Wangjun Liao

Metastasis-associated in colon cancer-1 upregulation predicts a poor prognosis of gastric cancer, and promotes tumor cell proliferation and invasion.

Metastasis‐associated in colon cancer‐1 (MACC1) is a newly identified oncogene, and little is known about its role in gastric cancer (GC). Our study was performed to investigate whether MACC1 influences the prognosis of GC patients and to explore the potential mechanisms involved. MACC1 expression was verified to be higher in GC tissues than in adjacent nontumorous tissues by Western blotting. A retrospective analysis of 361 GC patients (Stages I–IV) revealed that higher MACC1 expression was associated with more advanced disease, more frequent postoperative recurrence, more metastases and a higher mortality rate. The disease‐free survival of Stage I–III patients and overall survival of Stage‐IV patients were significantly worse when their tumors showed high MACC1 expression. To investigate the underlying mechanisms, MACC1 overexpression and downregulation were established in two GC cell lines (BGC‐823 and MKN‐28 cells). MACC1 overexpression significantly accelerated tumor growth and facilitated metastasis in athymic mice. MACC1 also promoted the proliferation, migration and invasion of both GC cell lines. Moreover, gastric MACC1 mRNA expression levels were significantly correlated with markers of the epithelial‐to‐mesenchymal transition (EMT) in patients with GC. MACC1 overexpression upregulated mesenchymal–epithelial transition factor and induced changes to markers of EMT, whereas silencing of MACC1 reversed all these changes. These findings provide some novel insights into the role of MACC1, a gene that contributes to a poor prognosis of GC by promoting tumor cell proliferation and invasion as well as the EMT.

XB130 promotes proliferation and invasion of gastric cancer cells

BackgroundXB130 has been reported to be expressed by various types of cells such as thyroid cancer and esophageal cancer cells, and it promotes the proliferation and invasion of thyroid cancer cells. Our previous study demonstrated that XB130 is also expressed in gastric cancer (GC), and that its expression is associated with the prognosis, but the role of XB130 in GC has not been well characterized.MethodsIn this study, we investigated the influence of XB130 on gastric tumorigenesis and metastasis in vivo and in vitro using the MTT assay, clonogenic assay, BrdU incorporation assay, 3D culture, immunohistochemistry and immunofluorescence. Western blot analysis was also performed to identify the potential mechanisms involved.ResultsThe proliferation, migration, and invasion of SGC7901 and MNK45 gastric adenocarcinoma cell lines were all significantly inhibited by knockdown of XB130 using small hairpin RNA. In a xenograft model, tumor growth was markedly inhibited after shXB130-transfected GC cells were implanted into nude mice. After XB130 knockdown, GC cells showed a more epithelial-like phenotype, suggesting an inhibition of the epithelial-mesenchymal transition (EMT) process. In addition, silencing of XB130 reduced the expression of p-Akt/Akt, upregulated expression of epithelial markers including E-cadherin, α-catenin and β-catenin, and downregulated mesenchymal markers including fibronectin and vimentin. Expression of oncoproteins related to tumor metastasis, such as MMP2, MMP9, and CD44, was also significantly reduced.ConclusionsThese findings indicate that XB130 enhances cell motility and invasiveness by modulating the EMT-like process, while silencing XB130 in GC suppresses tumorigenesis and metastasis, suggesting that it may be a potential therapeutic target.

Long non-coding RNA MALAT1 promotes gastric cancer tumorigenicity and metastasis by regulating vasculogenic mimicry and angiogenesis.

MALAT1 is an oncogenic long non-coding RNA that has been found to promote the proliferation of many malignant cell types and non-malignant human umbilical vein endothelial cells (HUVECs). However, the functions of MALAT1 in vasculogenic mimicry (VM) and angiogenesis and the potential mechanisms responsible have not yet been investigated in any malignancy. Here, in situ hybridization and CD31/periodic acid-Schiff double staining of 150 gastric cancer (GC) clinical specimens revealed that MALAT1 expression was tightly associated with densities of VM and endothelial vessels. MALAT1 knockdown markedly reduced GC cell migration, invasion, tumorigenicity, metastasis, and VM, while restricting HUVEC angiogenesis and increasing vascular permeability. Moreover, MALAT1 was found to regulate expression of VE-cadherin, β-catenin, MMPs 2 and 9, MT1-MMP, p-ERK, p-FAK, and p-paxillin, which have been established as classical markers of VM and angiogenesis and components of associated signaling pathways. Consistent with this, the p-ERK inhibitors U0126 and PD98059 both effectively blocked GC cell VM. In conclusion, MALAT1 can promote tumorigenicity and metastasis in GC by facilitating VM and angiogenesis via the VE-cadherin/β-catenin complex and ERK/MMP and FAK/paxillin signaling pathways.

Elevated Orai1 and STIM1 expressions upregulate MACC1 expression to promote tumor cell proliferation, metabolism, migration, and invasion in human gastric cancer.

