The WNT5A/ROR2 signaling pathway in pancreatic ductal adenocarcinoma (PDAC).

PURPOSE: WNT5A/ROR2 signaling pathway has been involved in many human cancers. Its role in pancreatic ductal adenocarcinoma (PDAC) has not been clarified yet. The purpose of this study was to determine the prognostic value of WNT5A expression in conjunction with the ROR2 expression in the same PDAC human tissues. METHODS: We retrospectively analyzed by immunohistochemistry the WNT5A and ROR2 expression in117 paraffin-embedded PDAC specimens following surgical pancreatic resection. The prognostic value of WNT5A and ROR2 was assessed using Kaplan-Meier survival curves and multivariate Cox regression models. RESULTS: High ROR2 expression was detected in 65.8% (77/117) of PDAC tumors, in 28.2% (33/117) in tumor-stroma, and in 71.1% (65/90) of normal pancreatic tissue. High WNT5A expression was found in 76.9% (90/117) of tumors, in 59.0% (69/117) of tumor-stroma, and in 83.0% (73/88) of normal pancreatic tissue. Spearman's correlation coefficiency demonstrated weak association between ROR2 and WNT5A expression in tumor (r=0.184; p=0.047), and no association in stroma (r=0.036; p=0.699). Multivariate analysis showed that regional lymph node invasion and differentiation were independent prognostic factors of survival, while ROR2- and WNT5A expression were not. CONCLUSIONS: Variable expression patterns for ROR2 and WNT5A were demonstrated in PDAC and normal pancreatic tissues suggesting a role for WNT5A/ROR2 signalling pathway, not only in PDAC but also in the normal pancreatic tissue during inflammation. The lack of prognostic significance for ROR2 and WNT5A expression in our cohort, either alone or in subgroup analysis, underlines the complexity of their role in PDAC, which is highly dependent on the different molecular receptor-ligand tissue contexts.

The Emerging Role of Sfrp5 and Wnt5a in the Pathogenesis of Obesity: Implications for a Healthy Diet and Lifestyle.

In recent decades, the prevalence of obesity has risen dramatically worldwide among all age groups. Obesity is characterized by excess fat accumulation and chronic low-grade inflammation. The adipose tissue functions as a metabolically active endocrine organ secreting adipokines. A novel duo of adipokines, the anti-inflammatory secreted frizzled-related protein 5 (Sfrp5) and the proinflammatory wingless type mouse mammary tumor virus (MMTV) integration site family member 5A (Wnt5a), signal via the non-canonical Wnt pathway. Recent evidence suggests that Sfpr5 and Wnt5a play a key role in the pathogenesis of obesity and its metabolic complications. This review summarizes the current knowledge on the novel regulatory system of anti-inflammatory Sfrp5 and pro-inflammatory Wnt5a, and their relation to obesity and obesity-related complications. Future studies are required to investigate the potential role of Sfrp5 and Wnt5a as biomarkers for monitoring the response to lifestyle interventions and for predicting the development of cardiometabolic risk factors. These adipokines may also serve as novel therapeutic targets for obesity-related disorders.

Wnt5a-induced M2 polarization of tumor-associated macrophages via IL-10 promotes colorectal cancer progression.

BACKGROUND: Tumor-associated macrophages (TAMs) in the tumor microenvironment influence tumor initiation, invasion and metastasis. Several studies have shown that Wnt5a is mainly expressed in the tumor stroma, especially in TAMs. However, whether Wnt5a regulates the polarization and biological function of TAMs in colorectal cancer (CRC) is incompletely understood. METHODS: Immunofluorescence staining was performed to detect CD68 and Wnt5a expression in colorectal tissues from patients (63 CRC specimens VS 20 normal tissues). RT-qPCR, flow cytometry, ELISA and inhibitors were carried out to explore the role of Wnt5a in the polarization of TAMs. Clone formation and transwell assays were performed to determine the effects of Wnt5a-treated macrophages on tumor proliferation, migration and invasion in vitro. Finally, a xenograft model was applied to confirm the effects of Wnt5a+ TAMs on CRC tumorigenesis. RESULTS: We found that high Wnt5a+CD68+/CD68+ TAMs ratio was significantly associated with poor prognosis in CRC patients and Wnt5a+ TAM was an M2-like TAM subtype. Subsequently, we found that Wnt5a induced macrophages to secrete IL-10, which then acted as an autocrine cytokine to induce M2 polarization of these macrophages. IL-10 neutralizing antibody completely reversed the pro-M2 effect of Wnt5a. Mechanistically, the CaKMII-ERK1/2-STAT3 pathway was required for Wnt5a-mediated IL-10 expression in macrophages. Furthermore, Wnt5a-induced M2 macrophages promoted CRC cells proliferation, migration and invasion; knockdown of Wnt5a in TAMs significantly impaired the pro-tumor functions of TAMs. CONCLUSIONS: Our data indicate that Wnt5a could induce M2 polarization of TAMs by regulating CaKMII-ERK1/2-STAT3 pathway-mediated IL-10 secretion, ultimately promoting tumor growth and metastasis of CRC.

