Oral squamous cell carcinoma cell-derived GM-CSF regulates PD-L1 expression in tumor-associated macrophages through the JAK2/STAT3 signaling pathway.

Previous study reported that gastric cancer-derived granulocyte-macrophage colony-stimulating factor (GM-CSF) could mediate neutrophil activation and induce PD-L1 expression through JAK2/STAT3 signaling pathway. Moreover, this pathway in various cancers could also regulate PD-L1 expression of tumor cells. Therefore, our study aimed to investigate whether the JAK2/STAT3 pathway regulates PD-L1 expression in tumor-associated macrophages (TAMs) in oral squamous cell carcinoma (OSCC), which can help us achieve further understanding of immune escape mechanisms in OSCC. We induced human monocytes THP-1 into M0, M1, and M2 macrophages, and applied them to common medium and tumor-conditioned medium, the latter was collected from two types of OSCC cell line. Western blot and RT-PCR were used to detect PD-L1 expression and activation of JAK2/STAT3 pathway in macrophages under various conditions. We found that GM-CSF in tumor-conditioned medium from OSCC cells increased PD-L1 expression in M0 macrophages in a time-dependent manner. Moreover, both GM-CSF neutralizing antibody and JAK2/STAT3 pathway inhibitor AG490 could inhibited its up-regulation. In the meantime, we confirmed that GM-CSF indeed acted through JAK2/STAT3 pathway by measuring phosphorylation of key proteins in this pathway. Therefore, we concluded that OSCC cell-derived GM-CSF was able to up-regulate PD-L1 expression in TAMs through JAK2/STAT3 signaling pathway.

Long-term morphometric changes in the anterior alveolar bone in adolescents and adults after space closure: A retrospective study.

OBJECTIVES: To analyse the morphometric changes in the anterior alveolar bone of both the maxilla and mandible after space closure and retention for 18-36 mo in adults and adolescents. MATERIALS AND METHODS: Forty-two subjects with 4 first premolars extracted followed by retracting anterior teeth were included and divided into two age groups: adult group (4 males, 17 females, mean age: 23.67 ± 5.29 y, treatment duration: 27.95 mo, retention duration: 26.96 mo, ANB: 4.8 ± 2.1, U1-L1: 117.2 ± 9.2, U1-PP: 120.2 ± 7.2, L1-MP: 99.2 ± 5.3) and adolescent group (6 males, 15 females, mean age: 11.52 ± 1.21 y, treatment duration: 26.18 mo, retention duration: 25.79 mo, ANB: 5.2 ± 2.1, U1-L1: 116.0 ± 8.6, U1-PP: 119.8 ± 4.9, L1-MP: 99.7 ± 4.9). Alveolar bone height and thickness of anterior teeth in both groups were measured using cone beam computed tomography (CBCT) imaging performed at the pretreatment (T1), posttreatment (T2) and retention phases (T3). One-way repeated-measure ANOVAs were performed to evaluate the alveolar bone changes. Voxel-based superimpositions were performed to measure the amount of tooth movement. RESULTS: After orthodontic treatment, the lingual bone height and thickness of both arches and the labial bone height of the mandible decreased significantly in both age groups (P < .05). Most of the labial bone height and thickness of the maxilla in both groups remained unchanged (P > .05). After retention, the lingual bone height and thickness increased significantly in both age groups (P < .05). The amounts of increased height ranged from 1.08 to 1.64 mm in adults and from 0.78 to 1.21 mm in adolescents, and the amounts of increased thickness ranged from 0.23 mm to 0.62 mm in adults and from 0.16 mm to 0.36 mm in adolescents. Obvious movements of the anterior teeth during retention were not found (P > .05). CONCLUSIONS: Although lingual alveolar bone loss occurred in adolescents and adults during orthodontic treatment, continuous remodelling occurred in the later retention phase, which provides a reference for clinical treatment planning of bimaxillary dentoalveolar protrusion.

SET/PP2A signaling regulates macrophage positioning in hypoxic tumor regions by amplifying chemotactic responses.

