AlphaVbeta6 integrin promotes invasion of squamous carcinoma cells through up-regulation of matrix metalloproteinase-9

The integrin alphaVbeta6 is a fibronectin receptor, which is not detectable on normal epithelium but is neo-expressed in oral epithelial dysplasia and oral squamous-cell carcinoma (SCC), suggesting a role in promoting malignant behaviour and tumour progression. We used transfection and retroviral infection to create a panel of SCC cell lines expressing various levels of alphaVbeta6 to examine this possibility. We found that increased expression of alphaVbeta6 in malignant keratinocytes up-regulates MMP-9 and MMP-2 expression and promotes invasion in an MMP-9-dependent manner. Our results suggest a possible mechanism for the involvement of alphaVbeta6 in squamous carcinoma in vivo.     

Microtubule-dependent matrix metalloproteinase-2/matrix metalloproteinase-9 exocytosis: prerequisite in human melanoma cell invasion

Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that cleave and degrade a wide spectrum of extracellular matrix components. By enhancing turnover of extracellular matrix, MMP activity is also known to play a key role in tumor cell invasion. Because extracellular protease activity requires efficient release of these proteases to the cellular surface, we investigated storage, transport, and exocytosis of MMP-2 and MMP-9 in human melanoma cells using immunofluorescence, electrical, and biochemical techniques. Immunolabeling of melanoma cells with antibodies specific for MMP-2 and MMP-9 led to the identification of two distinct populations of small cytoplasmatic vesicles containing MMP-2 or MMP-9, respectively. In combination with alpha-tubulin-specific antibodies, both vesicle populations were found to be aligned along the microtubular network. Moreover, the molecular motor protein kinesin is shown to be localized on most of these vesicles, providing evidence that the identified vesicles are actively propelled along microtubules toward the plasma membrane. The functional relevance of these findings is demonstrated using low dosage (5.9 nmol/L) of paclitaxel to affect the microtubular function of melanoma cells. Although cell proliferation is not altered, paclitaxel treatment impairs secretion of MMP-2/MMP-9 and significantly reduces invasive activity in our new cell invasion assay. In conclusion, we demonstrate in melanoma cells that microtubule-dependent traffic of MMP-containing vesicles and exocytosis are critical steps for invasive behavior and therefore are potential targets for specific antitumor drugs.     

Relationship between expression of matrix metalloproteinase-2 and matrix metalloproteinase-9 and invasion ability of cervical cancer cells

Constitutive overexpression of matrix metalloproteinases (MMPs) is frequently observed in malignant tumors. MMPs are a family of zinc endopeptidases consisting of at least 20 different members. In particular, MMP-2 and MMP-9 are reported to be closely associated with invasion and metastasis in several cancers. We investigated whether expression of MMP-2 and MMP-9 is associated with invasion ability of seven cervical cancer cells by administration of o-phenanthroline as MMP inhibitor. In two cell lines, Siha and Caski, MMP-2 mRNA and protein were expressed at high levels. After treatment with o-phenanthroline, the rate of invasion in these two cell lines was significantly decreased. In contrast, in the other two cell lines, HT-3 and Caski, high levels of MMP-9 mRNA and protein were expressed but there was no decrease in the rate of invasion in these cells after treatment with o-phenanthroline. The data suggest that expression level of MMP-2 mRNA may regulate with invasion ability of cervical cancer.     