LncRNA LINP1 regulates acute myeloid leukemia progression via HNF4α/AMPK/WNT5A signaling pathway.

LncRNAs play critical roles in various pathophysiological and biological processes, such as protein translation, RNA splicing, and epigenetic modification. Indeed, abundant evidences demonstrated that lncRNA act as competing endogenous RNAs (ceRNAs) to participate in tumorigenesis. However, little is known about the underlying function of lncRNA in nonhomologous end joining (NHEJ) pathway 1 (LINP1) in pediatric and adolescent acute myeloid leukemia (AML). The expression of LINP1 was examined in AML patient samples by qRT-PCR. Cell proliferation was examined by CCK-8 and Edu assays. ß-Galactosidase senescence assay, mGlucose uptake assay, lactate production assay, and Gene Ontology (GO) analysis were performed for functional analysis. We found that LINP1 was significantly overexpressed in AML patients at diagnosis, whereas downregulated after complete remission (CR). Furthermore, knockdown of LINP1 expression remarkably suppressed glucose uptake and AML cell maintenance. Mechanistically, LINP1 was found to inhibit the glucose metabolism by suppressing the expression of HNF4a. Both LINP1 and HNF4a knockdown reduced the expression levels of AMPK phosphorylation and WNT5A, indicating for the first time that LINP1 strengthened the HNF4a-AMPK/WNT5A signaling pathway involved in cell glucose metabolism modulation and AML cell survival. Taken together, our results indicated that LINP1 promotes the malignant phenotype of AML cells and stimulates glucose metabolism, which can be regarded as a potential prognostic marker and therapeutic target for AML.

Oscillatory cortical forces promote three dimensional cell intercalations that shape the murine mandibular arch.

Multiple vertebrate embryonic structures such as organ primordia are composed of confluent cells. Although mechanisms that shape tissue sheets are increasingly understood, those which shape a volume of cells remain obscure. Here we show that 3D mesenchymal cell intercalations are essential to shape the mandibular arch of the mouse embryo. Using a genetically encoded vinculin tension sensor that we knock-in to the mouse genome, we show that cortical force oscillations promote these intercalations. Genetic loss- and gain-of-function approaches show that Wnt5a functions as a spatial cue to coordinate cell polarity and cytoskeletal oscillation. These processes diminish tissue rigidity and help cells to overcome the energy barrier to intercalation. YAP/TAZ and PIEZO1 serve as downstream effectors of Wnt5a-mediated actomyosin polarity and cytosolic calcium transients that orient and drive mesenchymal cell intercalations. These findings advance our understanding of how developmental pathways regulate biophysical properties and forces to shape a solid organ primordium.

Wnt5a contributes to dectin-1 and LOX-1 induced host inflammatory response signature in Aspergillus fumigatus keratitis.

Fungal keratitis causes devastating corneal ulcers which can result in significant visual impairment and even blindness. As a ligand that activates the non-canonical Wnt signaling pathways, Wnt5a triggers the production of important inflammatory chemokines and the chemotactic migration of neutrophils. In this study we aimed to characterize the role of Wnt5a production, in situ, in vivo and in vitro in response to fungal keratitis. Wnt5a expression in corneas of Aspergillus fumigatus (A. fumigatus) keratitis patients was determined by quantitative polymerase chain reaction (qRT-PCR) and immunofluorescence. In vivo and in vitro experiments were then performed in mouse models and THP-1 macrophages cell cultures infected with A. fumigatus, respectively. C57BL/6 mice were pretreated with siRNAs or neutralizing antibodies for dectin-1, LOX-1 and Wnt5a, or inhibitors of erk1/2 and JNK. Changes in Wnt5a expression were assessed by clinical evaluation, qRT-PCR, immunofluorescence, western blot and bioluminescence imaging system image acquisition. We confirmed that corneal Wnt5a expression increased with A. fumigatus keratitis in patients and a murine model. Wnt5a production was dependent on dectin-1 and LOX-1 expression with contributions by Erk1/2 and JNK pathways. Additionally, Wnt5a knockdown revealed decreased levels of MPO, lower neutrophil recruitment, and a higher fungal load in mouse models. Compared with controls, Wnt5a knockdown impaired pro-inflammatory cytokine IL-1ß production in response to A. fumigatus exposure. Wnt5a also produces dectin-1 and LOX-1 induced inflammatory signature via effective neutrophil recruitment and inflammatory cytokine production in response to A. fumigatus keratitis. These findings demonstrate that Wnt5a is a critical component of the antifungal immune response.

