Cancer/testis antigen LDHC promotes proliferation and metastasis by activating the PI3K/Akt/GSK-3β-signaling pathway and the in lung adenocarcinoma

The cancer/testis antigen lactate dehydrogenase-C4 (LDHC) is a specific isoenzyme of the LDH family that regulates invasion and metastasis in some malignancies; however, little is known regarding its role in progression of lung adenocarcinoma (LUAD). Thus, we investigated LDHC expression by immunohistochemistry, and analyzed its clinical significance in 88 LUAD specimens. The role and molecular mechanisms subserving LDHC in cellular proliferation, migration, and invasion were explored both in vitro and in vivo. As a result, we found that high LDHC expression was significantly correlated with clinicopathological features of aggressive LUAD and a poor prognosis. Overexpression of LDHC induced LUAD cells to produce lactate and ATP, increased their metastatic and invasive potential—, and accelerated xenograft tumor growth. We further demonstrated that overexpression of LDHC affected the expression of cell proliferation-related proteins (cyclin D1 and c-Myc) and epithelial-mesenchymal transition (EMT)-related proteins (MMP-2, MMP-9, E-cadherin, Vimentin, Twist, Slug, and Snail) both in vitro and in vivo. Finally, excessive activation of LDHC enhanced the phosphorylation levels of AKT and GSK-3β, revealing activation of the PI3K/Akt/GSK-3β oncogenic-signaling pathways. Treatment with a PI3K inhibitor reversed the effects of LDHC overexpression by inhibiting cellular proliferation, migration, and invasion, with diminished levels of p-Akt and p-GSK3β. PI3K inhibition also reversed cell proliferation-related and EMT-related proteins in LDHC-overexpressing A549 cells. In conclusion, LDHC promotes proliferation, migration, invasion, and EMT in LUAD cells via activation of the PI3K/Akt/GSK-3β pathway.     

Focal adhesion kinase mediates TGF-β1-induced renal tubular epithelial-to-mesenchymal transition in vitro

Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase which participates in many important cellular processes such as cell adhesion and migration. However, the role of FAK in renal tubular epithelial-to-mesenchymal transition (EMT) is still unknown. FAK was knocked down by transfection of specific small interfering RNA (siRNA) in cultured HK-2 cells, then the cells were stimulated with transforming growth factor-beta 1 (TGF-β1). The expression of FAK, α-smooth muscle actin (α-SMA),E-cadherin, Akt, matrix metallopeptidase-9 (MMP-9),tissue inhibitor of metalloproteinase-1 (TIMP-1), and collagen IV were detected by RT-PCR, Western blot and immunofluorescence methods, respectively. Cell migration was determined by transwell assay. The results suggest that the expression of FAK was up-regulated in HK-2 cells when incubated with TGF-β1(10 μg/l), which was accompanied by reduced expression of E-cadherin and increased expression of α-SMA. All these changes were restored by FAK siRNA. Akt phosphorylation was induced by the treatment with TGF-β1, which was blocked by FAK siRNA. TGF-β1-induced down-regulation of E-cadherin was recovered by a PI3K/Akt inhibitor, LY294002, without affecting the expression of FAK. Functionally, TGF-β1 induced an increase in MMP-9 expression, as well as decreased expression of TIMP-1 and collagen IV, which were all restored by the FAK siRNA transfection. In addition, FAK siRNA significantly reduced TGF-β1-induced cells migration. In conclusion, FAK may play a crucial role in mediating TGF-β1-induced EMT through the activation of Akt pathway.     

