Regulation Of Matrix Metalloproteinases Research Articles

Anti-IL-20 monoclonal antibody inhibited tumor growth in hepatocellular carcinoma

Interleukin (IL)-20 is a proinflammatory cytokine involved in rheumatoid arthritis, atherosclerosis, and osteoporosis. However, the role of IL-20 in hepatocellular carcinoma (HCC) is unclear. We explored the function of IL-20 in HCC. Tumor tissue samples were analyzed the expression of IL-20 and cyclin D1 by using immunohistochemistry staining and quantitative real-time polymerase chain reaction (qRT-PCR) analysis. To examine the role of anti-IL-20 monoclonal antibody (7E) in tumor growth, BALB/c mice was injected with ML-1 cells and treated with 7E. HCC tumor tissue expressed higher levels of IL-20 than did non-tumor tissue. High IL-20 expression in HCC was correlated with poor overall survival (relative risk:>3). IL-20 and cyclin D1 expression were also highly correlated in HCC patient specimens and 3 human HCC cell lines. IL-20 also increased cell proliferation and migration, and it regulated matrix metalloproteinase (MMP)-13, tumor necrosis factor (TNF)-α, cyclin D1, and p21WAF1 expression in ML-1 cells. 7E attenuated tumor growth in mice inoculated with ML-1 cells. The expression of cyclin D1, TNF-α, MMP-9, and vascular endothelial growth factor was significantly inhibited after 7E treatment. The findings of this study suggest that IL-20 plays a role in the tumor progression of HCC.

Hypoxia promotes invasion of retinoblastoma cells in vitro by upregulating HIF-1α/MMP9 signaling pathway.

The aim of this study is to investigate the effect of hypoxia-inducible transcription factors-1α (HIF-1α)/matrix metalloproteinase 9 (MMP9) signaling pathway on hypoxia triggered invasion in retinoblastoma cell line HXO-RB44. HXO-RB44 cells were cultured under hypoxia conditions for 24 h. The effect of hypoxia on invasion ability of HXO-RB44 cells was monitored with transwell invasion assay; the mRNA and protein expression levels of HIF-1α and MMP9 were detected by Real-time polymerase chain reaction (RT-PCR) and Western blot; luciferase assay was performed to assess the MMP9 regulation by HIF-1α, and HIF-1α regulation by hypoxia. Furthermore, HIF-1α and MMP9 siRNA were used to investigate the effect of HIF-1α/MMP9 signaling on hypoxia-induced cell invasion. The results demonstrated that hypoxia could promote HXO-RB44 cells invasion. The mRNA and protein level of HIF-1α and MMP9 were upregulated by hypoxia treatment, whereas HIF-1α and MMP9 siRNA could reverse these processes. Hypoxia promotes retinoblastoma cell line HXO-RB44 invasion by activating HIF-1α/MMP9 signaling pathway.

GV1001 Induces Apoptosis by Reducing Angiogenesis in Renal Cell Carcinoma Cells Both In Vitro and In Vivo

To investigate the anticancer effects of GV1001 and its biological mechanism of action in renal cell carcinoma (RCC). The effects of GV1001 on cell survival and apoptosis in RCC cells were examined in vitro using cell viability assay, fluorescence-activated cell sorting, and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. To evaluate the effect of GV1001 on migration, invasion, and angiogenesis, we used wound healing, invasion, endothelial cell tube formation assay, and western blot analysis. Furthermore, we used an RCC xenograft model with either phosphate buffered saline or GV1001 to confirm the anticancer effect of GV1001 in vivo. Tumor volume was monitored during treatment, and tumor weight was measured after animals were killed. Apoptosis and angiogenesis of the tumor tissue were assessed using hematoxylin and eosin staining, immunohistochemistry, and western blot analysis. GV1001 reduced cell viability and induced apoptosis in RCC cells in vitro. Furthermore, GV1001 suppressed the migration and invasion of RCC cells through regulation of matrix metalloproteinases and tissue inhibitors of metalloproteinases. In addition, GV1001 reduced angiogenesis via regulation of hypoxia-inducible factor 1α. In xenograft mouse model experiment, GV1001 reduced tumor growth and induced apoptosis. As in the in vitro results, GV1001 significantly reduced angiogenesis through regulation of hypoxia-inducible factor 1α in vivo. Our data demonstrated that GV1001 induced apoptosis through suppression of angiogenesis in RCCs both in vitro and in vivo, which suggests that GV1001 may be a potential therapeutic target for RCC.

