Expression Levels Of Notch1 Research Articles

Downregulation of SNHG1 suppresses cell proliferation and invasion by regulating Notch signaling pathway in esophageal squamous cell cancer.

Long non-coding RNAs (lncRNAs) exert important functions involved in tumorigenesis and cancer progression including esophageal squamous cell cancer (ESCC), however, the clinical role and underlying biological function of Small Nucleolar RNA Host Gene 1 (SNHG1) in ESCC is not well known. Quantitative Real-time polymerase chain reaction (QRT-PCR) was used to detect the SNHG1 expression levels in ESCC tissues and adjacent non-cancerous tissues. Chi-square test was used to evaluate the association between clinicopathological features and SNHG1 expression in ESCC patients, Kaplan-Meier curve and log rank test was performed to analyze the association between overall survival and SNHG1 expression. Cell proliferation and invasion was assessed by MTT assay, colony formation, and transwell cell invasion assays. Western blot were also performed to examine protein expression levels of E-cadherin, Vimentin and N-cadherin, Notch 1 and Hes-1. We demonstrated that lncRNA SNHG1 was significantly up-regulated in ESCC tissues compared with adjacent non-cancerous tissues in ESCC patients. Meanwhile, increased lncRNA SNHG1 expression levels markedly correlated with lymph node metastasis, depth of invasion, TNM stage and reduced over survival time in ESCC patients. Furthermore, MTT assay, colony formation, transwell cell invasion, qRT-PCR and Western-blot assays demonstrated that knockdown of lncRNA SNHG1 could inhibit cell proliferation and cell invasion capacity and cell Epithelial-Mesenchymal Transition (EMT) phenomenon by up-regulation E-cadherin and down-regulating Vimentin and N-cadherin in ESCC cells. Besides, we demonstrated that knockdown of SNHG1 suppressed the Notch signaling pathway by reducing the Notch1 and Hes-1 expression levels in ESCC cells. These results indicated that lncRNA SNHG1 may be a potential predictor of prognosis in ESCC patients and a novel target for ESCC treatment.

Longitudinal study of esophageal mucosal damage after esophagectomy and gastric interposition: relationship between reflux-related mucosal injury and Notch signaling.

Esophagectomy with gastric interposition could serve as a good human reflux model to study the molecular pathogenesis of esophageal mucosal damage induced by gastroesophageal reflux. This study was to investigate the role of Notch signaling in reflux injury of esophageal mucosa. Patients undergoing Ivor-Lewis esophagectomy for early stage esophageal squamous cell carcinoma were included. Follow-ups were scheduled at 6, 18, 36 and 48 months postoperatively, including reflux symptom assessment, endoscopic and histological evaluation of esophageal mucosal damage. The expressions of Notch1 and its downstream target gene Hes1 were evaluated by real-time polymerase chain reaction (PCR) and immunohistochemistry (IHC). Forty-four out of 48 patients completed four follow-ups. Injuries of esophageal remnant confirmed by endoscopical and histological examinations were both more often with a longer postoperative period (P<0.05). The mRNA expression levels of Notch1 and Hes1 were decreased in a time-dependent manner after operation (P<0.001). Notch1 and Hes1 mRNA levels were significantly higher in normal squamous mucosa than in esophagitis, and higher in esophagitis than in metaplasia (P<0.05). Immunohistochemical study also demonstrated a similar protein expression pattern. Samples with endoscopic evidence of mucosal damage exhibited lower expression of Notch1 mRNA levels as compared to biopsies without visualized damage (P=0.035). This is the first longitudinal study on Notch signaling in human esophagectomy model, our preliminary findings suggest decreased Notch signaling might be involved in the development of mucosa damage caused by gastroesophageal reflux.

Interference of Notch 1 inhibits the proliferation and invasion of breast cancer cells: Involvement of the β‑catenin signaling pathway.

