miR-590-5p/Tiam1-mediated glucose metabolism promotes malignant evolution of pancreatic cancer by regulating SLC2A3 stability.

BACKGROUND: T lymphoma invasion and metastasis 1 (Tiam1) is a tumor related gene that specifically activates Rho-like GTPases Rac1 and plays a critical role in the progression of various malignancies. Glycolysis plays an important role in cancer progression, it is crucial for supplying energy and producing metabolic end products, which can maintain the survival of tumor cells. As yet, however, the mechanism of Tiam1 in glycolysis reprogramming of pancreatic cancer (PC) remains to be clarified. Here, we investigated the functional role of Tiam1 in PC cell proliferation, metastasis and glycolysis reprogramming. It is expected to provide a new direction for clinical treatment. METHODS: The clinical relevance of Tiam1 was evaluated in 66 patients with PC, the effect of Tiam1 on cell proliferation was detected via 5-Ethynyl-2'-deoxyuridine (EdU) and colony formation. The ability of cell migration was detected by the wound healing and Transwell. Quantitative real time polymerase chain reaction (qRT-PCR) and luciferase reporter gene experiments clarify the regulatory relationship of miR-590-5p inhibiting Tiam1. Detection of the molecular mechanism of Tiam1 regulating glucose metabolism reprogramming in PC by glucose metabolism kit. RNA sequencing and Co-Immunoprecipitation (CoIP) have identified glucose transporter protein 3 (SLC2A3) as a key downstream target gene for miR-590-5p/Tiam1. RESULTS: We found that Tiam1 expression increased in PC tissues and was associated with lymph node metastasis. The silencing or exogenous overexpression of Tiam1 significantly altered the proliferation, invasion, and angiogenesis of PC cells through glucose metabolism pathway. In addition, Tiam1 could interact with the crucial SLC2A3 and promote the evolution of PC in a SLC2A3-dependent manner. Moreover, miR-590-5p was found to exacerbate the PC cell proliferation, migration and invasion by targeting Tiam1. Furthermore, the reversing effects on proliferation, migration and invasion were found in PC cells with miR-590-5p/Tiam1 overexpression after applying glucose metabolism inhibition. CONCLUSIONS: Our findings demonstrate the critical role of Tiam1 in PC development and the miR-590-5p/Tiam1/SLC2A3 signaling pathway may serve as a target for new PC therapeutic strategies.

Granulocyte colony-stimulating factor protected against brain injury in a rat cerebral hemorrhage model by modulating inflammation.

Granulocyte colony-stimulating factor (G-CSF) has been reported to exert a protective effect against secondary brain damage, but the underlying mechanisms remain unknown. We explored the ability of G-CSF to protect the brain from injury in a rat autologous blood-induced model of intracerebral hemorrhage (ICH), with a special focus on the anti-inflammation effect. An ICH was induced in 8-week-old male rats by an infusion of autologous blood, and the rats were then randomly assigned to five treatment groups: sham, ICH, and ICH+ low-dose (25 µg/kg), middle-dose (50 µg/kg), and high-dose (75 µg/kg) G-CSF. We then evaluated the levels of brain inflammation-related genes and proteins. The levels of tumor-necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) mRNA increased between days 1 and 14 post-ICH, with the highest expression on day 3. These changes were rectified by G-CSF in a dose-dependent manner. At day 3 post-injury, an elevation of the nuclear factor-kappa B (NF-κB) p65 protein level and a reduction of the inhibitor of NF-κB alpha (IκBα) protein level were observed; G-CSF treatment exerted a beneficial effect on both protein expressions. The expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins were increased; these changes were rectified by the highest dose of G-CSF. The brain-protecting effects of G-CSF are likely to be attributable, at least in part, to attenuation of the TNF-α, IL-6, iNOS, and COX-2 expressions induced by NF-κB activation in the brain tissues of this autologous blood-induced ICH rat model.

PIAS3/SOCS1-STAT3 axis responses to oxidative stress in hepatocellular cancer cells.