Glycogen synthase kinase-3β mediated regulation of matrix metalloproteinase-9 and its involvement in oral squamous cell carcinoma progression and invasion.

Oral squamous cell carcinoma (OSCC)-related deaths mainly result from invasion of the tumor cells into local cervical lymph nodes. It has been reported that progressive basement membrane loss promotes the metastatic and invasive capacities of OSCCs. Matrix metalloproteinase-9 (MMP-9) is known to play a central role in tumor progression and invasion. However, the role of MMP-9 in OSCC invasion has so far remained paradoxical and little is known about its regulation. Here, we aimed to assess MMP-9 expression regulation and its activation by glycogen synthase kinase-3β during human OSCC progression and invasion. In the present study, 178 human OSCC samples, including 118 fresh samples (18 adjacent normal, 42 noninvasive and 58 invasive tumor samples) and 60 archival human tissue microarray (TMA) tongue cancer samples, were included. mRNA expression, protein expression, MMP-9/-2 activity, protein-protein interaction and Snail, c-Myc, β-catenin and TIMP1 expression were assessed using RT-PCR, immunohistochemistry, Western blotting, co-immunoprecipitation and gelatin zymography analyses, respectively. Wnt5a and LPA mediated MMP-9 regulation was assessed in OCSCC-derived SCC-9 cells exogenously expressing GSK3β (WT) or non phosphoryable GSK3β (S9A). We observed a progressive up-regulation/activation of MMP-9 at various stages of oral tumor progression/invasion. Positive correlations were observed between MMP-9 and c-Myc expression, MMP-9 and MMP-2 activity, MMP-9 and TIMP1 expression and MMP-9 activity and TIMP1-MMP-9 interaction. In contrast, a negative correlation between phosphorylated β-catenin and MMP-9 expression was observed. Conversely, we found that in oral tongue SCC MMP-9 expression was positively correlated with inactivation of GSK3 signaling. Finally, we found thatWnt5a and LPA mediated increased MMP-9 and decreased GSK3β activities in tongue SCC-derived SCC-9 cells. MMP-9 regulation by GSK3β was confirmed by using phosphoryable/regulatory GSK3β (WT construct) and not by non-phosphoryable GSK3β (S9A construct). Collectively, our results show that MMP-9 overexpression and activation are important events occurring during OSCC progression/invasion and that this overexpression/activation is regulated by c-Myc, active MMP-2 and inactive GSK3β mediated pathways.

VANGL2 interacts with integrin αv to regulate matrix metalloproteinase activity and cell adhesion to the extracellular matrix

Planar cell polarity (PCP) proteins are implicated in a variety of morphogenetic processes including embryonic cell migration and potentially cancer progression. During zebrafish gastrulation, the transmembrane protein Vang-like 2 (VANGL2) is required for PCP and directed cell migration. These cell behaviors occur in the context of a fibrillar extracellular matrix (ECM). While it is thought that interactions with the ECM regulate cell migration, it is unclear how PCP proteins such as VANGL2 influence these events. Using an in vitro cell culture model system, we previously showed that human VANGL2 negatively regulates membrane type-1 matrix metalloproteinase (MMP14) and activation of secreted matrix metalloproteinase 2 (MMP2). Here, we investigated the functional relationship between VANGL2, integrin αvβ3, and MMP2 activation. We provide evidence that VANGL2 regulates cell surface integrin αvβ3 expression and adhesion to fibronectin, laminin, and vitronectin. Inhibition of MMP14/MMP2 activity suppressed the cell adhesion defect in VANGL2 knockdown cells. Furthermore, our data show that MMP14 and integrin αv are required for increased proteolysis by VANGL2 knockdown cells. Lastly, we have identified integrin αvβ3 as a novel VANGL2 binding partner. Together, these findings begin to dissect the molecular underpinnings of how VANGL2 regulates MMP activity and cell adhesion to the ECM.