Breast cancer is one of the most common types of carcinoma in humans. The aim of the present study was to identify the role of Notch 1 in the proliferation and invasion of human breast cancer cells. Firstly, the levels of Notch 1 were determined by western blot analysis in breast cancer cell lines, and the results revealed that the expression levels of Notch 1 were markedly higher in MDA‑MB‑231 and MCF‑7 cells, and lower in MCF‑10A cells, compared with human mammary epithelial cells. An MTT assay was used to determine the viability of breast cancer cells. The optical density (OD)490 values were significantly decreased in Notch 1 short hairpin (sh)RNA‑transfected MCF‑7 and MDA‑MB‑231 cells, compared with the OD490 values in the negative control shRNA‑transfected cells. The MCF‑7 cells and MDA‑MB‑231 cells were also treated with increasing concentrations of MRK003, a Notch 1 inhibitor, for 24, 48 and 72h, respectively. The inhibition rate was gradually increased in the MRK003‑treated cells in a time‑ and dose‑dependent manner. The invasive ability of the cells was determined using a Transwell migration assay. The migration ability was significantly decreased in the Notch 1‑transfected MCF‑7 cells and MDA‑MB‑231 cells. The molecular mechanism was examined, and the knockdown of Notch 1 significantly decreased the expression levels of β‑catenin, matrix metalloproteinase (MMP)‑2 and MMP‑9, and was also correlated with the downregulation of β‑catenin in the nucleus. In conclusion, Notch1 was key in the progression of breast cancer, and knocking down the expression of Notch 1 significantly suppressed the proliferation and invasion of breast cancer cells. This provides novel clues for cancer therapy in human breast cancer.

Progressive and Prognosis Value of Notch Receptors and Ligands in Hepatocellular Carcinoma: A Systematic Review and Meta-analysis

Hepatocellular carcinoma (HCC) is not sensitive to radiotherapy and chemotherapy and experiences postoperative relapse extremely easy, which is the major cause of the high mortality rate. The Notch signaling pathway is expected to become a new target for the biological treatment of HCC. We searched databases for studies that evaluated the expression of Notch receptors and/or ligands in human HCC tissue. The search yielded 15 studies that enrolled 1643 patients. Compared with non-HCC tissues, Notch 1 was associated with a higher expression level (odds risk 1.59, 95% confidence interval 0.34 to 7.45), as well as Notch 3 (2.63, 0.69 to 10.02), Notch 4 (1.33, 0.74 to 2.38) and Jagged 1 (1.47, 0.23 to 9.53); however, Notch 2 showed the opposite result (0.60, 0.30 to 1.20). Larger tumor size (>5 cm), metastasis positive, and micro vascular invasion positive were features that were associated with over-expression in Notch 1 according to the clinicopathological features. The expression levels of Notch 1, 3, 4 and Jagged 1 were associated with higher expression in HCC tissues, while Notch 2 had the opposite result. This study is registered with PROSPERO (CRD42017055782).

Hematopoietic stem cell-derived exosomes promote hematopoietic differentiation of mouse embryonic stem cells in vitro via inhibiting the miR126/Notch1 pathway.

Cell-derived exosomes (EXs) can modulate target cell differentiation via microRNAs (miRs) that they carried. Previous studies have shown that miR126 is highly expressed in hematopoietic stem cells (HSCs) and plays a role in hematopoiesis via modulating the Notch pathway that participates in progenitors' cell fate decisions. In this study we investigated whether HSC-derived EXs (HSC-EXs) could affect the differentiation of mouse embryonic stem cells (ESCs) into HSCs. We prepared HSC-EXscon, HSC-EXssc and HSC-EXsmiR126 from control HSCs and the HSCs transfected with scramble control or miR126 mimics, respectively. HSC-EXs were isolated by ultracentrifugation and analyzed using nanoparticle tracking analysis. We incubated the collected EXs with mouse ESCs over a 10-d differentiation induction period, during which HSC-EXs and a Notch pathway activator (Jagged1, 100 ng/mL) were added to the cultures every 3 d. After the 10-d differentiation period, the expression levels of miR126, SSEA1, CD117, Sca1, Notch1 and Hes1 in ESCs were assessed. The generated HSCs were validated by flow cytometry using antibodies against HSC markers (CD117, CD34 and Sca1). Our results revealed that: (1) transfection with miR126 mimics significantly increased miR126 levels in HSC-EXsmiR126. (2) HSC-EX co-culture promoted mouse ESCs differentiation into HSCs with the most prominent effect found in the HSC-EXsmiR126 co-culture. (3) HSC differentiation was verified by reduced SSEA1 expression and increased CD117 and Sca1 expression. (4) All the effects caused by HSC-EXs were accompanied by significant reduction of Notch1 and Hes1 expression, thus inhibition of the Notch1/Hes1 pathway, whereas activation of Notch by Jagged1 abolished the effects of HSC-EXsmiR126. In conclusion, HSC-EXs promote hematopoietic differentiation of mouse ESCs in vitro by inhibiting the miR126/Notch1 pathway.