Tumor-associated macrophages (TAMs) are one of the main cellular components in the tumor microenvironment (TME). In many types of solid tumors, TAMs tend to accumulate in hypoxic areas and are intimately related to poor patient prognosis. However, the underlying mechanisms by which TAMs infiltrate hypoxic tumor regions remain unclear. In this study, we report that genetic deletion of SE translocation (SET) in myeloid cells inhibited the entry of TAMs into the hypoxic tumor region and abated their proangiogenic and immunosuppressive functions, ultimately inhibiting tumor growth. Mechanistically, in response to hypoxic tumor supernatant stimulation, SET in macrophages shuttled between the nucleus and cytoplasm via the PKC-CK2α signaling axis. Cytoplasmic retention of SET increased ERK and P38 signaling by inhibiting PP2A, which promoted TAM migration into the hypoxic area and polarization toward the M2 phenotype. Therefore, we conclude that SET modulates tumor immunity by acting as a key regulator of macrophage positioning and function in the tumor.

The Absence of STING Ameliorates Non-Alcoholic Fatty Liver Disease and Reforms Gut Bacterial Community.

Non-alcoholic fatty liver disease (NAFLD) is one of the primary causes of cirrhosis and a major risk factor for hepatocellular carcinoma and liver-related death. It has been correlated with changes in the gut microbiota, which promote its development by regulating insulin resistance, bile acid and choline metabolism, and inflammation. Recent studies suggested a controversial role of the stimulator of interferon genes (STING) in the development of NAFLD. Here, we showed that as an immune regulator, STING aggravates the progression of NAFLD in diet-induced mice and correlated it with the changes in hepatic lipid metabolism and gut microbiota diversity. After feeding wild-type (WT) and STING deletion mice with a normal control diet (NCD) or a high-fat diet (HFD), the STING deletion mice showed decreased lipid accumulation and liver inflammation compared with WT mice fed the same diet. In addition, STING specifically produced this hepatoprotective effect by inhibiting the activation of CD8+ T cells. The gut microbiota analysis revealed significant differences in intestinal bacteria between STING deletion mice and WT mice under the same diet and environmental conditions; moreover, differential bacterial genera were associated with altered metabolic phenotypes and involved in related metabolic pathways. Overall, our findings reveal the important regulatory role that STING plays in the progression of NAFLD. In addition, the change in intestinal microbiota diversity may be the contributing factor.

Dynamics of estrogen-induced ROS and DNA strand break generation in estrogen receptor α-positive breast cancer.

DNA repair machinery is involved in estrogen-dependent transactivation. Mounting evidence suggests that mechanisms underlying estrogen-induced DNA damage are complicated. To date estrogen-induced DNA oxidation and its impact on ERα-mediated transaction remains ambiguous. Herein, we found that the process of 17ß-estradiol (E2)-induced ROS production can be approximately divided into two phases according to responding time and generation mechanisms. The intracellular Ca2+ fluctuation and ERα-dependent transcription lead to temporospatially different oxidative DNA damage. Further, we demonstrate that DNA oxidation is dispensable for estrogen-responsive gene expression. Dynamics of estrogen-induced DNA strand break generation also show two-phase pattern and topoisomerase-mediated DNA stand breaks are essential in estrogen signaling. Collectively, our findings have provided new insights into oxidative DNA damage in estrogen signaling.

Crosstalk between macrophages and natural killer cells in the tumor microenvironment.

The tumor microenvironment (TME) is jointly constructed by a variety of cell types, including tumor cells, immune cells, fibroblasts, and epithelial cells, among others. The cells within the TME interact with each other and with tumor cells to influence tumor development and progression. As the most abundant immune cells in the TME, macrophages regulate the immune network by not only secreting a large amount of versatile cytokines but also expressing a series of ligands or receptors on the surface to interact with other cells directly. Due to their strong plasticity, they exert both immunostimulatory and immunosuppressive effects in the complex TME. The major effector cells of the immune system that directly target cancer cells include but are not limited to natural killer cells (NKs), dendritic cells (DCs), macrophages, polymorphonuclear leukocytes, mast cells, and cytotoxic T lymphocytes (CTLs). Among them, NK cells are the predominant innate lymphocyte subsets that mediate antitumor and antiviral responses. The activation and inhibition of NK cells are regulated by cytokines and the balance between activating and inhibitory receptors. There is an inextricable regulatory relationship between macrophages and NK cells. Herein, we systematically elaborate on the regulatory network between macrophages and NK cells through soluble mediator crosstalk and cell-to-cell interactions. We believe that a better understanding of the crosstalk between macrophages and NKs in the TME will benefit the development of novel macrophage- or NK cell-focused therapeutic strategies with superior efficacies in cancer therapy.