Molecular Characterization and In-Silico Analysis of the Tissue Inhibitor of Metalloproteinases-3 (TIMP-3) Gene of Canine Mammary Tumor.

Mammary tumors are the second most common tumors (after skin tumors) in female dogs (Canis lupus familiaris). Tissue Inhibitor of Metlloproteinases-3 (TIMP-3) is a matrix associated endogenous inhibitor of Matrix Metalloproteinases (MMPs). Cancer metastasis occurs as a result of imbalance between MMPs and TIMPs. TIMP-3 is involved significantly in regulation of MMPs as well as progression of canine mammary tumor. The present study was conducted to identify the structural and functional relationship between TIMP-3 and MMP which can aid in identifying the role of these proteins in canine mammary tumor. Molecular characterization of TIMP-3 protein was done by molecular biology techniques such as gene cloning and sequencing. The homology based model of TIMP-3 protein was created and verified with a variety of available computational techniques as well as molecular dynamics simulation. The results indicated that predicted TIMP-3 protein structure of Canis lupus familiaris was reliable and more stable. The docking of TIMP-3 protein with MMP-2 and MMP-9 represents conformational structure of these two proteins which interact with each other but if misled canresult in the progression of tumor in canine. The three dimensional structure of TIMP-3 was generated and its interactions with MMP-2 and MMP-9, demonstrates the role of key binding residues. Until now, no structural details were available for canine TIMP-3 proteins, hence this study will broaden the horizon towards understanding the structural and functional aspects of this proteins in canine.

Toxicological effects of NCKU-21, a phenanthrene derivative, on cell growth and migration of A549 and CL1-5 human lung adenocarcinoma cells.

BackgroundChemotherapy insensitivity continues to pose significant challenges for treating non-small cell lung cancer (NSCLC). The purposes of this study were to investigate whether 3,6-dimethoxy-1,4,5,8-phenanthrenetetraone (NCKU-21) has potential activity to induce effective toxicological effects in different ethnic NSCLC cell lines, A549 and CL1-5 cells, and to examine its anticancer mechanisms.MethodsMitochondrial metabolic activity and the cell-cycle distribution were analyzed using an MTT assay and flow cytometry in NCKU-21-treated cells. NCKU-21-induced cell apoptosis was verified by Annexin V-FITC/propidium iodide (PI) double-staining and measurement of caspase-3 activity. Western blotting and wound-healing assays were applied to respectively evaluate regulation of signaling pathways and cell migration by NCKU-21. Molecular interactions between target proteins and NCKU-21 were predicted and performed by molecular docking. A colorimetric screening assay kit was used to evaluate potential regulation of matrix metalloproteinase-9 (MMP-9) activity by NCKU-21.ResultsResults indicated that NCKU-21 markedly induced cytotoxic effects that reduced cell viability via cell apoptosis in tested NSCLC cells. Activation of AMP-activated protein kinase (AMPK) and p53 protein expression also increased in both NSCLC cell lines stimulated with NCKU-21. However, repression of PI3K-AKT activation by NCKU-21 was found in CL1-5 cells but not in A549 cells. In addition, increases in phosphatidylserine externalization and caspase-3 activity also confirmed the apoptotic effect of NCKU-21 in both NSCLC cell lines. Moreover, cell migration and translational levels of the gelatinases, MMP-2 and MMP-9, were obviously reduced in both NSCLC cell lines after incubation with NCKU-21. Experimental data obtained from molecular docking suggested that NCKU-21 can bind to the catalytic pocket of MMP-9. However, the in vitro enzyme activity assay indicated that NCKU-21 has the potential to increase MMP-9 activity.ConclusionsOur results suggest that NCKU-21 can effectively reduce cell migration and induce apoptosis in A549 and CL1-5 cells, the toxicological effects of which may be partly modulated through PI3K-AKT inhibition, AMPK activation, an increase in the p53 protein, and gelatinase inhibition.