Increased Extracellular Vesicles Mediate WNT5A Signaling in Idiopathic Pulmonary Fibrosis.

Rationale: Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease characterized by lung epithelial cell injury, increased (myo)fibroblast activation, and extracellular matrix deposition. Extracellular vesicles (EVs) regulate intercellular communication by carrying a variety of signaling mediators, including WNT (wingless/integrated) proteins. The relevance of EVs in pulmonary fibrosis and their potential contribution to disease pathogenesis, however, remain unexplored.Objectives: To characterize EVs and study the role of EV-bound WNT signaling in IPF.Methods: We isolated EVs from BAL fluid (BALF) from experimental lung fibrosis as well as samples from IPF, non-IPF interstitial lung disease (ILD), non-ILD, and healthy volunteers from two independent cohorts. EVs were characterized by transmission electron microscopy, nanoparticle tracking analysis, and Western blotting. Primary human lung fibroblasts (phLFs) were used for EV isolation and analyzed by metabolic activity assays, cell counting, quantitative PCR, and Western blotting upon WNT gain- and loss-of-function studies.Measurements and Main Results: We found increased EVs, particularly exosomes, in BALF from experimental lung fibrosis as well as from patients with IPF. WNT5A was secreted on EVs in lung fibrosis and induced by transforming growth factor-ß in primary human lung fibroblasts. The phLF-derived EVs induced phLF proliferation, which was attenuated by WNT5A silencing and antibody-mediated inhibition and required intact EV structure. Similarly, EVs from IPF BALF induced phLF proliferation, which was mediated by WNT5A.Conclusions: Increased EVs function as carriers for signaling mediators, such as WNT5A, in IPF and thus contribute to disease pathogenesis. Characterization of EV secretion and composition may lead to novel approaches to diagnose and develop treatments for pulmonary fibrosis.

Coordinated directional outgrowth and pattern formation by integration of Wnt5a and Fgf signaling in planar cell polarity.

Embryonic morphogenesis of a complex organism requires proper regulation of patterning and directional growth. Planar cell polarity (PCP) signaling is emerging as a crucial evolutionarily conserved mechanism whereby directional information is conveyed. PCP is thought to be established by global cues, and recent studies have revealed an instructive role of a Wnt signaling gradient in epithelial tissues of both invertebrates and vertebrates. However, it remains unclear whether Wnt/PCP signaling is regulated in a coordinated manner with embryonic patterning during morphogenesis. Here, in mouse developing limbs, we find that apical ectoderm ridge-derived Fgfs required for limb patterning regulate PCP along the proximal-distal axis in a Wnt5a-dependent manner. We demonstrate with genetic evidence that the Wnt5a gradient acts as a global cue that is instructive in establishing PCP in the limb mesenchyme, and that Wnt5a also plays a permissive role to allow Fgf signaling to orient PCP. Our results indicate that limb morphogenesis is regulated by coordination of directional growth and patterning through integration of Wnt5a and Fgf signaling.

A newly identified lncRNA MAR1 acts as a miR-487b sponge to promote skeletal muscle differentiation and regeneration.

