It has been shown that in hereditary and most sporadic colon tumours, components of the <em>Wnt</em> pathway are mutated. The <em>Wnt</em> target MET has been implicated in the development of colon cancer. Here, we show that overexpression of wild-type or a constitutively activated form of MET in colon epithelial cells leads to increased transformation irrespective of <em>Wnt</em> signalling. Fetal human colon epithelial cells without aberrant <em>Wnt</em> signalling were transfected with wild-type or mutated MET constructs. Expression of these constructs leads to increased phosphorylation of MET and its downstream targets PKB and MAPK. Upon stimulation with HGF, the expression of E-cadherin is downregulated in wild-type MET-transfected cells, whereas cells expressing mutated MET show low E-cadherin levels independent of stimulation with ligand. This implies a higher migratory propensity of these cells. Furthermore, fetal human colon epithelial cells expressing the mutated form of MET have colony-forming capacity in soft agar, while cells expressing wild-type MET show an intermediate phenotype. Subcutaneous injection of mutated MET-transfected cells in nude mice leads to the formation of tumours within <em>12</em> days in all mice injected. At this time point, mock-transfected cells do not form tumours, while wild-type MET-transfected cells form subcutaneous tumours in one out of five mice. We thus show that MET signalling can lead to increased transformation of colon epithelial cells independent of <em>Wnt</em> signalling and in this way could play an essential role in the onset and progression of colorectal cancer.

BACKGROUND

This study was to explore the expression profile of endometrial carcinoma (EC) and identify the potential molecular mechanism and therapeutic targets.

METHODS

Differentially expressed genes (DEGs) and differentially expressed miRNAs (DEMs) were identified in EC using mRNA and miRNA sequencing data released by the Cancer Genome Atlas database; then, gene function and pathway of DEGs were analyzed based on the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway databases; finally, the transcription factors (TFs) latently regulating the DEGs and DEMs were predicted and a TF-miRNA-Gene network was then established to summarize the regulatory links between TFs, DEMs and DEGs.

RESULTS

One thousand five hundred and forty two upregulated and 1,885 downregulated DEGs, 34 upregulated and <em>12</em> downregulated DEMs were identified. The principal DEGs-enriched functions were cell differentiation, cell migration, and cell surface receptor signaling pathway. The DEGs-enriched cell signaling pathways including the MAPK, <em>Wnt</em> signaling pathway, and the p53 signaling pathway. As shown in the TF-miRNA-Gene network, TFs such as CPBP and GKLF, miRNAs such as miR-141-3p and miR-130b-3p, regulated most of DEGs and DEMs.

CONCLUSIONS

These results may contribute to the study of the molecular mechanism and therapeutic targets in EC, and facilitate the discovery of new biomarkers for screening, diagnosis and monitoring.

The herb <i>Radix Ophiopogonis</i> (RO) has been used effectively to treat nasopharyngeal carcinoma (NPC) as an adjunctive therapy. Due to the complexity of the traditional Chinese herbs, the pharmacological mechanism of RO's action on NPC remains unclear. To address this problem, an integrative approach bridging proteome experiments with bioinformatics prediction was employed. First, differentially expressed protein profile from NPC serum samples was established using isobaric tag for relative and absolute quantification (iTRAQ) coupled 2-D liquid chromatography (LC)-MS/MS analysis. Second, the RO putative targets were predicted using Traditional Chinese Medicines Integrated Database and known therapeutic targets of NPC were collected from Drugbank and OMIM databases. Then, a network between RO putative targets and NPC known therapeutic targets was constructed. Third, based on pathways enrichment analysis, an integrative network was constructed using DAVID and STRING database in order to identify potential candidate targets of RO against NPC. As a result, we identified 13 differentially expressed proteins from clinical experiments compared with the healthy control. And by bioinformatics investigation, <em>12</em> putative targets of RO were selected. Upon interactions between experimental and predicted candidate targets, we identified three key candidate targets of RO against NPC: VEGFA, TP53, and HSPA8, by calculating the nodes' topological features. In conclusion, this integrative pharmacology-based analysis revealed the anti-NPC effects of RO might be related to its regulatory impact via the PI3K-AKT signaling pathway, the <em>Wnt</em> signaling pathway, and the cAMP signaling pathway by targeting VEGFA, TP53, and HSPA8. The findings of potential key targets may provide new clues for NPC's treatments with the RO adjunctive therapy.