Group IB secretory phospholipase A2 promotes matrix metalloproteinase-2-mediated cell migration via the phosphatidylinositol 3-kinase and Akt pathway

Secretory phospholipase A(2) (sPLA(2)), abundantly expressed in various cells including fibroblasts, is able to promote proliferation and migration. Degradation of collagenous extracellular matrix by matrix metalloproteinase (MMP) plays a role in the pathogenesis of various destructive disorders, such as rheumatoid arthritis, tumor invasion, and metastasis. Here we show that group IB PLA(2) increased pro-MMP-2 activation in NIH3T3 fibroblasts. MMP-2 activity was stimulated by group IB PLA(2) in a dose- and time-dependent manner. Consistent with MMP-2 activation, sPLA(2) decreased expression of type IV collagen. These effects are due to the reduction of tissue inhibitor of metalloproteinase-2 (TIMP-2) and the activation of the membrane type1-MMP (MT1-MMP). The decrease of TIMP-2 levels in conditioned media and the increase of MT1-MMP levels in plasma membrane were observed. In addition, treatment of cells with decanoyl Arg-Val-Lys-Arg-chloromethyl ketone, an inhibitor of pro-MT1-MMP, suppressed sPLA(2)-mediated MMP-2 activation, whereas treatment with bafilomycin A1, an inhibitor of H(+)-ATPase, sustained MMP-2 activation by sPLA(2). The involvement of phosphatidylinositol 3-kinase (PI3K) and Akt in the regulation of MMP-2 activity was further suggested by the findings that PI3K and Akt were phosphorylated by sPLA(2). Expression of p85alpha and Akt mutants, or pretreatment of cells with LY294002, a PI3K inhibitor, attenuated sPLA(2)-induced MMP-2 activation and migration. Taken together, these results suggest that sPLA(2) increases the pro-MMP-2 activation and migration of fibroblasts via the PI3K and Akt-dependent pathway. Because MMP-2 is an important factor directly involved in the control of cell migration and the turnover of extracellular matrix, our study may provide a mechanism for sPLA(2)-promoted fibroblasts migration.     

Effect of IL-17A on the Migration and Invasion of NPC Cells and Related Mechanisms

In carcinogenesis, inflammasomes may play contradictory roles through facilitating anti-tumor immunity or inducing oncogenic factors. Their function in cancer remains poorly characterized. In this study, we explored the effect of interleukin-17A (IL-17A) on the migration and invasion activity of nasopharyngeal carcinoma (NPC) cell lines and account for related mechanisms. Our results revealed that exogenous IL-17A promoted cell migration and invasion significantly in both NPC-039 and CNE-2Z cell lines. In addition, the expression of matrix metalloproteinase-2 (MMP-2)/-9 and Vimentin could be elevated by IL-17A stimulation; meanwhile the expression of E-cadherin was decreased. The results also show that IL-17A could activate the p38 signaling pathway in IL-17A-stimulated NPC-039 and CNE-2Z cell lines. Combining treatment with a p38 inhibitor (SB203580) resulted in decreased invasion capabilities of NPC-039 and CNE-2Z cell lines. SB203580 also inhibited the expression of MMP-2/-9 and increased the expression of E-cadherin in IL-17A-stimulated NPC-039 and CNE-2Z cell lines. IL-17A also could activate NF-κB in NPC-039 and CNE-2Z cell lines. In summary, our data show that IL-17A promote the cell migration and invasion of NPC cells. The effect of IL-17A on cell migration and invasion may be mediated via regulation of the expression of MMP-2/-9 and epithelial-mesenchymal transition (EMT) via p38-NF-κB signaling pathway. Thus, IL-17A or its related signaling pathways may be a promising target for preventing and inhibiting NPC metastasis.     

Hepatocyte growth factor enhances proteolysis and invasiveness of human nasopharyngeal cancer cells through activation of PI3K and JNK

The hepatocyte growth factor (HGF) receptor, Met, is frequently overexpressed in nasopharyngeal cancer (NPC). Here, we showed for the first time that human NPC cells with high Met expression were more sensitive to the cell motility and invasion effect of HGF. The downregulation of Met by small interfering RNA decreased tumor cell invasion/migration. HGF significantly increased matrix metalloproteinase-9 production. This was inhibited by blocking phosphatidylinositide 3-kinase (PI3K) and c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase signaling pathways. We also demonstrated that PI3K induced activation of JNK, with Akt as a potential point of this cross-talk. These results provide new insights into the molecular mechanism responsible for NPC progression and metastasis.     

