMs fetuses. Further analyses of PRKCA showed lower protein expression level in ARMs group, which was opposite to miR-92a-2-5p. In vitro experiments revealed that overexpression of miR-92a-2-5p or knockdown of PRKCA can down-regulate PRKCA, up-regulate and facilitate nuclear localization of β-catenin, increase apoptosis and decrease proliferation of IEC-6. Taken together, these findings suggest that aberrantly high expression of miR-92a-2-5p potentially contribute to ARMs by inhibiting proliferation and enhancing apoptosis of intestinal cells via negatively regulating PRKCA/β-catenin., Competing Interests: Declaration of competing Interest The authors declares that they have no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Introduction: Predictors of non-response in mismatch repair deficiency cancers are poorly understood. Upregulation of the canonical Wnt pathway has been associated with decreased immune cell infiltration in many cancer types. The relationship between Wnt/β-catenin pathway activation and the programmed death-ligand 1 axis in endometrial cancer remains poorly characterized. This study evaluates β-catenin expression in a well characterized cohort of endometrial cancers by mismatch repair status and programmed death-ligand 1 expression., Methods: Whole sections of formalin-fixed, paraffin embedded tissue from 23 Lynch syndrome-associated carcinomas, 20 mutL homolog-1 (MLH1) promoter hypermethylated carcinomas, and 19 mismatch repair intact carcinomas were evaluated. Immunohistochemistry staining for β-catenin and programmed death-ligand 1 was performed on all cases. Programmed death-ligand 1 expression was scored in both the tumor and the peri-tumoral immune compartment. Tumor staining was classified as positive when membranous (programmed death-ligand 1) staining was present in ≥1% of tumor cells. Immune stromal staining was scored as positive when ≥5% of peritumoral and intratumoral immune cells (including lymphocytes and macrophages) showed reactivity., Results: Six tumors (6/62, 9.7%) demonstrated nuclear expression of β-catenin (4 were Lynch syndrome-associated, 1 was MLH1 methylated, 1 was mismatch repair intact). The majority of tumors with nuclear β-catenin expression demonstrated concomitant tumoral programmed death-ligand 1 expression (5/6, 83.3%) and were more likely to demonstrate tumoral programmed death-ligand 1 expression compared to tumors without nuclear β-catenin expression (83.3% vs 39.3%, p=0.04). Both tumoral and immune cell expression of programmed death-ligand 1 was statistically significantly associated with mismatch repair deficient tumors., Discussion: Tumors demonstrating nuclear β-catenin expression were more likely to express tumoral programmed death-ligand 1 staining than tumors without nuclear β-catenin expression. Nuclear β-catenin expression could be a potential predictive biomarker for non-response to immune checkpoint inhibition in mismatch repair deficient tumors. Nuclear β-catenin expression status should be considered as a translational endpoint in future clinical trials of immune checkpoint inhibition in endometrial cancer., Competing Interests: Competing interests: None declared., (© IGCS and ESGO 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: The tumor-initiating function of long non-coding RNA (lncRNA), zinc finger protein multitype 2 antisense RNA 1 (ZFPM2-AS1) was reported in lung cancer, yet the relevance of ZFPM2-AS1 in retinoblastoma (RB), a malignancy representing 2.5% to 4% incidence of cancers among children, has not been elucidated. Thus, we attempted to assess the effect of ZFPM2-AS1 and underlying mechanism in RB progression., Methods: First, comparing the differentially expressed lncRNAs in normal retinal tissues as well as RB tissues, the target lncRNA ZFPM2-AS1 was screened out. We then assayed the ZFPM2-AS1 expression in three RB cell lines, and carried out methylthiazol tetrazolium (MTT), transwell assays, and flow cytometric analyses to examine the role of si-ZFPM2-AS1 on cell behaviors. Following online database predication, the correlations between ZFPM2-AS1 and microR-515 (miR-515) or homeobox A1 (HOXA1) were corroborated by dual-luciferase reporter gene assays. Quantitative real-time PCR along with Western blot assays was fulfilled to ascertain the expression of relevant genes., Results: ZFPM2-AS1 was significantly overexpressed in RB tissues and cell lines, and ZFPM2-AS1 silencing curtailed the growth and metastasis of RB cells both in vitro and in vivo. Bioinformatic websites and dual-luciferase reporter gene assays disclosed that ZFPM2-AS1 might perform as a competing endogenous RNA for miR-515 and positively correlate with HOXA1 to activate the Wnt/β-catenin signaling pathway., Conclusion: Altogether, these data demonstrated that ZFPM2-AS1 interacted with HOXA1 to promote RB development via mediating miR-515, establishing a promising therapeutic biomarker for RB and prognosis.

