MMP-9 Pathway Research Articles

N-acetylcysteine attenuated sepsis-induced cardiac depression: down-regulation of MMP-2 pathway in mice

During sepsis,the inflammatory responses mediate myocardium injury,including LV dysfunction and cardiac pathophysiological changes. To understanding the pathway of sepsis in cardiac depression,we tested the hypothesis that N-acetylcysteine attenuated sepsis-induced cardiac depression through down-regulation of MMP-2 pathway. Adult (4 to 6 months) male Albino-Webster mice,and their weights ranged from 25 to 30 gm,were pre-treated with N-acetylcysteine ip following cecal ligation and puncture (CLP). Left ventricle (LV) function was assessed using a micro-catheter system. MCP-1 and cytokines mediator’s in plasma and myocardium were analyzed by enzyme-linked immunosorbent assay (ELISA). Further,the cardiac MMP2 measured by qRT-PCR and the pathological changes and cells injuries in the myocardium were examined using hematoxylin and eosin staining. CLP mice displayed worse LV function. The exaggerated cardiac depression in CLP mice was associated with higher levels of MCP-1 and cytokines in plasma and myocardium together with greater cardiac levels of cTn-I and MMP2. Neutralization of sepsis by NAC resulted in improved LV function and greater reductions in inflammatory mediators,MMP2 and myocardium injury. Taken together,NAC improved LV function following sepsis through down-regulation of MMP2 and serve as a potential therapeutic in cardiac endotoxemia.

Downregulated expression of hepatoma-derived growth factor inhibits migration and invasion of prostate cancer cells by suppressing epithelial-mesenchymal transition and MMP2, MMP9.

Hepatoma-derived growth factor (HDGF) is commonly over-expressed and plays critical roles in the development and progression in a variety of cancers. It has previously been shown that HDGF is overregulated in prostate cancer cells compared to normal prostate cells, which is correlated with cellular migration and invasion of prostate cancer. Here, the molecular mechanisms of HDGF in prostate cancer is investigated. It is shown that HDGF knockdown reduces prostate cancer cellular migration and invasion in both androgen-sensitive LNCaP cells and androgen-insensitive DU145 and PC3 cells. Furthermore, Western blot analysis reveals that HDGF knockdown inhibits epithelial-mesenchymal transition (EMT) of prostate cancer cells by upregulation of protein E-cadherin and downregulation of proteins N-cadherin, Vimentin, Snail and Slug. In addition, mechanistic studies reveal that proteins MMP2 and MMP9 are down-regulated. In conclusion, our data suggested that HDGF knockdown inhibits cellular migration and invasion in vitro of prostate cancer via modulating epithelial-mesenchymal transition (EMT) signaling pathway, as well as MMP2 and MMP9 signaling pathway. These results supported that HDGF is a relevant protein in the progression of prostate cancer and may serve as a potentially therapeutic target for prostate cancer as well as its downstream targets.

Baicalin Attenuates Blood-Brain Barrier Disruption and Hemorrhagic Transformation and Improves Neurological Outcome in Ischemic Stroke Rats with Delayed t-PA Treatment: Involvement of ONOO--MMP-9 Pathway.

Tissue plasminogen activator (t-PA) has a restrictive therapeutic window within 4.5h after ischemic stroke with the risk of hemorrhagic transformation (HT) and neurotoxicity when it is used beyond the time window. In the present study, we tested the hypothesis that baicalin, an active compound of medicinal plant, could attenuate HT in cerebral ischemia stroke with delayed t-PA treatment. Male Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 4.5h and then continuously received t-PA infusion (10mg/kg) for 0.5h and followed by 19-h reperfusion. Baicalin (50, 100, 150mg/kg) was administrated via femoral vein at 4.5h after MCAO cerebral ischemia. Delayed t-PA infusion significantly increased the mortality rate, induced HT, blood-brain barrier (BBB) damage, and apoptotic cell death in the ischemic brains and exacerbated neurological outcomes in cerebral ischemia-reperfusion rats at 24h after MCAO cerebral ischemia. Co-treatment of baicalin significantly reduced the mortality rates, ameliorated the t-PA-mediated BBB disruption and HT. Furthermore, baicalin showed to directly scavenge peroxynitrite and inhibit MMP-9 expression and activity in the ischemic brains with the delayed t-PA treatment. Baicalin had no effect on the t-PA fibrinolytic function indicated by t-PA activity assay. Taken together, baicalin could attenuate t-PA-mediated HT and improve the outcomes of ischemic stroke treatment possibly via inhibiting peroxynitrite-mediated MMP-9 activation.