The members of the <em>Wnt</em> glycoprotein family are important in embryogenesis and adult tissue homeostasis, and deletion of <em>WNT</em>-4 gene in mice leads to improper development of many organs including the adrenals. The objective of this study was to investigate the expression of <em>WNT</em>-4 gene in human adrenals and adrenocortical tumors. The <em>WNT</em>-4 mRNA expression (analyzed by quantitative real-time RT-PCR) was significantly higher in Conn's adenomas (p<0.01) and lower in Cushing's adenomas, virilizing carcinomas and fetal adrenals (p<0.05) compared with normal adult adrenals. <em>WNT</em>-4 mRNA expression was clearly upregulated by ACTH and 8-bromo-cAMP (8-BrcAMP) in primary cultures of normal adult adrenocortical cells, but downregulated by 8-BrcAMP and <em>12</em>- O-tetradecanoylphorbol-13-acetate (TPA) in human NCI-H295R adrenocortical carcinoma cells. Angiotensin II tended to increase <em>WNT</em>-4 mRNA expression at 24 hours and decreased it at 48 hours time point in both cell culture types. The abundant <em>WNT</em>-4 mRNA expression in Conn's adenomas and its hormonal regulation in adrenocortical cells suggest a role for <em>WNT</em>-4 in human adrenocortical function.

A proteomic analysis of procyanidin B(2) isolated from cocoa against oxidized low-density lipoprotein-induced lipid-laden macrophage formation was performed. Of approximately 400 detected proteins, <em>12</em> were differentially expressed as a result of B(2) treatment. They were subsequently identified by liquid chromatography-electrospray ionization-tandem mass spectrometry and the SWISS-PROT database. Further reverse transcriptase-polymerase chain reaction and Western blot analysis revealed that B(2) strongly inhibited arachidonic acid inflammatory reactions, apoptosis, and their coupled mitogen-activated protein kinase and NF-kappaB pathways. To highlight proteins or genes with similar expressed patterns and similarly biological function induced by B(2) in lipid-laden macrophages, a cluster and Kyoto Encyclopedia of Genes and Genomes pathway analysis were performed. The data were mapped to multiple pathways. Further validation of the bioinformatic results revealed that activation of <em>Wnt</em> signaling may contribute to the cardioprotection of B(2). The differentially expressed genes and proteins mentioned above induced by B(2) are through regulating nuclear transcription factors, activating peroxisome proliferator-activated receptor-gamma and inhibiting AP-1 mRNA expressions. These in vitro data help to interpret the beneficial effects of B(2) in reducing the risk of atherosclerosis after consumption of flavonoid-rich foods. Many differentially expressed genes induced by B(2) help to uncover novel targets and may help to target disease interactions in atherosclerosis in the future.

The appropriate spatial and temporal regulation of canonical <em>Wnt</em> signaling is vital for eye development. However, the literature often conflicts on the distribution of canonical <em>Wnt</em> signaling in the eye. Here, using a sensitive mouse transgenic reporter line, we report a detailed re-evaluation of the spatiotemporal dynamics of canonical <em>Wnt</em> signaling in the developing eye. Canonical <em>Wnt</em> activity was dynamic in the optic vesicle and later in the retina, while it was absent from the ectodermal precursors of the lens and corneal epithelium. However, later in corneal development, canonical <em>Wnt</em> reporter activity was detected in corneal stroma and endothelium precursors as they form from the neural crest, although this was lost around birth. Interestingly, while no canonical <em>Wnt</em> signaling was detected in the corneal limbus or basal cells at any developmental stage, it was robust in adult corneal wing and squamous epithelial cells. While canonical <em>Wnt</em> reporter activity was also absent from the postnatal lens, upon lens injury intended to model cataract surgery, it upregulated within <em>12</em> h in remnant lens epithelial cells, and co-localized with alpha smooth muscle actin in fibrotic lens epithelial cells from 48 h post-surgery onward. This pattern correlated with downregulation of the inhibitor of canonical <em>Wnt</em> signaling, Dkk3. These data demonstrate that canonical <em>Wnt</em> signaling is dynamic within the developing eye and upregulates in lens epithelial cells in response to lens injury. As canonical <em>Wnt</em> signaling can collaborate with TGFβ to drive fibrosis in other systems, these data offer the first evidence in a lens-injury model that canonical <em>Wnt</em> may synergize with TGFβ signaling to drive fibrotic posterior capsular opacification (PCO).

Emerging evidence has suggested that aberrant expression of micro (mi)RNAs contributes to the development of alcoholic liver injury (ALD). However, miRNA profiles distinguishing different stages of ALD have not yet been reported. The present study was designed to investigate the unique miRNA expression patterns at different stages of ALD in a rat model and analyze the gene functions and pathways of dysregulated miRNA‑targeted genes. Using microarray and stem‑loop quantitative polymerase chain reaction analyses, 16 miRNAs were identified as upregulated and 13 were identified as downregulated in an alcoholic steatohepatitis (ASH) group compared with the control group, while five miRNAs were identified to be upregulated and eight were identified to be downregulated in the alcoholic fatty liver (AFL) group as compared with the control group. Following further confirmation by Significance Analysis of Microarray and prediction by Prediction Analysis of Microarray, 8 and <em>12</em> types of miRNA were screened as molecular signatures in distinguishing AFL and ASH, respectively, from normal rat liver. In addition, several miRNA‑target pairs were predicted by computer‑aided algorithms (Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses using the Database for Annotation, Visualization and Integrated Discovery platform) and these genes may be involved in cancer signaling pathways, the <em>Wnt</em> signaling pathway and other signaling pathways. These results may provide novel miRNA targets for diagnosis and therapeutic intervention at different stages of ALD.