Tenascin-C stimulates glioma cell invasion through matrix metalloproteinase-12

The capacity of glioma cells to invade extensively within the central nervous system is a major cause of the high morbidity rate of primary malignant brain tumors. Glioma cell invasion involves the attachment of tumor cells to extracellular matrix (ECM), degradation of ECM components, and subsequent penetration into adjacent brain structures. These processes are accomplished in part by matrix metalloproteinases (MMP) within a three-dimensional milieu of the brain parenchyma. As the majority of studies have used a two-dimensional monolayer culture system, we have used a three-dimensional matrix of collagen type I gel to address glioma-secreted proteases, ECM, and invasiveness of glioma cells. We show that in a three-dimensional collagen type I matrix, the presence of tenascin-C, commonly elevated in high-grade gliomas, increased the invasiveness of glioma cells. The tenascin-C-mediated invasiveness was blocked by metalloproteinase inhibitors, but this did not involve the gelatinases (MMP-2 and MMP-9) commonly implicated in two-dimensional glioma growth. A thorough analysis of 21 MMPs and six members of a disintegrin and metalloproteinase domain showed that MMP-12 was increased in gliomas by tenascin-C in three-dimensional matrix. Furthermore, examinations of resected specimens revealed high MMP-12 levels in the high-grade glioblastoma multiforme tumors. Finally, a function-blocking antibody as well as small interfering RNA to MMP-12 attenuated the tenascin-C-stimulated glioma invasion. These results identify a new factor, MMP-12, in regulating glioma invasiveness through interaction with tenascin-C.     

LMO2蛋白在前列腺基质细胞中介导的IL-11、FGF-9旁分泌促进前列腺癌细胞增殖与侵袭

背景与目的：前列腺癌多发生于前列腺外周带,前列腺增生多发于前列腺移行带。前列腺疾病的带性差异机制可能与前列腺组织微环境有关。该研究的前期研究提示,不同区带来源的前列腺基质细胞对上皮细胞的作用存在明显差异,基因芯片筛查发现LMO2蛋白在前列腺外周带基质细胞高表达与前列腺癌发生、发展密切相关。该研究旨在分析前列腺基质细胞LMO2基因的表达对前列腺癌细胞系增殖、侵袭能力的影响及其机制。方法：分别应用慢病毒过表达载体和短发卡RNA(shRNA)建立过表达和低表达LMO2的前列基质细胞,利用实时荧光定量聚合酶链反应(real-time lfuorescent quantitative polymerase chain reaction,RTFQ-PCR)、蛋白[质]印迹法(Western blot)分别检测LMO2 mRNA和蛋白的表达;将不同处理的前列腺基质细胞分别同PC-3细胞共培养,利用CCK-8检测PC-3的增殖能力,利用基质胶侵袭实验检测PC-3的侵袭能力;利用生物素标记的人蛋白抗体芯片检测过表达LMO2的前列腺基质细胞条件培养基中蛋白因子表达变化。结果：成功建立过表达及低表达LMO2的前列腺基质细胞;CCK-8实验及基质胶实验提示,与过表达LMO2的前列腺WPMY-1基质细胞共培养后,PC-3细胞的增殖和侵袭能力增强;与低表达LMO2的CAFs细胞共培养后,PC-3细胞的增殖和侵袭能力降低;蛋白芯片检测发现过表达LMO2后,前列腺外周带基质细胞分泌白介素-11(interleukin-11, IL-11)和成纤维细胞生长因子-9(ifbroblast grouth factor-9,FGF-9)增多。结论：LMO2基因在前列腺外周基质细胞中的高表达可能与前列腺癌的发生、发展有关;过表达LMO2的前列腺基质细胞通过旁分泌IL-11、FGF-9等细胞因子促进前列腺癌细胞增殖与侵袭。     

Platelet-derived growth factor-D promotes ovarian cancer invasion by regulating matrix metalloproteinases 2 and 9

Objective: Platelet-derived growth factor-D (PDGF-D) can enhance invasion and metastasis in several human malignancies, though little is known about its functions in ovarian cancer. Methods: In this study, we detected expression of PDGF-D in ovarian cancer tissues and cell lines by qRT-PCR, immunohistochemistry and western blotting, investigating the influences on cellular proliferation, invasion and apoptosis by upregulating its expression. Results: 79.5% (62/78) of ovarian cancer samples proved to be PDGF-D positive, in contrast to just 38.5%(30/78) in their adjacent non-cancer tissues (p<0.001). Moreover, we found high levels of PDGF-D were correlated with lymph node metastasis (p=0.025) and positive cancer cells in abdominal washings/ascites (p=0.042). In vitro, upregulation of PDGF-D enhanced the invasiveness of SKOV3 cells (p<0.01), but had no impact on cellular proliferation or apoptosis. Furthermore, expression of matrix metalloproteinases 2/9 (MMP2 and MMP9) was positively related with PDGF-D, indicating their involvement in the invasion and metastasis of ovarian cancer. Conclusions: Our findings proved that PDGF-D could promote ovarian cancer invasion by upregulating MMPs, which might be a potential target for ovarian cancer treatment.     

