Haematopoietic stem cell induction by somite-derived endothelial cells controlled by meox1.

Haematopoietic stem cells (HSCs) are self-renewing stem cells capable of replenishing all blood lineages. In all vertebrate embryos that have been studied, definitive HSCs are generated initially within the dorsal aorta (DA) of the embryonic vasculature by a series of poorly understood inductive events. Previous studies have identified that signalling relayed from adjacent somites coordinates HSC induction, but the nature of this signal has remained elusive. Here we reveal that somite specification of HSCs occurs via the deployment of a specific endothelial precursor population, which arises within a sub-compartment of the zebrafish somite that we have defined as the endotome. Endothelial cells of the endotome are specified within the nascent somite by the activity of the homeobox gene meox1. Specified endotomal cells consequently migrate and colonize the DA, where they induce HSC formation through the deployment of chemokine signalling activated in these cells during endotome formation. Loss of meox1 activity expands the endotome at the expense of a second somitic cell type, the muscle precursors of the dermomyotomal equivalent in zebrafish, the external cell layer. The resulting increase in endotome-derived cells that migrate to colonize the DA generates a dramatic increase in chemokine-dependent HSC induction. This study reveals the molecular basis for a novel somite lineage restriction mechanism and defines a new paradigm in induction of definitive HSCs.

The role of WNT10B in normal prostate gland development and prostate cancer.

BACKGROUND: WNT signaling is implicated in embryonic development, and in adult tissue homeostasis, while its deregulation is evident in disease. This study investigates the unique roles of canonical WNT10B in both normal prostate development and prostate cancer (PCa) progression. METHODS: Organ culture and rat ventral prostates (VPs) were used to study Wnt10b ontogeny and growth effect of WNT10B protein. PB-SV40 LTag rat VPs were utilized for Wnt expression polymerase chain reaction (PCR) array and immunohistochemistry. Human localized PCa tissue microarrays (TMAs) were investigated for differential WNT10B expression. Human RNA-seq data sets were queried for differential expression of WNT10B in metastatic and localized PCa. Knockdown of WNT10B in PC3 cells was utilized to study its effects on proliferation, stemness, epithelial to mesenchymal transition (EMT), and xenograft propagation. RESULTS: Wnt10b expression was highest at birth and rapidly declined in the postnatal rat VP. Exogenous WNT10B addition to culture developing VPs decreased growth suggesting an antiproliferative role. VPs from PB-SV40 LTag rats with localized PCa showed a 25-fold reduction in Wnt10b messenger RNA (mRNA) expession, confirmed at the protein level. Human PCa TMAs revealed elevated WNT10B protein in prostate intraepithelial neoplasia compared with normal prostates but reduced levels in localized PCa specimens. In contrast, RNA-seq data set of annotated human PCa metastasis found a significant increase in WNT10B mRNA expression compared with localized tumors suggesting stage-specific functions of WNT10B. Similarly, WNT10B mRNA levels were increased in metastatic cell lines PC3, PC3M, as well as in HuSLC, a PCa stem-like cell line, as compared with disease-free primary prostate epithelial cells. WNT10B knockdown in PC3 cells reduced expression of EMT genes, MMP9 and stemness genes NANOG and SOX2 and markedly reduced the stem cell-like side population. Furthermore, loss of WNT10B abrogated the ability of PC3 cells to propagate tumors via serial transplantation. CONCLUSIONS: Taken together, these results suggest a dual role for WNT10B in normal development and in PCa progression with opposing functions depending on disease stage. We propose that decreased WNT10B levels in localized cancer allow for a hyperproliferative state, whereas increased levels in advanced disease confer a stemness and malignant propensity which is mitigated by knocking down WNT10B levels. This raises the potential for WNT10B as a novel target for therapeutic intervention in metastatic PCa.

Expression Analysis of ACSL5 and Wnt2B in Human Congenital Pulmonary Airway Malformations.