Mesothelin enhances invasion of ovarian cancer by inducing MMP-7 through MAPK/ERK and JNK pathways

Ovarian cancer has one of the highest mortalities in malignancies in women, but little is known of its tumour progression properties and there is still no effective molecule that can monitor its growth or therapeutic responses. MSLN (mesothelin), a secreted protein that is overexpressed in ovarian cancer tissues with a poor clinical outcome, has been previously identified to activate PI3K (phosphoinositide 3-kinase)/Akt signalling and inhibit paclitaxel-induced apoptosis. The present study investigates the correlation between MSLN and MMP (matrix metalloproteinase)-7 in the progression of ovarian cancer, and the mechanism of MSLN in enhancing ovarian cancer invasion. The expression of MSLN correlated well with MMP-7 expression in human ovarian cancer tissues. Overexpressing MSLN or ovarian cancer cells treated with MSLN showed enhanced migration and invasion of cancer cells through the induction of MMP-7. MSLN regulated the expression of MMP-7 through the ERK (extracellular-signal-regulated kinase) 1/2, Akt and JNK (c-Jun N-terminal kinase) pathways. The expression of MMP-7 and the migrating ability of MSLN-treated ovarian cancer cells were suppressed by ERK1/2- or JNK-specific inhibitors, or a decoy AP-1 (activator protein 1) oligonucleotide in in vitro experiments, whereas in vivo animal experiments also demonstrated that mice treated with MAPK (mitogen-activated protein kinase)/ERK- or JNK-specific inhibitors could decrease intratumour MMP-7 expression, delay tumour growth and extend the survival of the mice. In conclusion, MSLN enhances ovarian cancer invasion by MMP-7 expression through the MAPK/ERK and JNK signal transduction pathways. Blocking the MSLN-related pathway could be a potential strategy for inhibiting the growth of ovarian cancer.     

沉默脂肪细胞增强子结合蛋白2通过PI3K/Akt途径抑制肝癌细胞增殖和迁移

脂肪细胞增强子结合蛋白2(AEBP2)作为多梳抑制复合物2(PRC2)的组成蛋白质,参与多种肿瘤细胞的增殖和迁移,然而其在肝癌中的作用尚不清楚。本研究基于UALCAN和Kaplan-Meier Plotter数据库分析发现,AEBP2在肝癌组织中高表达,并且与患者的不良预后呈正相关。实时荧光定量PCR和蛋白质印迹结果证实,AEBP2在肝癌细胞中的表达高于正常肝细胞。在HepG2和Huh-7细胞中转染AEBP2 siRNA,平板克隆、CCK-8、流式细胞术、划痕愈合和Transwell结果显示,沉默AEBP2可以抑制肝癌细胞增殖、迁移和侵袭,并促进细胞凋亡(P＜0.05)。免疫荧光检测和蛋白质印迹结果显示,沉默AEBP2能够抑制肝癌细胞上皮-间质转化(EMT)(P＜0.05)。生物信息学分析结果表明,AEBP2参与调控PI3K/Akt信号通路。蛋白质印迹结果证实,沉默AEBP2能下调PI3K、p-AKT (S473)、mTOR、MMP-2和MMP-9的蛋白质表达水平(P＜0.05)。此外,沉默AEBP2对HepG2细胞迁移和侵袭的影响可被PI3K/Akt通路激动剂胰岛素样生长因子1(IGF-1)部分逆转(P＜0.01)。综上所述,AEBP2可能通过调节PI3K/Akt途径促进肝癌细胞增殖和迁移。本研究为AEBP2在肝癌中的作用提供理论依据。     

Inhibition of Autophagy Potentiated the Antitumor Effect of Nedaplatin in Cisplatin-Resistant Nasopharyngeal Carcinoma Cells