ORAI calcium release-activated calcium modulator 1 (Orai1)- and stromal interacting molecule 1 (STIM1)-mediated store-operated Ca(2+) entry (SOCE) have been increasingly implicated in tumor progression; however, its role in gastric cancer (GC) is not well elucidated. We aimed to determine whether SOCE influences GC prognosis and elucidate the underlying mechanisms. Orai1 and STIM1 expressions were higher in GC tissues compared to adjacent non-tumor tissues according to RT-PCR and western blotting. Higher Orai1 and/or STIM1 expression was associated with more advanced disease, more frequent recurrence, and higher mortality rates in our study of 327 GC patients. The disease-free survival rates of Stage I-III patients and the overall survival rates of Stage IV patients were significantly worse when the tumors had high Orai1 and/or STIM1 expressions. Orai1 and/or STIM1 knockdown caused significantly reduced tumor growth and metastasis in athymic mice. Orai1 and/or STIM1 knockdown lowered the proliferation, metabolism, migration, and invasion of two GC cell lines. Also, Orai1 and/or STIM1 knockdown changed the markers of the cell cycle and epithelial-mesenchymal transition (EMT). These effects were reversed by metastasis-associated in colon cancer-1 (MACC1) overexpression. In summary, the composite molecules of SOCE suggest a poor prognosis for GC by promoting tumor cell proliferation, metabolism, migration, and invasion by targeting MACC1.

Development of trastuzumab-resistant human gastric carcinoma cell lines and mechanisms of drug resistance

Trastuzumab has been successfully employed for the treatment of Her-2-positive gastric cancer. However, there are problems with both primary and secondary resistance to trastuzumab. In this study, we employed the human gastric carcinoma cell line NCI-N87 with high Her-2 expression to create trastuzumab-resistant NCI-N87/TR cells by stepwise exposure to increasing doses of trastuzumab. Western blotting and Real-time PCR were conducted to detect protein and gene levels. Compared with NCI-N87 cells, the expression of P-IGF-1R and P-AKT proteins was significantly increased in NCI-N87/TR cells (both P = 0.000), while PTEN gene and protein expression showed a significant decrease (both P = 0.000). In addition, mutations of the PTEN gene were detected at exons 5, 7, and 8. The sensitivity of NCI-N87/TR cells to trastuzumab was increased by transfection with the PTEN gene, or by incubation with a PI3K inhibitor (LY294002) or an IGF-IR inhibitor (AG1024), as well as siRNA targeting PI3K p110 or IGF-1R. Taken together, our findings showed that activation of the PI3K-AKT signaling pathway was one of the major mechanisms leading to resistance of NCI-N87/TR gastric cancer cells to trastuzumab, which was probably associated with PTEN gene down-regulation and mutation, as well as with over-activity of the IGF-1R signaling pathway.

Multivariate Analysis of Several Molecular Markers and Clinicopathological Features in Postoperative Prognosis of Hepatocellular Carcinoma

This study was designed to assess the impact of several molecular markers and clinicopathological characteristics on postoperative survival of patients with hepatocellular carcinoma (HCC). Postoperative clinical data of 64 patients with HCC were retrospectively analyzed. K‐ras, PIK3CA, and BRAF gene mutations in surgically resected specimens of the 64 patients with HCC were detected by pyrosequencing. H‐ras and XB130 protein expression was examined by immunohistochemistry. A Cox proportional hazards regression model was used for univariate and multivariate survival analyses of the clinical and pathological parameters. The mutation rates of K‐ras, PIK3CA, and BRAF genes in HCC were found to be 4.69%, 1.56%, and 0%, respectively. Positive expression rate of XB130 and H‐ras in HCC was 75.0% and 93.8%, respectively. Univariate analysis revealed that clinicopathological factors impacting postoperative prognosis of patients with HCC include clinical stage, tumor diameter, and postoperative transcatheter arterial embolization therapy for HCC. Meanwhile, multivariate analysis showed that clinical stage (relative risk [RR]: 6.420, P = 0.013) and tumor diameter (RR: 1.498, P = 0.014) were independent factors impacting postoperative survival of patients with HCC. These findings indicate that the clinical stage and tumor diameter are independent risk factors impacting postoperative survival of patients with HCC. Gene mutations of K‐ras and PIK3CA and protein expression of XB130 and H‐ras are not associated with the postoperative prognosis of patients with HCC. Anat Rec, 2012.

FGF23 promotes myocardial fibrosis in mice through activation of β-catenin

Fibroblast growth factor 23 (FGF23) has been reported to induce left ventricular hypertrophy, but it remains unclear whether FGF23 plays a role in cardiac fibrosis. This study is attempted to investigate the role of FGF23 in post-infarct myocardial fibrosis in mice. We noted that myocardial and plasma FGF23 and FGF receptor 4 were increased in mice with heart failure as well as in cultured adult mouse cardiac fibroblasts (AMCFs) exposed to angiotensin II, phenylephrine, soluble fractalkine. Recombinant FGF23 protein increased active β-catenin , procollagen I and procollagen III expression in cultured AMCFs. Furthermore, intra-myocardial injection of adeno-associated virus-FGF23 in mice significantly increased left ventricular end-diastolic pressure and myocardial fibrosis, and markedly upregulated active β-catenin, transforming growth factor β (TGF-β), procollagen I and procollagen III in both myocardial infarction (MI) and ischemia/reperfusion (IR) mice, while β-catenin inhibitor or silencing of β-catenin antagonized the FGF23-promoted myocardial fibrosis in vitro and in vivo. These findings indicate that FGF23 promotes myocardial fibrosis and exacerbates diastolic dysfunction induced by MI or IR, which is associated with the upregulation of active β-catenin and TGF-β.