Protective effects of taurine against renal ischemia/reperfusion injury in rats by inhibition of gelatinases, MMP-2 and MMP-9, and p38 mitogen-activated protein kinase signaling

Dysregulated expression of matrix metalloproteinases (MMPs) is closely associated with the pathogenesis of renal ischemia/reperfusion injury (I/R). The production of excessive reactive oxygen species (ROS) causes tissue damage. Increased ROS production causes activation of p38 mitogen-activated protein kinase (MAPK) signaling, which participates in gene regulation of MMPs, especially MMP-2 and MMP-9 (gelatinases). Taurine (2-aminoethanesulfonic acid) in mammalian cells functions in bile acid conjugation, maintenance of calcium homeostasis, osmoregulation, membrane stabilization, and antioxidation, antiinflammatory, and antiapoptotic action. We investigated the effects of taurine and the possible role of p38 MAPK signaling on regulation of MMP-2 and MMP-9 in a renal I/R injury model in rats. Rats were divided into three groups: sham, I/R, and I/R + taurine treated. After a right nephrectomy, I/R was induced by clamping the left renal pedicle for 1 h followed by 6 h reperfusion. Taurine was administered 45 min prior to induction of ischemia. Renal function was assessed by serum creatinine and blood urea nitrogen (BUN) levels. Tubule injury and structural changes were evaluated by light microscopy. Malondialdehyde (MDA) levels were analyzed by high performance liquid chromatography (HPLC). Superoxide dismutase (SOD) activity levels were measured using a colorimetric kit. mRNA expression of MMP-2 and MMP-9 was determined by real-time polymerase chain reaction. MMP-2 and MMP-9 activities were measured using a fluorimetric kit. Phosphorylated p38 (p-p38) and total p38 MAPK protein expressions were evaluated by western blot. Taurine pretreatment significantly attenuated renal dysfunction and histologic damage, such as renal tubule dilation and loss of brush borders. The pretreatment also decreased the MDA level and attenuated the reduction of SOD activity in the kidney during I/R. Taurine pretreatment also decreased significantly both MMP-2 and MMP-9 mRNA expression and MMP-9 activity induced by I/R. In addition, the activity of p38 MAPK signaling was down-regulated significantly by taurine administration. Inhibition of MMP-2 and MMP-9 expression and MMP-9 activity caused by taurine may be associated with suppression of p38 MAPK activation during I/R induced renal injury in rats. Therefore, taurine administration may prove to be a strategy for attenuating renal I/R injury.

Post-transcriptional Regulation of MMP16 and TIMP2 Expression via miR-382, miR-410 and miR-200b in Endometrial Cancer.

The post-transcriptional regulation of matrix metalloproteinases (MMPs) via microRNAs (miRNAs) has been recently described in numerous human malignancies. However, the exact mechanisms of miRNA-mediated MMPs deregulation in endometrial cancer (EC) remain unclear. Herein, we aimed to analyze the expression of MMP2, MMP16 and TIMP2 and identify miRNAs that modulate their expression. Protein expression was assessed by immunohistochemistry in formalin-fixed paraffin-embedded EC samples. Target prediction algorithms were applied to select miRNAs binding the 3'UTRs of MMP16 (miR-377, miR-382, miR-410, miR-200b) or TIMP2 (miR-200b), and their levels were measured by qPCR in laser capture-microdissected tissue fragments. Luciferase assays and western blotting were used to indicate individual miRNA- mRNA interactions. Overexpression of MMP2 and MMP16 in cancerous tissues corresponded to down-regulation of miR-377, miR-382 and miR-410, while decreased expression of TIMP2 was associated with miR-200b up-regulation. In vitro experiments confirmed direct regulation of MMP16 by miR-382 and miR-410, and TIMP2 by miR-200b in EC Ishikawa cells. We demonstrated novel mechanisms of miRNA-mediated regulation of MMPs activity in EC.

