TRAF4 enhances oral squamous cell carcinoma cell growth,invasion and migration by Wnt-β-catenin signaling pathway

Oral squamous cell carcinoma (OSCC) ranks as the fifth most common cancer worldwide with poor prognosis. Recently, tumor necrosis factor receptor-associated factor 4 (TRAF4) has attracted increasing attenuation due to its overexpression in certain cancers. However, its function and underlying mechanism in OSCC remains elusive. In this study, the high expression of TRAF4 mRNA and protein levels was noted in OSCC cell lines. Its overexpression with pcDNA3.1-TRAF4 vector transfection dramatically promoted cell proliferation and inhibited cell apoptosis, indicating a pivotal role of TRAF4 in OSCC cell growth. Simultaneously, TRAF4 elevation also increased cell invasion and migration. Mechanism analysis confirmed that TRAF4 up-regulation induced the expression of β-catenin and the downstream target molecules of cyclinD1, c-myc, Bcl-2, MMP-9 and MMP-2, indicating that TRAF4 could induce the activation of Wnt/β-catenin pathway. After pretreatment with β-catenin siRNA, the pathway was remarkably silenced. Simultaneously, cell growth, invasion and migration induced by TRAF4 were strikingly abrogated, suggesting that TRAF4 may promote OSCC cell growth, invasion and migration by Wnt/β-catenin pathway. Together, this study confirmed that TRAF4 acts as an oncogene for the development and progression of OSCC. Therefore, our study may support a promising therapeutic target for the treatment of OSCC.     

BDNF promotes human neural stem cell growth via GSK-3β-mediated crosstalk with the wnt/β-catenin signaling pathway

Brain-derived neurotrophic factor (BDNF) plays important roles in neural stem cell (NSC) growth. In this study, we investigated whether BDNF exerts its neurotrophic effects through the Wnt/β-catenin signaling pathway in human embryonic spinal cord NSCs (hESC-NSCs) in vitro. We found an increase in hESC-NSC growth by BDNF overexpression. Furthermore, expression of Wnt1, Frizzled1 and Dsh was upregulated, whereas GSK-3β expression was downregulated. In contrast, hESC-NSC growth was decreased by BDNF RNA interference. BDNF, Wnt1 and β-catenin components were all downregulated, whereas GSK-3β was upregulated. Next, we treated hESC-NSCs with 6-bromoindirubin-3′-oxime (BIO), a small molecule inhibitor of GSK-3β. BIO reduced the effects of BDNF upregulation/downregulation on the cell number, soma size and differentiation, and suppressed the effect of BDNF modulation on the Wnt signaling pathway. Our findings suggest that BDNF promotes hESC-NSC growth in vitro through crosstalk with the Wnt/β-catenin signaling pathway, and that this interaction may be mediated by GSK-3β.     

LncRNA MINCR activates Wnt/β-catenin signals to promote cell proliferation and migration in oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is a common type of malignant oral cancer with high recurrence. MYC-induced long non-coding RNA (MINCR) has been reported as a tumor suppressor in liver cancer and lung cancer. Whereas, it is unknown whether MINCR exerted function in OSCC progression. This study focused on its function and mechanism in OSCC. At first, the expression level of MINCR in OSCC tissues and cell lines as well as corresponding normal controls was evaluated by qRT-PCR assay. The relative high level of MINCR was observed in OSCC tissues and cell lines. The overall survival rate of OSCC patients with high or low level of MINCR was analyzed by using Kaplan-Meier method. In addition, functional assay revealed that MINCR knockdown significantly suppressed OSCC cell proliferation and invasion. Importantly, the effect of MINCR knockdown on Wnt/β-catenin signaling pathway was detected by luciferase reporter and western blot assays. It was found that MINCR knockdown obviously decreased the activity of Wnt/β-catenin pathway. Rescue assays were further used to validate the role of Wnt/β-catenin pathway in MINCR-mediated OSCC progression. The effects of MINCR knockdown on OSCC cell proliferation and migration were partly reversed by the activator of Wnt/β-catenin pathway (LiCl). Overall, our findings revealed that MINCR may be an oncogene in OSCC via modulation of Wnt/β-catenin pathway.     

