Expression Of Wnt10b Research Articles

Anti-inflammatory role of low-intensity pulsed ultrasound in inhibiting lipopolysaccharide-induced M1 polarization of RAW264.7 cells via Wnt2b/AXIN/β-catenin

Background Low-intensity pulsed ultrasound (LIPUS) is a special type of low-intensity ultrasound. In periodontal disease, LIPUS is applied as an adjuvant and non-invasive treatment. It has been reported that LIPUS significantly shifts the macrophage phenotype from M1 to M2, but the specific mechanism behind this shift is still unknown. Methods RAW264.7 cells were induced to M1/M2 polarization with lipopolysaccharide (LPS)/interleukin-4 (IL4). LIPUS was performed for 25 min two times, 24 h apart, at an intensity of 45 mW/cm2 to stimulate RAW264.7 cells. PolyA mRNA sequencing was conducted of both the LPS-induced RAW264.7 cells and the LPS-induced RAW264.7 cells with LIPUS treatment. The expression of Wnt2b in RAW264.7 cells was downregulated by siRNA. The macrophage surface markers and downstream inflammatory cytokines were detected using flow cytometry. The relative expression of proteins in the Wnt2b/AXIN/β-catenin pathway was assessed using reverse transcription real-time polymerase chain reaction (RT-qPCR) and Western blot. Results LIPUS reversed the M1 polarization of RAW264.7 cells, with decreased expression of CD80 and CD86. In addition, LIPUS enhanced the M2 polarization of RAW264.7 cells, with upregulated expression of CD163 and CD206. The polyA mRNA sequencing results indicated that the Wnt signaling pathway participated in the M1 polarization of LIPUS-treated RAW264.7. The results of the RT-qPCR showed a higher expression of Wnt2b in LIPUS-treated and M1- or M2-polarized RAW264.7 cells. Knocking down Wnt2b was shown to reverse the inhibitory effect of LIPUS on M1 polarization and increase the expression of CD80 and CD86. Wnt2b knockdown also regulated downstream AXIN, β-catenin, and inflammatory factors such as tumor necrosis factor alpha (TNFα) and interleukin-6 (IL6). Conclusions LIPUS plays an anti-inflammatory role by inhibiting LPS-induced M1 polarization of RAW264.7 cells in a Wnt2b/AXIN/β-catenin-dependent way. LIPUS may play a therapeutic role in periodontal diseases by inhibiting inflammation through the regulation of macrophage differentiation.

WNT7B promotes cancer progression via WNT/β-catenin signaling pathway and predicts a poor prognosis in oral squamous cell carcinoma

BackgroundWNT7B is a glycoprotein that plays a crucial role in tumorigenesis. This study aimed to investigate the role of WNT7B in oral squamous cell carcinoma (OSCC).MethodsBioinformatic databases, immunohistochemistry, a real-time polymerase chain reaction, western blot, and enzyme-linked immunosorbent assay were used to detect WNT7B expression in OSCC. The clinical and prognostic importance of WNT7B expression was evaluated. WNT7B expression was examined in oral leukoplakia and carcinoma induced by 4-nitroquinoline 1-oxide in mice. Loss- and gain-of-function analyses were performed to elucidate the role of WNT7B in OSCC cells. Subcutaneous tumor model was established to observe the effects of WNT7B on tumor growth. Co-Immunoprecipitation was used to explore the Frizzled receptors that WNT7B may bind to.ResultsWNT7B upregulated in OSCC and associated with lymph node metastasis, perineural invasion, and an unfavorable prognosis in patients with OSCC. A gradual increased in WNT7B expression during the malignant progression of OSCC. WNT7B promoted cell proliferation, migration, invasion, while silencing WNT7B abolished these effects. Knocking down the expression of WNT7B inhibits tumor growth in vivo. WNT7B functions by binding to the Frizzled 7 receptor and facilitates the nuclear translocation of β-catenin.ConclusionsWNT7B contributes to the progression of OSCC by modulating the WNT/β-catenin signaling pathway. These findings highlight the potential of WNT7B as a novel prognostic biomarker and promising therapeutic target for OSCC.