BACKGROUND: Skeletal muscle atrophy induced by either aging (sarcopenia) or mechanical unloading is associated with serious health consequences. Long non-coding RNAs (lncRNAs) are implicated as important regulators in numerous physiological and pathological processes. METHODS: Microarray analysis was performed to identify the differentially expressed lncRNAs in skeletal muscle between adult and aged mice. The most decreased lncRNA in aged skeletal muscle was identified. The C2C12 mouse myoblast cells were used to assess the biological function of the lncRNA in vitro. The target microRNA of lncRNA and the target protein of microRNA were predicted by bioinformatics analysis and validated in vitro. Furthermore, the biology function of the lncRNA in vivo was investigated by local overexpression or knockdown the lncRNA in skeletal muscle. The therapeutic effect of the lncRNA overexpression in age-related or mechanical unloading-induced muscle atrophy was also evaluated. RESULTS: We identified a novel lncRNA (muscle anabolic regulator 1, MAR1) which was highly expressed in mice skeletal muscle and positively correlated with muscle differentiation and growth in vitro and in vivo. We predicted and validated that microRNA-487b (miR-487b) was a direct target of MAR1. We also predicted and validated that Wnt5a, an important regulator during myogenesis, was a target of miR-487b in C2C12 cells. Our findings further demonstrated that enforced MAR1 expression in myoblasts led to derepression of Wnt5a. Moreover, MAR1 promoted skeletal muscle mass/strength and Wnt5a protein level in mice. Enforced MAR1 expression in mice attenuated muscle atrophy induced by either aging or unloading. CONCLUSIONS: The newly identified lncRNA MAR1 acts as a miR-487b sponge to regulate Wnt5a protein, resulting in promoting muscle differentiation and regeneration. MAR1 could be a novel therapeutic target for treating muscle atrophy induced by either aging or mechanical unloading.

Ibrutinib inhibition of ERBB4 reduces cell growth in a WNT5A-dependent manner.

Alterations in ERBB family members have been associated with many tumor malignancies. EGFR and ERBB2 have been extensively explored in clinical oncology and several drugs currently target them therapeutically. However, the significance of ERBB4 as a potential therapeutic target remains mostly unexplored, even though ERBB4 is overexpressed or mutated in many solid tumors. Using a unique functional protein microarray platform, we found that ibrutinib inhibits ERBB4 activity in the same nM range as its canonical target, BTK. Cell-based assays revealed that ibrutinib treatment inhibited cell growth and decreased phosphorylation of ERBB4 and downstream targets MEK and ERK in cancer cell lines with high levels of endogenous ERBB4. In vivo, ibrutinib-responsive mouse xenograft tumors showed decreased tumor volumes with ibrutinib treatment. Interestingly, global gene expression comparisons between responsive and non-responsive cells identified a signature featuring the WNT pathway that predicts growth responsiveness to ibrutinib. Non-responsive ERBB4-expressing cell lines featured elevated activity of the WNT pathway, through the overexpression of WNT5A. Moreover, inhibition of WNT5A expression led to an ibrutinib response in non-responsive cell lines. Our data show that inhibiting ERBB4 reduces cell growth in cells that have low WNT5A expression and reveal a link between the ERBB4 and WNT pathways.

WNT5A promotes migration and invasion of human osteosarcoma cells via SRC/ERK/MMP-14 pathway.

WNT5A, a representative ligand of activating several non-canonical WNT signal pathways, plays significant roles in oncogenesis and tumor inhibition. It has been shown that the non-receptor tyrosine kinase SRC is required for WNT5A-induced invasion of osteosarcoma cells. However, the precise molecular mechanism underlying WNT5A/SRC-mediated osteosarcoma cells invasion remains poorly defined. The study was designed to explore the role of ERK1/2 in WNT5A/SRC-induced osteosarcoma cells invasion and the downstream target of the SRC/ERK1/2 signalings. We found that WNT5A (100 ng/mL) remarkably stimulated migration and invasion of human osteosarcoma MG-63 cells, whereas inhibiting either SRC kinase activity by siRNA-mediated SRC silence or ERK1/2 phosphorylation by PD98059 treatment suppressed these effects, which suggested that the activation of SRC and ERK1/2 is essential for WNT5A-induced MG-63 cells migration and invasion. Furthermore, ERK1/2 phosphorylation induced by WNT5A was dramatically blocked by SRC siRNA. Additionally, our study further demonstrated that MMP-14 was upregulated after exposure to WNT5A in MG-63 cells, and the increased expression was blocked by SRC siRNA or PD98059. Collectively, these results indicate that WNT5A activates SRC/ERK1/2 signal pathway, leading to the upregulation of MMP-14 expression and MG-63 cells migration and invasion.

Bivalent Histone Codes on WNT5A during Odontogenic Differentiation.