MicroRNA‑455 inhibits cell proliferation and invasion of epithelial ovarian cancer by directly targeting Notch1.

Ovarian cancer (OC) is the fifth leading cause of cancer‑associated mortality among women in developed countries. Numerous microRNAs are aberrantly expressed in epithelial (E)OC, and are involved in EOC formation and progression. As such, microRNAs may be investigated as diagnostic and prognostic molecular biomarkers, and therapeutic targets for patients with EOC. For instance, microRNA‑455 (miR‑455) is abnormally expressed in various types of human cancer. However, details on the expression level, biological roles and underlying mechanism of miR‑455 in EOC remain unclear. In the present study, the expression of miR‑455 in EOC was detected, and its association with clinical characteristics was analysed. The functional roles and underlying mechanisms of miR‑455 in EOC were also investigated. The results revealed that miR‑455 expression in EOC tissues and cell lines was significantly downregulated. Furthermore, miR‑455 downregulation was correlated with tumor size, International Federation of Gynecology and Obstetrics stage and lymph node metastasis of patients with EOC. Restoration of miR‑455 expression efficiently inhibited cell proliferation and invasion invitro. Notch1 was identified as a direct target of miR‑455 in EOC. The mRNA and protein expression levels of Notch1 were higher in EOC tissues compared with in normal ovarian epithelial tissues. Spearman's correlation analysis indicated that miR‑455 expression was negatively correlated with the mRNA level of Notch1 in EOC tissues. Notch1 overexpression was able to restore the EOC cell proliferation and invasion abilities suppressed by miR‑455. The present study provided evidence that the dysregulation of the miR‑455/Notch1 signalling pathway may be essential for EOC occurrence and development. In addition, the results confirmed the tumor‑suppressive roles of miR‑455 in modulating EOC proliferation and invasion through regulation of Notch1 expression. Thus, miR‑455 may be a novel miRNA‑based therapeutic target to treat patients with EOC.

Potential role of the Jagged1/Notch1 signaling pathway in the endothelial-myofibroblast transition during BLM-induced pulmonary fibrosis.

Endothelial cell myofibroblast transition (EndoMT) is found during the process of bleomycin (BLM)-induced pulmonary fibrosis in rats, and plays a very important role in sustaining inflammation and collagen secretion. Moreover, some studies have suggested that the Notch1 signaling pathway may be involved in the expression of α-smooth muscle actin (α-SMA) in pulmonary microvascular endothelial cells (PMVECs), a protein marker of EndoMT. Therefore, we aimed to investigate the expression level of α-SMA and Notch1-related signaling molecules in PMVECs from BLM-induced rats and determine the relationship between the Notch1 signaling pathway and the expression of α-SMA in PMVECs. We found that the expression levels of α-SMA, Notch1, and Jagged1 were upregulated, while the expression levels of Dll4 were downregulated. Furthermore, there was a positive correlation between the expression of Jagged1 and the α-SMA proteins in PMVECs, and NF-κB was downregulated by decreasing the expression of Jagged1. In conclusion, the Jagged1/Notch1 signaling pathway is activated in PMVECs during the pathogenesis of BLM-induced pulmonary fibrosis in rats, and it may induce α-SMA expression via a non-canonical pathway involving NF-κB as the target molecule. The precise mechanism and the molecules involved in this signaling pathway need to be further elucidated.

Expression and association of VEGF-Notch pathways in infantile hemangiomas.