Introduction: Recent reports on gene expression profiling (GEP) show several genes associated with malignant progression of GIST. However, genes associated with malignant transformation have not been clarified. Here, we aimed to reveal distinct genes in aggressive malignant GIST, using comprehensive gene expression analysis., Materials and Methods: We investigated GEP obtained by microarrays for 43 gastric GISTs, which mostly harbored KIT and PDGFRA mutations and integrated clinicopathological risk information. RT-PCR and immunohistochemistry were performed for FZD7, a receptor of Wnt ligands., Results: GEP divided 43 gastric GISTs into two clusters. A cluster included seven of eight high-risk GISTs (88%) in modified NIH classification and was defined as high-risk cluster; the other cluster was defined as low-risk cluster. The number of probes with over 3-fold changes between the two clusters was 1,177, in which probes corresponding to 16 oncogenes were included. Genes involved in the Wnt signaling pathway were the most abundant among the 16 oncogenes. Focusing on 73 Wnt signaling pathway genes of the 21,578 probes, 12 upregulated and 5 downregulated genes were found in the high-risk cluster. Major cascade genes promoting the Wnt/β-catenin signaling pathway, including WNT11, FZD family, and DVL2, were upregulated in the high-risk cluster. SNAI1, SNAI2, and BIRC5, which are activated by this pathway and increase cell proliferation, were also upregulated. These gene expression alterations were consistent in the positive direction of this pathway. GISTs in high-risk cluster strongly expressed FZD7., Conclusion: Wnt/β-catenin signaling pathway may play an important role in malignant transformation of indolent GIST., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest., (Copyright © 2020 Elsevier Ltd, BASO ~ The Association for Cancer Surgery, and the European Society of Surgical Oncology. All rights reserved.)

Objective: The aim of this study was to investigate the role of micro ribonucleic acid (miR)-92a in the pathogenesis of oral squamous cell carcinoma (OSCC)., Materials and Methods: The relative expression level of miR-92a in OSCC cell lines and normal oral epithelial keratinocyte cell lines was detected via quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR). Subsequently, the effects of miR-92a silencing on proliferation and apoptosis of OSCC cells were determined via cell counting kit-8 (CCK-8) assay, flow cytometry, and colony formation assay, respectively. Whether miR-92a could bind to the 3'-untranslated region (3'-UTR) of target mRNA was detected using Dual-Luciferase reporter gene assay. Furthermore, changes in Wnt/β-catenin pathway-associated proteins were explored via Western blotting., Results: The expression of miR-92a in OSCC cell lines was significantly higher than that of normal oral epithelial keratinocyte cell lines (p<0.05). Silencing of miR-92a significantly inhibited proliferation and promoted apoptosis of OSCC cells (p<0.05). Luciferase reporter gene assay confirmed that miR-92a could bind to the 3'-UTR of Kruppel-like factor 4 (KLF4) gene. After miR-92a silencing, the expressions of Wnt/β-catenin pathway-associated proteins were remarkably down-regulated, including β-catenin, c-Myc, and Wnt3a (p<0.05)., Conclusions: Silencing of miR-92a inactivates the Wnt/β-catenin signaling pathway by targeting KLF4, thereby inhibiting proliferation and promoting apoptosis of OSCC cells. Our findings suggest that miR-92a may be a potential therapeutic target for OSCC patients.

