Differential transcriptional activation of matrix metalloproteinase-2 and membrane type-1 matrix metalloproteinase by experimental deep venous thrombosis and thrombin

ObjectiveResolution of deep venous thrombosis (DVT) is involved in the pathogenesis of postthrombotic syndrome. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes that are critical in angiogenesis and tissue remodeling. We hypothesized that MMP-2 and its membrane-bound activator membrane type-1 matrix metalloproteinase (MT1-MMP) expression would be expressed and activated during the resolution of DVT.MethodsDVT was generated by caval ligation in wild-type and MMP-2 transgenic reporter mice. Ligated and sham-operated (control) cavae were analyzed for MMP-2 transcription (β–galactosidase activity in MMP-2 reporter mice) and MT1-MMP mRNA by real-time polymerase chain reaction. MMP-2 activity was determined by zymography, and immunohistochemical staining for β–galactosidase and MT1-MMP protein was used to localize expression. Human umbilical vascular endothelial cells (HUVEC) were treated with 10 U/mL thrombin and MMP-2 and MT1-MMP mRNA levels and MMP-2 activity was determined.ResultsMMP-2 activity increased 71% (n = 5, P < .05) at day 8 in ligated vs control cavae by zymography. β-galactosidase activity showed a 1.2-fold (n = 8, P < .05) and 1.7-fold (n = 8, P < .05) induction in MMP-2 transcription at day 3 and day 8, respectively. No significant MT1-MMP gene induction was seen at day 3 in ligated vs control cavae, but MT1-MMP mRNA was upregulated 2.5-fold (n = 8, P < .05) in ligated cavae at day 8. Immunohistochemical staining localized MMP-2 and MT1-MMP expression to the vein wall and cellular infiltrates of the thrombus. Thrombin-treated HUVEC showed differential responses of MMP-2 and MT1-MMP. Zymography of conditioned media and cell lysates illustrated a 220% (152.6 ± 8.6 vs 69.445 ± 5.46 pixels/unit area, n = 5, P < .05) and 150% (74.1 ± 7.3 vs 49.2 ± 5.7 pixels/unit area, n = 5, P < .05) increases in MMP-2 activity respectively. MMP-2 mRNA levels were downregulated 30% (0.48 ± 0.023 vs 0.63 ± 0.035 copies of MMP-2 mRNA/copy GAPDH, n = 5, P < .05), whereas MT1-MMP message was upregulated 250% (0.147 ± 0.009 vs 0.059 ± 0.005 copies of MT1-MMP mRNA/copy GAPDH, n = 5, P < .05).ConclusionsResolution of DVT is associated with increased MMP-2 transcription and activity as well as MT1-MMP expression. Thrombin may mediate the increase in MT1-MMP noted in DVT. This is the first article studying MMP-2 and MT1-MMP transcription in DVT. These findings add DVT resolution to the class of inflammatory and fibrotic disorders in which transcriptional activation of the MT1-MMP/MMP-2 genes occurs and identify a potential therapeutic target to modulate this clinically relevant process.Clinical RelevancePostthrombotic syndrome remains a significant clinical problem after deep venous thrombosis (DVT), but the cellular and molecular mechanisms involved in thrombus resolution and vein wall fibrosis remain undefined. Matrix metalloproteinase (MMP) enzymes are critical to cell migration and matrix breakdown. We identify gene transcription and activity of two MMP isoforms, MMP-2 and MMP-14 (membrane type MMP 1, MT1-MMP) in the resolution phase of experimental DVT and in thrombin-treated endothelial cells. These studies define new proteases potentially important to resolution of DVT and development of postthrombotic syndrome

Families of maps Singularities and its Gauss maps

Abstract: This paper mainly studies the Singularities of smooth mapping. The singularities of the families of Gauss maps corresponding to the family of mappings are studied and the shape of these families and their singularities using mathematica program are illustrated and plotted. By changing the control parameters we find some singular points for the family which can be classified according to the famous theorems in singularity theory. Using the Hessian matrix we can obtain the singular points and singular set. Geometrically these singularities can be plotted but the classification of them can&apos;t be a valuable for all points. Using the terminology of level set which tell us the type of singular points like folds (level sets is start line), cusp (level set is semicubical parabola). In general there is no existance of some famous types. where ∆ is the discriminant set and it is a plan. Remark 4 The family of contours is given from z = k (constant), and the family of zero level set corresponding to k = 0 are given through the figurer

Metamorphosis of plasma turbulence-shear flow dynamics through a transcritical bifurcation

The structural properties of an economical model for a confined plasma turbulence governor are investigated through bifurcation and stability analyses. A close relationship is demonstrated between the underlying bifurcation framework of the model and typical behavior associated with low- to high-confinement transitions such as shear flow stabilization of turbulence and oscillatory collective action. In particular, the analysis evinces two types of discontinuous transition that are qualitatively distinct. One involves classical hysteresis, governed by viscous dissipation. The other is intrinsically oscillatory and non-hysteretic, and thus provides a model for the so-called dithering transitions that are frequently observed. This metamorphosis, or transformation, of the system dynamics is an important late side-effect of symmetry-breaking, which manifests as an unusual non-symmetric transcritical bifurcation induced by a significant shear flow drive.Comment: 17 pages, revtex text, 9 figures comprised of 16 postscript files. Submitted to Phys. Rev.

A Novel Intracellular Isoform of Matrix Metalloproteinase-2 Induced by Oxidative Stress Activates Innate Immunity

Experimental and clinical evidence has pinpointed a critical role for matrix metalloproteinase-2 (MMP-2) in ischemic ventricular remodeling and systolic heart failure. Prior studies have demonstrated that transgenic expression of the full-length, 68 kDa, secreted form of MMP-2 induces severe systolic failure. These mice also had unexpected and severe mitochondrial structural abnormalities and dysfunction. We hypothesized that an additional intracellular isoform of MMP-2, which affects mitochondrial function is induced under conditions of systolic failure-associated oxidative stress.Western blots of cardiac mitochondria from the full length MMP-2 transgenics, ageing mice and a model of accelerated atherogenesis revealed a smaller 65 kDa MMP-2 isoform. Cultured cardiomyoblasts subjected to transient oxidative stress generated the 65 kDa MMP-2 isoform. The 65 kDa MMP-2 isoform was also induced by hypoxic culture of cardiomyoblasts. Genomic database analysis of the MMP-2 gene mapped transcriptional start sites and RNA transcripts induced by hypoxia or epigenetic modifiers within the first intron of the MMP-2 gene. Translation of these transcripts yields a 65 kDa N-terminal truncated isoform beginning at M(77), thereby deleting the signal sequence and inhibitory prodomain. Cellular trafficking studies demonstrated that the 65 kDa MMP-2 isoform is not secreted and is present in cytosolic and mitochondrial fractions, while the full length 68 kDa isoform was found only in the extracellular space. Expression of the 65 kDa MMP-2 isoform induced mitochondrial-nuclear stress signaling with activation of the pro-inflammatory NF-κB, NFAT and IRF transcriptional pathways. By microarray, the 65 kDa MMP-2 induces an innate immunity transcriptome, including viral stress response genes, innate immunity transcription factor IRF7, chemokines and pro-apoptosis genes.A novel N-terminal truncated intracellular isoform of MMP-2 is induced by oxidative stress. This isoform initiates a primary innate immune response that may contribute to progressive cardiac dysfunction in the setting of ischemia and systolic failure