Solid pseudopapillary neoplasms of the pancreas (SPN) are rare tumors affecting mainly women. They show an activating mutation in CTNNB1, the gene for β-catenin, and consequently an overactivation of the Wnt/β-catenin pathway. This signaling pathway is implied in the pathogenesis of various aggressive tumors, including pancreatic adenocarcinomas (PDAC). Despite this, SPN are characterized by an unusually benign clinical course. Attempts to explain this lack of malignancy have led to the discovery of an aberrant expression of the transcription factor FLI1 in SPN.

In 42 primary pancreatic tumors the RNA-expression of the FLI1 targets DKK1, INPP5D, IGFBP3 and additionally two members of the Wnt/β-catenin pathway, namely BCL9 and BCL9L, was investigated using quantitative real time PCR. Expression of these genes was evaluated in SPN (n = 18), PDAC (n = 12) and the less aggressive intraductal papillary mucinous neoplasm IPMN (n = 12) and compared to normal pancreatic tissue. Potential differences between the tumor entities were evaluated using students t-test.

The results demonstrated a differential RNA-expression of BCL9L with a lack of expression in SPN (p < 0.001), RNA levels similar to normal tissue in IPMN and increased expression in PDAC (p < 0.04). Further, overexpression of the cyclin D1 inhibitor INPP5D in IPMN (p < 0.0001) was found. PDAC, on the other hand, showed the highest expression of IGFBP3 (p < 0.00001) with the gene still being significantly overexpressed in IPMN (p < 0.001). Nevertheless the difference in expression was significant between PDAC and IPMN (p < 0.05) and IGFBP3 RNA levels were significantly higher in PDAC and IPMN than in SPN (p < 0.0001 and p < 0.02, resp.).

This study demonstrates a significantly decreased expression of the β-catenin stabilizing gene BCL9L in SPN as a first clue to the possible reasons for the astonishingly benign behavior of this entity. In contrast, high expression of the gene was detected in PDAC supporting the connection between BCL9L expression and tumor malignancy in pancreas neoplasias. IPMN, accordingly, showed intermediate expression of BCL9L, but instead demonstrated a high expression of the cyclin D1 inhibitor INPP5D, possibly contributing to the better prognosis of this neoplasia compared to PDAC.

The use of high-throughput array technology is omnipresent in diverse areas specifically, early diagnosis of disease, discovery of infectious agents, search for biological markers and screening of potential drug candidates. Here, we integrated gene expression data with the network-based approach to identify novel genes that were playing central role in the network through interconnecting to a number of differentially expressed breast cancer genes. The 62 cancerous genes retrieved from the Breast Cancer Gene Database (BCGD) were mapped in the normalized data accessed from Stanford Microarray Database (SMD) to analyze their pattern. Interaction networks for each gene were constructed to understand the biology of the metastasis at systems level. The individual networks were fused together for the detection of interacting hubs, 38 novel genes were found to be deeply intermingled with the central hub node. Gene Ontology studies were made to depict the biology of the hub nodes not alone through gene ranking but by applying the Hyper geometric test with the Benjamini Hochberg False Discovery Rate (FDR) correction method at a significance level of 0.05. Analyzing p-values from the statistical test indicated that most of the novel genes were involved in the same biological function as the disordered genes like signal transducer, transcription regulator, enzyme binding, molecular transducer and receptor signaling protein activity and same pathway as MAPK signaling, Apoptosis, <em>Wnt</em> Signaling, ErbB signaling and Cell Cycle. Lastly, we identified 3 novel genes CHUK, INSR and CREBBP showing high connections with the <em>12</em> novel genes reported in literatures as well with the perturbed genes. As a result, these genes can be considered as significant finding in revealing the basis and pathways responsible for breast cancer.