Activation of the WNT7B/β-Catenin Pathway Initiates GLUT1 Expression and Promotes Aerobic Glycolysis in Colorectal Cancer Cells

Glucose is an important energy source for tumors, however the molecular mechanisms by which tumor cells regulate glucose uptake remain unclear. In this study, we aimed to investigate the regulation mechanism of the WNT7B/β-catenin pathway for glucose transporter 1 (GLUT1)-mediated glucose metabolism in colorectal cancer. Here, we found that WNT7B expression levels were significantly increased in colorectal cancer tissues and closely associated with the clinical stage and lymph node metastasis in patients with colorectal cancer. Next, we confirmed that WNT7B significantly increased the glucose consumption and lactic acid levels in SW480 cells by overexpressing WNT7B. Additionally, gene and protein levels of GLUT1 were increased in WNT7B-overexpressing SW480 cells. However, WNT7B knockdown reversed these effects. WNT7B also enhanced GLUT1-mediated cell proliferation, invasion, and migration. WNT7B overexpression inhibited the effect of glucose deprivation on apoptosis. The WNT/β-catenin signaling pathway inhibitor, LGK974, inhibited WNT7B secretion, leading to GLUT1 levels downregulation and promotion of cell apoptosis. Ectopic tumor xenograft model experiments revealed that WNT7B promoted tumor progression in mice. Overall, our results suggest that WNT7B promotes β-catenin entry into the nucleus to initiates GLUT1 transcription, increases glucose transport and consumption, and enhances aerobic glycolysis, thus promoting tumor progression in colorectal cancer cells.

Acute myeloid leukemia cells adhere to bone marrow and acquire chemoresistance by downregulating UNC5B expression.

Acute myeloid leukemia (AML) is a malignant tumor of the hematological system. Because of its characteristics of recurrence, refractory and chemoresistance, new therapeutic targets need to be identified. Adhesion and proliferation are characteristics of AML cells, and critical steps in inducing chemotherapy resistance. In this study, we reported that UNC5B inhibits AML cell bone marrow adhesion, inhibits AML cell proliferation and increases sensitivity to chemotherapy. Mechanistically, RNA sequencing (RNA-seq) and experimental results revealed that overexpression of UNC5B inhibits adhesion and proliferation signaling pathways and inhibits the expression of MPZL1, CLDN23, IGF2 and WNT7B. In conclusion, our findings suggest that UNC5B serves as a prognostic indicator and a potential therapeutic target for AML.

A pan-cancer analysis ofWnt family member 7Bin human cancers.

Previous studies have highlighted the crucial role of Wnt7B in the development of various cancers, including breast, pancreatic, and gastric cancers. However, research into the involvement of Wnt7B is often confined to specific tumor types, with a noticeable lack of comprehensive studies spanning multiple cancer forms. The potential of Wnt7B as a diagnostic or prognostic cancer biomarker has not been fully explored. In this study, we combined bioinformatics and immunohistochemistry analyses to examine the expression patterns and functions of Wnt7B in cancerous and adjacent noncancerous tissues across a range of tumors. Our data indicate that Wnt7B may serve as a novel prognostic biomarker and therapeutic target in certain cancers. We found significant upregulation of Wnt7B expression levels in the majority of cancer cases examined. Furthermore, Wnt7B can influence cancer prognosis by modulating the tumor microenvironment, immune cell infiltration, and tumor stemness, among other factors. Additionally, we examined the associations between anticancer drug sensitivity and Wnt7B expression, which could aid in the development of more precise clinical therapies.

Lineage tracing using Wnt2b-2A-CreERT2 knock-in mice reveals the contributions of Wnt2b-expressing cells to novel subpopulations of mesothelial/epicardial cell lineages during mouse development.

Mesothelial and epicardial cells give rise to various types of mesenchymal cells via epithelial (mesothelial)-to-mesenchymal transition during development. However, the genes controlling the differentiation and diversification of mesothelial/epicardial cells remain unclear. Here, we examined Wnt2b expression in the embryonic mesothelium and epicardium and performed lineage tracing of Wnt2b-expressing cells by using novel Wnt2b-2A-CreERT2 knock-in and LacZ-reporter mice. Wnt2b was expressed in mesothelial cells covering visceral organs, but the expression was restricted in their subpopulations. Wnt2b-expressing cells labeled at embryonic day (E) 10.5 were distributed to the mesothelium and mesenchyme in the lungs, abdominal wall, stomach, and spleen in Wnt2b2A-CreERT2/+;R26RLacZ/+ mice at E13.0. Wnt2b was initially expressed in the proepicardial organ (PEO) at E9.5 and then in the epicardium after E10.0. Wnt2b-expressing PEO cells labeled at E9.5 differentiated into a small fraction of cardiac fibroblasts and preferentially localized at the left side of the postnatal heart. LacZ+ epicardium-derived cells labeled at E10.5 differentiated into a small fraction of fibroblasts and smooth muscle cells in the postnatal heart. Taken together, our results reveal novel subpopulations of PEO and mesothelial/epicardial cells that are distinguishable by Wnt2b expression and elucidate the unique contribution of Wnt2b-expressing PEO and epicardial cells to the postnatal heart.

