Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously reported that two closely related protein kinase C (PKC) isoforms, PKC alpha and PKC beta I, had divergent effects on the growth and transformation of the same parental R6 rat embryo fibroblast cell line (Housey, G. M., Johnson, M. D., Hsiao, W.-L. W. O'Brian, C. A., Murphey, J. P., Kirschmeier, P., and Weinstein, I. B. (1988) Cell 52, 343-354; Borner, C., Filipuzzi, I., Weinstein, I. B., and Imber, R. (1991) Nature 353, 78-80). Whereas cells that overexpress PKC beta I lost anchorage dependence, grew to higher saturation densities, and generated small tumors when injected into nude mice, none of these properties were seen with cells that overexpress PKC alpha. In fact, the latter cells grew even slower and to lower saturation densities as compared to control cells. Here we investigate possible molecular mechanisms underlying the reciprocal effects of PKC alpha and PKC beta I. Overexpression of both isoforms enhanced 12-O-tetradecanoyl phorbol-13 acetate-induced expression of the growth regulatory genes c-jun, c-myc, and collagenase and enhanced feedback inhibition of epidermal growth factor receptor binding and cellular levels of diacylglycerol. However, the cells overexpressing PKC beta I differed from those overexpressing PKC alpha by displaying a decreased requirement for growth factors and by the production of a mitogenic factor. Thus, the basis for enhanced growth and transformation of cells overexpressing PKC beta I may be the establishment of an autocrine growth factor loop. These findings may be relevant to the roles of specific isoforms of PKC in carcinogenesis and tumor growth.
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PMID:Two closely related isoforms of protein kinase C produce reciprocal effects on the growth of rat fibroblasts. Possible molecular mechanisms. 781 23

The highly metastatic amelanotic C8161 human melanoma line was found to exhibit complete dominance of its undifferentiated and metastatic phenotype in multiple somatic cell hybridization studies designed to bypass the presence of potential tumor suppressor genes. In a three armed approach involving somatic cell fusions of C8161 with recipient lines of greater differentiation, different lineage, and different tumorigenicity status, the metastatic and undifferentiated phenotype of C8161 was promiscuously dominant. In somatic cell hybrids produced between the C8161 and a group of non-metastatic human melanoma lines which exhibited melanocyte differentiation markers including S100, HMB-45, NKI/C3, and melanin, the fusions were uniformly metastatic and undifferentiated. In somatic cell hybrids of C8161 and MCF-7 the fusions exhibited an estrogen independent and unresponsive, estrogen receptor (ER) negative, and highly metastatic phenotype. In fusions between C8161 and HMS-1, an immortalized 'benign' human myoepithelial line which produced an abundant extracellular matrix (ECM) and high levels of protease and angiogenic inhibitors including maspin, tissue inhibitor of metalloproteinase-1 (TIMP-1), alpha1-antitrypsin (alpha1-AT), protease nexin II (PN-II), thrombospondin-1 and soluble basic fibroblast growth factor (bFGF) receptors, the hybrids showed complete absence of matrix, absent maspin expression, markedly decreased protease inhibitor and angiogenic inhibitor production, high levels of proteases and angiogenic factors, and a highly metastatic phenotype. In our somatic cell fusions, the human-human hybrids represented true and complete fusions and not hybrid clones selected for by loss of dominant-acting growth suppressor genes. This finding was supported by detailed comparative genomic hybridization (CGH) studies, Q-banding karyotype analysis, and autofusions of representative clones. The purposeful creation of inherently unstable human-murine fusions between C8161 and B16-F1 where loss of putative suppressor loci would be expected, resulted in fusions exhibiting decreased growth and non-metastatic behavior with progressive chromosomal loss. Neither p53, nm23, DNA methyltransferase, activated ras, fibroblast growth factor-4 (FGF-4), or epidermal growth factor receptor (EGFR) mediated the acquisition of the metastatic or undifferentiated phenotype within the C8161-human fusions. These studies are the first studies ever to successfully transfer the complete metastatic phenotype by somatic cell fusion and support the presence of a new high level regulatory pathway(s) involving dominant trans-acting factors which act pleiotropically to regulate an undifferentiated and highly metastatic phenotype.
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PMID:Evidence of a dominant transcriptional pathway which regulates an undifferentiated and complete metastatic phenotype. 936 25