Lineage-committed differentiation is an essential biological program during odontogenesis, which is tightly regulated by lineage-specific genes. Some of these genes are modified by colocalization of H3K4me3 and H3K27me3 marks at promoter regions in progenitors. These modifications, named "bivalent domains," maintain genes in a poised state and then resolve for later activation or repression during differentiation. Wnt5a has been reported to promote odontogenic differentiation in dental mesenchyme. However, relatively little is known about the epigenetic modulations on Wnt5a activation during tooth development. Here, we investigated the spatiotemporal patterns of H3K4me3 and H3K27me3 marks in developing mouse molars. Associated H3K4me3 methylases (mixed-lineage leukemia [MLL] complex) and H3K27me3 demethylases (JMJD3 and UTX) were dynamically expressed between early and late bell stage of human tooth germs and in cultured human dental papilla cells (hDPCs) during odontogenic induction. Poised WNT5A gene was marked by bivalent domains containing repressive marks (H3K27me3) and active marks (H3K4me3) on promoters. The bivalent domains tended to resolve during inducted differentiation, with removal of the H3K27me3 mark in a JMJD3-dependent manner. When JMJD3 was knocked down in cultured hDPCs, odontogenic differentiation was suppressed. The depletion of JMJD3 epigenetically repressed WNT5A activation by increased H3K27me3 marks. In addition, JMJD3 could physically interact with ASH2L, a component of the MLL complex, to form a coactivator complex, cooperatively modulating H3K4me3 marks on WNT5A promoters. Overall, our study reveals that transcription activities of WNT5A were epigenetically regulated by the negotiated balance between H3K27me3 and H3K4me3 marks and tightly mediated by JMJD3 and MLL coactivator complex, ultimately modulating odontogenic commitment during dental mesenchymal cell differentiation.

Wnt5a promotes epithelial-to-mesenchymal transition and metastasis in non-small-cell lung cancer.

A recent study indicated that high Wnt5a expression is associated with poor prognosis in non-small-cell lung cancer (NSCLC) patients; however, the underlying mechanism was not clear yet. Immunohistochemistry and Western blotting were performed to examine the protein expression level in NSCLC tissues and cell lines. The role of Wnt5a in clone formation, invasiveness, migration, and epithelial-to-mesenchymal transition (EMT) of NSCLC cells was studied. Luciferase reporter assay was used to evaluate the Tcf/Lef transcriptional activity. For assessing the effects of Wnt5a on tumor growth and metastasis in vivo, A549 cells transfected with sh-Wnt5a were subcutaneously or orthotopically injected into nude mice. In NSCLC tissues, higher expression levels of Wnt5a and ROR2 were found, ß-Catenin was expressed exceptionally, and EMT was prompted. Wnt5a overexpression increased clone formation, migration, and invasion, as well as prompted EMT of NSCLC cell in vitro, whereas Wnt5a knockdown showed the absolutely reversed results. Wnt5a overexpression enhanced the Tcf/Lef transcriptional activity and elevated the nuclear ß-catenin level in NSCLC cells, without altering the ROR2 expression. We also demonstrated that si-ß-catenin antagonized Wnt5a overexpression nduced EMT and invasiveness. Besides, in vivo experiment showed that sh-Wnt5a significantly increased tumor volume and tumor weight, and prompted EMT in A549 tumor-bearing mice as compared with the control. No metastasis was found in the liver tissue after sh-Wnt5a-transfected cells were orthotopically injected into nude mice as compared with the control. In conclusion, Wnt5a promotes EMT and metastasis in NSCLC, which is involved in the activation of ß-catenin-dependent canonical Wnt signaling.

The secreted protein WNT5A regulates condylar chondrocyte proliferation, hypertrophy and migration.