Rosiglitazone alleviates lipopolysaccharide-induced inflammation in RAW264.7 cells via inhibition of NF-κB and in a PPARγ-dependent manner.

Rosiglitazone is a synthetic peroxisome proliferator-activated receptor (PPAR)γ agonist widely used for the treatment of type 2 diabetes. Recent studies have demonstrated that rosiglitazone displays anti-inflammatory effects. The present study aimed to investigate whether rosiglitazone alleviates decreases in RAW264.7 cell viability resulting from lipopolysaccharide (LPS)-induced inflammation, as well as exploring the underlying mechanism. A macrophage inflammatory injury model was established by treating RAW264.7 cells with 100 ng/ml LPS. Cells were divided into LPS and rosiglitazone groups with different concentrations. Cell viability was assessed by performing an MTT assay. The expression of inflammatory cytokines was detected by conducting enzyme-linked immunosorbent assays and reverse transcription-quantitative PCR. Nitric oxidesecretion was assessed using the Griess reagent system. The expression levels of key nuclear factor-κB pathway-associated proteins were detected via western blotting. Rosiglitazone alleviated LPS-induced decrease in RAW264.7 cell viability and inhibited inflammatory cytokine expression in a concentration-dependent manner. Rosiglitazone significantly inhibited LPS-induced upregulation of p65 phosphorylation levels and downregulated IκBα expression levels. However, rosiglitazone-mediated inhibitory effects were reversed by PPARγ knockdown. The results of the present study demonstrated that rosiglitazone significantly inhibited LPS-induced inflammatory responses in RAW264.7 macrophage cells, which was dependent on PPARγ activation and NF-κB suppression.

Comprehensive analysis of key lncRNAs in HCV-positive hepatocellular carcinoma.

INTRODUCTION: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Despite the therapeutic advances in HCC in the past few decades, the mortality rate of HCC is still high. Hepatitis C (HCV) infection is one of the major etiological risk factors of HCCs. However, the underlying mechanisms of HCV-induced hepatocarcinogenesis remain largely unclear. MATERIAL AND METHODS: Our study represented the comprehensive analysis of differentially expressed lncRNAs in HCV-positive HCC for the first time by analyzing the public dataset GSE17856. Co-expression network and gene ontology (GO) analysis revealed the functions of those differentially expressed lncRNAs. RESULTS: We identified 256 upregulated lncRNAs and 198 downregulated lncRNAs in HCV- positive HCC compared to the normal liver tissues. Co-expression network and GO analysis showed that these lncRNAs were involved in regulating metabolism, energy pathways, proliferation and the immune response. Seven lncRNAs (LOC341056, CCT6P1, PTTG3P, LOC643387, LOC100133920, C3P1 and C22orf45) were identified as key lncRNAs and co-expressed with more than 100 differentially expressed genes (DEGs) in HCV-related HCC. Kaplan-Meier analysis showed that higher expression levels of LOC643387, PTTG3P, LOC341056, CCT6P1 and lower expression levels of C3P1 and C22orf45 were associated with shorter survival time in the TCGA dataset. CONCLUSIONS: We believe that this study can provide novel potential therapeutic and prognostic biomarkers for HCV-positive HCC.

Association between TNF-α G-308A (rs1800629) polymorphism and susceptibility to chronic periodontitis and type 2 diabetes mellitus: A meta-analysis.