The metabolites of Bifidobacterium longum BB536 alleviate DHT-damaged human dermal papilla cells by activating WNT/β-catenin signaling

Probiotics have numerous biological functions and clinical studies have reported that consuming probiotic foods could promote hair growth and hair follicle health. Bifidobacterium longum BB536 was selected to investigate the possibility of treating hair loss with Cell-Free Supernatant (CFS) of B. longum BB536 and the alleviation mechanism of CFS on dihydrotestosterone (DHT)-damaged human dermal papilla cells (HDPCs). The results showed that HDPCs treated with 10% CFS of B. longum BB536 for 24 h could significantly promote cell proliferation, repair cell damage, accelerate the process of cell cycle and increase the secretion of VEGF. The CFS of B. longum BB536 reduced the apoptosis rate of DHT-damaged HDPCs. In addition, it could reduce the level of pro-inflammatory factors and improve the alkaline phosphatase (ALP) activity. Metabolites of B. longum BB536 may increase the protein expression of Wnt10b, β-catenin and LEF-1 by activating Wnt/β-catenin signaling pathway and inhibit the expression of DHT-induced negative feedback factors DKK1 and GSK-3β, which alleviates the damage of DHT on HDPCs, some protein expression levels such as LEF-1 are close to minoxidil (common drugs for hair loss). So, metabolites of B. longum BB536 can be used as new treatment for hair loss due to its safety. This study provides theoretical basis for probiotics to prevent androgenic alopecia.

WNT5B promotes the malignant phenotype of non-small cell lung cancer via the FZD3–DVL3–RAC1–PCP–JNK pathway

The WNT5B ligand regulates the non-canonical wingless-related integration site (WNT)–planar cell polarity (PCP) pathway. However, the detailed mechanism underlying the activity of WNT5B in the WNT–PCP pathway in non-small cell lung cancer (NSCLC) is unclear. In this study, we assessed the clinicopathological significance of WNT5B expression in NSCLC specimens. WNT5B-overexpression and -knockdown NSCLC cell lines were generated in vivo and in vitro, respectively. WNT5B overexpression in NSCLC specimens correlates with advanced tumor node metastasis (TNM) stage, lymph node metastasis, and poor prognosis in patients with NSCLC. Additionally, WNT5B promotes the malignant phenotype of NSCLC cells in vivo and in vitro. Interactions were identified among WNT5B, frizzled3 (FZD3), and disheveled3 (DVL3) in NSCLC cells, leading to the activation of WNT–PCP signaling. The FZD3 receptor initiates DVL3 recruitment to the membrane for phosphorylation in a WNT5B ligand-dependent manner and activates c-Jun N-terminal kinase (JNK) signaling via the small GTPase RAC1. Furthermore, the deletion of the DEP domain of DVL3 abrogated these effects. Overall, we demonstrated a novel signal transduction pathway in which WNT5B recruits DVL3 to the membrane via its DEP domain through interaction with FZD3 to promote RAC1–PCP–JNK signaling, providing a potential target for clinical intervention in NSCLC treatment.