Protocadherins are cell-cell adhesion molecules encoded by a large family of genes. Recent reports demonstrate recurrent silencing of protocadherin genes in tumors and provide strong arguments for their tumor supresor functionality. Loss of protocadherins may contribute to cancer development not only by altering cell-cell adhesion, that is a hallmark of cancer, but also by enhancing proliferation and epithelial mesenchymal transition of cells via deregulation of the <em>WNT</em> signaling pathway. In this study we have further corroborated our previous findings on the involvement of PCDH17 in laryngeal squamous cell carcinoma (LSCC). We used bisulfite pyrosequencing to analyze a cohort of primary LSCC tumors for alterations in PCDH17 promoter DNA methylation as an alternative gene inactivation mechanism to the homozygous deletions reported earlier. Moreover, we analyzed primary LSCC samples by immunohistochemistry for PCDH17 protein loss. We identified recurrent elevation of PCDH17 promoter DNA methylation in 32/81 (40%) primary tumors (P < 0.001) and therein hypermethylation of <em>12</em> (15%) cases in contrast to no tumor controls (n = 24) that were all unmethylated. Importantly, DNA demethylation by decitabine has restored low level PCDH17 expression in LSCC cell lines. In conclusion, we provide a mechanistic explanation of recurrently observed PCDH17 silencing in LSCC by demonstrating the role of promoter methylation in this process. In light of these findings and recent reports showing that PCDH17 methylation is detectable in serum of cancer patients we suggest that testing PCDH17 DNA methylation might serve as a potential biomarker in LSCC.

Although many studies have implicated the crosstalk between the <em>Wnt</em> and PKC signaling pathways in tumor initiation and progression, the molecular roles of PKC isoforms in the <em>Wnt</em> signaling pathway remain poorly understood. In this study, we explored the contribution of PKC isoforms to canonical and noncanonical <em>Wnt</em> signaling pathway in mediating cell migration and an epithelial-mesenchymal transition (EMT). When MCF-7 cells were treated with <em>12</em>-O-tetradecanoylphorbol-13-acetate (TPA) for up to 3 weeks, the effect of TPA on <em>Wnt</em> signaling pathway was dramatically different depending on the exposure time. The short term exposure (3 days) of MCF-7 cells to TPA exhibited significant induction of <em>Wnt</em>5a expression, along with the enhanced expression of PKC-α, to promote cell migration, which suggested that activation of noncanonical <em>Wnt</em> signaling pathway is associated with PKC-α. However, the chronic exposure (3 weeks) of cells to TPA completely suppressed <em>Wnt</em>5a expression and the expression of PKC-η and PKC-δ, whereas the expression of <em>Wnt</em>3a and PKC-θ were up-regulated to activate the canonical <em>Wnt</em> signaling pathway. Moreover, the loss of epithelial markers, including E-cadherin and GATA-3, suggested that chronic exposure of TPA stimulates EMT. Taken together, our data suggest that PKC-θ positively regulates the canonical <em>Wnt</em> signaling pathway, and that PKC-η and PKC-δ negatively modulate this signaling pathway.

OBJECTIVE

To investigate the role of Wnt/β-catenin signaling in aging of mesenchymal stem cells (MSCs) of rats.

METHODS

Serum samples were collected from young (8 ≈ 12 w) and aged (64 ≈ 72 w) SD rat. Four experiment groups were assigned: young rat serum (YRS), YRS+Wnt 3a, old rat serum (ORS) and ORS+DKK1 groups. Immunofluorescence and Western blotting were used to detect the expression of intracellular β-catenin. The senescence-associated changes were examined with SA-β-galactosidase staining. The proliferation ability was tested by MTT assays. The survived and apoptotic cells were determined by AO/EB staining. The expressions of γ-H2A. X and p53 protein were detected by immunofluorescence and Western blotting. RT-PCR was used to detect the expression of p53 and p21 mRNA.

RESULTS

Compared with the YRS group, the intracellular expression of β-catenin in the ORS group was significantly increased,especially in the nuclei of MSCs. After treatment of DKK1 in ORS, the γ-catenin expression was reduced. The number of SA-β-galactosidase positive MSCs was significantly higher in the YRS+Wnt 3a group than that in the YRS group (P<0.01), and the proliferative and survival ability of MSCs was significantly decreased in the YRS+Wnt 3a group. The number of SA-β-galactosidase positive MSCs in the ORS+DKK1 group was significantly decreased compared with that in ORS group (P <0.01), and the proliferative and survival ability of MSCs was significantly increased in the ORS+DKK1 group. The expression of γ-H2A.X, p53 and p21 was markedly increased in the ORS group than that in YRS group, however, after treatment with Wnt/β-catenin signaling inhibitor DKK1, the expression of γ-H2A.X, p53 and p21 was significantly decreased compared with that in the ORS group.

CONCLUSIONS

Results suggest that the Wnt/β-catenin signaling is activated in the MSCs cultured with ORS and excessive activation of Wnt/β-catenin signaling can promote MSCs aging. The DNA damage response and p53/p21 pathway may be main mediators of MSC aging induced by excessive activation of Wnt/β-catenin signaling.