As a model system for the identification of genes involved in the progression of human breast cancer, differential gene expression in cell lines MCF-7 and MCF-7ADR was investigated. The latter cell line is derived from the former. Cell line MCF-7 is estrogen receptor-positive, vimentin-negative and uninvasive in the Matrigel outgrowth assay and in the nude mouse, while MCF-7ADR is estrogen receptor-negative, hormone-resistant, vimentin-positive, invasive in the Matrigel outgrowth assay and in the nude mouse and resistant to adriamycin due to overexpression of glycoprotein gp170. We have shown that tumor progression in this model system is mediated by transcriptional regulation of mitochondria-related genes, proteases, transmembrane receptors and cell cycle-related gene proteins. Among the genes differentially regulated at the transcriptional level in the cell lines MCF-7 and MCF-7ADR are a new mitochondrial transcript, mitochondrial creatine kinase, matrix metalloproteinase-1, stromelysin-3, urokinase and its receptor, tissue factor, E-cadherin, epidermal growth factor receptor, transmembrane proteins Mat-8 and progression associated protein (PAP), cyclin E, cyclin-dependent kinase-2 and cell cycle inhibitory proteins p16, p21 and p27.
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PMID:Molecular analysis of two mammary carcinoma cell lines at the transcriptional level as a model system for progression of breast cancer. 951 94

In response to cutaneous injury, expression of collagenase-1 is induced in keratinocytes via alpha2beta1 contact with native type I collagen, and enzyme activity is essential for cell migration over this substratum. However, the cellular mechanism(s) mediating integrin signaling remain poorly understood. We demonstrate here that treatment of keratinocytes cultured on type I collagen with epidermal growth factor receptor (EGFR) blocking antibodies or a specific receptor antagonist inhibited cell migration across type I collagen and the matrix-directed stimulation of collagenase-1 production. Additionally, stimulation of collagenase-1 expression by hepatocyte growth factor, transforming growth factor-beta1, and interferon-gamma was blocked by EGFR inhibitors, suggesting a required EGFR autocrine signaling step for enzyme expression. Collagenase-1 mRNA was not detectable in keratinocytes isolated immediately from normal skin, but increased progressively following 2 h of contact with collagen. In contrast, EGFR mRNA was expressed at high steady-state levels in keratinocytes isolated immediately from intact skin but was absent following 2 h cell contact with collagen, suggesting down-regulation following receptor activation. Indeed, tyrosine phosphorylation of the EGFR was evident as early as 10 min following cell contact with collagen. Treatment of keratinocytes cultured on collagen with EGFR antagonist or heparin-binding (HB)-EGF neutralizing antibodies dramatically inhibited the sustained expression (6-24 h) of collagenase-1 mRNA, whereas initial induction by collagen alone (2 h) was unaffected. Finally, expression of collagenase-1 in ex vivo wounded skin and re-epithelialization of partial thickness porcine burn wounds was blocked following treatment with EGFR inhibitors. These results demonstrate that keratinocyte contact with type I collagen is sufficient to induce collagenase-1 expression, whereas sustained enzyme production requires autocrine EGFR activation by HB-EGF as an obligatory intermediate step, thereby maintaining collagenase-1-dependent migration during the re-epithelialization of epidermal wounds.
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PMID:Keratinocyte collagenase-1 expression requires an epidermal growth factor receptor autocrine mechanism. 1018 26

