Geraniol prevents Helicobacterium pylori-induced human gastric cancer signalling by enhancing peroxiredoxin-1 expression in GES-1 cells.

Helicobacter pylori (H. pylori), a gram-negative bacterial microbiological carcinogen, has been identified as the leading jeopardy feature for developing human gastric cancer (GC). As a result, inhibiting H. pylori growth has been identified as an effective and critical technique for preventing GC development. In this study, geraniol inhibits H. pylori-induced gastric carcinogen signalling in human gastric epithelial cells (GES-1). Geraniol prevents cytotoxicity, ROS and apoptosis in H. pylori-induced GES-1 cells. Furthermore, geraniol protects against H. -induced antioxidant depletion caused by malondialdehyde, damage of reactive DNA and nuclear fragmentation. Geraniol significantly reduced the expression of phosphorylated mitogen activated protein kinases (MAPKs) proteins such as p38 MAPK, extracellular signal-regulated kinase-1 (ERK1), c-Jun N-terminal kinase (c-JNK), tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and cyclooxygenase-2 (COX-2) in GES-1 infected with H. pylori. Furthermore, geraniol increased the antioxidant protein peroxiredoxin-1 (Prdx-1) in H. pylori-infected cells. Geraniol thus protects H. pylori-concomitant infection, and its resistance may be a possible method in preventing gastric cancer caused by H. pylori.

High-fat diet induced cyclophilin B enhances STAT3/lncRNA-PVT1 feedforward loop and promotes growth and metastasis in colorectal cancer.

High-fat diet (HFD) has been implicated to promote colorectal cancer (CRC). Recently, oncogene Cyclophilin B (CypB) is reported to be induced by cholesterol. However, the role of CypB in CRC carcinogenesis and metastasis associated with HFD remains unknown. In the present study, we showed that HFD-induced CypB enhances proliferation and metastasis through an inflammation-driven circuit, including Signal Transducer and Activator of Transcription 3 (STAT3)-triggered transcription of lncRNA-PVT1, and its binding with CypB that promotes activation of STAT3. CypB was found to be upregulated in CRC, which was correlated with elevated body mass index and poor prognosis. HFD induced CypB expression and proinflammatory cytokines in colon of mice. Besides, CypB restoration facilitated growth, invasion and metastasis in CRC cells both in vitro and in vivo. Moreover, RIP sequencing data identified lncRNA-PVT1 as a functional binding partner of CypB. Mechanistically, PVT1 increased the phosphorylation and nuclear translocation of STAT3 in response to IL-6, through directly interaction with CypB, which impedes the binding of Suppressors Of Cytokine Signalling 3 (SOCS3) to STAT3. Furthermore, STAT3 in turn activated PVT1 transcription through binding to its promoter, forming a regulatory loop. Finally, this CypB/STAT3/PVT1 axis was verified in TCGA datasets and CRC tissue arrays. Our data revealed that CypB linked HFD and CRC malignancy by enhancing the CypB/STAT3/PVT1 feedforward axis and activation of STAT3.

Retinoblastoma tumor suppressor gene 1 enhances 5-Fluorouracil chemosensitivity through SDF-1/CXCR4 axis by regulating autophagy in gastric cancer.