The vascular endothelial growth factor (VEGF) and Notch signaling pathways have been identified to be involved in the neovascularization, and angiogenesis of various tumor types. However, there is little data regarding their roles and association in infantile hemangiomas (IHs). In the present study, the significance and association of the VEGF-VEGF receptor (R)2, and δ like canonical Notch ligand 4 (Dll4)-Notch1 pathways in different clinical phases of IHs were investigated. Specimens in the proliferating phase (n=15) and involuting phase (n=12) were collected. Quantitative polymerase chain reaction was used to analyze the mRNA levels of VEGF, VEGFR2, Dll4 and Notch1. A further 61 paraffin-embedded IHs (26 in the proliferating group and 35 in the involuting group) specimens were collected to investigate the protein levels of Notch1 and Dll4 using immunohistochemistry, and then analyzed for the association between these factors and microvessel density (MVD). The relative expression levels of VEGF, VEGFR2 and Dll4 mRNA in the proliferating group were significantly higher compared with that in the involuting group. In addition, the relative levels of Notch1 mRNA were similar in the proliferating and involuting phases. Expression levels of VEGFR2 and Dll4 mRNA were positively correlated with Notch1 expression in the proliferating IHs. The relative expression of VEGFR mRNA was positively correlated with Dll4 in the proliferating and involuting groups. Immunohistochemical staining demonstrated that Notch1 and Dll4 protein were upregulated in the proliferating IHs compared with the control group. In the proliferating IHs, Notch1 and Dll4 protein expression levels were positively correlated with MVD, while in the involuting phase, only Dll4 was positively correlated with MVD. The expression levels of related factors in the VEGF and Notch pathways, and the associations among them suggest that they may be involved in the regulation of IH neovascularization and angiogenesis.

Disorder of the mevalonate pathway inhibits calcium-induced differentiation of keratinocytes.

Mutation of genes encoding the enzymes of the mevalonate pathway cause a variety of diseases, including skin disorders. Mutation of four genes in this pathway, including mevalonate kinase, phosphomevalonate kinase, mevalonate diphosphate decarboxylase and farnesyl diphosphate synthase, have demonstrated to be responsible for porokeratosis (PK). However, the pathogenesis of PK remains unclear. In the present study, specific enzyme inhibitors of the mevalonate pathway, including pravastatin (PRA), alendronate (ALD), farnesyl transferase inhibitor (FTI‑277) and geranylgeranyl transferase inhibitor (GGTI‑298), were used to investigate the effect on differentiation of keratinocytes (KCs). Western blotting demonstrated that PRA, ALD, FTI‑277 or GGTI‑298 alone, or in combination, inhibited the expression level of calcium‑induced differentiation maker involucrin (INV) in KCs. ALD and PRA induced greater inhibition of INV compared with FTI‑277 and GGTI‑298 treatment. These inhibitors additionally influenced the expression levels of keratin1. Mechanistic studies revealed that treatment of cells with inhibitors decreased the expression levels of p53 and Notch1, and regulated activation of the mitogen activated protein kinase and phosphoinositide‑3‑kinase/protein kinase B signaling pathways. The results of the present study suggested that regulation of the mevalonate pathway may be necessary for differentiation of KCs, and the pathogenesis of disseminated superficial actinic PK.

A combination of valproic acid sodium salt, CHIR99021, E-616452, tranylcypromine, and 3-Deazaneplanocin A causes stem cell-like characteristics in cancer cells.

Many studies are based on the hypothesis that recurrence and drug resistance in lung carcinoma are due to a subpopulation of cancer stem-like cells (CSLCs) in solid tumors. Therefore it is crucial to screen for and recognize lung CSLCs. In this study, we stimulated non-small cell lung cancer (NSCLC) A549 cells to display stem cell-like characteristics using a combination of five small molecule compounds. The putative A549 stem cells activated an important CSLC marker, CD133 protein, as well multiple CSLC-related genes including ATP-binding cassette transporter G2 (ABCG2), C-X-C chemokine receptor type 4 (CXCR4), NESTIN, and BMI1. The A549 stem-like cells displayed resistance to the chemotherapeutic drugs etoposide and cisplatin, epithelial-to-mesenchymal transition properties, and increased protein expression levels of NOTCH1 and Hes Family bHLH Transcription Factor 1 (HES1). When A549 cells were pretreated with a NOTCH signaling pathway inhibitor before compound induction, expression of the NOTCH1 target gene HES1 was reduced. This demonstrated that the NOTCH signaling pathway in the putative A549 stem-like cells had been activated. Together, the results of our study showed that a combination of five small molecule agents could transform A549 cells into putative stem-like cells, and that these compounds could also elevate CD133 and ABCG2 protein expression levels in H460 cells. This study provides a convenient method for obtaining lung CSLCs, which may be an effective strategy for developing lung carcinoma treatments.