Expression of cytokeratin 34βE12 is a good indicator of tumor progression in esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma (ESCC) is considered one of the most aggressive cancers with poor prognosis. The high molecular weight cytokeratin 34βE12 (CK34βE12) is recognized by the antibody, that is expressed in the cytoplasm of epithelial basal cells, and has been considered as a potential marker for prostate cancer, breast cancer, and basaloid carcinoma of the lung. However, there are no clinicopathological studies investigating CK34βE12 expression at the invasive front of ESCC. In this study, we examined 170 surgically resected cases of ESCC to clarify the clinicopathological significance of CK34βE12 expression. CK34βE12 expression was found in 85.3% (145/170) of ESCC cases and was significantly correlated with lymph node metastasis (66.2% [96/145], P = 0.034), depth of tumor invasion (57.9% [84/145], P = 0.042), and differentiation (82.1% [119/145], P = 0.013). These results indicated that CK34βE12 expression is a good indicator of lymph node metastasis, depth of tumor invasion, and differentiation in case of ESCC.     

HPV-16 E6 promotes cell growth of esophageal cancer via downregulation of miR-125b and activation of Wnt/β-catenin signaling pathway

High-risk human papillomavirus (HPV) is a possible cause of esophageal cancer. However, the molecular pathogenesis of HPV-infected esophageal cancer remains unclear. The expression levels of some microRNAs including miR-125b have been negatively correlated with HPV infection, and miR-125b downregulation is associated with tumorigenesis. In addition, Wnt/β-catenin signaling pathway has been suggested to play an important role in esophageal cancer (EC). We examined miR-125b and Wnt/β-catenin signaling pathway in HPV-16 E6 promoted tumor progression in EC. HPV-16 E6 transfection decreased markedly the expression levels of miR-125b and promoted the colony formation in the Eca 109 and Kyse 150 cell lines, and restoration of miR-125b expression level antagonized the increased colony formation in HPV-16 E6 transfected cell lines. We also demonstrated that overexpression of E6 upregulated the Wnt/β-catenin signaling activity via modulating the multiple regulators including TLE1, GSK3β, and sFRP4. Overexpression of miR-125b restored the expression levels of these proteins. Expression of miR-125b was lower in HPV-16 E6 positive esophageal cancer tissues, and was negatively correlated with E6 mRNA levels. Our results indicate that HPV-16 E6 promotes tumorigenesis in EC via down-regulation of miR-125b, and this underlying mechanism may be involved in the activation of the Wnt/β-catenin signaling pathway.     

Down-regulated FSTL5 promotes cell proliferation and survival by affecting Wnt/β-catenin signaling in hepatocellular carcinoma

Follistatin-like 5 (FSTL5), a member of the follistatin family of genes, encodes a secretory glycoprotein. Previous study revealed that it might play a suppressive role in hepatocellular carcinoma (HCC). However, its clinical significances, biological functions and molecular mechanisms in HCC development are poorly understood. To gain insight to the functions of FSTL5 in HCC, We examined FSTL5 expression pattern in 117 HCC tissue samples. The results of immunohistochemical staining analysis showed that FSTL5 is more commonly down-regulated in HCC compared to adjacent tissues and further clinicopathological analysis showed that its expression level is closely correlated with tumor size, TNM stage, local infiltration and patient prognosis. Both gain function assays and recombinant human FSTL5 protein treatment assays in vitro revealed that over-expressing FSTL5 could inhibit the abilities of cancer cell proliferation and survival. Further, we found that those effects on HCC growth and survival are associated with Wnt/β-catenin signaling. Taken together, all of our results validate that FSTL5 plays a suppressive role in HCC and suggest that down-regulated FSTL5 could elevate abilities of growth and survival of HCC cells by activation of Wnt/β-catenin signaling.     