Long non-coding RNA DLX6-AS1 aggravates hepatocellular carcinoma carcinogenesis by modulating miR-203a/MMP-2 pathway

Long non-coding RNAs (lncRNAs) have been wildly verified to modulate multiple tumorigenesis, especially hepatocellular carcinoma (HCC). In present study, our team aims to investigate the role of lncRNA DLX6-AS1 in the HCC carcinogenesis. Results of early-stage experiments found that DLX6-AS1 expression level was up-regulated in 60 cases of HCC tissue samples compared with adjacent normal tissue. Moreover, the aberrant overexpression of DLX6-AS1 indicated the poor prognosis of HCC patients. Loss-of-function experiments revealed that DLX6-AS1 knockdown inhibited the proliferation, migration and invasion of HCC cells in vitro, and decreased the tumor growth in vivo. Bioinformatics analysis predicted that miR-203a potentially targeted DLX6-AS1 3′-UTR, suggesting the interaction between miR-203a and DLX6-AS1. Furthermore, miR-203a also targeted MMP-2 mRNA 3′-UTR, which was validated by luciferase reporter assay. Taken together, our study discovered the oncogenic role of DLX6-AS1 in clinical specimens and cellular experiments, showing the potential DLX6-AS1/miR-203a/MMP-2 pathway. This results and findings provide a novel insight for HCC tumorigenesis.

Glucose-derived AGEs enhance human gastric cancer metastasis through RAGE/ERK/Sp1/MMP2 cascade.

Advanced glycation end products (AGEs) have been reported to take part in many cancer processes. Whether AGEs contribute to gastric cancer (GC) course and the underlying mechanism are still unclear. Here, glucose-derived AGEs are detected to be accumulated in tumor tissues and blood of patients with GC. As the receptor for AGEs, RAGE is highly expressed in cancer tissues, and closely associated with the depth of cancer invasion, lymph node metastasis and TNM stage. Both in vivo and in vitro treatment of AGEs accelerate the tumor invasion and metastasis, with upregualtion of RAGE, Specificity Protein 1 (Sp1), and MMP2 protein expression, as well as enhancement of MMP2 activity. Either RAGE-blocking antibody or Sp1-knockdown can partially block the AGEs-induced effects. Moreover, AGEs increased the phosphorylation of ERK, and reducing the phosphorylation level of ERK by MEK1/2 inhibitor decreased the expression of Sp1. These results indicate that accumulation of glucose-derived AGEs may act as one of potential risk factors for GC progression and promote the invasion and metastasis of gastric cancer partially through the activation of RAGE/ERK/Sp1/MMP2 pathway.

P-REX1 amplification promotes progression of cutaneous melanoma via the PAK1/P38/MMP-2 pathway

P-REX1 (PIP3-dependent Rac exchange factor-1) is a guanine nucleotide exchange factor that activates Rac by catalyzing exchange of GDP for GTP bound to Rac. Aberrant up-regulation of P-REX1 expression has a role in metastasis however, copy number (CN) and function of P-REX1 in cutaneous melanoma are unclear. To explore the role of P-REX1 in melanoma, SNP 6.0 and Exon 1.0 ST microarrays were assessed. There was a higher CN (2.82-fold change) of P-REX1 in melanoma cells than in melanocytes, and P-REX1 expression was significantly correlated with P-REX1 CN. When P-REX1 was knocked down in cells by P-REX1 shRNA, proliferation, colony formation, 3D matrigel growth, and migration/invasiveness were inhibited. Loss of P-REX1 inhibited cell proliferation by inhibiting cyclin D1, blocking cell cycle, and increased cell apoptosis by reducing expression of the protein survivin. Knockdown of P-REX1 expression inhibited cell migration/invasiveness by disrupting P-REX1/RAC1/PAK1/p38/MMP-2 pathway. Assessment of patient tumors and disease outcome demonstrated lower distant metastasis-free survival among AJCC stage I/II/III patients with high P-REX1 expression compared to patients with low P-REX1 expression. These results suggest P-REX1 plays an important role in tumor progression and a potential theranostic target.