Numerous reports have shown an association between overexpression of the epidermal growth factor receptor (EGFR), and poor prognosis in head and neck squamous cell carcinomas (HNSCC), however, the underlying mechanisms are still unclear. In the present study, we set out to determine whether EGFR expression was associated with in vitro invasive capacity in a panel of four established and ten newly derived HNSCC lines. Ten of the cell lines expressed high levels of EGFR as determined by a ligand-binding assay and dot blot analysis, whereas the remaining four showed weak overexpression or normal levels of EGFR. The ability of cells to invade through Matrigel was found to be higher in the EGFR overexpressing cell lines (p < 0. 0001). Expression levels of matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-7, MMP-9, MMP-10, MMP-11, MMP-13, MT1-MMP) and tissue inhibitors of MMP (TIMP-1, TIMP-2) were evaluated by semiquantitative RT-PCR, substrate zymography and western blot. We found a strong positive correlation between EGFR levels and the expression of MMP-9 mRNA (r(2) = 0.95; p < 0.0001), MMP-9 enzyme activity (r(2) = 0.8099; p < 0.0001) and an inverse correlation with TIMP-1 (r(2) = 0.48; p = 0.0059). In six selected HNSCC lines, in vitro invasion was assayed in the presence of an anti-EGFR monoclonal antibody, ICR62. A significant reduction of invasion in four selected EGFR-overexpressing cell lines was found with 30 nM ICR62 (from 50% to 70%; p < 0.001) but there was no effect in two cell lines with normal EGFR levels. Our results show that the in vitro invasive phenotype of HNSCC lines correlates with high EGFR and MMP-9 expression, and it is therefore suggested that the EGFR signaling pathway might play an important role in the invasive behavior of HNSCC via specific upregulation of MMP-9 and downregulation of TIMP-1.
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PMID:Overexpression of epidermal growth factor receptor in human head and neck squamous carcinoma cell lines correlates with matrix metalloproteinase-9 expression and in vitro invasion. 1076 Aug 16

Incubation of human dermal fibroblasts in keratinocyte-conditioned culture medium led to a 5.7-fold increase in the level of matrix metalloproteinase-1. Virtually all of the matrix metalloproteinase-1 - inducing activity could be related to agonists acting through members of the epidermal growth factor receptor family or to agonists acting through the interleukin-1 receptor. The same keratinocyte-conditioned medium also induced a modest increase in fibroblast proliferation (approximately 1.8-fold). Growth-stimulating activity could be attributed to epidermal growth factor receptor (but not interleukin-1 receptor) function. In fibroblasts exposed to keratinocyte-conditioned medium, mitogen-activated protein kinase signalling through both the extracellular signal-related kinase pathway and p38 pathway occurred. When recombinant epidermal growth factor or recombinant interleukin-1beta were used as a control, they induced mitogen-activated protein kinase signalling consistent with the combined effects of epidermal growth factor receptor - specific and interleukin-1 receptor - specific agonists in keratinocyte-conditioned medium. Recombinant epidermal growth factor stimulated both matrix metalloproteinase-1 induction and proliferation while recombinant interleukin-1beta stimulated matrix metalloproteinase-1 elaboration but not fibroblast growth. An inhibitor of extracellular signal-related kinase pathway signalling (U0126) blocked induction of matrix metalloproteinase-1 production induced by keratinocyte-conditioned medium (as well as by epidermal growth factor or interleukin-1beta), and also inhibited proliferation. A p38 signalling inhibitor (SB203580) blocked matrix metalloproteinase-1 elaboration induced by keratinocyte-conditioned medium or interleukin-1beta, but did not inhibit matrix metalloproteinase-1 elaboration or cell growth induced by epidermal growth factor. These data indicate that keratinocyte-fibroblast interactions are mediated by multiple stimulating agents acting on specific receptors to induce signalling through different mitogen-activated protein kinase pathways leading to altered expression of key biological functions.
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PMID:Keratinocyte stimulation of matrix metalloproteinase-1 production and proliferation in fibroblasts: regulation through mitogen-activated protein kinase signalling events. 1217 84