O-078 Deletion of interlukin-17A protects against ovarian aging via activating wnt10b/β-catenin signaling

Abstract Study question Is there any compelling evidence to suggest that the deletion of IL17A could potentially ameliorate ovarian aging? Summary answer IL17A is elevated in the follicular fluid of aged women. Knocking out Il17a in mice improves ovarian aging phenotypes by activating Wnt10b/β-catenin signaling. What is known already Inflammation, a hallmark of aging, negatively impacts female fertility. Interleukin 17 (IL-17) cytokines, including IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, and IL-17F, are signature cytokines of T helper 17 cells. IL-17A, the first-discovered and well-explored among them, has been predominantly characterized as a kind of proinflammatory cytokine and a promoter in the pathogenesis of chronic autoimmune diseases such as psoriasis and ankylosing spondylitis. Significant accumulation of CD4-CD8-double-negative T cells in aged mouse ovaries, identified as a crucial source of IL-17, was reported via single-cell RNA sequencing. However, much remains unknown about the IL17 and ovarian aging relationship. Study design, size, duration Follicular fluid was collected from 51 women (age≥35 years) and 37 controls (age <35 years) at the Southern Medical University Southern Hospital's Reproductive Center to measure IL17A levels from March 2022 to August 2023. Additionally, we generated Il-17a knockout (Il17-a KO) mice (42 KO mice and 49 controls) to elucidate the role of Il-17a in ovarian aging in vivo. Further, RNA-seq was employed to explore the mechanism involved in the Il17a-associated ovarian aging. Participants/materials, setting, methods Clinical data were compared from two infertile patient cohorts. IL17A levels in follicular fluid were quantified using ELISA. Pearson correlation analysis explored the relationship between IL17A levels and age, ovarian reserve, and clinical outcomes. We also assessed aging-related phenotypes in Il17a-KO and WT mice at 3, 8, 12, and 15 months of age. Fertility assessment, ovarian reserve, histological analysis, mason staining, western blot analysis, qRT-PCR, IVF, spindle formation, and RNA-seq were used in this study. Main results and the role of chance Elevated IL17A levels in follicular fluid from aged patients were observed and may be used as a sensitive biomarker for advanced age (P < 0.001). Deletion of Il17a not only enhanced fertility and ovarian reserve in mice but also ameliorated inflammation, the senescent associated secretory phenotype (SASP), senescence and fibrosis. RNA-seq analysis identified 705 differentially expressed genes, primarily enriched in the Wnt10b signaling pathway. Validation experiments suggested that IL-17A deficiency may improve ovarian aging by upregulating wnt10b expression, promoting β-catenin nuclear translocation, and activating downstream molecules (c-Myc, Cyclin D1, c-jun). Limitations, reasons for caution This study found a correlation between IL17A and aging. However, how knocking out IL17A promotes the increase of Wnt10b and other potential functions of Il17a in ovarian aging remain to be further explored. Wider implications of the findings This study shows new insights into the mechanisms of IL17A in ovarian aging and provides potential biomarkers and therapeutic targets for the treatment of ovarian aging. In the future, a larger sample of studies are needed to investigate the relationship between IL17A and aged women. Trial registration number not applicable

Purification and identification of novel osteogenic peptides from sea cucumber intestine hydrolysates and their pro-osteogenesis effects on MC3T3-E1 cells

The present study aimed to purify and identify osteogenic peptides from sea cucumber intestine hydrolysates (Mw＜3 kDa) and further evaluate their pro-osteogenesis properties on MC3T3-E1 cells. Two novel osteogenic peptides, RPQYPQYPS and LSFSPY, were purified and identified from the alcalase hydrolysate of sea cucumber intestine by gel filtration chromatography, ion exchange column chromatography, RP-HPLC, which were further sequenced by LC-MS/MS and synthesized with solid-phase synthesis method. Peptides RPQYPQYPS and LSFSPY significantly increased the proliferation of MC3T3-E1 cells by 38.7% and 32.8%. Alizarin red staining also confirmed that peptides RPQYPQYPS and LSFSPY significantly promoted MC3T3-E1 cell mineralization in a dose-dependent manner. Further, molecular docking results showed that RPQYPQYPS and LSFSPY had a higher affinity with integrin α5β1. Western blot results confirmed that RPQYPQYPS and LSFSPY significantly upregulated the expression of integrin downstream signals PI3K, ERK and Wnt10b. These findings provided evidence for the osteogenic peptides from sea cucumber intestines as a functional food ingredient.