Early Increased Bradykinin 1 Receptor Contributes to Hemorrhagic Transformation After Ischemic Stroke in Type 1 Diabetic Rats.

Hemorrhagic transformation (HT) is a major complication of ischemic stroke and further deteriorates neurological outcomes. Bradykinin 1 receptor (B1R) has been proven to mediate vasculo-toxicity in various experimental models. However, its role in the development of HT after stroke remains unclear. We detected the B1R expression in brain tissues with or without HT in a rat model of cerebral ischemia/reperfusion (I/R) with type 1 diabetes, showing higher B1R expression in the hemorrhagic areas than the ischemic tissues. Then, B1R agonist or antagonist was administrated intravenously just before reperfusion to investigate its effect on HT and the underlying molecular mechanism. Administration of low (300nmol/kg) or high (1μmol/kg) dose of B1R antagonist mitigated hemorrhage, improved neurobehavioral deficits, and preserved blood-brain-barrier (BBB) integrity after reperfusion for 8h whereas the 300nmol/kg of B1R agonist aggravated these outcomes, though only the high does of B1R antagonist affected the infarction volume. Extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation was increased by B1R activation but decreased by B1R inhibition, which mediated B1R toxicity on BBB disruption and ischemia-related HT. Furthermore, B1R activation facilitated the mRNA and protein expressions of MMP-9 in the hemorrhagic tissues, and these increases were blocked by both ERK inhibitor U0126 and NF-κB inhibitor PDTC. U0126 also remarkably decreased the B1R-induced NF-κB/p65 activation. We concluded that upregulated B1R may contribute to early HT after I/R in type 1 diabetic rats via ERK1/2/NF-κB/MMP-9 pathway. B1R inhibition could be an encouraging therapeutic strategy to withstand HT after ischemic stroke in diabetic patients.

Overexpression of Rsf-1 correlates with poor survival and promotes invasion in non-small cell lung cancer.

Rsf-1 (HBXAP) was recently reported to play roles in tumorigenesis and tumor progression. There have been many reports referred to Rsf-1 overexpression in various cancers and associated with the malignant behavior of cancer cells. However, the molecular mechanism of Rsf-1 in non-small cell lung cancer aggressiveness remains ambiguous. In the present study, we found that there was a significant association between Rsf-1 overexpression and poor overall survival (p=0.028) in lung cancer. Furthermore, knockdown of Rsf-1 expression in H1299 and H460 cells with high endogenous Rsf-1 expression inhibited cell migration and invasion and downregulated MMP2 expression and nuclear levels of NF-κB. NF-κB inhibitor could also block the effect of Rsf-1 in regulation of MMP2 expression. Further experiments demonstrated that Rsf-1 depletion restrained NF-κB reporter luciferase activity and downregulated bcl-2 and p-IκB protein level. In conclusion, we demonstrated that Rsf-1 was overexpressed in lung cancer and associated with poor survival. Rsf-1 regulated cell invasion through MMP2 and NF-κB pathway.

Increased NDRG1 expression attenuate trophoblast invasion through ERK/MMP-9 pathway in preeclampsia

ObjectiveThe aim of this study was to investigate the expression of N-myc downstream-regulated gene1(NDRG1)in the placentas of pregnancies complicated with early-onset and late-onset preeclampsia (PE) and its underlying mechanism on the pathophysiology of PE. MethodsThe expressions of NDRG-1 in placentas of pregnancies complicated with early-onset PE and late-onset PE were detected using immunohistochemistry, western blot assays and fluorescence quantitative PCR. The expressions of MMP-2, MMP-9 and ERK1/2 protein were detected by western blot analysis and cell invasion assay was performed using transwell chambers in NDRG1 silenced JEG-3 cells. ResultsCompared with the normal term pregnancies, the expression of both NDRG1 mRNA and protein were significantly high in placentas from PE, and the expression of NDRG1 in early-onset PE was higher than that in late-onset PE. In NDRG1-silenced JEG-3 cells, MMP-2, MMP-9 and phosphorylation of ERK1/2 protein increased obviously and the number of cells that penetrated the membrane increased. ConclusionUpregulation of NDRG1 is associated with impaired trophoblast invasion in PE by inhibition ERK/MMP-2 and MMP-9 Pathway.