Nedaplatin, a cisplatin analog, was developed to reduce the toxicity of cisplatin, whereas it can be cross-resistant with cisplatin in some circumstances. This study aimed to investigate the role of autophagy in nedaplatin induced cell death in cisplatin-resistant nasopharyngeal carcinoma cells. Here, we showed that HNE1/DDP and CNE2/DDP cells were resistant to nedaplatin-induced cell death with reduced apoptotic activity. Nedaplatin treatment resulted in autophagosome accumulation and increased expression of LC3-II, indicating the induction of autophagy by nedaplatin in HNE1/DDP and CNE2/DDP cells. Inhibition of autophagy by Bafilomycin A1 (Baf A1) and 3-Methyladenine (3-MA) remarkably enhanced the antitumor efficacy of nedaplatin in HNE1/DDP and CNE2/DDP cells, suggesting that the resistance to nedaplatin-induced cell death was caused by enhanced autophagy in nedaplatin-resistant NPC cells. Additionally, Baf A1 enhanced reactive oxygen species (ROS) generation and apoptosis induced by nedaplatin in HNE1/DDP cells. Mechanistically, nedaplatin treatment caused activation of ERK1/2 and suppression of Akt/mTOR signaling pathways. While inhibition of ERK1/2 by MEK1/2 inhibitor, U0126, could reduce the expression of LC3-II in nedaplatin-resistant NPC cells. Furthermore, suppression of ROS could inhibit nedaplatin-induced ERK activation in HNE1/DDP cells, indicating that ROS and ERK were involved in nedaplatin-induced autophagy. Together, these findings suggested that autophagy played a cytoprotective role in nedaplatin-induced cytotoxicity of HNE1/DDP and CNE2/DDP cells. Furthermore, our results highlighted a potential approach to restore the sensitivity of cisplatin-resistant nasopharyngeal cancer cells to nedaplatin in combination with autophagy inhibitors.     

MTA1 Overexpression Induces Cisplatin Resistance Innasopharyngeal Carcinoma by Promoting Cancer Stem Cells Properties

Themetastasis-associated gene 1 (MTA1) oncogene hasbeen suggested to be involved in the regulation of cancer progression. However, there is still no direct evidence that MTA1 regulates cisplatin (CDDP) resistance, as well as cancer stem cell properties. In this study, we found that MTA1 was enriched in CNE1/CDDP cells. Knock down of MTA1 in CNE1/CDDP cells reversed CSCs properties and CDDP resistance. However, ectopic expression of MTA1 in CNE1 cells induced CSCs phenotypes and CDDP insensitivity. Interestingly, ectopic overexpression of MTA1-induced CSCs properties and CDDP resistance were reversed in CNE1 cells after inhibition of PI3K/Akt by {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002. In addition, MTA1 expression and Akt activity in CNE1/CDDP cells was much higher than that in CNE1 cells. These results suggested that MTA1 may play a critical role in promoting CDDP resistance in NPC cells by regulatingcancer stem cell properties via thePI3K/Akt signaling pathway. Our findings suggested that MTA1 may be a potential target for overcoming CDDP resistance in NPC therapy.     

Activation of EGFR promotes squamous carcinoma SCC10A cell migration and invasion via inducing EMT-like phenotype change and MMP-9-mediated degradation of E-cadherin