LINC01605 promotes malignant phenotypes of cervical cancer via miR-149-3p/WNT7B axis

BackgroundLong non-coding RNAs (LncRNA) play a pivotal role in the progression of various malignancies. Despite recent identification as an oncogene associated with tumorigenesis. The precise role of LINC01605 in cervical cancer (CC) remains unclear. Therefore, the objective of this study was to investigate the influence of LINC01605 on proliferation and invasion of CC cells, while also exploring its potential underlying mechanisms. MethodsThe expression of LINC01605 in CC cell lines was analyzed using the TCGA database and qRT-PCR. Various assays, including CCK-8 and transwell analysis, were conducted on CC cells to assess the influence of LINC01605 on their proliferation, migration, and invasion capabilities. Bioinformatics and dual luciferase reporter gene assays were employed to analyze the target genes of LINC01605 and miR-149-3p. To further investigate the mechanism of action, transfection and investigation were performed using specific siRNA, miRNA mimics, or inhibitors. ResultsThe expression of LINC01605 exhibited a significant increase in CC cell lines, and this upregulation was associated with an unfavorable prognosis. Modulating the expression of LINC01605, either by down-regulating or up-regulating it, exerted suppressive or stimulatory effects on the growth and invasion of HeLa and Siha cells. LINC01605 functioned as a competitive endogenous RNA (ceRNA) for miR-149-3p, with WNT7B being identified as a target gene of miR-149-3p. The involvement of LINC01605 in CC development is facilitated by its ability to regulate the expression of WNT7B through sequestering miR-149-3p. ConclusionOur study demonstrates that LINC01605 acts as a competitive endogenous RNA in modulating the effects of WNT7B on the proliferation and invasion of CC cells by sequestering miR-149-3p. This research provides novel insights into the involvement of LINC01605 in the advancement of CC.

WNT5B drives osteosarcoma stemness, chemoresistance and metastasis.

Treatment for osteosarcoma, a paediatric bone cancer with no therapeutic advances in over three decades, is limited by a lack of targeted therapies. Osteosarcoma frequently metastasises to the lungs, and only 20% of patients survive 5 years after the diagnosis of metastatic disease. We found that WNT5B is the most abundant WNT expressed in osteosarcoma tumours and its expression correlates with metastasis, histologic subtype and reduced survival. Using tumor-spheroids to model cancer stem-like cells, we performed qPCR, immunoblotting, and immunofluorescence to monitor changes in gene and protein expression. Additionally, we measured sphere size, migration and forming efficiency to monitor phenotypic changes. Therefore, we characterised WNT5B's relevance to cancer stem-like cells, metastasis, and chemoresistance and evaluated its potential as a therapeutic target. In osteosarcoma cell lines and patient-derived spheres, WNT5B is enriched in stem cells and induces the expression of the stemness gene SOX2. WNT5B promotes sphere size, sphere-forming efficiency, and cell proliferation, migration, and chemoresistance to methotrexate (but not cisplatin or doxorubicin) in spheres formed from conventional cell lines and patient-derived xenografts. In vivo, WNT5B increased osteosarcoma lung and liver metastasis and inhibited the glycosaminoglycan hyaluronic acid via upregulation of hyaluronidase 1 (HYAL1), leading to changes in the tumour microenvironment. Further, we identified that WNT5B mRNA and protein correlate with the receptor ROR1 in primary tumours. Targeting WNT5B through inhibition of WNT/ROR1 signalling with an antibody to ROR1 reduced stemness properties, including chemoresistance, sphere size and SOX2 expression. Together, these data define WNT5B's role in driving osteosarcoma cancer stem cell expansion and methotrexate resistance and provide evidence that the WNT5B pathway is a promising candidate for treating osteosarcoma patients. WNT5B expression is high in osteosarcoma stem cells leading to increased stem cell proliferation and migration through SOX2. WNT5B expression in stem cells increases rates of osteosarcoma metastasis to the lungs and liver in vivo. The hyaluronic acid degradation enzyme HYAL1 is regulated by WNT5B in osteosarcoma contributing to metastasis. Inhibition of WNT5B with a ROR1 antibody decreases osteosarcoma stemness.