Calcitonin is expressed in the epithelium of endometrium, and modulates zonula adherens junctions which are composed of cadherin-catenins complex during the implantation window. Trophoblastic cells which have complex interaction with the epithelial cells of endometrium during implantation were demonstrated to have calcitonin receptors. Mechanism of action of calcitonin on trophoblastic cells has not yet been elucidated. Therefore, it was aimed to determine the effects of calcitonin on the expressions of β-catenin and phospho-β-catenin in a dose depended manner under the influence of progesterone and estrogen hormones (P + E) by using JAR cell line through the immunocytochemical and Western blot analyses. Moreover, adherens junctions (AJs) were ultrastructurally investigated to assess the involvement of cadherin-catenin complex in accordance with the changes in the specified parameters. Immunocytochemical analysis showed that only 10 nM calcitonin treated group had increased expression of membranous β-catenin compared to the control group, while there was decreased expression of β-catenin in the nucleus of all the experimental groups. Cytoplasmic expressions of the phospho-β-catenin decreased in all experimental groups compared to the control group while the decrease in the nuclear expression was remarkable in the groups treated with P + E, and P + E + 250 nM calcitonin. Western blot analysis showed that total β-catenin and phospho-β-catenin expressions were not significantly different. Ultrastructural analysis showed that increase in the number of AJs was noticeable in the group treated with 10 nM calcitonin. Overall, the localization and expression levels of β-catenin and phospho-β-catenin suggest that calcitonin could show its effects through the non-canonical pathway in the trophoblastic cells., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest. The authors alone are responsible for the content and writing of the paper., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Frizzled-2 plays an important role in maintaining normal hepatic cell functionality. This study aimed to investigate the role of inhibition of Frizzled-2 in protecting rat liver BRL-3A cells from Hypoxia/Reoxygenation (H/R). In vitro H/R hepatic cell model was established by culturing BRL-3A cells under H/R condition. Frizzled-2 siRNA was transfected into BRL-3A cells to inhibit Frizzled-2 signaling. Wnt5a and Frizzled-2 were significantly increased in BRL-3A cells upon H/R treatment. H/R treatment induced cell cytotoxicity, the early apoptosis rate and the intracellular Ca 2+ level in BRL-3A cells while silencing frizzled-2 gene decreased the H/R induced cell cytotoxicity, apoptosis and intracellular Ca 2+ level. In vivo mice study further showed the up-regulation of Frizzled-2/Wnt 5 pathway and cleaved Caspase-3 expression in liver tissues under ischemia and reperfusion injury (IRI). In summary, inhibition of Frizzled-2 by its siRNA may protects BRL-3A cells by attenuating the H/R induced cell cytotoxicity and apoptosis., (Copyright © 2019. Publicado por Elsevier España, S.L.U.)