The participation of STAT3 and its upstream inhibitors, PIAS3 and SOCS1, in the oxidative response of hepatocellular carcinoma (HCC) cells was uncertain. Here, the expression of PIAS3 and SOCS1 in HCC tissues and cell lines was explored, and we sought to determine whether oxidative stress epigenetically regulated PIAS3 and SOCS1 expression and STAT3 activation in HCC cells. The expression of PIAS3 and SOCS1 was markedly decreased in HCC cell lines and tissues compared to normal hepatic cells and tissues. In HCC patients, low PIAS3 and SOCS1 expression were associated with poor survival. Oxidative stress induced by H2O2 in HepG2 cells was indicated by low antioxidant levels and high protein carbonyl content. Moreover, oxidative stress in HepG2 cells contributed to reduced proliferation but increased apoptosis, migration, and invasion capacity, which might be counteracted by antioxidants, such as tocopheryl acetate (TA). PIAS3 and SOCS1 expression was markedly decreased, while STAT3 was activated in HepG2 cells in response to H2O2 exposure. Co-treatment with antioxidant TA effectively increased the expression of PIAS3 and SOCS1, but it dephosphorylated STAT3 in H2O2-treated cells. PIAS1 or SOCS1 overexpression in HepG2 cells after H2O2 treatment restored cell viability and anti-oxidative responses and decreased apoptosis, migration, and invasion ability, and dephosphorylated STAT3 levels. Co-administration of the STAT3 activator, colivelin, partially abolished the effect of PIAS3 and SOCS1 overexpression in these processes. Therefore, oxidative stress in HCC cells may improve their migration and reduce proliferation through STAT3 activation through the repression of PIAS3 and SOCS1 expression.

Role of Dipeptidyl Peptidase-4 in Atherosclerotic Cardiovascular Disease in Humans and Animals with Chronic Stress.

Exposure to psychosocial stress is a risk factor for cardiovascular disease, including vascular atherosclerosis-based cardiovascular disease (ACVD). Dipeptidyl peptidase-4 (DPP-4) is a complex enzyme that acts as a membrane-anchored cell surface exopeptidase. DPP-4 is upregulated in metabolic and inflammatory cardiovascular disorders. DPP-4 exhibits many physiological and pharmacological functions by regulating its extremely abundant substrates, such as glucagon-like peptide-1 (GLP-1). Over the last 10 years, emerging data have demonstrated unexpected roles of DPP-4 in extracellular and intracellular signaling, immune activation, inflammation, oxidative stress production, cell apoptosis, insulin resistance, and lipid metabolism. This mini-review focuses on recent novel findings in this field, highlighting a DPP-4-mediated regulation of GLP-1-dependent and -independent signaling pathways as a potential therapeutic molecular target in treatments of chronic psychological stress-related ACVD in humans and animals.

Exogenous pancreatic kininogenase protects against renal fibrosis in rat model of unilateral ureteral obstruction.

Tissue kallikrein has protective function against various types of injury. In this study, we investigated whether exogenous pancreatic kininogenase (PK) conferred renoprotection in a rat model of unilateral ureteral obstruction (UUO) and H2O2-treated HK-2 cells in vitro. SD rats were subjected to UUO surgery, then PK (7.2 U/g per day, ip) was administered for 7 or 14 days. After the treatment, rats were euthanized; the obstructed kidneys were harvested for further examination. We found that PK administration significantly attenuated interstitial inflammation and fibrosis, and downregulated the expression of proinflammatory (MCP-1, TLR-2, and OPN) and profibrotic (TGF-ß1 and CTGF) cytokines in obstructed kidney. UUO-induced oxidative stress, closely associated with excessive apoptotic cell death and autophagy via PI3K/AKT/FoxO1a signaling, which were abolished by PK administration. We further showed that PK administration increased the expression of bradykinin receptors 1 and 2 (B1R and B2R) mRNA and the production of NO and cAMP in kidney tissues. Coadministration with either B1R antagonist (des-Arg9-[Leu8]-bradykinin) or B2R antagonist (icatibant) abrogated the renoprotective effects of PK, and reduced the levels of NO and cAMP in obstructed kidney. In H2O2-treated HK-2 cells, addition of PK (6 pg/mL) significantly decreased ROS production, regulated the expression of oxidant and antioxidant enzymes, suppressed the expression of TGF-ß1 and MCP-1, and inhibited cell apoptosis. Our data demonstrate that PK treatment protects against the progression of renal fibrosis in obstructed kidneys.