Uterine fibroids are the most common gynecological tumors affecting women in their reproductive age. Despite this high incidence the pathogenesis of fibroids is widely unsolved. Whereas formerly only imbalances in hormonal levels were considered to account for tumor development, the identification of genetic changes likely to affect myometrial stem cell reservoirs provided a novel approach to fibroid genesis. Here, we identified a certain subset of cells by the surface marker CD24 with increased abundance in fibroids compared with myometrial tissue. Fibroid cells expressing CD24 shared certain features of immature or progenitor-like cells such as quiescence, reduced expression of smooth muscle differentiation markers and elevated expression of genes involved in the wingless-type (<em>WNT</em>)-pathway such as beta-catenin. In addition, a positive correlation between CD24 and wingless-type family member 4 (<em>WNT</em>4) expression was observed in uterine fibroids with mediator subcomplex <em>12</em> gene (MED<em>12</em>) mutations. Our findings suggest that cells highly expressing CD24 represent a type of immature smooth muscle progenitor cells. Their accumulation might be driven by disturbed differentiation processes caused by genetic changes possibly involving MED<em>12</em> mutations or high mobility group AT-hook (HMGA)2 rearrangements.

The lymphoid enhancer-binding factor-1 (LEF1) belongs to a family of regulatory proteins that share homology with the high mobility group protein-1 (HMG1). The LEF1 gene is a mediator in the canonical <em>Wnt</em>-signalling pathway required for morphogenesis of early mammary gland during embryogenesis. Here we describe the molecular characterisation of the porcine LEF1 gene and its association with number of teats and inverted teats in experimental and commercial populations. The 2357-bp cDNA sequence contains an 1197-bp open reading frame encoding a protein of 398 amino acids. The porcine LEF1 protein shares high identity with LEF1 in other mammalian species. The LEF1 gene contains <em>12</em> exons and maps to pig chromosome 8 (SSC8). We identified two single nucleotide polymorphisms (SNPs), a T1351C transition and an A1666C transversion, in the 3' end of LEF1. Associations of the SNP A1666C with presence of inverted teats (P<or= 0.01), total number of teats (P <or= 0.01) and total number of inverted teats (P <or= 0.01) were highly significant; SNP T1351C showed near significance with total number of inverted teats (P <or= 0.1) in the experimental DUMI population. SNP T1351C was significantly associated with total number of inverted teats (P= 0.04) and close to significance with affected teats (P = 0.06) in commercial populations. Haplotype analysis confirmed the tendency towards association with affected teats (P = 0.06) in the experimental DUMI population. The function, position, and associations shown here promote LEF1 as a candidate gene for number of teats and in particular for presence and number of inverted teats.

This study investigated whether various dietary fats affected the <em>Wnt</em> signaling pathway of preneoplastic lesions of colon mucosa in acrylamide (ACR)-treated rats. Male Sprague-Dawley rats were given intraperitoneal injections of ACR at a dose of 5 mg/kgbw and diets supplemented with 10% corn, olive, beef, or fish oil for 8 weeks; and then rats were still fed with diets supplemented with 10% oil for other 40 weeks. Aberrant crypt foci (ACF) were examined at <em>12</em> weeks post-ACR-exposure. At 48 weeks, normal appearing colon mucosal proliferation and apoptosis were evaluated by 5-bromo-2'-deoxyuridine (BrdU) incorporation and percentages of fragmented DNA, respectively; the expressions of beta-catenin, cyclin D1, <em>Wnt</em>2, <em>Wnt</em>3, and <em>Wnt</em>5a of normal appearing colon mucosa were analyzed by Western blot analysis. Results from this study showed that long-term exposure of rats to dietary corn oil and beef tallow enhanced ACF formation in ACR rats. In contrast, olive and fish oil weakened the ACF formation. Dietary corn oil and beef tallow increased BrdU incorporation, expression of cytosolic beta-catenin and cyclin D1; and decreased apoptosis in the colon mucosa of ACR rats. ACR rats fed beef tallow showed increased expressions of <em>Wnt</em>2 and <em>Wnt</em>3. ACR rats fed corn oil showed increased expressions of <em>Wnt</em>5a. These findings suggest that long-term high intake of corn oil and beef tallow enhanced, whereas olive and fish oil weakened cell proliferation through <em>Wnt</em> signaling, which might contribute to promoting effects in preneoplastic lesions of colon mucosa.