Bcl-w promotes gastric cancer cell invasion by inducing matrix metalloproteinase-2 expression via phosphoinositide 3-kinase, Akt, and Sp1

Given a previous report that Bcl-w is expressed in gastric cancer cells, particularly in those of an infiltrative morphology, we investigated whether Bcl-w expression influences the invasiveness of gastric cancer cells. To accomplish this, Bcl-w was overexpressed in adherent types of gastric adenocarcinoma cell lines, and this was found to result in an increase in their migratory and invasive potentials. These effects were not induced when Bcl-2 was overexpressed in the same cell types. Consistently, Bcl-w, but not Bcl-2, overexpression increased matrix metalloproteinase-2 (MMP-2) expression, and synthetic or natural inhibitors of MMP-2 abolished Bcl-w-induced cell invasion. Bcl-w overexpression also activated phosphoinositide 3-kinase (PI3K), Akt, and Sp1, and the blocking effects of each of these components using pharmacologic inhibitors, dominant-negative mutants, or small interfering RNA abolished the ability of Bcl-w to induce MMP-2 and cell invasion. The inhibition of PI3K/Akt signaling also prevented Sp1 activation. Overall, our data suggest that Bcl-w, which was previously shown to enhance gastric cancer cell survivability, also promotes their invasiveness by inducing MMP-2 expression via the sequential actions of PI3K, Akt, and Sp1.     

Correlation between expression of matrix metalloproteinase-2, matrix metalloproteinase-9 and angiogenesis in gastric cancer

Objective： To investigate the relationship between matrix metalloproteinase-2 （MMP-2）, MMP-9 as well as microvessel density （MVD） and their clinicopathological features in gastric cancer. Methods： The expression of MMP-2, MMP-9 and MVD were detected by immunohistochemistry SP method on 65 cases of gastric cancer tissue and 32 adjacent gastric mucosa. Results： The positive expression of MMP-2, MMP-9 and MVD in cancer group were significantly higher than those of adjacent mucosa group （P〈0.01）. The positive expression of MM P-2 and MMP-9 had closely correlation with clinical features of lymphatic metastasis, pathological type and stages （P〈0.05）, and the high level of CD 34 had correlation with metastasis and stages （P〈0.01）. The positive expression of MMP-2 and MMP-9 showed significant correlation with the level of MVD. Prognosis was mostly affected by lymphatic metastasis and stages of clinical features. The low cum survival rate was showed in the groups of positive expression of MMP-2, MMP-9 and high level of MVD. Conclusion： MMP-2, MMP-9 and MVD play an important role in metastases, invasion and prognosis of gastric cancer, which is a valuable maker to evaluate the prognosis.     

塔斯品碱对A549细胞MMP-2与MMP-9表达的影响

目的:探讨塔斯品碱对A549细胞基质金属蛋白酶MMP-2、-9表达的影响,阐明塔斯品碱抑制肿瘤血管生成的作用机制。方法:采用免疫细胞化学SABC法和RT-PCR法检测MMP-2和MMP9在A549细胞中的表达。结果:塔斯品碱对A549细胞中MMP-2、-9蛋白及其mRNA的表达均显示一定的抑制作用并有剂量依赖关系(P<0.05)。结论:塔斯品碱抑制A549细胞中与血管生成密切相关的基质金属蛋白酶MMP-2、-9的表达,有可能用于肿瘤抗血管治疗。     

Nicotinamide N-methyltransferase induces cellular invasion through activating matrix metalloproteinase-2 expression in clear cell renal cell carcinoma cells