Increasingly, there is the need to analyze gene expression in tumor tissues and correlate these findings with clinical outcome. Because there are few tissue banks containing enough frozen material suitable for large-scale genetic analyses, methods to isolate and quantify messenger RNA (mRNA) from formalin-fixed, paraffin-embedded tissue sections are needed. Recovery of RNA from routinely processed biopsies and quantification by the polymerase chain reaction (PCR) has been reported; however, the effects of formalin fixation have not been well studied. We used a proteinase K-salt precipitation RNA isolation protocol followed by TaqMan quantitative PCR to compare the effect of formalin fixation for 24, 48, and 72 hours and for 1 week in normal (2), oral epithelial dysplasia (3), and oral squamous cell carcinoma (4) specimens yielding 9 fresh and 36 formalin-fixed samples. We also compared mRNA and protein expression levels using immunohistochemistry for epidermal growth factor receptor (EGFR), matrix metalloproteinase (MMP)-1, p21, and vascular endothelial growth factor (VEGF) in 15 randomly selected and routinely processed oral carcinomas. We were able to extract RNA suitable for quantitative reverse transcription (RT) from all fresh (9/9) and formalin-fixed (36/36) specimens fixed for differing lengths of time and from all (15/15) randomly selected oral squamous cell carcinoma. We found that prolonged formalin fixation (>48 h) had a detrimental effect on quantitative RT polymerase chain reaction results that was most marked for MMP-1 and VEGF but less evident for p21 and EGFR. Comparisons of quantitative RT polymerase chain reaction and immunohistochemistry showed that for all markers, except p21, there was good correlation between mRNA and protein levels. p21 mRNA was overexpressed in only one case, but protein levels were elevated in all but one tumor, consistent with the established translational regulation of p21. These results show that RNA can be reliably isolated from formalin-fixed, paraffin-embedded tissue sections and can produce reliable quantitative RT-PCR data. However, results for some markers are adversely affected by prolonged formalin fixation times.
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PMID:Effect of duration of fixation on quantitative reverse transcription polymerase chain reaction analyses. 1221 16

Human skin is exposed to solar ultraviolet radiation. Ultraviolet radiation damages human skin and results in an old and wrinkled appearance, called photoaging. We have previously reported that molecular mechanisms by which ultraviolet light causes photoaging involve activation of growth factor and cytokine receptors in keratinocytes and dermal cells. They lead to downstream signal transduction through activation of mitogen-activated protein kinase (extracellular signal-regulated kinase, c-jun N-terminal protein kinase, and p38) pathways. These signaling pathways converge in the nucleus of cells to form an activated complex of transcription factor activator protein 1 (cFos/cJun), which induces matrix metalloproteinases that degrade skin connective tissue. In addition to cell surface receptor activation, generation of reactive oxygen species by ultraviolet radiation is believed to be critical in triggering mitogen-activated protein kinase pathways. We investigated the ability of (i) ultraviolet irradiation to generate reactive oxygen species in human skin in vivo; and (ii) genistein, which possesses both tyrosine kinase inhibitory and antioxidant activities, and n-acetyl cysteine, which can be converted into the endogenous antioxidant glutathione, to impair responses to ultraviolet light that eventuate in photoaging in human skin in vivo. Ultraviolet irradiation caused a rapid and significant increase in hydrogen peroxide levels in human skin in vivo. Pretreatment of human skin with genistein inhibited ultraviolet-induced epidermal growth factor receptor tyrosine kinase activity, whereas n-acetyl cysteine did not. Genistein inhibited ultraviolet induction of both extracellular signal-regulated kinase and cJun N-terminal protein kinase activities. n-Acetyl cysteine inhibited extracellular signal-regulated kinase but not cJun N-terminal protein kinase activation. Both genistein and n-acetyl cysteine prevented ultraviolet induction of cJun protein. Consistent with this, genistein and n-acetyl cysteine blocked ultraviolet induction of cJun-driven enzyme, collagenase. Neither genistein nor n-acetyl cysteine acted as sunscreens as they had no effect on ultraviolet-induced erythema. These data indicate that compounds similar to genistein and n-acetyl cysteine, which possess tyrosine kinase inhibitory and/or antioxidant activities, may prevent photoaging.
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PMID:Topical N-acetyl cysteine and genistein prevent ultraviolet-light-induced signaling that leads to photoaging in human skin in vivo. 1271 90