Inhibitors of the enzymatic activity of alanyl-aminopeptidases severely affect growth and typical functions of human peripheral T cells both in vitro and in vivo. The most prominent changes observed include the activation of cellular signal transduction pathways such as MAP kinases Erk1/2 or the <em>Wnt</em>-pathway, a decrease of production and release of "pro-inflammatory" cytokines (IL-2, IL-<em>12</em>) and, most importantly, an induction of expression and release of the immunosuppressive cytokine, TGF-beta1. Similar effects on T cell proliferation and function have been observed in response to inhibition of DPIV, which is strongly suggestive of a functional synergism of APN and DPIV. In support of this hypothesis evidence is provided showing that the simultaneous application of inhibitors of DPIV and APN further enhances the anti-inflammatory and immunosuppressive effects provoked by the inhibition of APN or DPIV alone. Therefore, the simultaneous inhibition of these enzymes represents a promising strategy for the pharmacological therapy of T cell mediated diseases such as autoimmune disease, inflammation, allergy, and allograft rejection.

The aim of the present study was to examine the molecular factors associated with the prognosis of colon cancer. Gene expression datasets were downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus databases to screen differentially expressed genes (DEGs) between colon cancer samples and normal samples. Survival‑related genes were selected from the DEGs using the Cox regression method. A co‑expression network of survival‑related genes was then constructed, and functional clusters were extracted from this network. The significantly enriched functions and pathways of the genes in the network were identified. Using Bayesian discriminant analysis, a prognostic prediction system was established to distinguish the positive from negative prognostic samples. The discrimination efficacy of the system was validated in the GSE17538 dataset using Kaplan‑Meier survival analysis. A total of 636 and 1,892 DEGs between the colon cancer samples and normal samples were screened from the TCGA and GSE44861 dataset, respectively. There were 155 survival‑related genes selected. The co‑expression network of survival‑related genes included 138 genes, 534 lines (connections) and five functional clusters, including the signaling pathway, cellular response to cAMP, and immune system process functional clusters. The molecular function, cellular components and biological processes were the significantly enriched functions. The peroxisome proliferator‑activated receptor signaling pathway, <em>Wnt</em> signaling pathway, B cell receptor signaling pathway, and cytokine‑cytokine receptor interactions were the significant pathways. A prognostic prediction system based on a 65‑gene signature was established using this co‑expression network. Its discriminatory effect was validated in the TCGA dataset (P=3.56e‑<em>12</em>) and the GSE17538 dataset (P=1.67e‑6). The 65‑gene signature included kallikrein‑related peptidase 6 (KLK6), collagen type XI α1 (COL11A1), cartilage oligomeric matrix protein, wingless‑type MMTV integration site family member 2 (WNT2) and keratin 6B. In conclusion, a 65‑gene signature was screened in the present study, which showed a prognostic prediction effect in colon adenocarcinoma. KLK6, COL11A1, and WNT2 may be suitable prognostic predictors for colon adenocarcinoma.

Induction of functional CTLs is one of the major goals for vaccine development and cancer therapy. Inflammatory cytokines are critical for memory CTL generation. <em>Wnt</em> signaling is important for CTL priming and memory formation, but its role in cytokine-driven memory CTL programming is unclear. We found that <em>wnt</em> signaling inhibited IL-<em>12</em>-driven CTL activation and memory programming. This impaired memory CTL programming was attributed to up-regulation of eomes and down-regulation of T-bet. <em>Wnt</em> signaling suppressed the mTOR pathway during CTL activation, which was different to its effects on other cell types. Interestingly, the impaired memory CTL programming by <em>wnt</em> was partially rescued by mTOR inhibitor rapamycin. In conclusion, we found that crosstalk between <em>wnt</em> and the IL-<em>12</em> signaling inhibits T-bet and mTOR pathways and impairs memory programming which can be recovered in part by rapamycin. In addition, direct inhibition of <em>wnt</em> signaling during CTL activation does not affect CTL memory programming. Therefore, <em>wnt</em> signaling may serve as a new tool for CTL manipulation in autoimmune diseases and immune therapy for certain cancers.

Treatment of postmenopausal osteoporosis with teriparatide parathyroid hormone amino terminal 1-34 increases bone formation and improves bone microarchitecture. A possible modulator of action is periostin. In vitro experiments have shown that periostin might regulate osteoblast differentiation and bone formation through Wnt signaling. The effect of teriparatide on periostin is not currently known.

To determine the effect of teriparatide treatment on circulating levels of periostin and other regulators of bone formation and investigate how changes in periostin relate to changes in bone turnover markers, regulators of bone formation, and bone mineral density (BMD).

Twenty women with osteoporosis; a 2-year open-label single-arm study.

Teriparatide 20 µg was administered by subcutaneous injection daily for 104 weeks. Periostin, sclerostin, and Dickkopf-related protein 1, procollagen type I N-terminal propeptide (PINP), and C-telopeptide of type I collagen were measured in fasting serum collected at baseline (two visits) and then at weeks 1, 2, 4, 12, 26, 52, 78, and 104. BMD was measured at the lumbar spine, total hip, and femoral neck using dual energy x-ray absorptiometry.