The ratio of 1/3 linoleic acid to alpha linolenic acid is optimal for oligodendrogenesis of embryonic neural stem cells

During neural development, embryonic neural stem cells (eNSCs) differentiate toward glial, oligodendrocytic, and neuronal cells. Dysregulation of polyunsaturated fatty acids (PUFAs) induce a wide range of neurological and developmental disorders. In this study, we investigated the effect of various concentrations and ratios of linoleic acid (LA) and alpha linolenic acid (ALA), which belong respectively to omega-6 and omega-3 PUFAs, on the proliferation and differentiation of eNSCs.Results showed that low (25 and 50μM) or high (100 and 200μM) concentrations of ALA, but not LA, and the ratio of 1:3 of LA/ALA significantly increased neurospheres size, frequency and cell numbers, in comparison to controls. Moreover, low or high concentrations of ALA, but not LA, and different ratios of LA/ALA resulted in a significant increase in mRNA expression levels of Notch1, Hes1 and Ki-67, and the differentiation of eNSCs toward astrocytes (GFAP) and oligodendrocytes (MBP), but not neurons (β-III Tubulin), with the highest increase being for LA/ALA ratio of 1:3, in comparison to controls. These results demonstrate the importance of higher concentrations of ALA in enhancing proliferation and differentiation of eNSCs, which could be used in diet to help preventing neurodevelopmental syndromes, cognitive decline during aging, and various psychiatric disorders.

Effects of RUNX3 mediated Notch signaling pathway on biological characteristics of colorectal cancer cells.

This study investigated the effects of runt-related transcription factor3 (RUNX3) mediated Notch pathway on the biological behavior of colorectal cancer (CRC) SW260 cells. CRC tissues and para-carcinoma tissues were collected from 182 CRC patients who had undergone surgical treatment between January2008 and December2010. Immunohistochemical staining with streptavidin-peroxidase (SP) was used to detect RUNX3, Notch1 and Jagged1 expression levels. CRC SW260 cells were divided into the following groups: Control group, si-NC group, si-RUNX3 group, DAPT group, si-RUNX3+DAPT group, and si-NC+DAPT group. Expression levels of RUNX3, and Notch signaling related genes were measured by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and western blotting invitro. Besides, MTT, soft agar colony formation, AnnexinV-FITC/PI double staining and Transwell were performed to analyze the effects of RUNX3 on cell growth and metastasis. Lower positive expression rate of RUNX3 and higher positive expression rate of Notch1 and Jagged1 were observed in CRC tissues than those in normal adjacent tissues with a negative correlation, and the expression levels were associated with the differentiation degree, TNM staging, lymph node metastasis and tumor invasion depth (all P<0.05). RUNX3 expression was reduced in si-RUNX3 and si-RUNX3+DAPT group but the expression levels of Notch signaling related genes were markedly increased in si-RUNX3 group or decreased in DAPT and si-NC+DAPT group, as compared with those in the control group (all P<0.05). In addition, the proliferation, colony formation, migration and invasion abilities of SW260 cells were enhanced in si-RUNX3 group but were restricted in DAPT and si-NC+DAPT group, which was contrary to cell apoptosis (allP<0.05). RUNX3 contributes to attenuate the proliferation and metastasis of CRC cells, and promotes cell apoptosis through inhibition of Notch signaling pathway.

Interferon regulatory factor 4 (IRF4) is overexpressed in human non‑small cell lung cancer (NSCLC) and activates the Notch signaling pathway.