BACKGROUND: The objective of this study was to determine acyl-CoA synthetase 5 (ACSL5) and Wnt2B expression patterns in human congenital pulmonary airway malformations (CPAMs) and to identify the possible roles of ACSL5 and Wnt2B in the pathogenesis of CPAM. METHODS: Expression of ACSL5 and Wnt2B was evaluated by immunohistochemical staining, Western blotting, and quantitative real-time polymerase chain reaction, which were performed on surgical specimens of CPAM and adjacent normal lung tissues as controls. RESULTS: Immunohistochemistry revealed that ACSL5 and Wnt2B immunopositive cells were predominantly detected in the mesenchymal cell nucleus, and there were lower expressions of ACSL5 and Wnt2B immunopositive cells in CPAM tissues than those in adjacent normal lung tissues. Western blotting and quantitative real-time polymerase chain reaction showed that ACSL5 and Wnt2B protein and mRNA expressions were significantly decreased in CPAM tissues as compared to the adjacent normal lung tissues (P < 0.05). In addition, there was a reduced level of ACSL5 relative to that of Wnt2B. CONCLUSIONS: The decreased ACSL5 and Wnt2B expressions correlated with aberrations in pulmonary development and in the pathogenesis of CPAM, so downregulation of ACSL5 and Wnt2B could play an important role in the development of bronchial-alveolar structures in CPAM.

Flow cytometer analysis of cell apoptosis of endometrial carcinoma with Wnt10b.

The aim of this study is to analyze the cell apoptosis of endometrial carcinoma (EC) with Wnt10b by Fluorescence Activated Cell Sorting (FACS) technology. AN3CA cell lines and Ishikawa-H-12 cell lines were taken as the in-vitro cell models to observe the influence of Wnt10b on key factors of Wnt signal pathway. Methyl thiazolyl tetrazolium (MTT) was applied for the detection of cell proliferation while FACS was used for the detection of cell apoptosis. Data were analyzed using statistical software SPSS14.0. After the overexpression of Wntl0b in AN3CA cells, the apoptosis rate dropped significantly compared with the two control groups (p < 0.05); while the apoptosis rate increased significantly compared with the control groups (p < 0.01) after Wntl0b knock-off in Ishikawa3-H-12 cells. In normal endometrium, Wnt10b gene expression was negative, while that in EC cells was positive. It can be concluded that Wnt10b gene can promote EC cell proliferation and inhibit its apoptosis.

Expression of Wnt3a, Wnt10b, ß-catenin and DKK1 in periodontium during orthodontic tooth movement in rats.

OBJECTIVE: To investigate the expression of Wnt3a, Wnt10b, ß-catenin and DKK1 in the periodontal ligament (PDL) during orthodontic tooth movement (OTM) in rats. MATERIALS AND METHODS: Nickel-titanium closed-coil springs were used to deliver an initial 50 g mesial force to the left maxillary first molars in 30 rats. The force was kept constant for 1, 3, 5, 7, 10 and 14 days until the animals were sacrificed. The right maxillary molars without force application served as control. Paraffin-embedded sections of the upper jaws were prepared for histological and immunohistochemical analyses to detect Wnt3a, Wnt10b, ß-catenin and DKK1 expression in PDL. RESULTS: Wnt3a, Wnt10b, ß-catenin and DKK1 were expressed on both the ipsilateral and contralateral sides of PDL in each group. After the application of orthodontic force, the expression of ß-catenin and DKK1 was initially increased and then decreased on both sides, with maximal levels of expression at day 7 and day 10, respectively. On the compression side, Wnt3a and Wnt10b levels started to increase at day 5, while on the tension side, these two molecules began to increase at day 1. Furthermore, the expression levels of Wnt3a, Wnt10b, and ß-catenin were much stronger on the tension side than on the compression side at any of the observation points, while DKK1 level was much higher on the compression side. CONCLUSION: Wnt3a, Wnt10b, ß-catenin and DKK1 expression may be related to the periodontal tissue remodeling following the application of an orthodontic force in rats. These observations suggest that the Wnt/ß-catenin signaling pathway may play a crucial role in periodontal tissue remodeling during OTM.

WNT5A modulates cell cycle progression and contributes to the chemoresistance in pancreatic cancer cells.