Due to lack of effective biomarkers for early diagnosis, most patients are diagnosed with advanced gastric cancer and have lower survival rates. 5-Fluorouracil (5-FU) is one of commonly used drugs for chemotherapy of gastric cancer, but drug resistance limits the wide application of agents. Retinoblastoma tumor suppressor gene 1 (RB1) is a key regulator in the progression of various human cancers, including gastric cancer. However, the effects of RB1 on chemosensitivity and the underlying mechanisms in gastric cancer (GC) are not clear. In this study, expressions of RB1 in GC cell lines were evaluated by RT-qPCR and western blot assay. CCK-8 was applied to examine the effect of 5-FU on cell viability. Meanwhile, IC50 values were calculated. The drug-resistance protein MDR1 and autophagy-related proteins were detected by western blot assay. Flow cytometry was used to detect cell cycle. The results showed that RB1 expressions were downregulated in GC cell lines and had significant differences between 5-FU resistance cell lines (SNU-620/5-FU and NUGC-3/5-FU) and non-resistance cell lines (SNU-620 and NUGC-3). Overexpression of RB1 enhanced 5-FU sensitivity of GC cells and caused cell cycle arrest in the S phase. Meanwhile, autophagy-related proteins were downregulated. Mechanistically, SDF-1/CXCR4 participated in the regulation of RB1 on cell autophagy. Autophagy activator, SDF-1 treatment and CXCR4 activation reversed the promoted effects of RB1 on 5-FU sensitivity in GC cells. In conclusion, our data revealed that RB1 was downregulated in GC cell lines. RB1 overexpression enhanced 5-FU chemosensitivity in GC cells by regulating cell autophagy via SDF-1/CXCR4 pathway. RB1 might serve as a promising therapeutic target of GC.

Suppression of FAK by nexrutine inhibits gastric cancer progression.

AIM: Nexrutine, an herbal extract of Phellodendron amurense, has been found to play a tumor-suppressive role in many cancers. However, its role in the pathogenesis of gastric cancer remains unclear. MATERIALS AND METHODS: Gastric cancer cells (SGC-7901 and MGC-803) were treated with nexrutine, and cell proliferation, invasion and apoptosis were analyzed. And the MGC-803 cells-derived xenograft mouse models were fed pelleted diet containing 600 mg/kg nexrutine for 21 days after inoculation. Mechanically, we focused on the influences of nexrutine on the levels and activation of STAT3 and NF-κB as well as their upstream regulator FAK. Additionally, we further verified whether nexrutine affected the proliferation, invasion and apoptosis of gastric cancer cells via FAK by upregulating FAK expression before nexrutine treatment. KEY FINDINGS: We found nexrutine inhibited the viability, invasion, and expression levels of PCNA, CyclinD1 and Bcl-2, promoted the apoptosis and Bax expression, decreased levels of STAT3, phospho-STAT3, NF-κB p65, phospho-p65, FAK and phospho-FAK in gastric cancer cells. Overexpression of FAK reversed the impacts of nexrutine on the levels of STAT3, phospho-STAT3, NF-κB p65, phospho-p65, as well as the malignant characteristics of gastric cancer cells. Moreover, nexrutine suppressed tumor volumes and weights, and decreased expression and phosphorylation of FAK, STAT3 and NF-κB p65 in vivo. SIGNIFICANCE: Nexrutine inhibited the malignant progression of gastric cancer via negatively regulating STAT3/NF-κB signaling pathway by suppressing FAK expression and activation.

miR-665 promotes the progression of gastric adenocarcinoma via elevating FAK activation through targeting SOCS3 and is negatively regulated by lncRNA MEG3.

Studies have found that miR-665 acted as a tumor suppressor or an oncogene in different malignancies. miR-665 expression was elevated in gastric adenocarcinoma tissues; however, its role and mechanism in this disease are not fully clarified. The expression of miR-665 and its target gene was detected in human gastric adenocarcinoma tissues and cells. Moreover, we analyzed the effects of miR-665 on the proliferation, migration, and epithelial-mesenchymal transition (EMT) of gastric adenocarcinoma cells as well as tumor growth in vivo. The mechanisms of miR-665 in gastric adenocarcinoma were investigated by using molecular biology techniques. We found miR-665 was upregulated and suppressor of cytokine signaling 3 (SOCS3) was downregulated in gastric adenocarcinoma tissues and cells. Elevated miR-665 was positively correlated with tumor size, lymph node metastasis, invasion depth, TNM stage, and poor differentiation in gastric adenocarcinoma patients. Overexpression of miR-665 promoted, whereas knockdown of miR-665 suppressed the proliferation, migration, and EMT of gastric adenocarcinoma cells. Furthermore, we demonstrated that miR-665 functioned through targeting SOCS3, followed by activation of the FAK/Src signaling pathway in gastric adenocarcinoma cells. miR-665 antagomir inhibited tumor growth as well as the activation of the FAK/Src pathway but increased SOCS3 expression in nude mice. In addition, miR-665 expression was negatively regulated by long noncoding RNA maternally expressed gene 3 (MEG3). In conclusion, miR-665 acted as an oncogene in gastric adenocarcinoma by inhibiting SOCS3 followed by activation of the FAK/Src pathway and it was negatively modulated by MEG3. miR-665 may be a promising therapeutic target for the treatment of gastric adenocarcinoma.