Rhein and rhubarb similarly protect the blood-brain barrier after experimental traumatic brain injury via gp91phox subunit of NADPH oxidase/ROS/ERK/MMP-9 signaling pathway.

Oxidative stress chiefly contributes to the disruption of the BBB following traumatic brain injury (TBI). The Chinese herbal medicine rhubarb is a promising antioxidant in treating TBI. Here we performed in vivo and in vitro experiments to determine whether rhubarb and its absorbed bioactive compound protected the BBB after TBI by increasing ZO-1 expression through inhibition of gp91phox subunit of NADPH oxidase/ROS/ERK/MMP-9 pathway. Rats were subjected to the controlled cortical impact (CCI) model, and primary rat cortical astrocytes were exposed to scratch-wound model. The liquid chromatography with tandem mass spectrometry method showed that rhein was the compound absorbed in the brains of CCI rats after rhubarb administration. The wet-dry weights and Evans blue measurements revealed that rhubarb and rhein ameliorated BBB damage and brain edema in CCI rats. Western blots showed that rhubarb and rhein downregulated GFAP in vitro. RT-PCR, immunohistochemistry, Western blot and dichlorodihydrofluorescein diacetate analysis indicated that rhubarb prevented activation of gp91phox subunit of NADPH oxidase induced ROS production, subsequently inhibited ERK/MMP-9 pathway in vivo and in vitro. Interestingly, rhein and rhubarb similarly protected the BBB by inhibiting this signaling cascade. The results provide a novel herbal medicine to protect BBB following TBI via an antioxidative molecular mechanism.

MicroRNA-892b influences proliferation, migration and invasion of bladder cancer cells by mediating the p19ARF/cyclin D1/CDK6 and Sp-1/MMP-9 pathways.

Cancers often utilize microRNAs to suppress tumor suppressor genes, thus facilitating their potential for growth and invasion. In the present study, we report the novel findings that miR-892b inhibits proliferation, migration and invasion of bladder cancer cells. The basal expression level of miR‑892b was significantly lower in 3different bladder cancer cell lines than in normal human urothelial cells. Transfection of miR-892b mimics to bladder cancer cells resulted in dose‑dependent growth arrest. Flow cytometric analysis of the cell cycle showed that miR-892b-transfected bladder cancer cells were subject to arrest in the G1phase, which was due to the downregulation of cyclinD1 and CDK6 followed by upregulation of p19ARF. In addition, overexpression of miR-892b impeded the migration and invasion of EJ cells. Expression of MMP-9 in EJ cells was blocked by transfection of miR-892b; the effect was regulated, at least in part, by activation of the Sp-1 transcription factor. Overall, we verified that miR-892b regulates the p19ARF/cyclinD1/CDK6 and Sp-1/MMP-9 signaling networks in bladder cancer cells and may provide a treatment option for advanced-stage bladder cancers.

Abstract 1844: MDM2 overexpression promotes angiogenesis and metastasis through dysregulation of MMP-9 and TIMP-1 pathway