Melatonin promotes proliferation of Inner Mongolia cashmere goat hair follicle papilla cells through Wnt10b

The study demonstrated that melatonin (MT) can induce the development of secondary hair follicles in Inner Mongolian cashmere goats through the Wnt10b gene, leading to secondary dehairing. However, the mechanisms underlying the expression and molecular function of Wnt10b in dermal papilla cells (DPC) remain unknown. This research aimed to investigate the impact of MT on DPC and the regulation of Wnt10b expression, function, and molecular mechanisms in DPC. The findings revealed that MT promotes DPC proliferation and enhances DPC activity. Co-culturing DPC with overexpressed Wnt10b and MT showed a significant growth promotion. Subsequent RNA sequencing (RNA-seq) of overexpressed Wnt10b and control groups unveiled the regulatory role of Wnt10b in DPC. Numerous genes and pathways, including developmental pathways such as Wnt and MAPK, as well as processes like hair follicle morphogenesis and hair cycle, were identified. These results suggest that Wnt10b promotes the growth of secondary hair follicles in Inner Mongolian cashmere goats by regulating crucial factors and pathways in DPC proliferation.

Bone canonical Wnt signaling is downregulated in type 2 diabetes and associates with higher advanced glycation end-products (AGEs) content and reduced bone strength.

Type 2 diabetes (T2D) is associated with higher fracture risk, despite normal or high bone mineral density. We reported that bone formation genes (SOST and RUNX2) and advanced glycation end-products (AGEs) were impaired in T2D. We investigated Wnt signaling regulation and its association with AGEs accumulation and bone strength in T2D from bone tissue of 15 T2D and 21 non-diabetic postmenopausal women undergoing hip arthroplasty. Bone histomorphometry revealed a trend of low mineralized volume in T2D (T2D 0.249% [0.156-0.366]) vs non-diabetic subjects 0.352% [0.269-0.454]; p=0.053, as well as reduced bone strength (T2D 21.60 MPa [13.46-30.10] vs non-diabetic subjects 76.24 MPa [26.81-132.9]; p=0.002). We also showed that gene expression of Wnt agonists LEF-1 (p=0.0136) and WNT10B (p=0.0302) were lower in T2D. Conversely, gene expression of WNT5A (p=0.0232), SOST (p<0.0001), and GSK3B (p=0.0456) were higher, while collagen (COL1A1) was lower in T2D (p=0.0482). AGEs content was associated with SOST and WNT5A (r=0.9231, p<0.0001; r=0.6751, p=0.0322), but inversely correlated with LEF-1 and COL1A1 (r=-0.7500, p=0.0255; r=-0.9762, p=0.0004). SOST was associated with glycemic control and disease duration (r=0.4846, p=0.0043; r=0.7107, p=0.00174), whereas WNT5A and GSK3B were only correlated with glycemic control (r=0.5589, p=0.0037; r=0.4901, p=0.0051). Finally, Young's modulus was negatively correlated with SOST (r=-0.5675, p=0.0011), AXIN2 (r=-0.5523, p=0.0042), and SFRP5 (r=-0.4442, p=0.0437), while positively correlated with LEF-1 (r=0.4116, p=0.0295) and WNT10B (r=0.6697, p=0.0001). These findings suggest that Wnt signaling and AGEs could be the main determinants of bone fragility in T2D.

Bone canonical Wnt signaling is downregulated in type 2 diabetes and associates with higher advanced glycation end-products (AGEs) content and reduced bone strength

Type 2 diabetes (T2D) is associated with higher fracture risk, despite normal or high bone mineral density. We reported that bone formation genes (SOST and RUNX2) and advanced glycation end-products (AGEs) were impaired in T2D. We investigated Wnt signaling regulation and its association with AGEs accumulation and bone strength in T2D from bone tissue of 15 T2D and 21 non-diabetic postmenopausal women undergoing hip arthroplasty. Bone histomorphometry revealed a trend of low mineralized volume in T2D (T2D 0.249% [0.156–0.366]) vs non-diabetic subjects 0.352% [0.269–0.454]; p=0.053, as well as reduced bone strength (T2D 21.60 MPa [13.46–30.10] vs non-diabetic subjects 76.24 MPa [26.81–132.9]; p=0.002). We also showed that gene expression of Wnt agonists LEF-1 (p=0.0136) and WNT10B (p=0.0302) were lower in T2D. Conversely, gene expression of WNT5A (p=0.0232), SOST (p&lt;0.0001), and GSK3B (p=0.0456) were higher, while collagen (COL1A1) was lower in T2D (p=0.0482). AGEs content was associated with SOST and WNT5A (r=0.9231, p&lt;0.0001; r=0.6751, p=0.0322), but inversely correlated with LEF-1 and COL1A1 (r=–0.7500, p=0.0255; r=–0.9762, p=0.0004). SOST was associated with glycemic control and disease duration (r=0.4846, p=0.0043; r=0.7107, p=0.00174), whereas WNT5A and GSK3B were only correlated with glycemic control (r=0.5589, p=0.0037; r=0.4901, p=0.0051). Finally, Young’s modulus was negatively correlated with SOST (r=−0.5675, p=0.0011), AXIN2 (r=−0.5523, p=0.0042), and SFRP5 (r=−0.4442, p=0.0437), while positively correlated with LEF-1 (r=0.4116, p=0.0295) and WNT10B (r=0.6697, p=0.0001). These findings suggest that Wnt signaling and AGEs could be the main determinants of bone fragility in T2D.