FOXO1 deletion in keratinocytes improves diabetic wound healing through MMP9 regulation

Keratinocyte migration is a key aspect of re-epithelialization during wound healing. Matrix metalloproteinase 9 (MMP9) contributes to this process and deficiencies in the MMP9 lead to impaired healing. Inappropriate expression of MMP9 also contributes to impaired re-epithelialization. Previously we demonstrated that FOXO1 was activated in wound healing but to higher levels in diabetic wounds. To address mechanisms of impaired re-epithelialization we examined MMP9 expression in vivo in full thickness dermal scalp wounds created in experimental K14.Cre+.Foxo1L/L mice with lineage-specific Cre recombinase deletion of floxed FOXO1 and compared the results to control littermates. MMP9 was induced during wound healing but at a significantly higher level in diabetic compared to normal wounds. FOXO1 deletion substantially blocked this increase. By chromatin immunoprecipitation FOXO1 was shown to bind to the MMP9 promoter, FOXO1 overexpression increased MMP9 transcriptional activity and increased MMP9 expression stimulated by high glucose was blocked by FOXO1 deletion or FOXO1 knockdown. We also show for the first time that high glucose impairs keratinocyte migration by inducing high levels of MMP9 expression and establish that it involves FOXO1. Thus, FOXO1 drives high levels of MMP9 expression in diabetic wound healing, which represents a novel mechanism for impaired re-epithelization in diabetic wounds.

Kruppel-like factor 4 regulates matrix metalloproteinase and aggrecanase gene expression in chondrocytes.

Kruppel-like factor 4 (KLF4) is a zinc finger transcription factor that plays crucial roles during the development and maintenance of multiple organs. We and others have previously shown that KLF4 is involved in bone modeling and remodeling but roles played by KLF4 during skeletogenesis are still not fully understood. Here, we show that KLF4 is expressed in the epiphyseal growth plate and articular chondrocytes. Most articular chondrocytes expressed KLF4 in embryos but it localized only in a subset of superficial zone cells in postnatal mice. When KLF4 was overexpressed in chondrocytes in vitro, it severely repressed chondrocytic gene expressions. Global gene expression profiling of KLF4-transduced chondrocytes revealed matrix degrading proteinases of the matrix metalloproteinase and disintegrin and metalloproteinase with thrombospondin-1 domain families within the group of upregulated genes. Proteinase induction by KLF4 was alleviated by Trichostatin A treatment suggesting the possible involvement of epigenetic mechanisms on proteinase induction by KLF4. These results indicate the possible involvement of KLF4 in physiological and pathological aspects during cartilage development and maintenance.

Phosphorylation of STAT3 at Tyr705 regulates MMP-9 production in epithelial ovarian cancer.

Ovarian cancer’s poor progression is closely associated with overexpression of matrix metalloproteinase 9 (MMP-9), which belongs to the class of enzymes believed to be involved in the degradation of extracellular matrix. However, the mechanisms underlying regulation of MMP-9 are not completely understood. STAT (signal transducer and activator of transcription) family of transcription factors is well known to be engaged in diverse cellular functions. Activation of STAT3 has been observed in a number of cancers, promoting tumorigenesis and metastasis via transcriptional activation of its target genes. In this study, we tested our hypothesis that STAT3 regulates MMP-9 gene expression in epithelial ovarian cancer. Using epithelial ovarian cancer cell lines as in vitro model, we show an abundance of phosphorylated STAT3 at Tyr705 (p-STAT3) in SKOV3 cell line. We further show that MMP-9 gene promoter was significantly enriched by p-STAT3, and IL-6 treatment led to a significant increase of MMP-9 at mRNA and protein levels, in addition to an association of p-STAT3 with MMP-9 gene. By using luciferase reporter assay, we determined that the STAT3 DNA responsive element of MMP-9 was sufficient to regulate transcriptional activity of a heterologous promoter. These results suggest that the phosphorylation of STAT3 regulates MMP-9 production in ovarian cancer, which might be responsible for its invasiveness and metastasis.