Abstract MDM2 is a multifunctional oncogene that has been associated with highly prevalent and aggressive cancers such as colon and prostate. MDM2 promotes a negative regulation of p53 activities and thus can prevent several of p53 protective effects including DNA repair, cell cycle control and apoptosis. Several pharmacological blockers that target the N-terminal domain of MDM2 leading to a disruption of MDM2-p53 interaction are being developed. For instance, nutlin-3 is the prototype of this family and has been shown to confine the tumor growth in many cancer types. Recently, it has been confirmed that MDM2 also promotes metastasis and angiogenesis through activation of number of mediators such VEGF and HIF-1α. However, mounting evidence suggests that MDM2 has p53 independent functions. Therefore, understanding the mechanisms by which MDM2 promotes the metastatic ability of tumors might provide an invaluable therapeutic target and a potential prognostic biomarker. Particularly, MMP-9/TIMP-1 dysregulation has been known to be associated with cancer progression and metastasis. This system is involved in the digestion of extracellular matrix and thus a release of several sequestered cytokines and growth factors such as TGFβ1. Among the release cytokines, TGFβ1 has been proven to be involved in promoting the MMP-9 expression and consequent metastatic events. Therefore, we hypothized that MDM2 promotes angiogenesis and metastasis through dysregulation of TIMP-1/MMP-9 pathway. Hence, the goal of our study was to evaluate the influence of MDM2 on tumor angiogenesis and metastatic ability through dysregulation of TIMP-1/MMP-9 pathway. To test our hypothesis, LNCaP prostate cancer and MDM2-transfected LNCaP-MST cell lines were used. On the other hand, nutlin-3 at the dose of 20 μM for 24 hours were used to examine the influence of MDM2 in regulating MMP-9/TIMP-1 in LNCaP and LNCaP-MST cells. Utilizing qPCR and western blotting techniques, we obtained data which suggest that MDM2 positively regulates the expression of MMP-9 and negatively regulates TIMP-1. We observed that MMP-9 overexpressed in LNCaP-MST, and this overexpression was reversed after nutlin-3 treatment in LNCaP-MST by 0.40 fold. Conversely, the results showed a complete abolishment of TIMP-1 protein level in LNCaP-MST, and nutlin-3 does not seem to impact TIMP-1 level. The data suggest that MDM2 positively impacts MMP-9 in a p53 dependent fashion as opposed to TIMP-1, which is negatively regulated by MDM2 independent of p53. In the same connection, TGFβ1 overexpression was reversed by 0.51 fold in LNCaP-MST after nutlin-3 treatment as shown by the qPCR results. Altogether, this project would provide a significant insight into understanding cancer metastasis processes with respect to MDM2 activity (The financial support from the Royal Dames of Cancer Research Inc., Ft. Lauderdale is gratefully acknowledged). Citation Format: Ali Alaseem, Thiagarajan Venkatesan, Priya Dondapati, Saad Alobid, Appu Rathinavelu. MDM2 overexpression promotes angiogenesis and metastasis through dysregulation of MMP-9 and TIMP-1 pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1844.

P21WAF1 Is Required for Interleukin-16-Induced Migration and Invasion of Vascular Smooth Muscle Cells via the p38MAPK/Sp-1/MMP-9 Pathway.

Interleukin-16 (IL-16) is a lymphocyte chemoattractant factor well known for its role in immune responses, but its role in vascular disease is unknown. Here, we explored the novel physiological function of IL-16 in vascular smooth muscle cells (VSMCs). The expression of IL-16 and its receptor CD4 was observed in VSMCs. Treatment with IL-16 enhanced the migration and invasion by VSMCs without altering the proliferative potential. IL-16 induced MMP-9 expression via the binding activity of transcription factors NF-κB, AP-1, and Sp-1 motifs in VSMCs. Among the relevant signaling pathways examined, only p38MAPK phosphorylation was significantly stimulated in IL-16-treated VSMCs. Treatment with p38MAPK inhibitor SB203580 prevented the IL-16-induced migration and invasion of VSMCs. SB203580 treatment inhibited the MMP-9 expression and activation of Sp-1 binding in IL-16-treated VSMCs, and siRNA knockdown of CD4 expression blocked the induction of migration, invasion, p38MAPK phosphorylation, MMP-9 expression, and Sp-1 binding activation stimulated by IL-16. The IL-16 induced cell-cycle-inhibitor p21WAF1 expression in VSMCs, but had no effect on the expression levels of other cell-cycle negative regulators. Finally, blockage of p21WAF1 function with specific siRNA abolished the IL-16-induced elevation of migration, invasion, p38MAPK phosphorylation, MMP-9 expression, and Sp-1 binding activation in VSMCs. Taken together, p21WAF1 was required for the induction of p38MAPK-mediated MMP-9 expression via activation of the Sp-1 binding motif, which led to migration and invasion of VSMCs interacting with IL-16/CD4. These results could provide that IL-16 is a new target in the treatment of vascular diseases such as atherosclerosis and re-stenosis.

IGF-1 Regulates the Extracellular Level of Active MMP-2 and Promotes Müller Glial Cell Motility.