BACKGROUND AND OBJECTIVE: Although the association between tumor necrosis factor-α (TNF-α) G-308A (rs1800629) polymorphism and chronic periodontitis (CP), chronic periodontitis with type 2 diabetes mellitus (DP) is assumed, results of this association have been contradictory. The aim of this study was to assess the relationship between rs1800629 polymorphism and CP/DP susceptibility. METHODS: We searched for studies on PubMed, Web of Science, MEDLINE, Chinese National Infrastructure, and WanFang databases. Study selection was performed using specific inclusion and exclusion criteria and fulfilled the PECO (participant, exposure, comparison, and outcome) format. The relationship between rs1800629 polymorphism and CP/DP susceptibility was evaluated by the effect summary odds ratio (OR) and 95% confidence intervals (CIs). Allele, dominant, and recessive genetic models were computed to assess the strength of the association. RESULTS: A total of 25 case-control studies were included in the analysis. In the Asian population, TNF-α rs1800629 polymorphism was found to be significantly associated with CP in the overall analyses and for all genetic contrasts, while no significant risks were found among Caucasian populations for all genetic contrasts. The TNF-α rs1800629 polymorphism was also associated with increased DP risk in Asians under the fixed-effects model, but not in the recessive comparison. CONCLUSION: The meta-analysis suggested that TNF-α rs1800629 polymorphism might affect the risk of CP and DP, particularly in individuals of Asian descent.

Obesity-Induced Upregulation of ZBTB7A Promotes Lipid Accumulation through SREBP1.

OBJECTIVE: Nonalcoholic fatty liver disease (NAFLD) is among the most common chronic liver diseases. However, the pathogenesis of NAFLD is not still unclear. This study aims at evaluating the role of zinc finger and BTB domain-containing 7A (ZBTB7A) in NAFLD. METHODS: Western blotting, real-time reverse transcription PCR (RT-PCR), and immunohistochemistry were submitted to evaluate the level of ZBTB7A in the high fatty diet- (HFD-) induced NAFLD mouse model. In vitro, the expression of ZBTB7A was assessed in oleic acid- (OA-) induced HepG2 cells with western blotting and RT-PCR. The luciferase reporter assay was used to estimate the effect of ZBTB7A on the SREBP1 and NF-κB, and the ChIP assay was subjected to evaluate the direct binding to the SREBP1 promoter. Oil Red staining was used to detect lipid accumulation, and the ELISA was used to verify the levels of TG, T-CHO, and MDA. ZBTB7A was knocked down with siRNA, and RT-PCR was performed to analyze the lipogenesis-, fatty acid transporter-, and oxidation metabolism-related genes expression. The levels of ZBTB7A in primary hepatocyte, Kupffer, and hepatic stellate cells (HSCs) were tested by RT-PCR. RESULTS: The upregulation of ZBTB7A expression was assessed in NAFLD mice, and ZBTB7A expression was positively correlated with TNFα, IL-6, TG, T-CHO, and MDA. ZBTB7A was highly expressed in the hepatocytes. In vitro, OA-induced ZBTB7A expression and ZBTB7A expression were closely associated with SREBP1c. ZBTB7A could activate the promoter activity of SREBP1 and activate NF-κB activity. Interestingly, the direct binding of ZBTB7A in the SREBP1 promoter was acquired in HepG2 cells. Inhibition of ZBTB7A expression could attenuate OA-induced lipid accumulation, inhibit the expression of the lipogenesis-related genes and fatty acid transporter genes, and promote the expression of oxidation metabolism-related genes. CONCLUSION: ZBTB7A plays a significant role in the development process of NAFLD, and obesity-induced upregulation of ZBTB7A promotes lipid accumulation through activation of SREBP1 and NF-κB. ZBTB7A may be a potential novel target for the therapy of NAFLD.

The role of MAPK signaling pathway in formation of EMT in oral squamous carcinoma cells induced by TNF-α.

To study the role of MAPK signaling pathway in the development of Epithelial-mesenchymal transition in oral squamous cell carcinoma induced by inflammatory factor TNF-α. After the action of TNF-α, the expression of JNK, ERK, P38 in MAPK signaling pathway increased and the expression of E-cadherin, Claudin1 decreased significantly compared to the normal control group. After the addition of corresponding inhibitor, the expression of JNK, ERK, P38 decreased and the expression of E-cadherin, Claudin1 increased compared with TNF-α group. TNF-α regulated the role of EMT in promoting the invasion and metastasis of oral squamous carcinoma cells through MAPK signaling pathway.

TNF-Alpha Promotes Invasion and Metastasis via NF-Kappa B Pathway in Oral Squamous Cell Carcinoma.