MicroRNA-92a-1-5p increases CDX2 by targeting FOXD1 in bile acids-induced gastric intestinal metaplasia.

BACKGROUND AND AIMS: Gastric intestinal metaplasia (IM) is common in the gastric epithelium of patients with chronic atrophic gastritis. CDX2 activation in IM is driven by reflux of bile acids and following chronic inflammation. But the mechanism underlying how bile acids activate CDX2 in gastric epithelium has not been fully explored. METHODS: We performed microRNA (miRNA) and messenger RNA (mRNA) profiling using microarray in cells treated with bile acids. Data integration of the miRNA/mRNA profiles with gene ontology (GO) analysis and bioinformatics was performed to detect potential miRNA-mRNA regulatory circuits. Transfection of gastric cancer cell lines with miRNA mimics and inhibitors was used to evaluate their effects on the expression of candidate targets and functions. Immunohistochemistry and in situhybridisation were used to detect the expression of selected miRNAs and their targets in IM tissue microarrays. RESULTS: We demonstrate a bile acids-triggered pathway involving upregulation of miR-92a-1-5p and suppression of its target FOXD1 in gastric cells. We first found that miR-92a-1-5p was increased in IM tissues and induced by bile acids. Moreover, miR-92a-1-5p was found to activate CDX2 and downstream intestinal markers by targeting FOXD1/FOXJ1 axis and modulating activation of nuclear factor kappa B (NF-κB) pathway. Furthermore, these effects were found to be clinical relevant, as high miR-92a-1-5p levels were correlated with low FOXD1 levels and high CDX2 levels in IM tissues. CONCLUSION: These findings suggest a miR-92a-1-5p/FOXD1/NF-κB/CDX2 regulatory axis plays key roles in the generation of IM phenotype from gastric cells. Suppression of miR-92a-1-5p and restoration of FOXD1 may be a preventive approach for gastric IM in patients with bile regurgitation.

Gastrin induces multidrug resistance via the degradation of p27Kip1 in the gastric carcinoma cell line SGC7901.

Multidrug resistance (MDR) is one of the major reasons for the failure of chemotherapy-based gastric carcinoma (GC) treatments, hence, biologically based therapies are urgently needed. Gastrin (GAS), a key gastrointestinal (GI) hormone, was found to be involved in tumor formation, progression, and metastasis. In this study, quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical staining analysis revealed a high level of expression of GAS in drug-insensitive GC tissues (P<0.01) and similar results were revealed in GC cell lines SGC7901 and its multidrug-resistant variants SGC7901/VCR and SGC7901/ADR. We constructed a eukaryotic expression vector pCDNA3.1(+)/GAS for GAS overexpression and recombinant lentiviral vectors for specific siRNA (siGAS). Transfection of pCDNA3.1(+)/GAS increased (P<0.05) while transfection of siGAS (P<0.05) and co-treated with paclitaxel (TAX) and vincristine (VCR) combination (TAX-VCR) decreased (P<0.01) the cell viability of SGC7901, SGC7901/VCR and SGC7901/ADR. Apoptosis rates of SGC7901/VCR and SGC7901/ADR were reduced by pCDNA3.1(+)/GAS and increased by siGAS (P<0.05). The apoptosis rates of SGC7901/VCR, SGC7901/ADR and SGC7901 were all upregulated (P<0.01) when cells were co-treated with a combination of siGAS and TAX-VCR. Additionally, siGAS significantly downregulated the expression of Bcl-2 and multidrug-resistant associate protein (MRP1) and P-glycoprotein (Pgp) (P<0.05) in SGC7901/VCR and SGC7901/ADR cells. Moreover, GAS overexpression in SGC7901 cells significantly inhibited p27Kip1 expression but increased phosphorylation levels of p27Kip1 on Thr (187) and Ser (10) sites (P<0.05), as well as increasing nuclear accumulation of S-phase kinase-associated protein 2 (Skp2) and cytoplasmic accumulation of the Kip1 ubiquitination-promoting complex (KPC) (P<0.05). Silencing of Skp2 blocked the promoting effects of pCDNA3.1(+)/GAS on viability, the expression of MRP1 and Pgp and the inhibitory effects of pCDNA3.1(+)/GAS on apoptosis. In conclusion, we suggest that GAS contributes to the emergence of MDR of SGC7901 cells via the degradation of p27Kip1.