Periostin levels increased by 6.6% [95% confidence interval (CI), -0.4 to 13.5] after 26 weeks of teriparatide treatment and significantly by 12.5% (95% CI, 3.3 to 21.0; P < 0.01) after 52 weeks. The change in periostin correlated positively with the change in the lumbar spine BMD at week 52 (r = 0.567; 95% CI, 0.137 to 0.817; P < 0.05) and femoral neck BMD at week 104 (r = 0.682; 95% CI, 0.261 to 0.885; P < 0.01).

Teriparatide therapy increases periostin secretion; it is unclear whether this increase mediates the effect of the drug on bone.

Human amniotic epithelial cells (hAECs) have recently been recognized as a potential source of stem cells. The present study was designed to investigate the effects of mechanical stretch on the osteogenic differentiation of hAECs. As it has been previously reported that the physical environment is an important factor in maintaining the phenotype and functionality of differentiated cells, mechanical stretch was use to mimic the mechanical environment in the present study, with the following parameters: 5% elongation of the hAECs at a frequency of 0.5 Hz, with evaluation at 2, 6, <em>12</em> and 24 h time points. The osteogenic differentiation process of the hAECs followed by mechanical stimulation was evaluated by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR), western blotting and immunocytochemistry. Additionally, in a parallel study, a runt‑related transcription factor 2 (Runx2)/core binding factor α 1 (Cbfa1)‑specific short hairpin RNA (shRNA) plasmid vector and a scrambled shRNA control plasmid was constructed for transfection into the hAECs prior to mechanical stimulation. The cultured hAECs exhibited a cobblestone‑shaped epithelial‑like phenotype and were positive for stage‑specific embryonic antigen‑4, cytokeratin‑19, cluster of differentiation 44 and octamer‑binding protein 4, as detected by flow cytometry, western blotting or confocal microscopy. The qPCR and western blotting data demonstrated that the mRNA and protein expression levels of Runx2/Cbfa1, alkaline phosphatase and osteocalcin were upregulated compared with the control group following stretching and they peaked at <em>12</em> h. These results indicated that the osteogenic differentiation of the hAECs was induced by mechanical stimuli. Additionally, the mRNA and protein expression levels of β‑catenin and cyclin D were increased significantly following stretching; however, they were decreased following Runx2/Cbfa1‑shRNA transfection as observed by RT‑qPCR and western blotting. These results suggested that the <em>Wnt</em>/β‑catenin pathway may have an important role in mechanical stretch‑induced osteogenic differentiation of the hAECs. Furthermore, the combination of stretch and osteogenic induction medium had synergistic effects on the osteogenic differentiation. The results of the present study demonstrated that mechanical stimuli have an important role in osteogenic differentiation of hAECs via the <em>Wnt</em>/β‑catenin signalling pathway, which may be a potential therapeutic strategy in bone regenerative medicine.

The very-low-density lipoprotein receptor (VLDLR) negatively regulates <em>Wnt</em> signaling. VLDLR has two major alternative splice variants, VLDLRI and VLDLRII, but their biological significance and distinction are unknown. Here we found that most tissues expressed both VLDLRI and VLDLRII, while the retina expressed only VLDLRII. The shed soluble VLDLR extracellular domain (sVLDLR-N) was detected in the conditioned medium of retinal pigment epithelial cells, interphotoreceptor matrix, and mouse plasma, indicating that ectodomain shedding of VLDLR occurs endogenously. VLDLRII displayed a higher ectodomain shedding rate and a more potent inhibitory effect on <em>Wnt</em> signaling than VLDLRI in vitro and in vivo O-glycosylation, which is present in VLDLRI but not VLDLRII, determined the differential ectodomain shedding rates. Moreover, the release of sVLDLR-N was inhibited by a metalloproteinase inhibitor, TAPI-1, while it was promoted by phorbol <em>12</em>-myristate 13-acetate (PMA). In addition, sVLDLR-N shedding was suppressed under hypoxia. Further, plasma levels of sVLDLR-N were reduced in both type 1 and type 2 diabetic mouse models. We concluded that VLDLRI and VLDLRII had differential roles in regulating <em>Wnt</em> signaling and that decreased plasma levels of sVLDLR-N may contribute to <em>Wnt</em> signaling activation in diabetic complications. Our study reveals a novel mechanism for intercellular regulation of <em>Wnt</em> signaling through VLDLR ectodomain shedding.