The transcription factor, interferon regulatory factor 4 (IRF4), serves an essential role in the regulation of immune responses, and has been reported to act as a diagnostic and prognostic marker for various hematological malignancies. The present study aimed to investigate whether IRF4 could exert effects on human non-small cell lung cancer (NSCLC) and to explore the underlying mechanism. The mRNA and protein expression of IRF4 was detected in NSCLC tissues using reverse-transcription quantitative polymerase chain reaction and western blotting, respectively. In the in vitro experiment, IRF4 expression was knocked down or overexpressed using lentivirus in human lung adenocarcinoma A549 and lung squamous cell carcinoma LC-AI cell lines. Cell proliferation and colony number were analyzed using MTT and colony formation assays, respectively. The expression levels of IRF4 mRNA and protein were significantly higher in NSCLC tissues (n=54) compared with that in adjacent non-tumor tissues. Similarly, the expression levels of Notch1 and Notch2 mRNA were significantly higher in NSCLC tissues. Furthermore, the expression level of IRF4 mRNA was positively correlated with the levels of Notch1 and Notch2 mRNA in NSCLC tissues. Consequently, using NSCLC cell lines, it was demonstrated that the knockdown of IRF4 expression significantly reduced the cell proliferation rate and colony formation, whereas IRF4-overexpression significantly increased them. Notably, the IRF4 knockdown significantly decreased the expression levels of Notch1 and Notch2 mRNA, and phosphorylated protein kinase B (AKT), whereas IRF4 overexpression resulted in the opposite. The results of the present study indicate that IRF4 is overexpressed and serves as a tumor promoter in human NSCLC, at least partially, through activating the Notch-Akt signaling pathway.

Inhibition of Notch1 protects against IL-1β-induced inflammation and cartilage destruction in temporomandibular chondrocytes.

Temporomandibular disorder (TMD) is a complex and multifactorial disease, with inflammatory response and cartilage destruction usually associated with its etiology. Notch1 is a type I transmembrane receptor, which is critical in determining the growth, differentiation and survival of various cell types. However, its role and mechanism in interleukin (IL)‑1β‑stimulated temporomandibular chondrocytes remain to be elucidated. In the present study, chondrocytes were isolated from Sprague‑Dawley rats and stimulated by IL‑1β at a concentration of 10ng/ml, and the expression of Notch1 was inhibited by small interfering RNA. The expression levels of Notch1 and Notch1 intracellular domain (NICD) were analyzed using reverse transcription‑quantitative polymerase chain reaction and western blot analyses. Matrix metalloproteinases (MMPs) and the tissue inhibitor of metalloproteinases (TIMPs) were assessed using western blot analysis, and nuclear factor‑κB (NF‑κB) was also analyzed. Compared with the control, IL‑1β stimulation increased the expression of Notch1 and NICD. In addition, IL‑1β stimulation increased the secretion of tumor necrosis factor‑α and IL‑6, and the expression of intercellular adhesion molecule 1 and inducible nitric oxide synthase. By contrast, inhibition of Notch1 reduced these inflammatory responses by suppressing the nuclear translocation and phosphorylation of NF‑κB p65. The inhibition of Notch1 also reduced the expression of MMPs and increased the expression of TIMP‑1. These data suggested that the inhibition of Notch1 prevented the IL‑1β‑induced inflammatory response partly by inhibiting the NF‑кB signaling pathway in temporomandibular chondrocytes. The inhibition of Notch1 suppressed cartilage destruction by modulating the balance between MMPs and TIMP‑1, therefore, inhibition of Notch1 may be a mechanism for the treatment of TMD.

NANOG overexpression and its correlation with stem cell and differentiation markers in meningiomas of different WHO grades.

NANOG, as a key regulator of pluripotency and acting synergistically with other factors, has been described as a crucial transcription factor in various types of cancer. In meningiomas the expression of this marker has not yet been described. With our study, we aimed to identify and localize NANOG and other possible markers of pluripotency, stem cell properties and differentiation in meningioma tissue, to elucidate a possible effect on tumorigenesis. The gene expression levels of NANOG (NANOG1 and NANOGP8), SOX2, OCT4, KLF4, ABCG2, CMYC, MSI1, CD44, NOTCH1, NES, SALL4B, TP53, and EPAS1 were quantitatively examined using RT-qPCR in 33 surgical specimens of low- (WHO grade I) as well as in high-grade (WHO grade II/III) meningiomas with dural tissue as reference. Immunofluorescence co-localization analysis following confocal fluorescence microscopy for NANOG, OCT4, SOX2, Nestin, KI-67, and CD44 was also performed. There was a significant overexpression of NANOG, MSI1, and EPAS1 and a downregulation of NES in all examined tumors. Subgroup analysis (WHO grade I versus grade II/III) revealed differences in the expression of NANOG, CD44, and MSI1. We found 1% NANOG-positive (NANOG+) cells in low-grade and 2% in grade II/III meningiomas co-expressing the other mentioned markers in various compositions. In particular, NANOG+ cells expressing SOX2 and OCT4 were successfully identified (26% low-grade versus 20% high-grade). Our data reveal an overexpression of NANOG and other markers of pluripotency and stemness in meningiomas. Such potentially pluripotent "stem cell-like" cells may have an impact on tumorigenesis and progression in human meningiomas.