Involvement of Wnt/β-catenin signaling in the development of neuropathic pain

Despite tremendous research effort in the field, our current understanding of the molecular mechanisms underlying neuropathic pain is still incomplete. In the present study, our objective was to elucidate the involvement of the Wnt/β-catenin signaling pathway in the development of neuropathic pain. We showed that Wnt/β-catenin signaling is activated in the spinal cord dorsal horn after partial sciatic nerve ligation (PSL). Expression of Wnt3a, a prototypic Wnt ligand that activates the Wnt/β-catenin pathway, was also upregulated in the dorsal horn. We then tested the effect of intrathecal administration of XAV939, a Wnt/β-catenin signaling inhibitor, and found that this treatment effectively attenuated the induction of neuropathic pain. Conversely, intrathecal administration of Wnt3a to the lumbar spinal cord of naïve animals triggered the development of allodynia. These results suggest a critical involvement of the Wnt/β-catenin pathway in the development of neuropathic pain. Moreover, we also found that PSL-induced microglial activation was significantly suppressed by intrathecal administration of XAV939 treatment. Because it was revealed that Wnt3a treatment triggered brain-derived neurotrophic factor (BDNF) release from microglial cells in vitro, it is possible that Wnt3a upregulation in the dorsal horn leads to the activation of microglial cells, then triggers BDNF secretion that is responsible for the establishment of neuropathic pain. Further studies will be needed for the comprehensive understanding of the roles of Wnt/β-catenin signaling in the development of neuropathic pain.     

Knockout of TLR4 promotes fracture healing by activating Wnt/β-catenin signaling pathway

ObjectivesThe aim of this study was to investigate the effect of Toll like receptor 4 (TLR4) on fracture healing.MethodsThe open tibial fracture models in TLR4 knockout (TLR4^−/−) and wild type (WT) C57BL-6 J mice were established. The radiological examination, tartrate-resistant acid phosphatase (TRAP) staining, Micro-CT scan and biological torsion test were performed on 7, 14 and 21 days after operation. Enzyme Linked Immunosorbent Assay (ELISA) kit was used to detect the expression levels of tumor necrosis factor-α (TNF-α), interleukin-1 beta (IL-1β) and interleukin 6 (IL-6). Western blotting was used to detect the expression of β-catenin, Wingless-type MMTV integration site family, member 4 and 5B (Wnt4 and Wnt5B), proliferating cell nuclear antigen (PCNA) and bone morphogenetic protein-2 (BMP-2) of the callus tissue obtained from mice.ResultsTLR4 knockout promoted fracture healing, reduced the number of osteoclasts, increased bone callus volume (BV) and callus mineralized volume fraction (BV/TV%) (P < 0.05), increased the maximum torque and torsional stiffness of callus (P < 0.05), reduced TNF-α, IL-1β and IL-6 expression (P < 0.01), and increased the expression levels of β-catenin, Wnt4, Wnt5B, PCNA and BMP-2 (P < 0.01).ConclusionTLR4 knockout reduced inflammatory and promoted fracture healing by activating Wnt/β-catenin signaling pathway.     

A novel oncogene TRIM63 promotes cell proliferation and migration via activating Wnt/β-catenin signaling pathway in breast cancer

The development of breast cancer is still a relatively unclear biological process, and there is currently no consensus on the occurrence of breast cancer and the process of tumor metastases. This study was to reveal a correlation between TRIM63 and the development of breast cancer. In this study, we found that the expression of TRIM63 was significantly increased in breast cancer tissues and closely related to pathological differentiation and TNM stage of breast cancer. Overexpression of TRIM63 could significantly promote proliferation and migration of breast cancer cells, while TRIM63 knockdown significantly inhibited the proliferation and migration of breast cancer cells. In addition, TRIM63 could activate Wnt/β-catenin signaling pathway in breast cancer cells. Further study found that TRIM63 could regulate β-catenin degradation by promoting GSK3β phosphorylation. Our study revealed that TRIM63, as an oncogene, involved in breast cancer progression by activating the Wnt/β-catenin signaling pathway, suggesting that the potential applicability of TRIM63 as a target for breast cancer treatment.     