Astragaloside attenuates lipopolysaccharide-induced cell apoptosis in human gingiva cells via MAPK signaling pathway.

SUBJECTS: The aim of this study was to research the antiapoptotic effect of astragaloside, the principal component of Astragalus membranaceus (Fisch) Bge, in human gingiva cells induced by lipopolysaccharide (LPS). METHODS: According to the treatment, human gingiva cells were divided into five groups, including (1) control group without drug treatment; (2) imitating group, treated with LPS (10 µg·mL-1 ) alone; (3) low group, treated with LPS and 50 µmol·L -1 astragaloside; (4) medium group, treated with LPS and 100 µmol·L -1 astragaloside; and (5) high group, treated with LPS and 150 µmol·L -1 astragaloside. Cell proliferation and apoptosis were investigated using MTT assay and flow cytometry, respectively. Apoptosis and mitogen-activated protein kinase associated proteins were determined using Western blot analysis. RESULTS: LPS significantly suppressed the proliferation of human gingiva cells, but astragaloside obviously attenuated this change with a dose-dependent manner. LPS significantly promoted the apoptosis of human gingiva cells, but astragaloside treatments significantly attenuated this change with a dose-dependent manner. In addition, LPS could significant upregulated the expression of P-p38, P-JNK, Bax, and caspase-3. CONCLUSION: Astragaloside preformed a promising antiapoptotic role in apoptosis of human gingiva cells induced by LPS. This finding might provide us with a novel therapeutic method in tooth protection.

Geraniol and lupeol inhibit growth and promote apoptosis in human hepatocarcinoma cells through the MAPK signaling pathway.

OBJECTIVES: Hepatocarcinoma is one of the most lethal cancers, leading to a 5-year survival rate as low as 30% due to recurrence and metastasis. The treatment of liver cancer includes surgery and medication, of which, the former is more effective. However, surgical resection is applicable in less than 40% of patients. Therefore, it is imperative to find effective medication options for liver cancer therapy. METHODS: In this study, we found that two natural products, geraniol and lupeol, had antiproliferative and proapoptotic effects on the hepatocarcinoma cell lines SMMC7721 and HepG2. We also detected a lower expression level of Bcl-2 and upregulation of BAX and caspase in the presence of geraniol and lupeol. RESULTS: Furthermore, geraniol or lupeol also altered the phosphorylation level of extracellular signal-regulated protein kinase, P38, and c-Jun NH2-terminal kinases, suggesting involvement in mitogen-activated protein kinase signaling. CONCLUSIONS: This study provided direct evidence to support the effect of geraniol and lupeol in hepatocarcinoma cell growth and apoptosis, which indicated the potential application of these two natural products in anti-liver cancer therapy.

Long non-coding RNA activated by TGF-ß expression in cancer prognosis: A meta-analysis.

BACKGROUND: Recently, long non-coding RNA activated by transforming growth factor beta (TGF-ß) (lncRNA ATB) was shown to be useful in cancer prognosis, however, its prognostic value in human cancer has been inconsistent. Our study aimed to explore the prognostic role of lncRNA ATB expression in cancer prognosis. METHODS: PubMed, Embase, and Cochrane Library databases were thoroughly searched to retrieve studies focusing on the prognostic role of lncRNA ATB expression in cancer, and meta-analysis was performed. RESULTS: A total of 15 studies were included into this meta-analysis. High lncRNA ATB expression was significantly related to shorter overall survival (OS) (HR = 2.44, 95%CI = 1.98-3.01, P < 0.01), recurrence-free survival (RFS) (HR = 1.85, 95%CI = 1.42-2.40, P < 0.01), disease-free survival (DFS) (HR = 3.61, 95%CI = 2.45-5.33, P < 0.01), and progression-free survival (PFS) (HR = 2.97, 95%CI = 2.12-4.16, P < 0.01) when compared with low lncRNA ATB expression in cancer. Moreover, Patients with high lncRNA ATB expression tended to have worse tumor differentiation (P < 0.01), more advanced clinical stage (P < 0.01), deeper tumor invasion (P < 0.01), earlier distant metastases (P = 0.02), lymph node metastases (P = 0.04), and vascular invasion (P < 0.01) when compared with those with low lncRNA ATB expression. CONCLUSIONS: High lncRNA ATB expression was significantly associated with worse prognosis in cancer. LncRNA ATB expression could be used as a prognostic biomarker for human cancer.

