Matrix metalloproteinase-mediated degradation of extracellular matrix is a crucial event for invasion and metastasis of malignant cells. The expressions of matrix metalloproteinases (MMPs) are regulated by different cytokines and growth factors. VEGF, a potent angiogenic cytokine, induces invasion of ovarian cancer cells through activation of MMPs. Here, we demonstrate that invasion and scattering in SKOV-3 cells were induced by VEGF through the activation of p38 MAPK and PI3K/AKT pathways. VEGF induced the expression of MMP-2, MMP-9, and MMP-13 and hence regulated the metastasis of SKOV-3 ovarian cancer cells, and the activities of these MMPs were reduced after inhibition of PI3K/AKT and p38 MAPK pathways. Interestingly, VEGF induced expression of ETS-1 factor, an important trans-regulator of different MMP genes. ETS-1 bound to both MMP-9 and MMP-13 promoters. Furthermore, VEGF acted through its receptor to perform the said functions. In addition, VEGF-induced MMP-9 and MMP-13 expression and in vitro cell invasion were significantly reduced after knockdown of ETS-1 gene. Again, VEGF-induced MMP-9 and MMP-13 promoter activities were down-regulated in ETS-1 siRNA-transfected cells. VEGF enriched ETS-1 in the nuclear fraction in a dose-dependent manner. VEGF-induced expression of ETS-1 and its nuclear localization were blocked by specific inhibitors of the PI3K and p38 MAPK pathways. Therefore, based on these observations, it is hypothesized that the activation of PI3K/AKT and p38 MAPK by VEGF results in ETS-1 gene expression, which activates MMP-9 and MMP-13, leading to the invasion and scattering of SKOV-3 cells. The study provides a mechanistic insight into the prometastatic functions of VEGF-induced expression of relevant MMPs.
"found that SAS cells stimulated with P. gingivalis exhibited sustained activation of Ets1 and phosphorylation of HSP27, while proMMP9 production was inhibited by knockdown of Ets1 or HSP27. Our findings are consistent with several previous reports, as proMMP9 expression was shown to be regulated by ERK1/2-Ets1 and p38/ HSP27 in a number of cell lines (Hansen et al., 2001; Liu et al., 2005; Ghosh et al., 2012), and knockdown of ERK1/2 and Ets1 decreased proMMP9 mRNA expression in response to TGFβ1 (Liu et al., 2005). Moreover, cancer cell migration and invasion were reported to be inhibited by knockdown of Ets1 and HSP27 (Hahne et al., 2005; Shin et al., 2005), and p38 deficiency inhibited HSP27 and MMP9 expression (Kumar et al., 2010). "
[Show abstract][Hide abstract] ABSTRACT: Recent epidemiological studies have revealed a significant association between periodontitis and oral squamous cell carcinoma (OSCC). Furthermore, matrix metalloproteinase 9 (MMP9) is implicated in the invasion and metastasis of tumor cells. We examined the involvement of Porphyromonas gingivalis, a periodontal pathogen, in OSCC invasion through induced expression of proMMP and its activation. proMMP9 was continuously secreted from carcinoma SAS cells, while P. gingivalis infection increased proenzyme expression and subsequently processed it to active MMP9 in culture supernatant, which enhanced cellular invasion. In contrast, Fusobacterium nucleatum, another periodontal organism, failed to demonstrate such activities. The effects of P. gingivalis were observed with highly invasive cells, but not with the low invasive type. P. gingivalis also stimulated proteinase-activated receptor 2 (PAR2) and enhanced proMMP9 expression, which promoted cellular invasion. P. gingivalis mutants deficient in gingipain proteases failed to activate MMP9. Infected SAS cells exhibited activation of ERK1/2, p38, and NF-kB, and their inhibitors diminished both proMMP9-overexpression and cellular invasion. Together, our results show that P. gingivalis activates the ERK1/2-Ets1, p38/HSP27, and PAR2/NFκB pathways to induce proMMP9 expression, after which the proenzyme is activated by gingipains to promote cellular invasion of OSCC cell lines. These findings suggest a novel mechanism of progression and metastasis of OSCC associated with periodontitis.