OBJECTIVE: Our previous study showed that WNT5A, a member of the noncanonical WNT pathway, is involved in interleukin-1beta induced matrix metalloproteinase expression in temporomandibular joint (TMJ) condylar chondrocytes. The purpose of this study is to further explore the roles of WNT5A in cartilage biology of the TMJ. METHODS: An early TMJ osteoarthritis-like rat model was constructed by a mechanical method (steady mouth-opening). The gene and protein levels of WNT5A during the condylar cartilage changes were measured. Effects of WNT5A on chondrocyte proliferation, hypertrophy and migration were analyzed after WNT5A gain or loss of function in vitro. A c-Jun N-terminal kinase (JNK) inhibitor SP600125 was used to evaluate the involvement of JNK pathway in these effects of WNT5A. The expression and transcription activity of cell cycle regulators c-MYC and Cyclin D1 were examined to determine the mechanism behind WNT5A regulation of chondrocyte proliferation. RESULTS: WNT5A was significantly upregulated in the condylar cartilage of rats in the early TMJ osteoarthritis-like model. Activating WNT5A facilitated condylar chondrocyte proliferation, hypertrophy and migration. Conversely, inhibiting WNT5A activity in chondrocytes decreased their proliferation, hypertrophy and migration. Blockage of the JNK pathway by its inhibitor, SP600125, impaired these effects of WNT5A on chondrocytes. WNT5A regulated both the expression and transcriptional activity of c-MYC and Cyclin D1 in chondrocytes, both of which were upregulated in condylar cartilage of the rat early TMJ osteoarthritis. CONCLUSION: WNT5A regulates condylar chondrocyte proliferation, hypertrophy and migration. These findings provide new insights into the role of WNT5A signaling in TMJ cartilage biology and its potential in future therapy for TMJ degenerative diseases.

Porcupine-dependent Wnt signaling controls stromal proliferation and endometrial gland maintenance through the action of distinct WNTs.

Wnt signaling has been shown to be important in orchestrating proper development of the female reproductive tract. In the uterus, six members of the Wnt family are expressed in the neonatal endometrium and deletion of individual Wnt genes often leads to similar phenotypes, suggesting an interaction of these genes in uterine development and function. Furthermore, Wnts may have complementary functions, which could mask the identification of their individual functional role in single gene deletions. To circumvent this issue, we have generated a deletion of the Porcupine homolog within the female reproductive tract using progesterone receptor-Cre mice (PgrCre/+); preventing Wnt secretion from the producing cells. We show that Porcupine-dependent Wnt signaling, unlike previously reported, is dispensable for postnatal gland formation but is required for post-pubertal gland maintenance as well as for stromal cell proliferation. Furthermore, our results demonstrate that WNT7a is sufficient to restore post-pubertal endometrial gland formation. Although WNT5a did not restore gland formation, it rescued stromal cell proliferation; up-regulating several secreted factors including Fgf10 and Ihh. Our results further elucidate the roles of Wnt signaling in uterine development and function as well as provide an ideal system to address individual Wnt functions in the uterus.

Noncanonical WNT-5A signaling impairs endogenous lung repair in COPD.

Chronic obstructive pulmonary disease (COPD) is a leading cause of death worldwide. One main pathological feature of COPD is the loss of functional alveolar tissue without adequate repair (emphysema), yet the underlying mechanisms are poorly defined. Reduced WNT-ß-catenin signaling is linked to impaired lung repair in COPD; however, the factors responsible for attenuating this pathway remain to be elucidated. Here, we identify a canonical to noncanonical WNT signaling shift contributing to COPD pathogenesis. We demonstrate enhanced expression of noncanonical WNT-5A in two experimental models of COPD and increased posttranslationally modified WNT-5A in human COPD tissue specimens. WNT-5A was increased in primary lung fibroblasts from COPD patients and induced by COPD-related stimuli, such as TGF-ß, cigarette smoke (CS), and cellular senescence. Functionally, mature WNT-5A attenuated canonical WNT-driven alveolar epithelial cell wound healing and transdifferentiation in vitro. Lung-specific WNT-5A overexpression exacerbated airspace enlargement in elastase-induced emphysema in vivo. Accordingly, inhibition of WNT-5A in vivo attenuated lung tissue destruction, improved lung function, and restored expression of ß-catenin-driven target genes and alveolar epithelial cell markers in the elastase, as well as in CS-induced models of COPD. We thus identify a novel essential mechanism involved in impaired mesenchymal-epithelial cross talk in COPD pathogenesis, which is amenable to therapy.

Niche WNT5A regulates the actin cytoskeleton during regeneration of hematopoietic stem cells.