EGFR is a potent stimulator of invasion and metastasis in head and neck squamous cell carcinomas (HNSCC). However, the mechanism by which EGFR may stimulate tumor cell invasion and metastasis still need to be elucidated. In this study, we showed that activation of EGFR by EGF in HNSCC cell line SCC10A enhanced cell migration and invasion, and induced loss of epitheloid phenotype in parallel with downregulation of E-cadherin and upregulation of N-cadherin and vimentin, indicating that EGFR promoted SCC10A cell migration and invasion possibly by an epithelial to mesenchymal transition (EMT)-like phenotype change. Interestingly, activation of EGFR by EGF induced production of matrix metalloproteinase-9 (MMP-9) and soluble E-cadherin (sE-cad), and knockdown of MMP-9 by siRNA inhibited sE-cad production induced by EGF in SCC10A. Moreover, both MMP-9 knockdown and E-cadherin overexpression inhibited cell migration and invasion induced by EGF in SCC10A. The results indicate that EGFR activation promoted cell migration and invasion through inducing MMP-9-mediated degradation of E-cadherin into sE-cad. Pharmacologic inhibition of EGFR, MEK, and PI3K kinase activity in SCC10A reduced phosphorylated levels of ERK-1/2 and AKT, production of MMP-9 and sE-cad, cell migration and invasion, and expressional changes of EMT markers (E-cadherin and N-cadherin) induced by EGF, indicating that EGFR activation promotes cell migration and invasion via ERK-1/2 and PI3K-regulated MMP-9/E-cadherin signaling pathways. Taken together, the data suggest that EGFR activation promotes HNSCC SCC10A cell migration and invasion by inducing EMT-like phenotype change and MMP-9-mediated degradation of E-cadherin into sE-cad related to activation of ERK-1/2 and PI3K signaling pathways.     

4-Hydroxynonenal enhances MMP-2 production in vascular smooth muscle cells via mitochondrial ROS-mediated activation of the Akt/NF-kappaB signaling pathways

4-Hydroxynonenal (HNE) accumulates at atherosclerotic lesions, but its role in the progression of atherosclerosis is not clear. Considering the role of matrix metalloproteinases (MMP) in plaque destabilization, we investigated the mechanism by which HNE induces MMP production in vascular smooth muscle cells (VSMC). VSMC stimulated by HNE (1.0 microM) produced enzymatically active MMP-2 with an increased promoter activity, which was abolished by mutation of the NF-kappaB binding site in the promoter region. The increased NF-kappaB activity with subsequent MMP-2 production by HNE was significantly attenuated by transfection with Akt siRNA as well as by pretreatment with the PI3K/Akt inhibitors LY294002 (10 microM) and SH-5 (1.0 microM). The phosphorylation of Akt occurred as early as 5 min in VSMC exposed to HNE and was markedly attenuated by inhibition of mitochondrial reactive oxygen species (ROS). Furthermore, the impact of mitochondrial ROS on HNE-induced Akt phosphorylation with subsequent MMP-2 production was also demonstrated in mitochondrial function-deficient VSMC, as well as in cells transfected with manganese superoxide dismutase. Taken together, these results suggest that HNE enhances MMP-2 production in VSMC via mitochondrial ROS-mediated activation of the Akt/NF-kappaB signaling pathways.     

Down-Regulation of Gab1 Inhibits Cell Proliferation and Migration in Hilar Cholangiocarcinoma

Hilar cholangiocarcinoma is a highly aggressive malignancy originating from the hilar biliary duct epithelium. Due to few effective comprehensive treatments, the prognosis of hilar cholangiocarcinoma is poor. In this study, immunohistochemistry was first used to detect and analyze the expression of Gab1, VEGFR-2, and MMP-9 in hilar cholangiocarcinoma solid tumors and the relationships to the clinical pathological features. Furthermore, Gab1 and VEGFR-2 siRNA were used to interfere the hilar cholangiocarcinoma cell line ICBD-1 and then detect the PI3K/Akt signaling pathway, MMP-9 levels and malignant biological behaviors of tumor cells. The data showed that 1. Gab1, VEGFR-2, and MMP-9 were highly expressed and positively correlated with each other in hilar cholangiocarcinoma tissues, which were related to lymph node metastasis and differentiation. 2. After Gab1 or VEGFR-2 siRNA interference, PI3K/Akt pathway activity and MMP-9 levels were decreased in ICBD-1 cells. At the same time, cell proliferation decreased, cell cycle arrested in G1 phase, apoptosis increased and invasion decreased. These results suggest that the expression of Gab1, VEGFR-2, and MMP-9 are significantly related to the malignant biological behavior of hilar cholangiocarcinoma. Gab1 regulates growth, apoptosis and invasion through the VEGFR-2/Gab1/PI3K/Akt signaling pathway in hilar cholangiocarcinoma cells and influences the invasion of tumor cells via MMP-9.     