Prognostic implications of ezrin and phosphorylated ezrin expression in non-small cell lung cancer.

BACKGROUND: The cytoskeletal organizer ezrin is a member of the ezrin-radixin-moesin (ERM) family and plays important roles in not only cell motility, cell adhesion, and apoptosis, but also in various cell signaling pathways. Phosphorylation at Thr-567 and Tyr-353 are key regulatory events in the transition of the dormant to active form of ezrin. This study investigated the prognostic implications of ezrin and phosphorylated ezrin (p-ezrin) expression in non-small cell lung carcinoma (NSCLC). METHODS: Ezrin and p-ezrin protein expressions were examined by immunohistochemistry in 150 NSCLC and adjacent non-tumor tissues and 14 normal lung tissues. qRT-PCR was used to determine ezrin mRNA expression levels in fresh tissues. The correlations between overexpression of ezrin and p-ezrin and the clinicopathological features of NSCLC were analyzed. The survival rates were calculated by the Kaplan-Meier method for 108 NSCLC cases. RESULTS: Ezrin and ezrinThr-567 proteins showed cytosolic and membranous staining patterns; however, ezrinTyr-353 protein only showed cytosolic staining. Ezrin and p-ezrin were significantly upregulated in NSCLC compared with the normal counterparts. Increased ezrin, ezrinThr-567, and ezrinTyr-353 levels were correlated with the late stage and poor differentiation of NSCLC. However, only ezrinThr-567 was correlated with the presence of lymph node metastasis. In regard to survival, only ezrinThr-567 was related with the overall survival time of patients with NSCLC, and both ezrin and ezrinThr-567 were associated with shortened survival time for patients with early stage NSCLC. CONCLUSIONS: Ezrin and p-ezrin, especially ezrinThr-567, may prove to be useful as a novel prognostic biomarker of NSCLC.

Correlation between HER-2/neu(erbB-2) expression level and therapeutic effect of combination treatment with HERCEPTIN and chemotherapeutic agents in gastric cancer cell lines.

INTRODUCTION: Although advanced gastric cancer has many limitations and response rate is marginal in chemotherapy. Overexpression of human epidermal growth factor receptor 2(HER-2/neu) gene and its protein are associated with increased cell division and a high rate of tumor growth and have been reported in several malignancies. Especially, approximately 30% of breast cancer patients have overexpression of HER-2/neu protein and the overexpression metastasize faster, induces resistance of the chemotherapy and down-regulate function of estrogen receptor. Recombinant humanized anti-HER2 antibody (Herceptin) inhibits proliferation of HER-2/neu overexpressing tumor cells and the use of that in combination in metastatic breast cancer have increased cytotoxicity of chemotherapeutic agents. METHODS: We evaluated the expression of HER-2/neu protein in gastric cell lines by FACS and then comparing the cytotoxicity in chemotherapeutics (doxorubicin, cisplatin, paclitaxel, 5-FU) alone and in combination with Herceptin according to the expression of HER-2/neu protein by MTT assay. RESULTS: 1. NCI-N87 (88%) gastric cancer cell line and SK-BR-3 (89%) breast cancer cell line with strong positivity of HER-2/neu expression. YBC-2 (55%) and YBC-3 (48%) gastric cancer cell line with intermediated, weak positivity respectively. Negative control U-87 MG (6%) brain cancer cell line were showed low expression of HER-2/neu. 2. Cell growth was dose-dependently inhibited in HER-2/neu positive, control cell line SK-BR-3 by Herceptin treatment but not observed in HER-2/neu negative control cell line U-87 MG. Effective growth inhibition was not observed in gastric cancer cell lines with single treatment of Herceptin, all cell lines observed the dose-dependent growth inhibition to chemotherapeutic agents (doxorubicin, cisplatin, paclitaxel and 5-FU). 3. Combination of Herceptin with doxorubicin observed synergistic effects in all cancer cell lines except YBC-3, combination of Herceptin with cisplatin observed NCI-N87 and SK-BR-3 and combination of Herceptin with paclitaxel observed synergistic effects in YBC-2. Combination of Herceptin with 5-FU observed antagonistic effects in all cancer cell lines. CONCLUSIONS: According to HER-2/neu expression level, effect of anti-cancer agents was observed differently in combination of Herceptin with chemotherapeutic agents. This suggests that HER-2/neu expression level can be applied standard of combination drug selection in combination of Herceptin With chemotherapeutic agents in gastric cancer.