Cell fate specification defines the earliest steps towards a distinct cell lineage. Neural crest, a multipotent stem cell population, is thought to be specified from the ectoderm, but its varied contributions defy canons of segregation potential and challenges its embryonic origin. Aiming to resolve this conflict, we have assayed the earliest specification of neural crest using blastula stage chick embryos. Specification assays on isolated chick epiblast explants identify an intermediate region specified towards the neural crest cell fate. Furthermore, low density culture suggests that the specification of intermediate cells towards the neural crest lineage is independent of contact mediated induction and Wnt-ligand induced signaling, but is, however, dependent on transcriptional activity of β-catenin. Finally, we have validated the regional identity of the intermediate region towards the neural crest cell fate using fate map studies. Our results suggest a model of neural crest specification within a restricted epiblast region in blastula stage chick embryos., Competing Interests: Declaration of competing interest Authors declare no competing financial interest., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Imbalance between inflammatory Th17 cells and immunosuppressive regulatory T cells (Treg) contributes to the progression of chronic obstructive pulmonary disease (COPD). We aims to investigate roles and mechanisms of secreted frizzled-related protein 2 (sFRP2) in airway inflammation and Th17/Treg differentiation in COPD. sFRP2 was significantly upregulated in the serum of patients with COPD and in human bronchial epithelial (HBE) cells that were exposed to cigarette smoke extract (CSE). sFRP2 was negatively correlated with FEV1/FVC. CSE increased IL-6 and TNF-α in HBE cells, which was reversed by sFRP2 silencing. CSE exposure elevated the percentage of Th17 in CD3 + CD8 - cells while reduced the percentage of Treg in CD4 + CD25 + cells. Knockdown of sFRP2 in peripheral blood mononuclear cells (PBMCs) attenuated Th17 differentiation and induced Treg differentiation. CSE suppressed the expression of β-catenin and Cyclin D1 in PBMCs while knockdown of sFRP2 markedly reversed the inhibitory effects of CSE. Wnt/β-catenin inhibition by Dickkopf-1 reversed the inhibitory effect of si-sFRP2 on the production of inflammatory cytokines and imbalance between Th17 and Treg cells caused by CSE. CSE induced sFRP2 potentiated airway inflammation and disturbed Th17/Treg homeostasis by inhibiting β-catenin., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Purpose: Long non-coding RNAs (lncRNAs) play important roles in cancer, but the effects of lncRNA-ATB on the proliferation and apoptosis of non-small cell lung cancer (NSCLC) cells remain unclear. Therefore, the present study was conducted to explore the role of this lnc-RNA in NSCLC., Methods: The NSCLC NCI-H838 cell line cultured in vitro were used as the objects of study. First, the expressions of lncRNA-ATB, miR-200a and β-catenin in cells were detected. Then, the expression of lncRNA-ATB was knocked down using siRNA, and the effects of low expression of lncRNA-ATB on miR-200a/β-catenin pathway and apoptosis were studied., Results: Compared with normal lung epithelial cells BEAS-2B, NCI-H838 cells had a significantly increased level of lncRNA-ATB (p<0.01), a significantly decreased level of miR-200a (p<0.01) and also a significantly increased level of β-catenin (p<0.01). After knockdown of lncRNA-ATB using si-ATB, the expression level of miR-200a was significantly increased, while that of β-catenin was significantly decreased. Besides, si-ATB remarkably increased the expression of Bcl-2 (p<0.01), and reduced the expressions of cleaved-caspase3 and Cytochrome C (p<0.01) and the apoptosis. In addition, the miR-200a mimic lowered obviously the expression of β-catenin (p<0.05) and reduced apoptosis., Conclusions: This study suggests that lncRNA-ATB promotes the apoptosis of NSCLC cells through inhibiting the expression of miR-200a and reversely promoting the expression of β-catenin.

Because posttraumatic inflammation contributes to the progression of neuron degeneration, attenuating inflammation is important for reducing neural degeneration. Sirtuin 1 (SIRT1) has been shown to play a critical role in the chronic diseases, such as neurodegenerative diseases and aging. However, the role that SIRT1 plays in regulating neuroinflammation in spinal cord injuries (SCIs) remains unclear. In this study, we investigate the effect of SIRT1 on the SCI model and on lipopolysaccharide (LPS)-treated primary microglia using a pharmacological intervention (SRT1720, an agonist of SIRT1). Results showed that SIRT1 levels gradually decreased in spinal cord until the fourth week after SCI, while the level of 8-hydroxy-2'-deoxyguanosine increased. SIRT1 was negatively correlated with the expression of β-catenin following SCI. The administration of SRT1720 significantly improved number of neurons and the Basso, Beattie, and Bresnahan score after SCI. The number of ionizing calcium-binding adaptor molecule 1 (Iba1)-positive microglia, levels of β-catenin and NF-kB p65, and proinflammatory cytokines [tumor necrosis factor alpha and interleukin (IL) 12] decreased significantly after SRT1720 treatment, while IL-10 increased after SCI. Furthermore, both SIRT1 and SRT1720 significantly inhibited β-catenin gene and protein expression; β-catenin transcriptional activity also decreased in a dose-dependent manner following SIRT1 treatment of LPS-treated microglia. These findings suggest that SIRT1 may have a neuroprotective effect by suppressing microglial activation via downregulation of the Wnt/β-catenin signal following SCI.