For growth stimulation of liver cells by hepatocyte growth factor (HGF) or transforming growth factor alpha (TGFalpha) via receptor tyrosine kinases, c-fos/c-jun has been considered a point of intersection for cross-talk between the different signal transduction pathways. Recent evidence strongly implicates translocation of pro-TGFalpha into the nucleus as an important step preceding the initiation of hepatic DNA synthesis. We asked whether an active c-jun is required for the nuclear translocation of pro-TGFalpha and its stimulatory effect on DNA synthesis. For this purpose we used mice with c-jun inactivated post partum in hepatocytes by the Cre-loxP recombination system (c-jun(Deltaliver)). Nuclear fractions from control and c-jun(Deltaliver) mouse livers contained TGFalpha as pro-form of 17 kDa and the epidermal growth factor receptor (erbb-1) with molecular weights of 170 and 150 kDa (truncated form). Hepatocytes were isolated by collagenase perfusion and cultivated. A lack of c-jun did not alter the apoptotic activity but significantly suppressed DNA synthesis in the cultured hepatocytes. In control and c-jun(Deltaliver) cells DNA synthesis was almost always associated with nuclear presence of pro-TGFalpha. 76.5 +/- 6.8% of hepatocytes with pro-TGFalpha positive nuclei and only 4.52 +/- 1.31% of hepatocytes with negative nuclei exhibited DNA replication. About 85% of the pro-TGFalpha positive nuclei also contained erbb-1. Treatment of cultures with mature TGFalpha or HGF elevated the frequency of pro-TGFalpha positive nuclei replicating DNA; HGF and TGFalpha-induced nuclear pro-TGFalpha and DNA synthesis significantly more in c-jun(Deltaliver) than in control hepatocytes. These results suggest that (i) a lack of c-jun suppresses basal rates of DNA replication in hepatocytes; (ii) c-jun deficient hepatocytes show a pronounced growth response towards HGF or TGFalpha; (iii) nuclear translocation of pro-TGFalpha together with erbb-1 and its association with DNA synthesis are independent of c-jun.
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PMID:Induction of DNA synthesis in primary mouse hepatocytes is associated with nuclear pro-transforming growth factor alpha and erbb-1 and is independent of c-jun. 1277 Oct 26

This study was undertaken to evaluate the effects of thiazolidinediones (TZD) on keratinocyte proliferation, motility, and matrix metalloproteinase (MMP) production. Rosiglitazone (a potent TZD) inhibited both proliferation and motility as well as elaboration of MMP-1 and MMP-9. Inhibition was obtained with keratinocytes in monolayer culture and human skin in organ culture. There were significant concentration-response differences in sensitivity of the three keratinocyte responses to treatment with rosiglitazone. In contrast to keratinocytes, dermal fibroblasts were resistant to the effects of rosiglitazone. Treatment of keratinocytes with rosiglitazone did not suppress epidermal growth factor receptor autophosphorylation, but inhibited signaling through the extracellular regulated kinase mitogen-activated protein kinase pathway without a concomitant effect on pathways that lead to c-jun activation. Pioglitazone, another TZD, also suppressed keratinocyte proliferation, although it was less effective than rosiglitazone. An experimental TZD (BP-1107) inhibited keratinocyte proliferation at a much lower concentration than either rosiglitazone or pioglitazone. Because enhanced keratinocyte motility and increased MMP production as well as increased keratinocyte proliferation are thought to contribute to the phenotype of psoriatic lesional skin, we propose that interference with these keratinocyte responses contributes to the previously reported antipsoriatic activity of TZD.
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PMID:Rosiglitazone inhibits proliferation, motility, and matrix metalloproteinase production in keratinocytes. 1496 1


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