BACKGROUND: Although there are many studies on the mechanism of chemoresistance in cancers, studies on the relations between WNT5A and chemoresistance in pancreatic cancer are rare. The present study was to examine the role of WNT5A in the regulation of cell cycle progression and in chemo-resistance in pancreatic cancer tissues and cell lines. METHODS: Fresh pancreatic cancer and paracarcinoma tissues were obtained from 32 patients. The expressions of WNT5A, AKT/p-AKT and Cyclin D1 were detected by immunohistochemistry, and the correlation between WNT5A expression and clinicopathological characteristics was analyzed. The relationship between WNT5A expression and gemcitabine resistance was studied in PANC-1 and MIAPaCa2 cell lines. The effect of WNT5A on the regulation of cell cycle and gemcitabine cytotoxicity were investigated. The associations among the expressions of p-AKT, Cyclin D1 and WNT5A were also analyzed in cell lines and the effect of WNT5A on restriction-point (R-point) progression was evaluated. RESULTS: WNT5A, p-AKT and Cyclin D1 were highly expressed in pancreatic cancer tissues, and the WNT5A expression was correlated with the TNM stages. In vitro, WNT5A expression was associated with gemcitabine chemoresistance. The percentage of cells was increased in G0/G1 phase and decreased in S phase after knockdown of WNT5A in PANC-1. WNT5A promoted Cyclin D1 expression through phosphorylation of AKT which consequently enhanced G1-S transition and gemcitabine resistance. Furthermore, WNT5A enhanced the cell cycle progression toward R-point through regulation of retinoblastoma protein (pRb) and pRb-E2F complex formation. CONCLUSIONS: WNT5A induced chemoresistance by regulation of G1-S transition in pancreatic cancer cells. WNT5A might serve as a predictor of gemcitabine response and as a potential target for tumor chemotherapy.

Downregulation of Wnt causes root resorption.

INTRODUCTION: There are multiple causes of external root resorption, but absent a disease state, it is most often observed when excessive physical force is used during orthodontic treatment. Even without mechanical stimulation, however, root resorption can still occur. The purpose of this study was to test whether Wnt signaling plays a role in pathologic root resorption, by conditionally deleting Wntless (Wls) from odontoblasts and osteoblasts and then evaluating the phenotypic effects on the maintenance of the root surface. METHODS: Ten (age, 1 month) and 20 (age, 3 months) OCN-Cre;Wls(fl/fl) mice and their wild-type littermates were evaluated using microcomputed tomography, histology, and immunohistochemistry. Phenotypic alterations in the alveolar bone, dentin, and cementum were characterized and quantified. RESULTS: In a genetic model of reduced Wnt signaling, we found that RANKL expression is upregulated, and osteoprotegerin expression is downregulated. This molecular disruption results in an increase in osteoclast activity, a decrease in osteoblast activity, and extensive, spontaneous root resorption. A genetic strain of mice in which Wnt signaling is elevated exhibits thicker cementum, whereas, even in the perinatal period, OCN-Cre;Wls(fl/fl) mice exhibit thinner cementum. CONCLUSIONS: Taken together, these data demonstrate that Wnts regulate cementum homeostasis, and that idiopathic cases of root resorption might have as their etiology a reduction in endogenous Wnt signaling.

Regulator of g protein signaling 3 modulates wnt5b calcium dynamics and somite patterning.

Vertebrate development requires communication among cells of the embryo in order to define the body axis, and the Wnt-signaling network plays a key role in axis formation as well as in a vast array of other cellular processes. One arm of the Wnt-signaling network, the non-canonical Wnt pathway, mediates intracellular calcium release via activation of heterotrimeric G proteins. Regulator of G protein Signaling (RGS) proteins can accelerate inactivation of G proteins by acting as G protein GTPase-activating proteins (GAPs), however, the possible role of RGS proteins in non-canonical Wnt signaling and development is not known. Here, we identify rgs3 as having an overlapping expression pattern with wnt5b in zebrafish and reveal that individual knockdown of either rgs3 or wnt5b gene function produces similar somite patterning defects. Additionally, we describe endogenous calcium release dynamics in developing zebrafish somites and determine that both rgs3 and wnt5b function are required for appropriate frequency and amplitude of calcium release activity. Using rescue of gene knockdown and in vivo calcium imaging assays, we demonstrate that the activity of Rgs3 requires its ability to interact with Galpha subunits and function as a G protein GAP. Thus, Rgs3 function is necessary for appropriate frequency and amplitude of calcium release during somitogenesis and is downstream of Wnt5 activity. These results provide the first evidence for an essential developmental role of RGS proteins in modulating the duration of non-canonical Wnt signaling.