Here, we show that the Wnt5a-haploinsufficient niche regenerates dysfunctional HSCs, which do not successfully engraft in secondary recipients. RNA sequencing of the regenerated donor Lin- SCA-1+ KIT+ (LSK) cells shows dysregulated expression of ZEB1-associated genes involved in the small GTPase-dependent actin polymerization pathway. Misexpression of DOCK2, WAVE2, and activation of CDC42 results in apolar F-actin localization, leading to defects in adhesion, migration and homing of HSCs regenerated in a Wnt5a-haploinsufficient microenvironment. Moreover, these cells show increased differentiation in vitro, with rapid loss of HSC-enriched LSK cells. Our study further shows that the Wnt5a-haploinsufficient environment similarly affects BCR-ABLp185 leukemia-initiating cells, which fail to generate leukemia in 42% of the studied recipients, or to transfer leukemia to secondary hosts. Thus, we show that WNT5A in the bone marrow niche is required to regenerate HSCs and leukemic cells with functional ability to rearrange the actin cytoskeleton and engraft successfully.

Wnt5a Drives an Invasive Phenotype in Human Glioblastoma Stem-like Cells.

Brain invasion by glioblastoma determines prognosis, recurrence, and lethality in patients, but no master factor coordinating the invasive properties of glioblastoma has been identified. Here we report evidence favoring such a role for the noncanonical WNT family member Wnt5a. We found the most invasive gliomas to be characterized by Wnt5a overexpression, which correlated with poor prognosis and also discriminated infiltrating mesenchymal glioblastoma from poorly motile proneural and classical glioblastoma. Indeed, Wnt5a overexpression associated with tumor-promoting stem-like characteristics (TPC) in defining the character of highly infiltrating mesenchymal glioblastoma cells (Wnt5aHigh). Inhibiting Wnt5a in mesenchymal glioblastoma TPC suppressed their infiltrating capability. Conversely, enforcing high levels of Wnt5a activated an infiltrative, mesenchymal-like program in classical glioblastoma TPC and Wnt5aLow mesenchymal TPC. In intracranial mouse xenograft models of glioblastoma, inhibiting Wnt5a activity blocked brain invasion and increased host survival. Overall, our results highlight Wnt5a as a master regulator of brain invasion, specifically TPC, and they provide a therapeutic rationale to target it in patients with glioblastoma. Cancer Res; 77(4); 996-1007. ©2016 AACR.

Can Wnt5a and Wnt non-canonical pathways really mediate adipocyte de-differentiation in a tumour microenvironment?

Wnt5a has been recently reported as a possible triggering factor of adipocyte de-differentiation into an adipocyte-derived fibroblast in the tumour microenvironment of pancreas cancer. The Wnt/ß-catenin pathway was described in processes involving de-differentiation and epithelial-mesenchymal transition but some Wnt family-belonging molecules exert an adipogenic role on adipocyte, while other ones, such as Wnt10b or Wnt3a, an anti-adipogenic role. Although this ability depends on the different tumoural microenvironments, it is intriguing to ascertain if some Wnt molecules, participating in the non-canonical pathway, may be targeted as fundamental factors able to trigger the desmoplastic reaction of peritumoural white adipose tissue.

A Possible Role for WNT5A Hypermethylation in Pediatric Acute Lymphoblastic Leukemia.

OBJECTIVE: WNT5A is one of the most studied noncanonical WNT ligands and is shown to be deregulated in different tumor types. Our aim was to clarify whether hypermethylation might be the cause of low WNT5A mRNA levels and whether we could restore this downregulation by reversing the event. MATERIALS AND METHODS: The expression of WNT5A mRNA was studied in a large acute lymphoblastic leukemia (ALL) patient group (n=86) by quantitative real-time PCR. The methylation status was detected by methylation-specific PCR (MSPCR) and bisulphate sequencing. In order to determine whether methylation has a direct effect on WNT5A expression, disease-representative cell lines were treated by 5'-aza-20-deoxycytidine. RESULTS: Here we designed a validation experiment of the WNT5A gene, which was previously examined and found to be differentially expressed by microarray study in 31 T-cell ALL patients. The expression levels were confirmed by quantitative real-time PCR and the expression levels were significantly lower in T-cell ALL patients than in control thymic subsets (p=0.007). MSPCR revealed that 86% of the patients were hypermethylated in the WNT5A promoter region. Jurkat and RPMI cell lines were treated with 5'-aza-20-deoxycytidine and WNT5A mRNA expression was restored after treatment. CONCLUSION: According to our results, WNT5A hypermethylation does occur in ALL patients and it has a direct effect on mRNA expression. Our findings show that epigenetic changes of WNT signaling can play a role in ALL pathogenesis and reversing methylation might be useful as a possible treatment of leukemia.