In ischemic proliferative retinopathies, Müller glial cells (MGCs) acquire migratory abilities. However, the mechanisms that regulate this migration remain poorly understood. In addition, proliferative disorders associated with enhanced activities of matrix metalloproteinases (MMPs) also involve insulin-like growth factor (IGF)-1 participation. Therefore, the main interest of this work was to investigate the IGF-1 effect on the extracellular proteolytic activity in MGCs. Cell culture supernatants and cell lysates of the human MGC line MIO-M1 stimulated with IGF-1 were analyzed for MMP-2 by zymographic and Western blot analysis. The MGCs' motility was evaluated by scratch wound assay. The MMP-2, β1-integrin, and focal adhesions were detected by confocal microscopy. The localization of active MMPs and actin cytoskeleton were evaluated by in situ zymography. The IGF-1 induced the activation of canonical signaling pathways through the IGF-1R phosphorylation. Culture supernatants showed a relative decrease in the active form of MMP-2, correlating with an increased accumulation of MMP-2 protein in the MGCs' lysate. The IGF-1 effect on MMP-2 was abolished by an IGF-1R blocking antibody, αIR3, as well as by the PI3-kinase inhibitor, LY294002. The IGF-1 increased the migratory capacity of MGCs, which was blocked by the GM6001 MMP inhibitor, LY294002 and αIR3. Finally, IGF-1 induced the intracellular distribution of MMP-2 toward cellular protrusions and the partial colocalization with β1-integrin and phospo-focal adhesion kinase signals. Gelatinase activity was concentrated along F-actin filaments. Taken together, these data indicate that IGF-1, through its receptor activation, regulates MGCs' motility by a mechanism that involves the MMP-2 and PI3K signaling pathway.

CRM197 in Combination With shRNA Interference of VCAM-1 Displays Enhanced Inhibitory Effects on Human Glioblastoma Cells.

CRM197 is a naturally nontoxic diphtheria toxin mutant that binds and inhibits heparin-binding epidermal growth factor-like growth factor. CRM197 serves as carrier protein for vaccine and other therapeutic agents. CRM197 also inhibits the growth, migration, invasion, and induces apoptosis in various tumors. Vascular cell adhesion molecule-1 (VCAM-1) is an important cell surface adhesion molecule associated with malignancy of gliomas. In this work, we aimed to investigate the role and mechanism of CRM197 combined with shRNA interference of VCAM-1 (shRNA-VCAM-1) on the migration, invasion, and apoptosis of glioblastoma cells. U87 and U251 human glioblastoma cells were treated with CRM197 (10 µg/ml) and shRNA interfering technology was employed to silence VCAM-1 expression. Cell viability, migration, invasiveness, and apoptosis were assessed with CCK8, Transwell and Annexin V-PE/7-AAD staining. Activation of cleaved caspase-3, 8, and 9, activity of matrix metalloproteinase-2/9 (MMP-2/9), and expression of phosphorylated Akt (p-Akt) were also checked. Results showed that CRM197 and shRNA-VCAM-1 not only significantly inhibited the cell proliferation, migration, invasion, but also promoted the apoptosis of U87 and U251 cells. Combined treatment of both displayed enhanced inhibitory effects on the malignant biological behavior of glioma cells. The activation of cleaved caspase-3, 8, 9 was promoted, activity of MMP-2 and MMP-9 and expression of p-Akt were inhibited significantly by the treatment of CRM197 and shRNA-VCAM-1 alone or in combination, indicating that the combination of CRM197 with shRNA-VCAM-1 additively inhibited the malignant behavior of human glioblastoma cells via activating caspase-3, 8, 9 as well as inhibiting MMP-2, MMP-9, and Akt pathway.

Qingre quyu granule stabilizes plaques through inhibiting the expression of tenascin-C in patients with severe carotid stenosis.