Endometriosis is an estrogen-dependent disease. The impaired estrogen and progesterone signaling over-activates the <em>Wnt</em>/β-catenin pathway in endometriosis patients, which can explain the increased invasion potency of endometrial cells derived from the endometrium of women with endometriosis. The regulatory effects of vitamin D on <em>Wnt</em>/β-catenin pathway were demonstrated by previous studies. According to gene prioritization method, among <em>Wnt</em> target genes, CD44 was in high ranking in relation to endometriosis. The aim of this study is to investigate the expression of CD44 in the endometrium of women with endometriosis and to study the effects of vitamin D on its expression. This prospective study was performed, during a <em>12</em> months period from December 2015 to November 2016, on healthy women as the control group (n = 14) and endometriosis patients (n = 34). The endometriosis patients randomly divided into two groups: One group treated according to the routine protocol and the other group, alongside the routine protocol, took 50,000 IU vitamin D weekly for <em>12</em>-14 weeks. Blood, endometrial fluid, and endometrial tissue samples were obtained from the control group and endometriosis groups before and after the intervention. We used in silico gene prioritization to study the relevance of CD44. The expression of CD44 was evaluated using the techniques of Western blot, real-time polymerase chain reaction, and ELISA. The eutopic endometrium of women with endometriosis in mid-secretory phase expressed significantly higher levels of CD44s, CD44V, and CD44v6. The concentration of soluble CD44 in the serum and endometrial fluid of endometriosis patients was higher than of healthy women. The expression level of CD44s, CD44V, and CD44v6 in the eutopic endometrium as well as the concentration of soluble CD44 in the endometrial fluid was decreased after modification of the circulating levels of 25(OH)D. It seems that the increased expression and extensive shedding of CD44 in eutopic endometrium play a role in the pathogenesis of endometriosis. Vitamin D can control and modify this process at least in part. We suggest more in vivo investigations on the therapeutic potency of vitamin D in endometriosis.

Recent studies have shown that microRNAs (miRNAs) are involved in the progression of colorectal cancer (CRC). The aim of this study is to identify a novel miRNA that especially relates to liver metastasis and to explore the underlying mechanism. Differentially expressed miRNAs were analyzed using microarray, in primary CRC tumors without metastasis (n=16), those with liver metastasis (n=<em>12</em>), and liver metastatic lesions (n=8). We found that miR-487b level decreased in liver metastatic lesions, and qRT-PCR confirmed the results in the validating cohort (n=134). Survival analysis indicated that high expression of miR-487b was associated with better prognosis. In vitro studies were also performed to investigate the functional significance of miR-487b in human CRC cell lines. miR-487b showed an inhibitory effect on cell proliferation and invasion of CRC cells. miR-487b downregulated KRAS and inhibited its downstream signal pathways, and the luciferase reporter assay revealed that miR-487b directly targeted LRP6, a receptor for <em>WNT</em>/β-catenin signaling. These findings showed that decrease in miR-487b was related with liver metastasis. Our data suggest a possibility that miR-487b may suppress metastasis of CRC progression through inhibition of KRAS.

OBJECTIVE

Periodontal tissue remodeling includes remodeling of alveolar bone, periodontal ligament, and cementum. Cementoblast plays a main role in repairing root resorption. Canonical Wnt/β-catenin signaling can promote the odontogenic differentiation in osteoblast. However, the mechanism on how the orthodontic force influences the function of cementoblast and the relationship between the canonical Wnt/β-catenin signaling and Runx2 of cementoblast are not yet known. The aim of this study is focus on this relationship.

METHODS

OCCM30 cementoblasts were subjected to mechanical strain by four-point bending system with tension stress for 0, 3, 6, and 12 h. They were pretreated with different concentrations of Dikkopf-1 (DKK1) for 48 h. Western blot analysis was performed to detect the β-catenin levels in the nucleus. Runx2 mRNA was observed by real-time quantitative polymerase chain reaction (RT-PCR). OCCM30 cementoblasts were then pretreated with 150 ng · mL(-1) DKK1 for 48 h and subjected to mechanical strain by FX4000T system with tension stress for 12 h. Western blot analysis was conducted to detect the β-catenin levels in the nucleus, and Runx2 mRNA was observed by RT-PCR.

RESULTS

OCCM30 cementoblasts had significantly higher Runx2 mRNA and β-catenin levels after being loaded with mechanical stress. The amount of Runx2 mRNA in OCCM30 cementoblasts was significantly decreased by DKK1. When OCCM30 cemento-blasts were pretreated with DKK1 without stress, their β-catenin level was significantly decreased by DKK1 and Wnt signaling was blocked. When they were not pretreated with stress, the β-catenin level with DKK1 was lower than that without DKK1. Without DKK1, the β-catenin level in OCCM30 cemento- blasts increased afterbeing loaded with mechanical stress. With DKK1, the β-catenin level in OCCM30 cementoblasts, which were loaded with mechanical stress, was higher than that without mechanical stress.

CONCLUSIONS

Cementoblasts had higher Runx2 mRNA expression under mechanical stress because of the Wnt/β-catenin signaling pathway effect.