Human cytomegalovirus infection promotes the stemness of U251 glioma cells.

Glioblastoma (GBM) are the most common and aggressive tumors of human brain. Recent studies showed that human cytomegalovirus (HCMV) can induce malignant transformation of tumor cells to maintain stemness. Transcription factor 5 (ATF5) is an anti-apoptotic protein that is highly expressed in malignant glioma. The aim of this study is to investigate the effect of HCMV infection on the stem cell makers of U251 cells. U251 cells were infected by AD169 HCMV strain (MOI = 1). The expression of stem cell makers (CD133, NES, Notch1) in infected U251 cells were compared with the expression in uninfected U251 cell to see the difference between them. Then, the changes of cell proliferation activity and the expression level of Notch intracellular domain (NICD), Notch1, ATF5, and IE protein were detected in the infected cells, and the expressions of Notch1 and NICD were increased. Cell proliferation assay showed that HCMV infection significantly increased the proliferation. These cells could form tumor spheres in non-adherent conditions. Consistent with these findings, the effect of silencing ATF5 on the proliferation of HCMV-infected U251 cells was also examined. The result shows that short interfering RNA-mediated ATF5 downregulation inhibited this process. These findings imply that HCMV infection may regulate ATF5 signaling pathway to increase cell malignant traits and maintain stemness. J. Med. Virol. 89:878-886, 2017. © 2016 Wiley Periodicals, Inc.

Prognostic roles of mRNA expression of notch receptors in non-small cell lung cancer.

Notch signalling is aberrantly activated in human non-small cell lung cancer (NSCLC). Nevertheless, the prognostic roles of mRNA expression of four Notch receptors in NSCLC patients remain elusive. In this report, we reported the prognostic roles of Notch receptors in a total of 1,926 NSCLC patients through “The Kaplan-Meier plotter” (KM plotter) database which is capable to assess the effect of 22,277 genes on survival of NSCLC patients. We found that mRNA high expression level of Notch1 was associated with better overall survival (OS) for all NSCLC patients, hazard ratio (HR) 0.78 (0.69-0.89), p=0.00019, better OS in adenocarcinoma (Ade) patients, HR 0.59 (0.46-0.75), p=1.5e-05, as well as in squamous cell carcinoma (SCC) patients, HR 0.78 (0.62-0.99), p=0.044. mRNA high expression levels of Notch2 and Notch3 were associated with worsen OS for all NSCLC patients, as well as in Ade, but not in SCC patients. mRNA high expression level of Notch4 was not found to be associated with to OS for all NSCLC patients. In addition, mRNA high expression levels of Notch2, Notch3, but Notch4 are significantly associated with the NSCLC patients who have different smoking status. Our results indicate that mRNA expression of Notch receptors may have distinct prognostic values in NSCLC patients. These results will benefit for developing tools to accurately predict the prognosis of NSCLC patients.

Dihydromyricetin-mediated inhibition of the Notch1 pathway induces apoptosis in QGY7701 and HepG2 hepatoma cells.