BACKGROUND Recent evidence reveals that the inflammatory microenvironment is associated with tumor migration, invasion, and metastasis. Tumor necrosis factor-α (TNF-α) play a vital role in regulation of the inflammatory process in tumor development. Nuclear factor-kappa B (NF-κB) is one of the key transcription factors which regulate processes in tumor promotion. The aim of this study was to explore the role of NF-κB on the invasion and migration of oral squamous cell carcinoma (OSCC). MATERIAL AND METHODS The IKKß and p65 mRNA and protein levels were determined by quantitative RT-PCR and western blot. Wound scratch healing assays and transwell migration assays were used to evaluate the effect of TNF-α and BAY11-7082 on the migration of the OSCC cell lines (HN4, HN6, and CAL27). RESULTS We observed a significant increase of the expression level of IKKß and p65 in OSCC cells from the experimental group at 24 h, 48 h, and 72 h after TNF-α stimulation. Invasion and metastasis of OSCC cells was obviously improved after the TNF-α stimulation. Invasion and metastasis ability of OSCC cells was inhibited in the suppression group, and no significant changes were observed in expression level of IKKß and p65 after the use of BAY11-7082. CONCLUSIONS Our results suggest that TNF-α enhances the invasion and metastasis ability of OSCC cells via the NF-κB signaling pathway.

Snail interacts with Id2 in the regulation of TNF-α-induced cancer cell invasion and migration in OSCC.

The inflammatory tumor microenvironment has been identified to play a pivotal role in tumor development and metastasis. Tumor necrosis factor-α (TNF-α) is one of the key cytokines that regulate the inflammatory processes in tumor promotion. In the current study, we treated three oral squamous cell carcinoma (OSCC) cell lines with TNF-α to study its role in inflammation-induced tumor progression. Here we show that TNF-α induces stabilization of the transcriptional repressor Snail and activates NF-κB pathway in the three OSCC cell lines. These activities resulted in the increased motility and invasiveness of three OSCC cell lines. In addition, upon dealing with TNF-α for the indicated time, three OSCC cell lines underwent epithelial-to-mesenchymal transition (EMT), in which they presented a fibroblast-like phenotype and had a decreased expression of epithelial marker (E-cadherin) and an increased expression of mesenchymal marker (vimentin). We further demonstrated that TNF-α can up-regulate the expression of Id2 while inducing an EMT in oral cancer cells. Finally, we showed that Id2 interacted with Snail which may constrain Snail-dependent suppression of E-cadherin. In conclusion, our study indicates that TNF-α induces Snail stabilization is dependent on the activation of NF-κB pathway and results in increasing cell invasion and migration in OSCC cells. Id2 may contribute to regulate the function of Snail during TNF-α-mediated EMT in OSCC. These findings have significant implications for inflammation-induced tumor promotion in OSCC.

Expression of E-cadherin and vimentin in oral squamous cell carcinoma.

The aim of the study is to determine the levels of E-cadherin, vimentin expression in tumor tissues from patients with oral squamous cell carcinoma (OSCC), and the relationship between the expression of E-cadherin, vimentin and epithelial-mesenchymal transition, in order to explore its values for predicting the invasion and metastasis of oral squamous cell carcinoma, short survival of patients in many types of cancer. E-cadherin and vimentin expression of 10 benign and 42 OSCC tumor tissues was examined by immunohistochemical staining. E-cadherin is positively expressed in normal oral mucosa epithelium, but vimentin expression is not found in normal oral mucosa epithelia; the E-cadherin and vimentin were expressed in 26 of 42 (61.9%) and 16 of 42 (38.1%), respectively. No statistically difference was found for E-cadherin and vimentin expression in patients with different age, gender and tumor location, E-cadherin and vimentin expression was significantly associated with lymph node metastasis and tissue location (P<0.05); E-cadherin expression was also significantly associated with tumor stage (P<0.05); there are significantly difference between infiltrative margin and central area in patients with oral squamous cell carcinoma for E-cadherin and vimentin positive expression (P<0.05). E-cadherin and vimentin positive expression was associated with tumor metastasis of oral squamous cell carcinoma. Our study preliminarily confirmed that EMT phenomenon is existed during the development of oral squamous cell carcinoma. Co-evaluation of E-cadherin and vimentin might be a valuable tool for predicting OSCC patient outcome.