Gastric Cancer Cell Proliferation and Survival Is Enabled by a Cyclophilin B/STAT3/miR-520d-5p Signaling Feedback Loop.

Molecular links between inflammation and cancer remain obscure despite their great pathogenic significance. The JAK2/STAT3 pathway activated by IL6 and other proinflammatory cytokines has garnered attention as a pivotal link in cancer pathogenesis, but the basis for its activation in cancer cells is not understood. Here we report that an IL6-triggered feedback loop involving STAT3-mediated suppression of miR-520d-5p and upregulation of its downstream target cyclophilin B (CypB) regulate the growth and survival of gastric cancer cells. In clinical specimens of gastric cancer, we documented increased expression of CypB and activation of STAT3. Mechanistic investigations identified miR-520d-5p as a regulator of CypB mRNA levels. This signaling axis regulated gastric cancer growth by modulating phosphorylation of STAT3. Furthermore, miR-520d-5p was identified as a direct STAT3 target and IL6-mediated inhibition of miR-520d-5p relied upon STAT3 activity. Our findings define a positive feedback loop that drives gastric carcinogenesis as influenced by H. pylori infections that involve proinflammatory IL6 stimulation. Cancer Res; 77(5); 1227-40. ©2016 AACR.

Loss of vinculin and membrane-bound ß-catenin promotes metastasis and predicts poor prognosis in colorectal cancer.

BACKGROUND: Loss of cell-cell adhesion is important for the development of cancer invasion and metastasis. Vinculin, a key adhesion-related protein, can affect metastasis and prognosis in several tumours. Here, we determined the biological roles of vinculin in the metastasis of colorectal cancer (CRC) and evaluated its clinical significance as a potential disease biomarker. METHODS: The expression level of vinculin in CRC cell lines and tissues was measured using Real-Time PCR and western blotting. Moreover, vinculin function was analysed using Transwell assays and in vivo metastasis assays in gain- and loss-of-function experiments. Furthermore, the impact of vinculin together with membrane-bound ß-catenin on the prognosis of 228 CRC patients was investigated by immunohistochemistry. Additionally, the expression of epithelial-mesenchymal transition (EMT) indicators was verified by immunohistochemistry in CRC tissues obtained from these patients. RESULT: Vinculin expression was found to be significantly downregulated in highly metastatic CRC cell lines and metastatic tissues. Both in vitro and in vivo experiments showed that vinculin suppressed invasion, migration and metastasis in CRC cells and that this suppression could be attenuated by silencing ß-catenin. Moreover, the expression of vinculin and membrane-bound ß-catenin were positively correlated in CRC tissues, and lack of vinculin expression emerged as an independent prognostic factor in patients with CRC. Finally, the loss of vinculin and membrane-bound ß-catenin was associated with node metastasis, organ metastasis and expression of EMT indicators. CONCLUSION: Our results suggest that vinculin may play specific roles in the EMT and metastasis of CRC and that loss of vinculin could be used as a prognostic factor for CRC.