Nicotinamide N-methyltransferase (NNMT) was recently identified as one clear cell renal cell carcinoma (ccRCC)-associated gene by analyzing full-length complementary DNA-enriched libraries of ccRCC tissues. The aim of this study is to investigate the potential role of NNMT in cellular invasion. A strong NNMT expression is accompanied with a high invasive activity in ccRCC cell lines, and small interfering RNA-mediated NNMT knockdown effectively suppressed the invasive capacity of ccRCC cells, whereas NNMT overexpression markedly enhanced that of human embryonic kidney 293 (HEK293) cells. A positive correlation between the expression of NNMT and matrix metallopeptidase (MMP)-2 was found in ccRCC cell lines and clinical tissues. The treatment of blocking antibody or inhibitor specific to MMP-2 significantly suppressed NNMT-dependent cellular invasion in HEK293 cells. Furthermore, SP-1-binding region of MMP-2 promoter was found to be essential in NNMT-induced MMP-2 expression. The specific inhibitors of PI3K/Akt signaling markedly decreased the binding of SP1 to MMP-2 promoter as shown by chromatin immunoprecipitation assay. We also demonstrated that PI3K/Akt pathway plays a role in NNMT-dependent cellular invasion and MMP-2 activation. Moreover, short hairpin RNA-mediated knockdown of NNMT expression efficiently inhibited the growth and metastasis of ccRCC cells in non-obese diabetic severe combined immunodeficiency mice. Taken together, the present study suggests that NNMT has a crucial role in cellular invasion via activating PI3K/Akt/SP1/MMP-2 pathway in ccRCC.     

Cannabinoids inhibit glioma cell invasion by down-regulating matrix metalloproteinase-2 expression

Cannabinoids, the active components of Cannabis sativa L. and their derivatives, inhibit tumor growth in laboratory animals by inducing apoptosis of tumor cells and impairing tumor angiogenesis. It has also been reported that these compounds inhibit tumor cell spreading, but the molecular targets of this cannabinoid action remain elusive. Here, we evaluated the effect of cannabinoids on matrix metalloproteinase (MMP) expression and its effect on tumor cell invasion. Local administration of Delta(9)-tetrahydrocannabinol (THC), the major active ingredient of cannabis, down-regulated MMP-2 expression in gliomas generated in mice, as determined by Western blot, immunofluorescence, and real-time quantitative PCR analyses. This cannabinoid-induced inhibition of MMP-2 expression in gliomas (a) was MMP-2-selective, as levels of other MMP family members were unaffected; (b) was mimicked by JWH-133, a CB(2) cannabinoid receptor-selective agonist that is devoid of psychoactive side effects; (c) was abrogated by fumonisin B1, a selective inhibitor of ceramide biosynthesis; and (d) was also evident in two patients with recurrent glioblastoma multiforme. THC inhibited MMP-2 expression and cell invasion in cultured glioma cells. Manipulation of MMP-2 expression by RNA interference and cDNA overexpression experiments proved that down-regulation of this MMP plays a critical role in THC-mediated inhibition of cell invasion. Cannabinoid-induced inhibition of MMP-2 expression and cell invasion was prevented by blocking ceramide biosynthesis and by knocking-down the expression of the stress protein p8. As MMP-2 up-regulation is associated with high progression and poor prognosis of gliomas and many other tumors, MMP-2 down-regulation constitutes a new hallmark of cannabinoid antitumoral activity.     