Studies on anti-angiogenesis of ginsenoside structure modification HRG in vitro

This study investigates the anti-angiogenic effect of 3β, 12β, 20(S)-trihydroxy dammarane-3-O-β-d-glucopyranosyl(1–2)-β-d-glucopyranoside(HRG), a new chemical compound obtained by structure modification on Ginseng saponins Rg3, associated with the regulation of matrix metalloproteinases(MMPs) and its upstream signal-regulated molecule of vascular endothelial growth factor(VEGF) and basic fibroblast growth factor(b-FGF) in vitro, which plays an critical role in angiogenesis during the process of carcinoma. In our study, to investigate the anti-angiogenesis effect of HRG in HUVECs, we utilized cell proliferation assay, tube formation assay, wound-healing assay, Semi-quantitative reverse transcription PCR, and Western blot assay. Our results demonstrated that HRG plays a major role in the regulation of proliferation, migration and tube formation of HUVECs by suppressing the expression of VEGF and b-FGF in both transcriptional and post-transcriptional levels. In addition, the expression of MMP-2 and MMP-9, which were related to the ECM degradation, were down-regulated after administration of HRG as well. Overall, our results revealed that HRG strongly inhibited the process of angiogenesis and shows better effectiveness than Rg3.

Metalloproteinases as mediators of inflammation and the eyes: molecular genetic underpinnings governing ocular pathophysiology.

There are many vision threatening diseases of the eye affecting millions of people worldwide. In this article, we are summarizing potential role of various matrix metalloproteinases (MMPs); the Zn (2+)-dependent endoproteases in eye health along with pathogenesis of prominent ocular diseases such as macular degeneration, diabetic retinopathy, and glaucoma via understanding MMPs regulation in affected patients, interactions of MMPs with their substrate molecules, and key regulatory functions of tissue inhibitor of metalloproteinases (TIMPs) towards maintaining overall homeostasis.

Novel STAT binding elements mediate IL-6 regulation of MMP-1 and MMP-3

Dynamic remodelling of the extracellular matrix (ECM) is a key feature of cancer progression. Enzymes that modify the ECM, such as matrix metalloproteinases (MMPs), have long been recognised as important targets of anticancer therapy. Inflammatory cytokines are known to play a key role in regulating protease expression in cancer. Here we describe the identification of gamma-activated site (GAS)-like, signal transducer and activator of transcription (STAT) binding elements (SBEs) within the proximal promoters of the MMP-1 and MMP-3 genes, which in association with AP-1 components (c-Fos or Jun), bind STAT-1 in a homodimer like complex (HDLC). We further demonstrate that MMP expression and binding of this complex to SBEs can either be enhanced by interleukin (IL)-6, or reduced by interferon gamma (IFN-γ), and that IL-6 regulation of MMPs is not STAT-3 dependent. Collectively, this data adds to existing understanding of the mechanism underlying cytokine regulation of MMP expression via STAT-1, and increases our understanding of the links between inflammation and malignancy in colon cancer.

EMMPRIN overexpression in SVZ neural progenitor cells increases their migration towards ischemic cortex

Stimulation of endogenous neurogenesis and recruitment of neural progenitors from the subventricular zone (SVZ) neurogenic site may represent a useful strategy to improve regeneration in the ischemic cortex. Here, we tested whether transgenic overexpression of extracellular matrix metalloproteinase inducer (EMMPRIN), the regulator of matrix metalloproteinases (MMPs) expression, in endogenous neural progenitor cells (NPCs) in the subventricular zone (SVZ) could increase migration towards ischemic injury. For this purpose, we applied a lentivector-mediated gene transfer system. We found that EMMPRIN-transduced progenitors exhibited enhanced MMP-2 activity in vitro and showed improved motility in 3D collagen gel as well as in cortical slices. Using a rat model of neonatal ischemia, we showed that EMMPRIN overexpressing SVZ cells invade the injured cortical tissue more efficiently than controls. Our results suggest that EMMPRIN overexpression could be suitable approach to improve capacities of endogenous or transplanted progenitors to invade the injured cortex.

Abstract 3891: Chemoresistant cancer stem cells undergo gene changes that drive tumor recurrence