Objective: This clinical study aimed to explore the expression of miR-195-5p and Yes-associated protein (YAP) in gastric cancer (GC) and their relevant apoptosis mechanism., Patients and Methods: Thirty GC patients treated in our hospital were enrolled as a patient group and 30 normal people who underwent physical examination during the same period were enrolled as a normal group. The purchased GC cells and human gastric mucous membrane cells were used to transfect miR-195-5p-mimics, miR-NC, si-YAP, and si-RNA into MKN45 and SGC7901 cells. qRT-PCR was used to detect the expression of miR-195-5p and YAP in samples, and WB was used to detect the expression of YAP, Caspase-3, Caspase-9, β-catenin, c-myc, and cyclin Dl. CCK-8, the transwell, and the flow cytometry were used to detect cell proliferation, invasion, and apoptosis. The dual fluorescent enzyme reporter was used to determine the relationship between miR-195-5p and YAP., Results: MiR-195-5p was lowly expressed in the tissues and serum of patients, but YAP was highly expressed, and the area under the miR-195-5p and YAP curves was more than 0.9. MiR-195-5p and YAP were associated with tumor diameter, TNM stage, lymph node metastasis, and differentiation in GC patients. The overexpression of miR-195-5p and the inhibition of YAP expression can inhibit cell proliferation and invasion and promote apoptosis. WB assay showed that the overexpression of miR-195-5p and the inhibition of YAP could inhibit the expression of β-catenin, c-myc, and cyclin D1 protein and promote the expression of Caspase-3, Caspase-9 protein. The dual fluorescent enzyme reporter revealed that there was a targeting relationship between miR-195-5p and YAP., Conclusions: The overexpression of miR-195-5p can inhibit YAP-mediated Wnt/β-catenin signaling pathway and promote cell apoptosis, so it may be a potential therapeutic target for GC.

Chromosome 8 open reading frame 4 (C8orf4) is an activator of Wnt signaling pathway, and participates in the tumorigenesis and progression of many tumors. The expression levels of C8orf4 and β-catenin were assessed via immunohistochemical staining in 100 cervical squamous cell carcinoma (CSCC) tissues, 50 high-grade squamous intraepithelial lesions (HSILs), 50 low-grade squamous intraepithelial lesions (LSILs), and 50 normal cervical tissues. Bisulfite sequencing polymerase chain reaction analysis was used to examine the methylation status of the C8orf4 locus in CSCC and normal cervical tissues. The expression rates of C8orf4 and β-catenin were significantly higher in CSCCs or HSILs than in LSILs or normal cervical tissues (P < .05). C8orf4 expression was positively correlated with the poor differentiation of CSCCs (P = .009), and with aberrant expression of β-catenin in CSCCs (P = .002) and squamous intraepithelial lesions (P < .001). The methylation rate of C8orf4 in CSCCs was significantly lower than that in normal cervical tissues (P = .001). The Cancer Genome Atlas genomics data also confirmed that the mRNA expression of C8orf4 was positively associated with the copy number alteration of C8orf4 (correlation coefficient = 0.213, P < .001), and negatively correlated with the methylation level of C8orf4 (correlation coefficient = -0.408, P < .001). In conclusion, the expressions of C8orf4 and β-catenin were synergistically increased in CSCCs and HSILs and higher than those in LSILs and normal cervical tissues. The methylation level of C8orf4 is decreased in CSCCs and is responsible for the increased expression of C8orf4.

Aim: To explore the feasibility of human placenta extracellular matrix (HPECM) hydrogel in restoring the hair-inductive capacity of high-passaged (P8) dermal papilla cells (DPCs) for hair follicle regeneration. Materials & methods: HPECM hydrogel was prepared following decellularization and enzymatic solubilization treatment. DPCs isolated from human scalp were cultured in 2D and 3D environments. The hair-inductive ability of DPCs was assessed by quantitative RT-PCR, immunofluorescence staining, immunoblotting and patch assay. Results: DPCs (P8) formed spheres when cultured on the HPECM hydrogel. The expression levels of Versican, ALP, and β -catenin were restored in the DP spheres. HPECM hydrogel-cultured DP spheres co-grafted with newborn mouse epidermal cells regenerated new hair follicle. Conclusion: HPECM hydrogel successfully restores the hair-inductive capacity of high-passaged DPCs.