Inductive effect of phytoglycoprotein (38 kDa) on G0/G1 arrest and apoptosis in diethylnitrosamine-treated ICR mice

Preeclampsia complicates 5–10 % of pregnancies and is a leading cause of maternal/fetal morbidity and mortality. Although the cause is unknown, the reduced migration/invasion of extravillous trophoblasts is generally regarded as a key feature of preeclampsia genesis. The present study examined the expression of activator protein-2α (AP-2α), tissue inhibitor of metalloproteinase 2 (TIMP-2), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), and E-cadherin in severe preeclamptic placentas and normal placentas using real-time PCR and immunohistochemistry. The expression levels of AP-2α, TIMP-2, and E-cadherin were elevated, while MMP-2 and MMP-9 levels were decreased in severe preeclamptic placentas when compared with normal placentas. To explore the underlying molecular mechanisms, BeWo cells were transfected with an AP-2α-expression construct as well as a siRNA against AP-2α. The over-expression of AP-2α decreased the invasive abilities of BeWo cells. AP-2α induction was followed by the induction of TIMP-2 and E-cadherin and a significant reduction of MMP-2 and MMP-9. Whereas in AP-2α-silencing BeWo cells, we observed the decreased expression of TIMP-2 and E-cadherin and the increased expression of MMP-2 and MMP-9. We presume that AP-2α may suppress trophoblast invasion by repression of MMP-2 and MMP-9 and up-regulation of E-cadherin, thus leading to shallow placentation in severe preeclampsia.     

Comparison of doxycycline and minocycline in the inhibition of VEGF-induced smooth muscle cell migration

Smooth muscle cell migration plays an important role during angiogenesis and vascular remodeling. In this study, we examined the effects of doxycycline and minocycline on vascular endothelial growth factor (VEGF)-induced human aortic smooth muscle cell (HASMCs) migration, and explored the mechanisms in which doxycycline or minocycline inhibit HASMC migration. We demonstrated that both doxycycline and minocycline attain consistent anti-angiogenic effects in the inhibition of HASMC migration via a different signal pathway (p<0.05). This effect is through attenuating VEGF-induced matrix metalloproteinase-9 (MMP-9) activity (p<0.05). Doxycycline could increase tissue inhibitors of metalloproteinases-1 (TIMP-1) expression while minocycline down-regulated PI3K/Akt phosphorylation in HASMC. Our study suggests that doxycycline has a stronger ability to inhibit MMP secretion in HASMC by up-regulating endogenous MMPs inhibitor TIMP-1, while minocycline implements anti-angiogenic effect through inhibiting HASMC migration by down-regulating PI3K/Akt pathway.     

HGF and c-Met Interaction Promotes Migration in Human Chondrosarcoma Cells

Chondrosarcoma is a type of highly malignant tumor with a potent capacity for local invasion and causing distant metastasis. Chondrosarcoma shows a predilection for metastasis to the lungs. Hepatocyte growth factor (HGF) has been demonstrated to stimulate cancer proliferation, migration, and metastasis. However, the effect of HGF on migration activity of human chondrosarcoma cells is not well known. Here, we found that human chondrosarcoma tissues demonstrated significant expression of HGF, which was higher than that in normal cartilage. We also found that HGF increased the migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. c-Met inhibitor and siRNA reduced HGF-increased cell migration and MMP-2 expression. HGF treatment resulted in activation of the phosphatidylinositol 3′-kinase (PI3K)/Akt/PKCδ/NF-κB pathway, and HGF-induced expression of MMP-2 and cell migration was inhibited by specific inhibitors or siRNA-knockdown of PI3K, Akt, PKCδ, and NF-κB cascades. Taken together, our results indicated that HGF enhances migration of chondrosarcoma cells by increasing MMP-2 expression through the c-Met receptor/PI3K/Akt/PKCδ/NF-κB signal transduction pathway.