Toll-like receptor-2 induced inflammation causes local bone formation and activates canonical Wnt signaling.

It is well established that inflammatory processes in the vicinity of bone often induce osteoclast formation and bone resorption. Effects of inflammatory processes on bone formation are less studied. Therefore, we investigated the effect of locally induced inflammation on bone formation. Toll-like receptor (TLR) 2 agonists LPS from Porphyromonas gingivalis and PAM2 were injected once subcutaneously above mouse calvarial bones. After five days, both agonists induced bone formation mainly at endocranial surfaces. The injection resulted in progressively increased calvarial thickness during 21 days. Excessive new bone formation was mainly observed separated from bone resorption cavities. Anti-RANKL did not affect the increase of bone formation. Inflammation caused increased bone formation rate due to increased mineralizing surfaces as assessed by dynamic histomorphometry. In areas close to new bone formation, an abundance of proliferating cells was observed as well as cells robustly stained for Runx2 and alkaline phosphatase. PAM2 increased the mRNA expression of Lrp5, Lrp6 and Wnt7b, and decreased the expression of Sost and Dkk1. In situ hybridization demonstrated decreased Sost mRNA expression in osteocytes present in old bone. An abundance of cells expressed Wnt7b in Runx2-positive osteoblasts and ß-catenin in areas with new bone formation. These data demonstrate that inflammation, not only induces osteoclastogenesis, but also locally activates canonical WNT signaling and stimulates new bone formation independent on bone resorption.

Role of Wnt5b-Ror1 signaling in the proliferation of pancreatic ductal adenocarcinoma cells.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most refractory cancers with the worst prognosis. Although several molecules are known to be associated with the progression of PDAC, the molecular mechanisms underlying the progression of PDAC remain largely elusive. The Ror-family receptors, Ror1 and Ror2, which act as a receptor(s) for Wnt-family ligands, particularly Wnt5a, are involved in the progression of various types of cancers. Here, we show that higher expression of Ror1 and Wnt5b, but not Ror2, are associated with poorer prognosis of PDAC patients, and that Ror1 and Wnt5b are expressed highly in a type of PDAC cell lines, PANC-1 cells. Knockdown of either Ror1 or Wnt5b in PANC-1 cells inhibited their proliferation significantly in vitro, and knockout of Ror1 in PANC-1 cells resulted in a significant inhibition of tumor growth in vivo. Furthermore, we show that Wnt5b-Ror1 signaling in PANC-1 cells promotes their proliferation in a cell-autonomous manner by modulating our experimental setting in vitro. Collectively, these findings indicate that Wnt5b-Ror1 signaling might play an important role in the progression of some if not all of PDAC by promoting proliferation.

Significance of the Wnt signaling pathway in coronary artery atherosclerosis.

The progression of coronary atherosclerosis is an active and regulated process. The Wnt signaling pathway is thought to play an active role in the pathogenesis of several cardiovascular diseases; however, a better understanding of this system in atherosclerosis is yet to be unraveled. In this study, real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting were used to quantify the expression of Wnt3a, Wnt5a, and Wnt5b in the human coronary plaque, and immunohistochemistry was used to identify sites of local expression. To determine the pathologic significance of increased Wnt, human vascular smooth muscle cells (vSMCs) were treated with Wnt3a, Wnt5a, and Wnt5b recombinant proteins and assessed for changes in cell differentiation and function. RT-PCR and Western blotting showed a significant increase in the expression of Wnt3a, Wnt5a, Wnt5b, and their receptors in diseased coronary arteries compared with that in non-diseased coronary arteries. Immunohistochemistry revealed an abundant expression of Wnt3a and Wnt5b in diseased coronary arteries, which contrasted with little or no signals in normal coronary arteries. Immunostaining of Wnt3a and Wnt5b was found largely in inflammatory cells and myointimal cells. The treatment of vSMCs with Wnt3a, Wnt5a, and Wnt5b resulted in increased vSMC differentiation, migration, calcification, oxidative stress, and impaired cholesterol handling. This study demonstrates the upregulation of three important members of canonical and non-canonical Wnt signaling pathways and their receptors in coronary atherosclerosis and shows an important role for these molecules in plaque development through increased cellular remodeling and impaired cholesterol handling.