To investigate the therapeutic effects of Qingre Quyu Granule (QQG) on the patients with severe carotid stenosis, and to explore the mechanism of it. Ninety-six patients with severe carotid stenosis were enrolled in the study and were classified into a QQG group (n=48) and a control group (n=48) randomly using consecutively numbered envelopes. The patients in the QQG group were given QQG and Western medicine, those in the control group were given Western medicine merely, the course of treatment was 16 weeks. All patients went through endarterectomy after treatment. Plaques were subjected to the analysis of CD3, CD68, soluble intercellular adhesion molecule 1 (ICAM-1), matrix metalloprotease-9 (MMP-9), CD40L, tenascin-C, and collagen content lipid content by immunohistochemistry or polarized light analysis. By the end of experiment, the expressions of CD3, CD68, ICAM-1, MMP9, CD40L and tenascin-C on the plaques were statistically significant lower in the QQG group compared with the control group(P<0.01). The lipid content of the plaque was also significantly lower in the QQG group compared with the control group (P<0.01). The interstitial collagen in the tissue sections of the plaques was also significantly higher in the QQG group in comparison with the control group (P<0.01). QQG could stabilize carotid artery plaques through inhibiting pro-inflammation factors and restraining the tenascin-C and MMP9 pathway.

Cochinchina momordica seed suppresses proliferation and metastasis in human lung cancer cells by regulating multiple molecular targets.

Cochinchina Momordica Seed, which is the dried ripe seed of Momordica cochinchinensis (Lour.) Spreng, has been used as a mainly anticancer ingredient for many years in China. This study aims at investigating the roles of an ethanol-soluble extract of Cochinchina Momordica Seed (ECMS) in suppressing the proliferation and metastasis of human lung cancer cells, and further elucidating underlying molecular mechanisms. Our researches suggest that ECMS dose-dependently decreased the survival rates of A549 and H1299 cells, and inhibited the migration and invasion in A549 cells. ECMS-induced apoptosis was accompanied by up-regulation of p53, Bax and the down-regulation of Bcl-2, PI-3K/Akt signal pathway, and resulted in the dissipation of mitochondrial membrane potential (ΔΨm) and sequentially activated caspase-3 cascade. Pre-treated with specific inhibitors, LY294002 (PI-3K inhibitor) and BAY11-7082 (NF-κB inhibitor) could enhance the anti-proliferation effects of ECMS on A549 cells. Furthermore, ECMS could increase the level of E-cadherin and decrease of the level of STAT-3 and MMP-2, and scarcely affected the expression of VEGF, and resulted in the inhibition of migration and invasion. Pre-treated with specific inhibitors, WP1066 (STAT-3 inhibitor) and TIMP-2 (MMP-2 inhibitor) could enhance the inhibitory effects of ECMS on migration. In conclusion, the current data demonstrated ECMS inhibited the proliferation of A549 cells by inducing apoptosis, at least partly through the activation of p53 and inactivation of PI-3K/Akt signaling. STAT-3 and MMP-2 pathways may be partly involved in anti-metastasis activities of ECMS. Hence, ECMS might be a promising candidate for the therapy of the non-small cell lung cancer by regulating multiple molecular targets.

Ang-(1–7) exerts protective role in blood–brain barrier damage by the balance of TIMP-1/MMP-9

Cerebrovascular disease (CVD) ranks as the top three health risks, specially cerebral ischemia characterized with the damage of blood–brain barrier (BBB). The angiotensin Ang-(1–7) was proven to have a protective effect on cerebrovascular diseases. However, its role on blood–brain barrier and the underlying molecular mechanism remains unclear. In this study, Ang-(1–7) significantly relieved damage of ischemia reperfusion injury on blood–brain barrier in cerebral ischemia reperfusion injury (IRI) rats. Furthermore, its treatment attenuated BBB permeability and brain edema. Similarly, Ang-(1–7) also decreased the barrier permeability of brain endothelial cell line RBE4. Further analysis showed that Ang-(1–7) could effectively restore tight junction protein (claudin-5 and zonula occludens ZO-1) expression levels both in IRI-rats and hypoxia-induced RBE4 cells. Furthermore, Ang-(1–7) stimulation down-regulated hypoxia-induced matrix metalloproteinase-9 (MMP-9) levels, whose silencing with (matrix metalloproteinase-9 hemopexin domain) MMP9-PEX inhibitor significantly increased the expression of claudin-5 and ZO-1. Further mechanism analysis demonstrated that Ang-(1–7) might junction protein levels by tissue inhibitor of metalloproteinase 1 (TIMP1)–MMP9 pathway, because Ang-(1–7) enhanced TIMP1 expression, whose silencing obviously attenuated the inhibitor effect of Ang-(1–7) on MMP-9 levels and decreased Ang-(1–7)-triggered increase in claudin-5 and ZO-1. Together, this study demonstrated a protective role of Ang-(1–7) in IRI-induced blood–brain barrier damage by TIMP1–MMP9-regulated tight junction protein expression. Accordingly, Ang-(1–7) may become a promising therapeutic agent against IRI and its complications.