KHC-4 is a 2-phenyl-4-quinolone analogue that exhibits anticancer activity. Aberrant activation of β-catenin signaling contributes to prostate cancer development and progression. Therefore, targeting β-catenin expression could be a useful approach to treating prostate cancer. We found that KHC-4 can inhibit β-catenin expression and its signaling pathway in DU145 prostate cancer cells. Treatment with KHC-4 decreased total β-catenin expression and concomitantly decreased β-catenin levels in both the cytoplasm and nucleus of cells. KHC-4 treatment also inhibited β-catenin expression and that of its target proteins, PI3K, AKT, GSK3β and TBX3. We monitored the stability of β-catenin with the proteasomal inhibitor, MG132, in DU145 cells and found that MG132 reversed KHC-4-induced proteasomal β-catenin degradation. We verified CDK1/β-catenin expression in KHC-4 treated DU145 cells. We found that roscovitine treatment reversed cell proliferation by arresting the cell cycle at the G2/M phase and β-catenin expression caused by KHC-4 treatment. We suggest that KHC-4 inhibits β-catenin signaling in DU145 prostate cancer cells.

Y-box binding protein-1 (YB-1), an important transcription and translation regulator protein, is known to increase cancer cell invasiveness and spreading. Here, we report its role in breast cancer, particularly in mediating cell invasion in triple-negative breast cancer (TNBC). YB-1 stable knockdown (shYB-1) significantly reduced the invasive potential of MDA-MB-231 TNBC cells in 2D and 3D (spheroid) cultures. Whole proteome mass spectrometry analysis showed an enrichment of cell adhesion and cell to matrix interaction proteins, notably, matrix metalloproteinase-1 (MMP1) and beta-catenin (CTNNB1), which are known to play critical roles in cancer metastasis. shYB-1 cells exhibited substantial downregulation of MMP1 and CTNNB1 mRNA and protein expression, with reduced MMP1 enzyme activity. YB-1 was also observed to bind to the promoter of MMP1 and overexpression of MMP1 plasmid in shYB-1 cells increased cell invasion. Finally, analysis of tumour samples from the Gene Expression-Based Outcome for Breast Cancer Online (GOBO) database revealed that high gene expressions of YBX1, MMP1 and CTNNB1 predict for a significantly lower 10-year distant metastasis free survival. Altogether, this study shows that YB-1 mediates breast cancer invasion and metastasis via regulation of MMP1 and beta-catenin., (Copyright © 2019 Elsevier B.V. All rights reserved.)

IQ motif-containing GTPase-activating protein 1 (IQGAP1) is a scaffold protein that participates in several cellular functions, including cytoskeletal regulation, cell adhesion, gene transcription and cell polarization. IQGAP1 has been implicated in the tumorigenesis and progression of several human cancers. However, the role of IQGAP1 in pancreatic ductal adenocarcinoma (PDAC) is still unknown. We found that IQGAP1 expression was an independent prognostic factor for PDAC. IQGAP1 upregulation significantly promoted cell proliferation, migration, invasion and epithelial-mesenchymal transition (EMT), whereas IQGAP1 downregulation impaired its oncogenic functions. Overexpression of IQGAP1 increased the protein level of Dishevelled2 (DVL2) and enhanced canonical Wnt signaling as evidenced by increased DVL2 level, β-catenin transcriptional activity, β-catenin nuclear translocation and expression of the direct target genes of β-catenin (cyclin D1 and c-myc). In contrast, knockdown of IQGAP1 decreased the level of DVL2 and attenuated Wnt/β-catenin signaling. In vivo results revealed that IQGAP1 promoted tumor growth and metastasis. Co-immunoprecipitation studies demonstrated that IQGAP1 interacted with both DVL2 and β-catenin. Moreover, knockdown of DVL2 reversed IQGAP1-induced EMT. Our findings thus confirmed that IQGAP1 could be used as a potential target for PDAC treatment.