Correlation of human papilloma virus with oral squamous cell carcinoma in Chinese population.

Previous studies indicated that oral squamous cell carcinomas (OSCC) might be related to human papilloma virus (HPV) infection. However, the relationship between OSCC in a Chinese population and oral HPV infection is still unclear. In this study, we evaluate the relationship of OSCC with HPV infection in a Chinese population via a meta-analysis. The reports on HPV and OSCC in a Chinese population published between January, 1994, and October, 2015 were retrieved via CNKI/WANFANG/pubmed databases. According to the inclusion criteria, we selected 26 eligible case-control studies. After testing the heterogeneity of the studies by the Cochran Q test, the meta-analyses for HPV and HPV16 were performed using the random effects model. Quantitative meta-analyses showed that, compared with normal oral mucosa the combined odds ratio of OSCC with HPV infection were 1.98 (95% CI: 1.34-2.92). The test for overall effect showed that the P value was less than 0.05 (Z = 3.46). Forest plot analyses were seen in Figures 2 and 3. Publication bias and bias risk analysis using RevMan 5.3 software were measured indicators of the graphics of the basic symmetry. High incidences of HPV infection were found in the samples of Chinese OSCC. For the Chinese population, HPV infection elevates the risk of OSCC tumorigenesis.

Zinc dependent endopeptidases of matrix metalloproteinases-9 expressions is associated with tumor metastases of oral squamous cell carcinoma in Chinese population: a meta-analysis.

Matrix metalloproteinase-9 (MMP-9) is an important member of zinc dependent endopeptidases family and is considered to be involved in the invasion and metastasis of cancer cells. Many studies were published to assess the prognostic role of MMP-9 expression in patients with oral squamous cell Carcinoma, but the findings from those studies were inconsistent in Chinese population. We searched eligible studies in Pubmed, Embase, and Web of Science databases. Six studies with a total of 556 patients were finally included into the meta-analysis. The pooled odds ratios (OR) with the corresponding 95% confidence interval (95% CIs) for positive rate of MMP-9 were calculated by using meta-analysis. Overall, MMP-9 positive expression was associated with tumor metastases in patients with oral squamous cell Carcinoma (fixed-effects OR 4.24, 95% CI 2.25-7.99, P < 0.001; random-effects OR 4.35, 95% CI 2.31-8.21, P < 0.001). Our results indicated that MMP-9 expression is associated with tumor metastases in patients with oral squamous cell carcinoma, and patients with higher MMP-9 expression have less tumor metastases.

Cytochrome P450 3A1 mediates 2,2',4,4'-tetrabromodiphenyl ether-induced reduction of spermatogenesis in adult rats.

BACKGROUND: 2,2',4,4'-tetrabromodiphenyl ether (BDE47) is the dominant PBDE congener in humans, wildlife, and the environment. It has been reported to be metabolized by cytochrome P450 (CYP) enzymes. Still, the effects of BDE47 on spermatogenesis failure are attracting an increasing amount of attention. However, it is unclear whether CYP-mediated metabolism contributes to BDE47-induced reproductive toxicity. METHODOLOGY AND PRINCIPAL FINDINGS: The role of cytochrome P450 3A1 (CYP3A1) in the formation of oxidative metabolites of BDE47 and its induced spermatogenesis failure was investigated in SD rats. BDE47 significantly increased the expression and activity of CYP3A1 in rat liver, and 3-OH-BDE47, the major oxidative metabolite of BDE47, dose-dependently increased in rat liver, serum, and testis, which was aggravated by dexamethasone (DEX), an inducer of CYP3A1. Additionally, testicular 3-OH-BDE47 and reactive oxygen species (ROS) in seminiferous tubules increased especially when BDE47 was administered in combination with DEX, which was confirmed in GC-1 and GC-2 cells that 3-OH-BDE47 induced more ROS production and cell apoptosis via the upregulation of FAS/FASL, p-p53 and caspase 3. As a result, daily sperm production dose-dependently decreased, consistent with histological observations in giant cells and vacuolar spaces and increase in TUNEL-positive apoptotic germ cells. CONCLUSION: CYP3A1-mediated metabolic activation of BDE47 and the active metabolite 3-OH-BDE47 and consequent ROS played an important role in reduction of spermatogenesis by germ cell apoptosis. Our study helps provide new insights into the mechanism of reproductive toxicity of environmental chemicals.