Abstract Despite the efficacy of chemotherapy, resistant cells are thought to contribute to tumor recurrence. In vitro models of recurrence must mimic the complexity and heterogeneity of in vivo tumors and provide the longevity needed to capture tumor dormancy following chemotherapy. We have previously described a 3D in vitro model that simulates in vivo tumors containing cancer stem cell niches. Encapsulation of murine renal adenocarcinoma (RENCA) cells in a double layer of agarose that form spherical macrobeads allows for the development of 3D tumor colonies comprised of tumor-initiating and tumor-amplifying cells within the inner agarose matrix. In the current study, we used the RENCA macrobead in combination with chemotherapy to evaluate genetic regulation that allows tumor dormancy and drives recurrence. RENCA macrobeads treated with docetaxel (5 μg/ml) undergo a significant loss of cell mass and a period of latency followed by tumor recurrence. RNA isolated from docetaxel-resistant cells (Wk.6 post-treatment), recurrent tumors (Wk.20 post-treatment) and paired vehicle controls were characterized using whole-transcriptome profiling (GeneChip® WT Pico Kit with Transciptome Analysis Console; Affymetrix). At Wk.6 post-treatment, genes associated with a stem cell phenotype (Car12, Arrb1, Nos2, Aldh1l2) and members of the ABC1 family associated with multidrug resistance were enriched in docetaxel-resistant cells. Genes regulating the canonical Wnt signaling pathway, specifically LRP family members, were significantly up regulated. Pathway analysis revealed dysregulated focal adhesion-PI3K-Akt-mTor-signaling, including genes modulating integrin adhesion (Itga5, Itga7), ECM receptor interaction (Lama5, Lamb2, Lamc1) and actin cytoskeletal remodeling (Vcl). Supporting the quiescent nature of the docetaxel-resistant population, genes implicated in DNA replication (Dbf4, Cdc6, Mcm5), transcriptional initiation (Taf13, Gtf2h1, Polr2i) and tumor progression (Mmp3, Mmp13) were significantly down regulated. At Wk.20 post-treatment, up regulation of matrix metalloproteinases (Mmp13) and partial retention of stem cell (Nos2, Car12) and drug resistant (Abcb1) genes supports the evolution of regenerated tumors to a metastatic phenotype. Genes involved in fatty acid β-oxidation (Acsl5, Lipf), oxidative phosphorylation (ND3, ND6) and the electron transport chain (COX2) were down regulated in concert with up regulation of Me2 suggesting reduced flux through the TCA cycle and use of aerobic glycolysis in rapidly growing cells. This study further supports the RENCA macrobead as a model system to evaluate the biology of chemotherapy-resistant cells and tumor recurrence in a 3D microenvironment. These data are in line with the notion that chemotherapy-resistant cells exhibit a period of quiescence with a stem cell-like gene profile and experience an intrinsic transformation towards an invasive phenotype that drives tumor recurrence. Citation Format: Prithy C. Martis, Atira Dudley, Melissa A. Laramore, Hunter L. Gazda, Michael P. Markey, Barry H. Smith, Lawrence S. Gazda. Chemoresistant cancer stem cells undergo gene changes that drive tumor recurrence [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3891. doi:10.1158/1538-7445.AM2017-3891

Abstract 2225: Green tea polyphenols suppress tumor growth and invasion by targeting matrix metalloproteinases, RECK and TIMP-3, in a mouse model implanted with prostate tumors

Abstract Green tea polyphenols (GTP) and its major constituent, epigallocatechin-3-gallate (EGCG) reactivate epigenetically silenced genes in cancer cells that reduces invasiveness and migration capabilities; however, the mechanisms whereby these effects occur are not well understood. RECK, a novel tumor suppressor and the tissue inhibitor of matrix metalloproteinase-3 (TIMP-3) genes negatively regulates matrix metalloproteinases (MMPs) and inhibits tumor invasion, angiogenesis and metastasis. In the present study, we demonstrate that GTP mediate epigenetic induction of RECK and TIMP-3 thereby playing a key role in suppressing invasiveness and gelatinolytic activity of MMP-2 and MMP-9 in vivo in an orthotopic implantation model of human prostate cancer. Athymic nude mice were implanted with human prostate cancer LNCaP tumor in the ventral prostate for 2 weeks and later treated DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (AZA), histone deacetylase inhibitor, trichostatin A (TSA) and histone methyltransferase inhibitor, 3-Deazaneplanocin A (DZNep) individually at 0.1 mg/kg body weight intraperitoneally at alternate days/week; and combination of AZA+TSA and DZNep+TSA at similar doses and times; whereas GTP was provided peroral by gavage at 7.5 and 15.0 mg/kg body weight freshly prepared in 100µl PBS. Treatment of mice with GTP resulted in marked decrease in tumor growth and its local invasion in dose dependent manner, compared to treatment with epigenetic inhibitors and their combination after 8 weeks of intervention. GTP treatment significantly reduced serum levels of MMP-2, MMP-9 and VEGF, compared to treatment with epigenetic inhibitors alone. Combination of AZA+TSA exhibited similar effect which was equivalent to the lower dose of GTP treatment. Furthermore, GTP treatment significantly reduced EZH2 and H3K27me3 and class I HDAC protein levels in tumors. GTP partially reversed the hypermethylation status of RECK and TIMP3 gene and significantly enhanced their protein expression in the tumor tissue. Inhibition of VEGF, MMP-2 and MMP-9 levels were also noted after GTP treatment in tumor tissue in dose-dependent manner. Our findings suggest that induction of RECK and TIMP-3 are key epigenetic events modulated by GTP that results in suppression of matrix degradation and angiogenesis to delay prostate cancer invasion and its subsequent progression. Citation Format: Eswar Shankar, Natarajan Bhaskaran, Rajnee Kanwal, Sanjay Gupta. Green tea polyphenols suppress tumor growth and invasion by targeting matrix metalloproteinases, RECK and TIMP-3, in a mouse model implanted with prostate tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2225. doi:10.1158/1538-7445.AM2017-2225