EPO gene expression promotes proliferation, migration and invasion via the p38MAPK/AP-1/MMP-9 pathway by p21WAF1 expression in vascular smooth muscle cells

The use of recombinant human erythropoietin (rHuEpo) can lead to hypertrophy and hyperplasia, and has induced the proliferation of vascular smooth muscle cells (VSMCs). The effect of the EPO gene in the migration and invasion of VSMCs remains unclear. In this study, overexpression of the EPO gene increased the DNA synthesis and phosphorylation of ERK1/2 and p38MAPK in VSMCs. In addition, EPO gene expression induced the migration and invasion of VSMCs via the expression of MMP-9 by the activation of NF-κB and AP-1 binding. A blockade of p38MAPK by specific p38MAPK inhibitor SB203580 led to a suppression of the increased DNA synthesis, migration, and invasion of VSMCs that was induced by the EPO gene. SB203580 treatment blocked the increased expression of MMP-9 through the binding activity of AP-1. Transfection of the EPO gene with VSMCs was associated with the up-regulation of cyclin D1/CDK4, cyclin E/CDK2, and p21WAF1, and with the down-regulation of p27KIP1. The specific suppression of p21WAF1 expression by siRNA rescued the enhancement of DNA synthesis via the phosphorylation of p38MAPK and the increase in migration and invasion through AP-1-mediated MMP-9 expression in EPO gene transfectants. These novel findings demonstrate that p21WAF1 regulates the proliferation, migration and invasion of VSMC induced by EPO gene.

Molecular Mechanisms of EML in AML1/ETO+ AML and Its Targeting Treatments

Our previous study has been reported that AML1/ETO positive patients with highly expressed of APP were much easier to extramedullary infiltration, and got poor prognosis£¨followed-up median 35 ( 6-96 ) months, RFS in the high expression APP group was significantly lower than the low expression group £¬40.0% vs 80.0%£¬P = 0.001). In vitro study, we constructed a cell model kasumi-1 which was consistent with AML1/ETO positive and high expressed of APP gene. The cell migration was significantly reduced after interferce of APP expression. This study was designed to investigate the molecule mechanism of extramedullary leukemia (EML) in kasumi-1 cell model and to invent a strategy for treatment in clinic.In this study, we found p-ERK, c-Myc and MMP-2 were significantly decreased after APP expression knockdown in kasumi-1 cell. Meanwhile, we added the inhibitors to block p-ERK and c-Myc expression. The results showed that protein expression of p-ERK and c-Myc was significantly decreased after p-ERK inhibitor performance, which was proportional to the time and concentration. c-Myc and MMP-2 protein expression was significantly reduced after c-Myc inhibitor was used, but p-ERK didn’t change (Fig.1B). So, we concluded that APP might regulated the AML cell migration via APP/p-ERK/c-Myc/MMP-2 pathway.Also, we found that kasumi-1 cell was resistant to adriamycin (ADM) and Ara-C, which meant APP may be related with drug resistance. So, we detected cell surviving fraction after ADM and Ara-C performance via MTT assay. The results showed that there was no difference in control group and siAPP group. But, when compared with controls groups, cell surviving fraction in siEZH2 group was significantly decreased after ADM and Ara-C performance respectively. Furthermore, we found protein expression of EZH2 was significantly reduced after LBH589 treatment in cell culture. So, we concluded that LBH589 or SiEZH2 could increase sensitivity of kasumi-1 cell to ADM and Ara-C.In sum, in AML1/ETO positive leukemia cells, we first report that APP gene regulates cell migration via APP/p-ERK/c-Myc/MMP-2 pathway and EZH2 gene induces drug resistantence. Interference or blocking of EZH2 and APP expression may be helpful in treating AML1/ETO positive leukemia. DisclosuresNo relevant conflicts of interest to declare.