"Ets-1 upregulation appears to associate specifically with more advanced, invasive tumors in breast and ovarian carcinomas [17-22], and is positively correlated with the enhanced metastatic potential of numerous cancers [17,23-26]. Indeed, there are many well-established target genes for Ets-1 that are closely linked to cancer progression, particularly mediators of extracellular matrix degradation, cancer cell migration and angiogenesis [16,25,27-31]. Thus, the consequences of Ets-1 overexpression are particularly relevant to the study of ovarian cancer as this type of malignancy is very difficult to detect, and is most commonly diagnosed at advanced stages of disease progression that include metastases. "
[Show abstract][Hide abstract] ABSTRACT: The Ets-1 proto-oncogene is frequently upregulated in cancer cells, with known involvement in cancer angiogenesis, metastasis, and more recently energy metabolism. In this study we have performed various bioinformatic analyses on existing microarray data to further clarify the role of Ets-1 in ovarian cancer, and validated these results with functional assays.
Functional pathway analyses were conducted on existing microarray data comparing 2008 and 2008-Ets1 ovarian cancer cells. Methods included over-representation analysis, functional class scoring and pathway topology, and network representations were visualized in Cytoscape. Oxidative stress regulation was examined in ovarian cancer cells by measuring protein expression and enzyme activity of glutathione peroxidases, as well as intracellular reactive oxygen species using dichlorofluorescin fluorescence. A stable Ets-1 knockdown MDA-MB-231 cell line was created using short hairpin RNA, and glycolytic dependence of these cells was measured following treatment with 2-deoxy-D-glucose and Hoechst nuclear staining to determine cell number. High-resolution respirometry was performed to measure changes in basal oxygen flux between MDA-MB-231 cells and MDA-Ets1KD variants.
Enrichments in oxidoreductase activity and various metabolic pathways were observed upon integration of the different analyses, suggesting that Ets-1 is important in their regulation. As oxidative stress is closely associated with these pathways, we functionally validated our observations by showing that Ets-1 overexpression resulted in decreased reactive oxygen species with increased glutathione peroxidase expression and activity, thereby regulating cellular oxidative stress. To extend our findings to another cancer type, we developed an Ets-1 knockdown breast cancer cell model, which displayed decreased glycolytic dependence and increased oxygen consumption following Ets-1 knockdown confirming our earlier findings.
Collectively, this study confirms the important role of Ets-1 in the regulation of cancer energy metabolism in ovarian and breast cancers. Furthermore, Ets-1 is a key regulator of oxidative stress in ovarian cancer cells by mediating alterations in glutathione antioxidant capacity.
"It is quite possible that βhCG/hCG induces the generation of a single molecule which in turn leads to the generation of the others via autocrine loops. Indeed, while both IL-8 and VEGF induce the production of MMP-2 and MMP-9 , , IL8 up-regulates VEGF  and VEGF has been shown to enhance IL-8 levels . Whether such a scenario is at work here is under investigation. "
[Show abstract][Hide abstract] ABSTRACT: Human chorionic gonadotropin (hCG) was initially thought to be made only during pregnancy, but is now known to also be synthesized by a variety of cancers and is associated with poor patient prognosis. Transgenic expression of βhCG in mice causes hyper-luteinized ovaries, a loss in estrous cyclicity and infertility, increased body weight, prolactinomas and mammary gland tumors. Strategies were devised to generate antibody responses against hCG to investigate whether reversal of the molecular processes driving tumorigenesis would follow. hCG-immunized transgenic mice did not exhibit increases in body weight or serum prolactin levels, and gross ovarian and pituitary morphology remained normal. While non-immunized transgenic animals demonstrated heightened levels of transcripts associated with pituitary tumorigenesis (HMG2A, E2F1, CCND1, PRL, GH, GAL, PTTG1, BMP4) and decreased levels of CDK inhibitors CDKN1B (p27), CDKN2A (p16) and CDKN2c (p18), immunization led to a reversal to levels found in non-transgenic animals. Serum derived from transgenic (but not non-transgenic) mice led to enhanced transcription as well as expression of VEGF, IL-8, KC (murine IL-8) and MMP-9 in tumor cells, effects not seen when sera derived from hCG-immunized transgenic mice was employed. As the definitive indication of the restoration of the reproductive axis, immunization led to the resumption of estrous cyclicity as well as fertility in transgenic mice. These results indicate that hCG may influence cancer pathogenesis and progression via several distinct mechanisms. Using a stringent in vivo system in which βhCG acts both a "self" antigen and a tumor-promoting moiety (putatively akin to the situation in humans), the data builds a case for anti-gonadotropin vaccination strategies in the treatment of gonadotropin-dependent or secreting malignancies that frequently acquire resistance to conventional therapy.
PLoS ONE 11/2012; 7(11):e51125. DOI:10.1371/journal.pone.0051125 · 3.23 Impact Factor
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