Objective: The objective of this study was to determine the prognostic and predictive impact of β-catenin, TCF21 and WISP1 expression in patients with squamous cell carcinomas of the head and neck who underwent primary radiotherapy or concomitant chemoradiotherapy., Study Design: Prospective cohort study., Setting: University hospital., Participants: Protein expression profiles of β-catenin, TCF21, WISP1 and p16 were determined by immunohistochemical analyses in tissue samples of 59 untreated patients. Expression was correlated with different outcome parameters., Main Outcome Measures: Impact of TNM classification, grading, sex, age, gender, type of therapy, response to therapy and p16 status on disease-specific (DSS) and disease-free survival (DFS)., Results: Patients with high expression of TCF21 were associated with significantly worse disease-specific survival (P = 0.005). In a multivariable analysis, TCF21 was a significant determinant of disease-specific survival. (HR 3.01; P = 0.036). Conversely, low expression of β-catenin (P = 0.025) and WISP1 (P = 0.037) revealed a better response to radiotherapy., Conclusion: Since data show that TCF21 is a prognostic factor for disease-specific survival and WISP1 and ß-catenin are predictive factors for clinical outcome after definitive radiotherapy, further studies are warranted to prove these preliminary but very promising findings., (© 2019 John Wiley & Sons Ltd.)

Prostate cancer (PCa) remains one of the leading causes of cancer-related deaths among males. The aim of the current study was to investigate the ability of microRNA-150 (miR-150) targeting transient receptor potential melastatin 4 (TRPM4) to mediate epithelial-mesenchymal transition (EMT), invasion, and metastasis through the β-catenin signaling pathway in PCa. Microarray analysis was performed to identify PCa-related differentially expressed genes, after which both the mirDIP and TargetScan databases were employed in the prediction of the miRNAs regulating TRPM4. Immunohistochemistry and RT-qPCR were conducted to determine the expression pattern of miR-150 and TRPM4 in PCa. The relationship between miR-150 and TRPM4 expression was identified. By perturbing miR-150 and TRPM4 expression in PCa cells, cell proliferation, migration, invasion, cycle, and apoptosis as well as EMT markers were determined accordingly. Finally, tumor growth and metastasis were evaluated among nude mice. Higher TRPM4 expression and lower miR-150 expression and activation of the β-catenin signaling pathway as well as EMT stimulation were detected in the PCa tissues. Our results confirmed TRPM4 as a target of miR-150. Upregulation of miR-150 resulted in inactivation of the β-catenin signaling pathway. Furthermore, the upregulation of miR-150 or knockdown of TRPM4 was observed to suppress EMT, proliferation, migration, and invasion in vitro in addition to restrained tumor growth and metastasis in vivo. The evidence provided by our study highlights the involvement of miR-150 in the translational suppression of TRPM4 and the blockade of the β-catenin signaling pathway, resulting in the inhibition of PCa progression.

Lack of blood or glucose supply is the most common pathological factor in the brain. To cope with such an energy stress, initiating programmed autophagic processes in neurons is required. However, the mechanisms controlling neuronal autophagy during starvation remain far from clear. Here, we report an essential role of 14-3-3γ in starvation-activated neuronal autophagic influx signaling and elucidate the underlying mechanism. Double-fluorescent immunostaining demonstrates that 14-3-3γ protein elevation is well co-localized with Beclin-1 and LC3 elevation in cortical neurons in ischemic brains. Starvation treatment activates autophagic influx and upregulates Beclin-1 and only the γ isoform of 14-3-3 in N2a cells and cultured cortical neurons. Suppressing overall 14-3-3 function by difopein overexpression or knocking-out the γ isoform of 14-3-3 is sufficient to abolish starvation-induced Beclin-1 induction and LC3 activation while overexpressing 14-3-3γ but no other 14-3-3 isoform significantly upregulate Beclin-1-LC3 signaling. Upon starvation, 14-3-3γ binds more p-β-catenin but less Beclin-1. Finally, overexpressing 14-3-3γ reactivates β-catenin-suppressed Beclin-1-LC3 signaling in neuronal cells. Taken together, our data reveal that starvation-induced 14-3-3γ is required for β-catenin-Beclin-1-LC3-autophagy in starved neurons in vitro and in vivo, which may provide insights in the treatment of neurologic diseases such as stoke.