Abstract 6953: Wnt2b deficiency exacerbates colitis and leads to increased tumorigenesis in mice

Abstract Colitis-associated cancer (CAC) is considered the most severe complication of inflammatory bowel disease (IBD). It has been demonstrated that WNT2B deficiency leads to severe intestinal impairment in humans and predisposes to colitis in mice. Evaluating a human cohort from The Cancer Genome Atlas (TCGA), we observed that low expression of WNT2B is associated with poor prognosis of patients with colon cancer. However, the functional roles of WNT2B in CAC development remain unclear, and we hypothesized that WNT2B is involved in the resistance to CAC development during long-term colitis. Thus, we model CAC development using Wnt2b deficient mice subjected to mutagenesis induction with 10 mg/kg Azoxymethane (AOM) and a long-term colitis protocol with three cycles of 2.5% Dextran Sodium Sulfate in the drinking water for three days, followed by a recovery period with autoclaved water. We observed that Wnt2b KO mice had increased susceptibility to colitis and more significant body weight changes than WT mice, especially in the first and second recovery cycles of colitis recovery. The maximum weight loss and disease activity index (DAI) were significantly higher in KO than in WT mice. Interestingly, Wnt2b KO mice developed more tumors, which were more extensive than those observed in WT. Moreover, the histopathological analysis showed that Wnt2b KO had the highest score for ulceration, aberrant crypt foci, and immune cell infiltration. Our data suggest a protective role for WNT2B, conferring resistance to tumor development during colitis. Furthermore, our data provides new insights into the functional roles of Wnt ligands in colon cancer. A better understanding of the factors involved in CAC etiology is critical for improving diagnosis and developing new targeted therapies. We believe WNT2B may emerge as one of these factors and offer novel targets for improved colorectal cancer management. Citation Format: Luiz Fernando Silva Oliveira, Amy E. O'Connell. Wnt2b deficiency exacerbates colitis and leads to increased tumorigenesis in mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6953.

WNT2B activates macrophages via NF-κB signaling pathway in inflammatory bowel disease.

Inflammation is a significant pathological manifestation of inflammatory bowel disease (IBD), yet its mechanism has remained unclear. Although WNT2B is enriched in the intestinal inflammatory tissue of IBD patients, the specific mechanism of WNT2B in the formation of intestinal inflammation remains unclear. This study was aimed to investigate whether macrophages expressing WNT2B can aggravate intestinal tissue inflammation. Samples were collected from both normal individuals and patients with IBD at multiple colon sites. Macrophages were identified using tissue immunofluorescence. IκB kinase (IKK)-interacting protein (IKIP), which interacts with WNT2B, was found by protein cross-linking and protein mass spectrometry. The expression of WNT2B, IKIP, the NF-κB pathway, and downstream molecules were analyzed. An acute colitis model of C57BL/6J mice was established using an adeno-associated virus (AAV)-mediated WNT2B knockdown system and 3% dextran sulfate sodium (DSS). The degree of intestinal inflammation in mice was assessed upon WNT2B knockdown in macrophages. Macrophages expressing WNT2B were found to be enriched in the colitis tissues of IBD patients. WNT2B in macrophages activated the NF-κB pathway and enhanced the expression of downstream inflammatory cytokines. By competitively binding IKIP, WNT2B reduced the binding of IKIP to IKKβ and promoted the activation of the NF-κB pathway. Using an AAV-mediated WNT2B knockdown system, WNT2B expression in intestinal macrophages was suppressed, leading to a reduction in intestinal inflammation. WNT2B activated the NF-κB pathway and enhanced the expression of downstream inflammatory cytokines by competitively binding to IKIP, potentially contributing to colon inflammatory injury in IBD.