Omentin-1 prevents cartilage matrix destruction by regulating matrix metalloproteinases

Matrix metalloproteinases (MMPs) play a crucial role in the degradation of the extracellular matrix and pathological progression of osteoarthritis (OA). Omentin-1 is a newly identified anti-inflammatory adipokine. Little information regarding the protective effects of omentin-1 in OA has been reported before. In the current study, our results indicated that omentin-1 suppressed expression of MMP-1, MMP-3, and MMP-13 induced by the proinflammatory cytokine interleukin-1β (IL-1β) at both the mRNA and protein levels in human chondrocytes. Importantly, administration of omentin-1 abolished IL-1β-induced degradation of type II collagen (Col II) and aggrecan, the two major extracellular matrix components in articular cartilage, in a dose-dependent manner. Mechanistically, omentin-1 ameliorated the expression of interferon regulatory factor 1 (IRF-1) by blocking the JAK-2/STAT3 pathway. Our results indicate that omentin-1 may have a potential chondroprotective therapeutic capacity.

Tangeretin alters neuronal apoptosis and ameliorates the severity of seizures in experimental epilepsy-induced rats by modulating apoptotic protein expressions, regulating matrix metalloproteinases, and activating the PI3K/Akt cell survival pathway

PurposeEpilepsy is complex neural disarray categorized by recurring seizures. Despite recent advances in pharmacotherapies for epilepsy, its treatment remains a challenge due to the contrary effects of the drugs. As a result, the identification of novel anti-epileptic drugs (AEDs) with neuroprotective properties and few side effects is of great value. Thus, the present study assessed the treatment effects of tangeretin using a rat model of pilocarpine-induced epilepsy. Materials and MethodsSeparate groups of male Wistar rats received oral administrations of tangeretin at 50, 100, or 200mg/kg for 10 days and then, on the 10th day, they received an intraperitoneal injection of pilocarpine (30mg/kg). Subsequently, neuronal degeneration and apoptosis were assessed using Nissl staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay procedures. Additionally, the expressions of phosphatidylinositol-3-kinase (PI3K/Akt) pathway proteins, cleaved caspase-3, Bad, Bcl-2, Bcl-xL, and Bax were determined using Western blot analyses. ResultsTangeretin reduced the seizure scores and latency to first seizure of the rats and effectively activated the pilocarpine-induced suppression of PI3K/Akt signaling. Additionally, tangeretin effectively regulated the levels of apoptosis-inducing factor (AIF) in mitochondria as well as the expressions of apoptotic pathway proteins. Seizure-induced elevations in the activities and expressions of matrix metalloproteinases (MMPs)-2 and -9 were also modulated. ConclusionThe present results indicate that tangeretin exerted potent neuroprotective effects against pilocarpine-induced seizures via the activation of PI3K/Akt signaling and the regulation of MMPs.