Significance of NQO1 overexpression for prognostic evaluation of gastric adenocarcinoma.

NQO1 (NAD(P)H: quinone oxidoreductase, also known as DT-diaphorase) plays a prominent role in maintaining cellular homeostasis. NQO1 is abnormally elevated in many solid cancer types, including those of the adrenal gland, breast, colon, lung, ovary, and thyroid. However, little is known about the status of NQO1 in gastric adenocarcinoma (GAC). To investigate the clinicopathological significance of NQO1 expression in GAC, and thus evaluate its role as a potential prognostic marker, 203 cases of primary GAC, 31 of gastric dysplasia, and 53 of adjacent non-tumor tissues were selected for immunohistochemical staining of NQO1 protein. Correlations between NQO1 overexpression and clinicopathological characteristics were evaluated by χ(2) test and Fisher's exact test, while survival rates were calculated by Kaplan-Meier method. The relationship between prognostic factors and patient survival was analyzed by Cox proportional hazards model. Through these analyses it was found that the strongly positive rate of NQO1 protein in GAC was significantly higher than that in gastric dysplasia and adjacent non-tumor tissues. Analysis by qRT-PCR also confirmed that NQO1 mRNA levels were increased in GAC compared with those detected in either adjacent non-tumor tissues or normal gastric mucosa. Additionally, the NQO1 expression rate was positively correlated with tumor size, serosal invasion, tumor stage, and both disease-free survival and 5-year survival rates. Further analysis showed that although NQO1 was not an independent predictor of GAC, elevated expression of NQO1 could predict lower disease-free survival and 5-year survival times in late-stage patients. In conclusion, NQO1 plays an important role in the progression of GAC, and might be a potential, but not an independent, poor prognostic biomarker and therapeutic target of GAC.

Identification of differentially expressed genes in gastric cancer by high density cDNA microarray.

The identification of molecular markers for diagnosis, treatment, and prognosis is a significant issue in the management of patients with gastric cancer. We compared the expression profiles of 23 gastric cancers and 22 normal gastric tissues using cDNA microarrays. We divided the samples into two sets, 11 pairs as a training set and 12 unpaired gastric cancer and 11 unpaired normal gastric tissues as a test set. We selected significant genes in the training set and validated the significance of the genes in the test set. We obtained 238 classifier genes that showed a maximum cross-validation probability and clear hierarchical clustering pattern in the training set, and showed excellent class prediction probability in the independent test set. The classifier genes consisted of known genes related to the biological features of cancer and 28% unknown genes. We obtained genome-wide molecular signatures of gastric cancer, which provides preliminary exploration data for the pathophysiology of gastric cancer.

Application of mesenchymal stem cells as a vehicle to deliver replication-competent adenovirus for treating malignant glioma.

Although gene therapy was regarded as a promising approach for glioma treatment, its therapeutic efficacy was often disappointing because of the lack of efficient drug delivery systems. Mesenchymal stem cells(MSCs) have been reported to have a tropism for brain tumors and thus could be used as delivery vehicles for glioma therapy. Therefore, in this study, we attempted to treat glioma by using MSCs as a vehicle for delivering replication-competent adenovirus. We firstly compared the infectivity of type 3, type 5, and type 35 fiber-modified adenoviruses in MSCs. We also determined suitable adenovirus titer in vitro and then used this titer to analyze the ability of MSCs to deliver replication-competent adenovirus into glioma in vivo. Our results indicated that type 35 fiber-modified adenovirus showed higher infectivity than did naked type 3 or type 5 fiber-modified adenovirus. MSCs carrying replication-competent adenovirus significantly inhibited tumor growth in vivo compared with other control groups. In conclusion, MSCs are an effective vehicle that can successfully transport replication-competent adenovirus into glioma, making it a potential therapeutic strategy for treating malignant glioma.