Effect of embryo vitrification on postnatal mouse uterine development.

OBJECTIVE: To determine whether the process of vitrification affects the development of the female Müllerian duct. STUDY DESIGN: We examined the difference in morphology and gene expression of the developing uteri of 30 female mice from transferred vitrified embryos and 30 female mice from transferred fresh embryos. RESULTS: By employing histology, the data showed that there were no significant differences between the 2 groups of the same age. With the use of reverse transcription polymerase chain reaction and Western blotting, the data showed that there was no significant change in the expression of Wnt genes (Wnt4, Wnt5a, Wnt7a), beta-catenin/TCF target genes, and homeobox A10 (HOXA10) gene during uterine development in the vitrified group as compared with the control group. CONCLUSION: These data suggest that vitrifying preimplantation embryos may have no effects on morphology and gene expression of the uterus of offspring.

Gastric epithelial stem cells.

Advances in our understanding of stem cells in the gastrointestinal tract include the identification of molecular markers of stem and early progenitor cells in the small intestine. Although gastric epithelial stem cells have been localized, little is known about their molecular biology. Recent reports describe the use of inducible Cre recombinase activity to indelibly label candidate stem cells and their progeny in the distal stomach, (ie, the antrum and pylorus). No such lineage labeling of epithelial stem cells has been reported in the gastric body (corpus). Among stem cells in the alimentary canal, those of the adult corpus are unique in that they lie close to the lumen and increase proliferation following loss of a single mature progeny lineage, the acid-secreting parietal cell. They are also unique in that they neither depend on Wnt signaling nor express the surface marker Lgr5. Because pathogenesis of gastric adenocarcinoma has been associated with abnormal patterns of gastric differentiation and with chronic tissue injury, there has been much research on the response of stomach epithelial stem cells to inflammation. Chronic inflammation, as induced by infection with Helicobacter pylori, affects differentiation and promotes metaplasias. Several studies have identified cellular and molecular mechanisms in spasmolytic polypeptide-expressing (pseudopyloric) metaplasia. Researchers have also begun to identify signaling pathways and events that take place during embryonic development that eventually establish the adult stem cells to maintain the specific features and functions of the stomach mucosa. We review the cytologic, molecular, functional, and developmental properties of gastric epithelial stem cells.

WNT7B in fibroblastic foci of idiopathic pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a devastating interstitial pneumonia causing a loss of respiratory surface area due to a proliferative fibrotic response involving hyperplastic, hypertrophic, and metaplastic epithelium, cystic honeycomb change, septal expansion, and variable inflammation. Wnt (wingless) signaling glycoproteins are known to be involved in lung development and tissue repair, and are up-regulated in patients with IPF. Based on previous qRT-PCR data showing increased Wnt7B in lungs of IPF patients, a systematic, quantitative examination of its tissue site distribution was undertaken. METHODS: Tissue samples from the Lung Tissue Research Consortium (LTRC) of 39 patients diagnosed with mild to severe IPF/usual interstitial pneumonia (UIP) and 19 normal patients were examined for the immunolocalization of Wnt7B. RESULTS: In normal lung, moderate Wnt7B reactivity was confined to airway epithelium, smooth muscle of airways and vasculature, and macrophages. IPF lung showed strong Wnt7B reactivity in fibroblastic foci, dysplastic airway and alveolar epithelium, and in highly discrete subepithelial, basement membrane-associated regions. All reactive sites were sized and counted relative to specific microscopic regions. Those in the subepithelial sites were found in significantly greater numbers and larger relative area compared with the others. No reactive sites were present in normal patient controls. CONCLUSIONS: The results demonstrate Wnt7B to be expressed at high concentrations in regions of active hyperplasia, metaplasia, and fibrotic change in IPF patients. In this context and its previously established biologic activities, Wnt7B would be expected to be of potential importance in the pathogenesis of IPF.

Wnt/beta-catenin signaling is required for CNS, but not non-CNS, angiogenesis.