Inhibitory effect of ARHI on pancreatic cancer cells and NF-κB activity.

The aim of this study was to investigate the effect of aplasia ras homolog member I (ARHI) on proliferation, apoptosis and the cell cycle in the pancreatic cancer cell line PANC-1. The study also aimed to examine the effect of ARHI on the activity of the nuclear factor (NF)-κB and to determine whether ARHI acts as a tumor suppressor in the development of pancreatic cancer by inhibiting the activity of NF-κB. A pIRES2­EGFP­ARHI vector, constructed by reverse transcrition (RT)­PCR, was transiently transfected into the PANC-1 cells and analyzed for the expression of the ARHI protein by western blotting. A MTT assay was used to quantify cell proliferation, and apoptosis was analyzed by flow cytometry. The NF­κB signaling pathway, specifically the pathway using the nuclear phosphorylated p65 isoform, was analyzed by western blotting. Expression of the ARHI protein was detected by western blotting subsequent to the PANC-1 cells being transiently transfected with the pIRES2­EGFP­ARHI construct. Cell proliferation was strongly inhibited in the PANC-1 cells transfected with pIRES2­EGFP­ARHI. The cell cycle assays indicated an increase in the number of cells at the G0/G1 phase and a decrease in the cells at the S phase, but the difference was not significant (P>0.05). Time course studies also indicated a marked increase in the apoptotic index following transient transfection, as well as a gradual decrease in the expression of the nuclear phosphorylated p65 protein. ARHI acts as a tumor suppressor by downregulating the NF­κB signaling pathway, which results in the inhibition of cell proliferation, apoptosis and the cell cycle in the pancreatic tumor PANC-1 cell line.

The regulation of trefoil factor 2 expression by the transcription factor Sp3.

Trefoil factor family 2 (TFF2) participates in mucus stabilization and repair, apoptosis, and inflammatory responses. Previously published reports have indicated that several growth factors and basal transcription factors are associated with the expression of TFF2. However, the detailed mechanisms that regulate TFF2 expression are not fully understood. The present study was designed to assess the essential role of the transcription factor SP3 with respect to TFF2 expression. We first demonstrated that there was a negative correlation between the expression levels of SP3 and TFF2. Thus, in the examined cells, the overexpression of SP3 decreased the expression level of TFF2, whereas the inhibition of SP3 increased the expression level of TFF2. Moreover, we discovered two GC boxes in the TFF2 promoter and confirmed the specific binding of SP3 to this promoter. On the whole, this study indicated that Sp3 was a major regulator of TFF2 expression. This knowledge should contribute to our understanding of the role that is played by SP3 in the regulation of TFF2 expression.

The silencing of Pokemon attenuates the proliferation of hepatocellular carcinoma cells in vitro and in vivo by inhibiting the PI3K/Akt pathway.

Pokemon (POK erythroid myeloid ontogenic factor), which belongs to the POK protein family, is also called LRF, OCZF and FBI-1. As a transcriptional repressor, Pokemon assumes a critical function in cellular differentiation and oncogenesis. Our study identified an oncogenic role for Pokemon in human hepatocellular carcinoma (HCC). We successfully established human HepG2 and Huh-7 cell lines in which Pokemon was stably knocked down. We demonstrated that Pokemon silencing inhibited cell proliferation and migration. Pokemon knockdown inhibited the PI3K/Akt and c-Raf/MEK/ERK pathways and modulated the expression of various cell cycle regulators in HepG2 and Huh-7 cells. Therefore, Pokemon may also be involved in cell cycle progression in these cells. We confirmed that Pokemon silencing suppresses hepatocellular carcinoma growth in tumor xenograft mice. These results suggest that Pokemon promotes cell proliferation and migration in hepatocellular carcinoma and accelerates tumor development in an Akt- and ERK-signaling-dependent manner.