Genetic analysis of liver metastatic cell lines with different metastatic potential.

Metastasis is the major feature of malignant tumors that causes 90% of cancer deaths. Our laboratory has already established liver metastatic clones with YCC-16, isolated from the blood of a gastric cancer patient and expanded in vitro culture using a repeated orthotopic implantation method, and had reported biologic behaviour of the parental YCC-16, the orthotopic primary S1L0, and S1L1, S2L2 and S3L3 liver metastatic clones. Here, using these cell lines, we screened from chromosomal abnormalities using karyotype analysis and micro-CGH matching. There were 31 genes screened using PCA method which were functionally related to cell adhesion. Also, there were 23 genes selected which were related to the liver specific metastasis but excluded genes related to adhesion. There were 4 genes which demonstrated reduced or increased expression stepwise with passage. In conclusion, our results should contribute to exploring the mechanisms of liver metastasis by gastric cancer.

Cellular adjustment of gastric cancer for hepatic metastasis in successive orthotopic implantation model.

In order to examine the mechanisms of hepatic metastasis in gastric cancer, a repeated orthotopic implantation method was used in nude mice with YCC-16, which was isolated from the blood of a gastric cancer patient. This study compared the biological and cytogenetic phenotypes of the five cell lines, the parental YCC-16, the orthotopic primary S1L0 and the 3 subsequent liver metastatic clones of S1L1, S2L2 and S3L3. E-cadherin and DAG1 gene expression levels were measured using real-time PCR. The parental YCC-16 showed multiple metastases, whereas the liver metastatic clones metastasized to the liver only. The liver metastatic rate showed increased trend with subsequent passages (passageII: 2/5, 40%; passageIII: 3/5, 60%; passageIV: 4/5, 80%). Otherwise, the liver metastatic clones had phenotype with a higher motility than that of the cell line from the orthotopic primary tumor (S1L0), and clonogenecity increased with subsequent passages in liver metastatic cell lines. The five cell lines had similar and additional chromosomal abnormalities were found in the selected clones. The E-cadherin expression level was decreased in all of the five cell lines, which was comparable to the common chromosomal changes. YCC-16 presented the lowest DAG1 expression level while S1L0 presented the highest. In the liver metastatic clones, the DAG1 expression level increased gradually with passages. Both the genetic alteration and cellular adjustment to the microenvironment are required for hepatic metastasis in gastric cancer. This model offers an opportunity to study the mechanisms of a hepatic metastasis of gastric cancer at the genetic and cellular level.

Human papillomavirus genotyping by oligonucleotide microarray and p16 expression in uterine cervical intraepithelial neoplasm and in invasive carcinoma in Korean women.

For evaluating the diagnostic significance of p16(INK4A) over-expression in the uterine cervical intraepithelial neoplasm and in invasive carcinoma, human papillomavirus (HPV) was detected and genotyped by oligonucleotide microarray in archival tissues of 117 cervical specimens, including 47 invasive squamous cell carcinomas (SCC), 30 cases of cervical intraepithelial neoplasia (CIN), 20 adenocarcinomas, and 20 cases of non-neoplastic cervix. The expression of p16(INK4A) protein was immunohistochemically studied in these cases and in five HPV-positive and one HPV-negative cervical cancer cell lines. HPV was detected in 50% of CIN, 61.7% of SCC, and 45.5% of adenocarcinomas. p16(INK4A) expression was seen in all 20 cases of adenocarcinoma, 78.7% (37/47) of SCC, and 96.7% (29/30) of CIN, but not in any cases of the non-neoplastic cervix. There was no difference in p16(INK4A) expression between the HPV-positive and HPV-negative cervical lesions. All HPV-positive and -negative cervical cancer cell lines expressed p16(INK4A) protein. In conclusion, the presence of p16(INK4A) expression in cervical squamous and glandular epithelium indicates the existence of dysplasia or malignancy in the uterine cervix, regardless of HPV infection.