Despite the importance of CNS blood vessels, the molecular mechanisms that regulate CNS angiogenesis and blood-brain barrier (BBB) formation are largely unknown. Here we analyze the role of Wnt/beta-catenin signaling in regulating the formation of CNS blood vessels. First, through the analysis of TOP-Gal Wnt reporter mice, we identify that canonical Wnt/beta-catenin signaling is specifically activated in CNS, but not non-CNS, blood vessels during development. This activation correlates with the expression of different Wnt ligands by neural progenitor cells in distinct locations throughout the CNS, including Wnt7a and Wnt7b in ventral regions and Wnt1, Wnt3, Wnt3a, and Wnt4 in dorsal regions. Blockade of Wnt/beta-catenin signaling in vivo specifically disrupts CNS, but not non-CNS, angiogenesis. These defects include reduction in vessel number, loss of capillary beds, and the formation of hemorrhagic vascular malformations that remain adherent to the meninges. Furthermore, we demonstrate that Wnt/beta-catenin signaling regulates the expression of the BBB-specific glucose transporter glut-1. Taken together these experiments reveal an essential role for Wnt/beta-catenin signaling in driving CNS-specific angiogenesis and provide molecular evidence that angiogenesis and BBB formation are in part linked.

Wnt-5a signaling is correlated with infiltrative activity in human glioma by inducing cellular migration and MMP-2.

Wnts are secreted ligands that consist of 19 members in humans, regulate cell proliferation, differentiation, motility and fate in many stages including the embryonic stage and tumorigenesis. Wnts bind to cell surface receptors named Frizzleds and LRPs, and transduce their signals through ß-catenin-dependent and -independent intracellular pathways. Gliomas are one of the most common intracranial tumors. Gliomas exhibit a progression associated with widespread infiltration into surrounding neuronal tissues. However, the molecular mechanisms that stimulate the invasion of glioma cells are not fully understood. We established two cell lines from human glioma cases and analyzed the expression of all Wnt and Frizzled members in these cell lines and other well-known glioma cell lines by real-time PCR study. The mRNA of Wnt-5a and -7b and Frizzled-2, -6 and -7 were overexpressed in glioma cells. The elevation of Wnt-5a expression was most remarkable. Although Wnt-5a is reported to have oncogenic and antioncogenic activity in several cancers, the role of Wnt-5a signaling in human glioma cells remains unclear. Immunohistochemical study also revealed high expression of Wnt-5a in 26 (79%) of 33 human glioma cases. The positivity of Wnt-5a expression was correlated with the clinical grade. Knockdown of Wnt-5a expression suppressed migration, invasion and expression of matrix metalloproteinase-2 of glioma cells. Reciprocally, treatment with purified Wnt-5a ligand resulted in stimulation of cell migration and invasion. MMP-2 inhibitor suppressed the Wnt-5a-dependent invasion of U251 cells. These results suggested that Wnt-5a is not only a prognostic factor but also a therapeutic target molecule in gliomas for preventing tumor cell infiltration.

[Expression of Wnt5a in the terminal rectum of children with anorectal malformation].

OBJECTIVE: To study the expression of Wnt5a protein in the terminal rectum of children with anorectal malformation (ARM) and the possible association between Wnt5a and ARM. METHODS: Specimens were obtained from 20 children with ARM, 7 children with acquired rectovestibular fistula and 6 children with non-gastrointestinal tract disease (control group). The expression of Wnt5a protein in the terminal rectum was determined by immunohistochemistry and Western blot. RESULTS: Wnt5a was mainly expressed in the rectum of the myenteric nerve plexus, mucosal layer and submucosa in the control group. Compared with the control group, Wnt5a expression in the terminal rectum decreased significantly in the ARM group, and decreased more significantly in children with high ARM. The results of Western blot showed the expression of Wnt5a protein in the high, intermediate and low ARM groups were significantly lower than that in the acquired rectovestibular fistula and the control groups (P<0.01). The expression of Wnt5a protein in the high and the intermediate ARM groups were also lower than that in the low ARM group (P<0.01). There was no significant difference in the Wnt5a protein expression between the acquired rectovestibular fistula and the control groups. CONCLUSIONS: The expression of Wnt5a in the termina1 rectum decreases in children with ARM, suggesting Wnt5a may play an important role in the development of ARM.

Quantitative analyses reveal extracellular dynamics of Wnt ligands in Xenopus embryos.