Aberrant Wnt/β-catenin signaling is a characteristic feature of colorectal cancer (CRC), therefore, understanding the underlying mechanisms of aberrant Wnt/β-catenin signaling will improve the treatment outcome of CRC. Expression of MNX1 in paired fresh CRC tissues and corresponding adjacent normal tissues were examined by qPCR and Western blotting. The levels of MNX1 in paraffin-embedded CRC specimens were detected by immunohistochemistry (IHC). The role of MNX1 in growth and proliferation of CRC cells was evaluated by MTT and colony formation assay. Luciferase reporter analysis and western blotting were carried out to explore the influence of MNX1 on Wnt/β-catenin signaling. The results showed that expression of MNX1 is markedly upregulated in CRC tissues and positively correlated with level of Ki67, and overexpression of MNX1 significantly promotes the proliferation of CRC cells. Further study showed that ectopic expression of MNX1 activates the Wnt/β-catenin signaling and upregulates the expression of c-Myc and CCND1, the downstream genes of Wnt/β-catenin signaling. Therefore, MNX1 plays an indispensable role in promoting of human CRC progression and may represent a novel therapeutic target for CRC., (© 2019 International Federation for Cell Biology.)

We previously reported that genetic deletion of β-catenin in mouse corneal keratocytes resulted in precocious corneal epithelial stratification. In this study, to strengthen the notion that corneal keratocyte-derived Wnt/β-catenin signaling regulates corneal epithelial stratification during mouse development, we examined the consequence of conditional overexpression of a stabilized β-catenin mutant (Ctnnb1 ΔE3 ) in corneal keratocytes via a doxycycline (Dox)-inducible compound transgenic mouse strain. Histological analysis showed that conditional overexpression of Ctnnb1 ΔE3 in keratocytes inhibited corneal epithelial stratification during postnatal development. Unlike the corneal epithelium of the littermate controls, which consisted of 5-6 cell layers at postnatal day 21 (P21), the mutant corneal epithelium contained 1-2 or 2-3 cell layers after Dox induction from embryonic day 0 (E0) to P21 and from E9 to P21, respectively. X-gal staining revealed that Wnt/β-catenin signaling activity was significantly elevated in the corneal keratocytes of the Dox-induced mutant mice, compared to the littermate controls. Furthermore, RT-qPCR and immunostaining data indicated that the expression of Bmp4 and ΔNp63 was downregulated in the mutant corneas, which was associated with reduced corneal epithelial proliferation in mutant epithelium, as revealed by immunofluorescent staining. However, the expression of Krt12, Krt14 and Pax6 in the mutant corneas was not altered after overexpression of Ctnnb1 ΔE3 mutant protein in corneal keratocytes. Overall, mutant β-catenin accumulation in the corneal keratocytes inhibited corneal epithelial stratification probably through downregulation of Bmp4 and ΔNp63 in the corneal epithelium.

Purpose: The aim of this study is to explore the roles of β-catenin, decorin, septin-7, and S100A10 expression in colorectal cancer development., Methods: Twenty-five BALB/c mice were divided into five groups; four groups were administrated N,N-dimethylhydrazine for 0, 10, 15, and 20 weeks, and one group was administrated normal saline for 20 weeks. The colons were collected for histopathological analysis. Protein samples prepared from the frozen colon tissues of mice treated with N,N-dimethylhydrazine for the different time points were evaluated using the isobaric tags for relative and absolute quantification (iTRAQ) labeling technique coupled with the 2D liquid chromatography-tandem mass spectrometry analysis. Based on the proteomic analysis results, immunohistochemical staining of β-catenin, decorin, septin-7, and S100A10 was performed in paraffin-embedded mice colorectal tissue, and 53 cases of human hereditary polyposis colorectal cancer samples., Results: Colorectal cancer was observed in mice treated with N,N-dimethylhydrazine for 20 weeks, and adenomas were observed in mice subjected to the 10-, and 15-week treatments. Seventy-two differentially expressed proteins were involved in the development of cancer as per the iTRAQ and spectrometry analysis. In normal epithelium, adenoma, and cancer from human hereditary polyposis colorectal cancer, S100A10 expression (c2 = 100.989, P = 0.000) was highest in cancer, whereas decorin (c2 = 12.852, P = 0.002) and septin-7 (c2 = 66.519, P = 0.002) expressions were highest in the normal epithelium, which was confirmed via immunohistochemical staining., Conclusions: The subcellular localization of β-catenin and decorin, septin-7, and S100A10 expressions are associated with the development of colorectal cancer in mice after N,N-dimethylhydrazine treatment and in human hereditary polyposis colorectal cancers.