Loss of hSef promotes metastasis through upregulation of EMT in prostate cancer

We have previously reported that the negative signaling regulator Similar Expression to FGF (hSef) is downregulated in prostate cancer and its loss is associated with clinical metastasis. Here, we explored the mechanistic basis of this finding. We first confirmed our clinical observation by testing hSef manipulation in an in vivo metastasis model. hSef stable expressing cells (PC3M‐hSef) or empty vector controls (PC3M‐EV) were injected subcutaneously into the lateral thoracic walls of NOD‐SCID gamma mice and lungs were harvested at autopsy. In this model, 6/7 PC3M‐EV xenografts had definitive lung micro‐metastasis whilst only 1/6 PC3M‐hSef xenografts exhibited metastasis recapitulating the clinical scenario (p = 0.03). Gene expression studies revealed key perturbations in genes involved in cell motility and epithelial to mesenchymal transition (EMT) along with alterations in cognate signaling pathways. These results were validated in an EMT specific PCR array whereby hSef over‐expression and silencing reciprocally altered E‐Cadherin expression (p = <0.001) amongst other EMT markers. Immunohistochemistry of excised tumors from the xenografts also confirmed the effect of hSef in suppressing E‐Cadherin expression at the protein level. Phosphokinase arrays further demonstrated a role for hSef in attenuating signaling of not only ERK‐MAPK but also the JNK and p38 pathways as well. Taken together, these data suggest evidence that loss of hSef may be a critical event facilitating tumor dissemination of prostate cancer through alteration of EMT. Detection of downregulated hSef, along with other negative regulators, may therefore be a useful biomarker heralding a transition to a metastatic phenotype and warrants further exploration in this context.     

Protein arginine methyltransferase 7 promotes breast cancer cell invasion through the induction of MMP9 expression

Recent evidence points to the protein arginine methyltransferase (PRMT) family of enzymes playing critical roles in cancer. PRMT7 has been identified in several gene expression studies to be associated with increased metastasis and decreased survival in breast cancer patients. However, this has not been extensively studied. Here we report that PRMT7 expression is significantly upregulated in both primary breast tumour tissues and in breast cancer lymph node metastases. We have demonstrated that reducing PRMT7 levels in invasive breast cancer cells using RNA interference significantly decreased cell invasion in vitro and metastasis in vivo. Conversely, overexpression of PRMT7 in non-aggressive MCF7 cells enhanced their invasiveness. Furthermore, we show that PRMT7 induces the expression of matrix metalloproteinase 9 (MMP9), a well-known mediator of breast cancer metastasis. Importantly, we significantly rescued invasion of aggressive breast cancer cells depleted of PRMT7 by the exogenous expression of MMP9. Our results demonstrate that upregulation of PRMT7 in breast cancer may have a significant role in promoting cell invasion through the regulation of MMP9. This identifies PRMT7 as a novel and potentially significant biomarker and therapeutic target for breast cancer.     

ARHGEF40通过上皮间质转化促进结肠癌细胞侵袭

目的:研究鸟嘌呤核苷酸交换因子40(recombinant human rho guanine nucleotide exchange factor 40,ARHGEF40)对结肠癌细胞侵袭能力和上皮间质转化(epithelial-mesenchymal transition,EMT)标志蛋白表达的影响,探索ARHGE...     

Expression and clinical significance of matrix metalloproteinase-2 and matrix metalloproteinase-9 in oral squamous cell carcinoma

To successfully establish a metastasis from an invasive carcinoma, the first step involves the degradation of the underlying basement membrane, which is mainly made up of type IV collagen. Matrix metalloproteinases (MMPs)-2 and -9 are thought to play an important role in its degradation because of their ability to destroy this type of collagen. In order to evaluate the prognostic significance of these proteases, we studied the expression of MMP-2 and -9 in series of 68 OSCC by immunohistochemistry. Of the oral carcinomas, 28% (n = 19) expressed MMP-2, and 17.6% (n = 12) expressed MMP-9. MMP-2 immunoreactivity was significantly higher in patients with alcohol consumption (p = 0.028) (OR = 4), and in those younger than 60 years (p = 0.041). MMP-9 immunostaining showed statistically significant association with the tumor grade of differentiation (p = 0.019), the T-stage (p = 0.05), and also with the alcohol intake (p = 0.04) (OR = 7.67). In the present study, although not statistically significant, we observed that immunoexpression of MMP-2 and MMP-9 was stronger in patients with lymph node metastasis (OR = 1.65 and 2.29, respectively). In patients without regional lymph node metastasis, positive MMP-9 immunostaining was related to poor survival rates (p = 0.02; OR = 5.8). MMP-2 and -9 are involved in the invasion process of oral cancer, and MMP-9 is related to poor prognosis in the subset of patients without neck node metastasis. Ethanol could enhance the carcinogenetic process in oral cavity through its influence in the expression of MMP-2 and -9.