The mechanism of intercellular transport of Wnt ligands is still a matter of debate. To better understand this issue, we examined the distribution and dynamics of Wnt8 in Xenopus embryos. While Venus-tagged Wnt8 was found on the surfaces of cells close to Wnt-producing cells, we also detected its dispersal over distances of 15 cell diameters. A combination of fluorescence correlation spectroscopy and quantitative imaging suggested that only a small proportion of Wnt8 ligands diffuses freely, whereas most Wnt8 molecules are bound to cell surfaces. Fluorescence decay after photoconversion showed that Wnt8 ligands bound on cell surfaces decrease exponentially, suggesting a dynamic exchange of bound forms of Wnt ligands. Mathematical modeling based on this exchange recapitulates a graded distribution of bound, but not free, Wnt ligands. Based on these results, we propose that Wnt distribution in tissues is controlled by a dynamic exchange of its abundant bound and rare free populations.

Wnt Family Member 9b (Wnt9b) Is a New Sensitive and Specific Marker for Breast Cancer.

Confirming the tumor origin is often a diagnostic challenge in pathology and carries significant therapeutic impacts. Cytokeratin 7, estrogen receptor, and GATA binding protein 3 (GATA3) are well-established diagnostic markers frequently used to support a tumor's breast origin. However, their specificities still have room to improve. Many nonbreast tumors express cytokeratin 7 and estrogen receptor, and urothelial tumors frequently express GATA3. There is a practical need for a new breast lineage marker that is sensitive and specific. Wnt family member proteins play critical roles in embryo development, tissue homeostasis and tumor development through ß-catenin dependent and independent pathways. The current study evaluated Wnt9b and GATA3 expression in 163 primary breast cancers, 63 metastatic breast cancers, and 525 nonbreast epithelial tumors. The positive rates of Wnt9b and GATA3 in primary breast cancer were both 98.7%. The positive rates in metastatic breast cancer were 87.3% for Wnt9b and 96.8% for GATA3. For nonbreast tumors, including 64 cases of urothelial carcinoma, Wnt9b was negative in all except salivary gland carcinomas. The study demonstrated that Wnt9b is a breast cancer marker with similar sensitivity as GATA3 but with greater specificity than GATA3 and may ultimately become a useful diagnostic tool in routine surgical pathology practice.

Chewing tobacco may act as a risk factor for dysplastic transformation of squamous cells in Oral leukoplakia- A cytochemistry based approach.

The use of chewing tobacco is a severe risk factor for oral mucosa related diseases including cancer in India as well as USA, although its relationship with Oral Leukoplakia (OL) or related carcinogenicity is still not clear. This work chose two oncogenic pathway proteins- the Epidermal Growth Factor Receptor and the WNT pathway among leukoplakia patients and established their correlation with the individuals' tobacco chewing habit. 89 fresh patients with OL were selected for the work. The samples were classified based on the individual's tobacco chewing habit. The divided samples were then immunostained with antibodies for both of the EGFR as well as WNT pathway proteins. The samples were further classified based on their proliferation status and the expression of these oncoproteins was also observed. In order to compare the cytological data with histological data, 30 OL patients undergoing biopsy were chosen and immunohistological analysis was performed for the same pathways. Results showed overexpressing EGFR and WNT pathway proteins in all OL samples. Structurally atypic cells had a tendency to overexpress these oncoproteins. However the immunocytochemistry data could not confirm any positive effect of chewing tobacco on the OL's proliferative state. Statistical data from the immunfluorescence finally revealed the overexpression of both EGFR and WNT pathway proteins on the proliferative population establishing chewing tobacco as a positive risk factor for the onset of OL. Data from biopsy samples followed the same trend of protein expression seen in the cytological samples. Dysplastic zones showed huge overexpression of EGFR and WNT pathway proteins among tobacco chewers. In conclusion, this is the first time report showing the effect of chewing tobacco on the EGFR and WNT pathway in OL and its possible role as a potential risk factor for its proliferative type.

ß-catenin/Wnt signalling pathway in fibromatosis, metaplastic carcinomas and phyllodes tumours of the breast.