Knockdown of eIF3D inhibits breast cancer cell proliferation and invasion through suppressing the Wnt/β-catenin signaling pathway

eIF3D (eukaryotic translation initiation factor 3 subunit D) is one member of the eIF3 family and plays a critical role in translation initiation. Previous studies showed that it was involved in the development and progression of several tumors. However, the role of eIF3D in breast cancer and the underlying mechanism is still unclear. Therefore, this study set out to investigate the role of eIF3D in breast cancer. Our results demonstrated that eIF3D is up-regulated in breast cancer cells. Knockdown of eIF3D inhibited breast cancer cell proliferation and invasion. In addition, knockdown of eIF3D inhibited the expression of β-catenin, cyclin D1 and c-Myc in breast cancer cells. Taken together, our findings show that siRNA-eIF3D inhibits breast cancer cell proliferation and invasion through suppressing the Wnt/β-catenin signaling pathway. Therefore, eIF3D may be a good molecular target for the prevention and treatment of breast cancer.     

LncRNA CASC7 inhibits the progression of glioma via regulating Wnt/β-catenin signaling pathway

Increasing evidence reveal the important role of long non-coding RNAs (lncRNAs) in the initiation and progression of glioma. However, the role of lncRNA cancer susceptibility candidate 7 (CASC7) in glioma is largely unknown. At first, the expression level of CASC7 was tested in glioma tissues and cell lines by using qRT-PCR. We applied Kaplan-Meier method to analyze the correlation between the expression level between CASC7 expression and the overall survival rate of glioma patients. We found that CASC7 was downregulated in glioma tissues and cell liens and predicted poor prognosis for patients with glioma. To determine the involvement of CASC7 in the biological processes of glioma, we conducted gain or loss-of function assays in two glioma patients. We found that CASC7 suppressed glioma cell proliferation and induced glioma cell apoptosis. Mechanistically, the expression level of CASC7 was negatively correlated with the expression levels of core factors of Wnt/β-catenin signaling pathway in glioma cells. Moreover, TOP flash luciferase activity further revealed the negative effect of CASC7 on the activity of Wnt/β-catenin signaling pathway. Finally, rescue assays were carried out to determine that Wnt/β-catenin signaling pathway involved in CASC7-mediated glioma progression. Taken together, all research findings suggested that CASC7 inhibited the progression of glioma via regulating Wnt/β-catenin signaling pathway.     

Significance of E-cadherin, β-catenin, and vimentin expression as postoperative prognosis indicators in cervical squamous cell carcinoma

Although early-stage cervical cancer can be treated by surgery, distant metastases can be life threatening. It has been a challenge to identify reliable biomarkers as indicators of metastasis or poor prognosis. We investigated the prognostic impact of vimentin, E-cadherin, and β-catenin expression measured by immunohistochemistry staining in samples from 135 patients with clinical stage I or II cervical squamous cell cancer and in normal cervical tissues from 55 patients who underwent hysterectomy for reasons other than neoplasia. Down-regulation of E-cadherin and β-catenin was positively related to histologic differentiation (P < .001), metastasis (P < .001), and recurrence (P < .001), whereas up-regulation of vimentin was inversely related to histologic differentiation, metastasis, and recurrence (P < .0001, .020, and .000, respectively). In univariate Cox regression analysis, high expression of E-cadherin or β-catenin was a positive prognostic indicator for overall survival (P < .001 and P < .001, respectively), whereas high expression of vimentin was a negative indicator (P < .001). In multivariate Cox regression analysis, high expression of E-cadherin was a positive prognostic indicator for overall survival (P = .002), whereas high expression of vimentin was a negative indicator (P = .034). The expression of E-cadherin and vimentin was associated with survival, and the 2 proteins were independent prognostic factors in univariate and multivariate analyses. The combination of a decrease of E-cadherin and an increase in vimentin might be a valuable survival indictor in cervical squamous cell cancer.