Microbubble-Mediated Sonothrombolysis Improves Outcome After Thrombotic Microembolism-Induced Acute Ischemic Stroke

Background and Purpose— Microthrombi originating from disintegrated clots or formed in situ may account for the poor clinical improvement of acute ischemic stroke after recanalization therapy. We attempted to determine whether microbubble-mediated sonothrombolysis could dissolve platelet-rich and erythrocyte-rich microthrombi, thereby reducing their brain injury-causing potential. Methods— Platelet- and erythrocyte-rich microthrombosis were induced by periadventitial application of 5% ferric chloride or thrombin to mesenteric microvessels in 75 Sprague–Dawley rats. Acute ischemic stroke was induced by intracarotid injection of platelet- or erythrocyte-rich microthrombi in another 50 rats. Rats were randomly divided into control (CON), ultrasound (US), ultrasound and microbubble (US+MB), recombinant tissue-type plasminogen activator (r-tPA), and US+MB+r-tPA groups. The post-treatment mesenteric microvessel recanalization rates, cerebral infarct volumes, and neurological scores were determined. Results— The recanalization rates of platelet- and erythrocyte-rich microthrombi in mesenteric microvessels were higher (P<0.05), and the cerebral infarct volumes and neurological scores of rats with either microthrombi were lower in the US+MB group than in the CON group (P<0.01). The infarct volumes and neurological scores were greater in the r-tPA group than in the US+MB and US+MB+r-tPA groups after treatment of rats with platelet-rich microthrombi (P<0.05). In contrast, after treatment of rats with erythrocyte-rich microthrombi, the infarct volumes and neurological scores were similar in the r-tPA and US+MB groups, but smaller in the US+MB+r-tPA group (P<0.05). Conclusions— Microbubble-mediated sonothrombolysis improved the outcomes of microthrombi-induced acute ischemic stroke. Thus, this method may serve as an attractive adjunct to recanalization therapy for acute ischemic stroke.

LC3A‐Positive “Stone‐Like” Structures Predict an Adverse Prognosis of Gastric Cancer

Microtubule‐associated protein light chain 3 (LC3A) is a reliable marker of autophagy that displays three distinct patterns of immunohistochemical staining in solid tumors: diffuse cytoplasmic staining, juxtanuclear staining, and staining of “stone‐like” structures. These three patterns have a different prognostic significance in many solid tumors, but little is known about their influence in gastric cancer (GC). This study was a retrospective analysis of 188 GC patients from stages I to IV. The pattern of LC3A expression was examined in tumor and nontumor tissues by immunohistochemistry. Then, the association between the pattern of LC3A expression in GC and the prognosis was investigated by Kaplan‐Meier analysis and the Cox proportional hazards model. Two distinct patterns of LC3A immunostaining (diffuse cytoplasmic expression and “stone‐like” structures) were observed in GC tissues. LC3A‐positive “stone‐like” structures were found only in the tumors, and the number of such structures was correlated with both the tumor type and tumor stage. In addition, a high number of LC3A‐positive “stone‐like” structures was closely associated with an increased risk of recurrence after radical resection of stages I–III cancer (P < 0.001; HR = 0.205) and was associated with a lower overall survival rate for stage IV cancer (P < 0.001; HR = 0.364). Taken together, our data demonstrate that LC3A‐positive “stone‐like” structures can be used as an independent biomarker for an adverse prognosis of GC, suggesting that “stone‐like” structures are correlated with the malignancy of this disease. Anat Rec, 297:653–662, 2014.

Theranostic, pH-Responsive, Doxorubicin-Loaded Nanoparticles Inducing Active Targeting and Apoptosis for Advanced Gastric Cancer

This study developed a kind of magnetic-polymer nanocarrier with folate receptor-targeting and pH-sensitive multifunctionalities to carry doxorubicin (DOX) for treatment of advanced gastric cancer (AGC). Folate-conjugated, pH-sensitive, amphiphilic poly(β-aminoester) self-assembled with hydrophobic oleic acid-modified iron oxide nanoparticles, and the resulting hydrophobic interaction area is a reservoir for lipophilic DOX (F-P-DOX). Confocal microscopy illustrated that F-P-DOX treatment could keep higher DOX accumulation in cells than P-DOX (without folate conjugation), and therefore get a higher efficiency of DOX internalization at pH 6.5 than at pH 7.4. Electron microscope characterization and real-time polymerase chain reaction revealed cell apoptosis promoted by F-P-DOX. The better efficacy of F-P-DOX on GC than free DOX and P-DOX was determined by MTT assay and xenograft model. Moreover, the accumulation of F-P-DOX in the tumor site was detected by magnetic resonance imaging (MRI). All those observations suggest F-P-DOX could be a promising theranostic candidate for AGC treatment.