Wnt signalling pathway is known to have a critical role in carcinogenesis and in epithelial-to-mesenchymal transition. Upon Wnt activation, ß-catenin is translocated from the membrane to the cytoplasm and nucleus, where it interacts with transcriptional activators. It has been suggested that various spindle cell lesions of the breast may harbour Wnt pathway activation. Given that ß-catenin nuclear localization constitutes a good surrogate marker of Wnt canonical pathway activation, we have investigated the distribution of ß-catenin in spindle cell lesions of the breast and whether it could be employed in the differential diagnosis of these lesions. A total of 52 metaplastic breast carcinomas, eight fibromatoses and 23 phyllodes tumours were retrieved from our institutions' archives. We performed immunohistochemistry using two anti-ß-catenin antibodies. In all, three fibromatoses and 21 metaplastic breast carcinomas were subjected to CTNNB1 (ß-catenin encoding gene) mutation analysis by direct gene sequencing. A good correlation between the two antibodies was observed (Spearman's r>0.82, P<0.001). All fibromatoses and 23% of metaplastic breast carcinomas expressed nuclear ß-catenin. In fibromatosis, ß-catenin was more often diffusely expressed, whereas in metaplastic breast carcinomas, expression was more frequently focal. Membranous ß-catenin expression was significantly lower in spindle cell carcinomas than in other subtypes of metaplastic breast carcinomas. In phyllodes tumours, stromal cells of benign and malignant subtypes displayed nuclear ß-catenin expression in 94 and 57% of cases, respectively. No CTNNB1 mutation was identified in any of the 21 metaplastic carcinomas analysed, whereas the mutations 45S>S/P and 41T>T/A were found in samples of fibromatosis. In conclusion, ß-catenin nuclear expression is a common feature in fibromatoses and in the stromal component of phyllodes tumours, but may also be observed in metaplastic breast carcinomas. ß-catenin nuclear expression should not be used as a single marker to differentiate fibromatosis from other spindle cell tumours of the breast.

Evidence for in vivo nicotine-induced alveolar interstitial fibroblast-to-myofibroblast transdifferentiation.

Nicotine exposure alters normal homeostatic pulmonary epithelial-mesenchymal paracrine signaling pathways, resulting in alveolar interstitial fibroblast (AIF)-to-myofibroblast (MYF) transdifferentiation. Though this has been described under in vitro conditions, it is not known if the same phenomenon also takes place in vivo. A well-established rodent model of lung damage following perinatal nicotine exposure was used. By probing for the well-established markers of fibroblast differentiation (parathyroid hormone-related protein [PTHrP], peroxisome proliferator-activated receptor gamma [PPARgamma], adipocyte differentiation-related protein, alpha-smooth muscle actin, and fibronectin) at the mRNA, protein, and tissue levels, the authors provide the first in vivo evidence for nicotine-induced AIF-to-MYF transdifferentiation. In addition, these data also provide the first evidence for nicotine-induced up-regulation of Wnt signaling, accompanying the down-regulation of PTHrP/PPARgamma signaling in vivo following nicotine exposure during pregnancy. These data provide an integrated mechanism for in utero nicotine-induced lung damage and how it could permanently alter the "developmental program" of the developing lung by disrupting critically important epithelial-mesenchymal interactions. More importantly, these data are likely to provide specific interventions to augment the pulmonary mesenchymal lipogenic pathway to ameliorate nicotine-induced in utero lung injury.

Prognostic role of Wnt7a expression in ovarian carcinoma patients.

Wnt7a is a secreted glycoprotein that regulates normal cellular proliferation and differentiation as well as tumorigenesis and progression. The aim of this study was to investigate the expression and prognostic significance of Wnt7a in ovarian carcinoma. Wnt7a expression was immunohistochemically examined in normal ovaries (n=15), benign tumors (n=50) and ovarian carcinomas (n=78). The correlation of Wnt7a expression with clinicopathological parameters and survival was evaluated. Wnt7a expression was higher in ovarian carcinomas compared to normal ovaries and benign tumors (p<0.001 and p=0.001, respectively). Wnt7a positive expression was significantly correlated with serous subtype (p<0.001), elder age (p=0.017), advanced stage (p<0.001), high grade (p=0.001), a high degree of ascitic fluid volume (p=0.015) and high CA125 expression (p=0.025). Wnt7a was found to be a significant prognostic factor in univariate and multivariate analysis. High Wnt7a expression in ovarian cancer may be associated with poor prognosis.