AIMTo investigate whether Dihydromyricetin (DHM) inhibits cell proliferation and promotes apoptosis by downregulating Notch1 expression.METHODSThe correlation between Notch1 and Hes1 (a Notch1 target molecule) expression in hepatoma samples was confirmed by qRT-PCR. In addition, MTT assays, flow cytometry and TUNEL analysis showed that DHM possessed strong anti-tumor properties, evidenced not only by reduced cell proliferation but also by enhanced apoptosis in QGY7701 and HepG2 hepatocellular carcinoma (HCC) cells. The expressions of Notch1, Hes1, Bcl-2 and Bax were determined by Western blot.RESULTSAmong the tested samples (n = 64), the expression levels of Notch1 (75% of patients) and Hes1 (79.7% of patients) mRNA in tumor tissues were higher than in the normal liver tissues. There was a negative correlation between the expression of Notch1 and the degree of differentiation and positively correlated with the Alpha Fetal Protein concentration. The viability of HCC cells treated with DHM was significantly inhibited in a dose and time-dependent manner. Apoptosis was induced in HepG2 and QGY7701 cell lines following 24 h of DHM treatment. After treatment with DHM, the protein expression of Notch1 was downregulated, the apoptosis-related protein Bax was upregulated and Bcl2 was downregulated. Notch1 siRNA further enhanced the anti-tumor properties of DHM.CONCLUSIONNotch1 is involved in the development of HCC and DHM inhibits cell proliferation and promotes apoptosis by down-regulating the expression of Notch1.

KLF2 inhibits cell growth via regulating HIF-1α/Notch-1 signal pathway in human colorectal cancer HCT116 cells.

The transcription factor Krüppel-like factor 2 (KLF2) has been shown to function as a tumor suppressor and regulate biological processes of cancer cells, such as cell growth, cell apoptosis and angiogenesis. However, the function and mechanism of KLF2 in colorectal cancer (CRC) is still unknown. In the present study, we show that the expression of KLF2 is diminished in a cohort of CRC cell lines. Also, KLF2 overexpression remarkably inhibits HCT116 and SW480 cell survival and proliferation. Moreover, cell death detection ELISA plus assay showed that KLF2 overexpression increased HCT116 cell proliferation. Caspase-3/7 activity also increased in HCT116 cells transfected with PcDNA3.1-KLF2. Further studies showed that KLF2 significantly suppresses the expression of Notch-1 and is dependent on the decline of the HIF-1α level. Most importantly, silencing Notch-1 expression or HIF-1α level both impair the action of KLF2 overexpression in CRC cells. Collectively, we demonstrated that KLF2 mediates CRC cell biological processes including cell growth and apoptosis via regulating the HIF-1α/Notch-1 signal pathway. These results indicated that KLF2 plays an important role in CRC and provided novel insight on the function of KLF2 in tumor progression.

Endosulfan inhibits proliferation through the Notch signaling pathway in human umbilical vein endothelial cells

Our previous research showed that endosulfan triggers the extrinsic coagulation pathway by damaging endothelial cells and causes hypercoagulation of blood. To identify the mechanism of endosulfan-impaired endothelial cells, we treated human umbilical vein endothelial cells (HUVECs) with different concentrations of endosulfan, with and without an inhibitor for Notch, N-[N-(3, 5-difluorophenacetyl)-1-alanyl]S-Phenylglycinet-butylester (DAPT, 20 μM), or a reactive oxygen species (ROS) scavenger, N-Acetyl-l-cysteine (NAC, 3 mM), for 24 h. The results showed that endosulfan could inhibit cell viability/proliferation by increasing the release of lactate dehydrogenase (LDH), arresting the cell cycle in both S and G2/M phases, and inducing apoptosis in HUVECs. We also found that endosulfan can damage microfilaments, microtubules, and nuclei; arrest mitosis; remarkably increase the expressions of Dll4, Notch1, Cleaved-Notch1, Jagged1, Notch4, Hes1, and p21; and significantly induce ROS and malondialdehyde production in HUVECs. The presence of DAPT antagonized the above changes of cycle arrest, proliferation inhibition, and expressions of Dll4, Notch1, Cleaved-Notch1, Hes1, and p21 caused by endosulfan; however, NAC could attenuate LDH release; ROS and malondialdehyde production; apoptosis; and the expression levels of Dll4, Notch1, Cleaved-Notch1, Notch4, and Hes1 induced by endosulfan. These results demonstrated that endosulfan inhibited proliferation through the Notch signaling pathway as a result of oxidative stress. In addition, endosulfan can damage the cytoskeleton and block mitosis, which may add another layer of toxic effects on endothelial cells.