Differential regulation of plasminogen activation in normal keratinocytes and SCC-4 cells by fibroblasts (original) (raw)
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2000
In vivo tumor progression in mice with targeted deficiencies in urokinase-type plasminogen activator (UPA ؊/؊ ) and its inhibitor, plasminogen activator inhibitor-1 (PAI-1 ؊/؊ ), was studied using a fibrosarcoma tumor model. Murine T241 fibrosarcoma cells were s.c. implanted into three groups of mice with the following genotypes, wild-type (WT), UPA ؊/؊ , and PAI-1 ؊/؊ . A significantly diminished primary tumor growth in UPA ؊/؊ and PAI-1 ؊/؊ mice occurred, relative to WT mice. Tumors in UPA ؊/؊ and PAI-1 ؊/؊ mice displayed lower proliferative and higher apoptotic indices and displayed a different neovascular morphology, as compared with WT mice. These results are consistent with the decreased growth rates of this tumor in these gene-deleted mice. Immunohistochemical analyses of the tumors revealed a decrease in vascularity and vascular endothelial growth factor expression only in tumors in PAI-1 ؊/؊ mice. Analyses of the relative extents of corneal angiogenesis in these same animals, induced by basic fibroblast growth factor, corroborated the resistance of PAI-1 ؊/؊ mice to neovascularization. The results obtained suggest that the host fibrinolytic system plays an important role in tumor growth in this model. Alterations in host expression of components of this system may alter tumor growth and dissemination by affecting the balance between tumor cell death and proliferation, as well as extracellular matrix changes needed for invasiveness and angiogenesis.
Cancer research, 2000
In vivo tumor progression in mice with targeted deficiencies in urokinase-type plasminogen activator (UPA ؊/؊ ) and its inhibitor, plasminogen activator inhibitor-1 (PAI-1 ؊/؊ ), was studied using a fibrosarcoma tumor model. Murine T241 fibrosarcoma cells were s.c. implanted into three groups of mice with the following genotypes, wild-type (WT), UPA ؊/؊ , and PAI-1 ؊/؊ . A significantly diminished primary tumor growth in UPA ؊/؊ and PAI-1 ؊/؊ mice occurred, relative to WT mice. Tumors in UPA ؊/؊ and PAI-1 ؊/؊ mice displayed lower proliferative and higher apoptotic indices and displayed a different neovascular morphology, as compared with WT mice. These results are consistent with the decreased growth rates of this tumor in these gene-deleted mice. Immunohistochemical analyses of the tumors revealed a decrease in vascularity and vascular endothelial growth factor expression only in tumors in PAI-1 ؊/؊ mice. Analyses of the relative extents of corneal angiogenesis in these same animals, induced by basic fibroblast growth factor, corroborated the resistance of PAI-1 ؊/؊ mice to neovascularization. The results obtained suggest that the host fibrinolytic system plays an important role in tumor growth in this model. Alterations in host expression of components of this system may alter tumor growth and dissemination by affecting the balance between tumor cell death and proliferation, as well as extracellular matrix changes needed for invasiveness and angiogenesis.
Induction of p53 by Urokinase in Lung Epithelial Cells
Journal of Biological Chemistry, 2005
Urokinase plasminogen activator (uPA) is a serine protease that catalyzes the conversion of plasminogen to plasmin. The plasminogen/plasmin system includes the uPA, its receptor, and its inhibitor (plasminogen activator inhibitor-1). Interactions between these molecules regulate cellular proteolysis as well as adhesion, cellular proliferation, and migration, processes germane to the pathogenesis of lung injury and neoplasia. In previous studies, we found that uPA regulates cell surface fibrinolysis by regulating its own expression as well as that of the uPA receptor and plasminogen activator inhibitor-1. In this study, we found that uPA alters expression of the tumor suppressor protein p53 in Beas2B airway epithelial cells in both a time-and concentration-dependent manner. These effects do not require uPA catalytic activity because the amino-terminal fragment of uPA lacking catalytic activity was as potent as two chain active uPA. Single chain uPA also enhanced p53 expression to the same extent as intact two chain active uPA and the amino-terminal fragment. Pretreatment of cells with anti-1 integrin antibody blocked uPA-induced p53 expression. uPA-induced p53 expression occurs without increased p53 mRNA expression. However, uPA induced oncoprotein MDM2 in a concentration-dependent manner. uPA-induced p53 expression does not require activation of tyrosine kinases. Inactivation of protein-tyrosine phosphatase SHP-2 inhibits both basal and uPA-induced p53 expression. Plasmin did not alter uPA-mediated p53 expression. The induction of p53 expression by exposure of lung epithelial cells to uPA is a newly recognized pathway by which urokinase may influence the proliferation of lung epithelial cells. This pathway could regulate pathophysiologic alterations of p53 expression in the setting of lung inflammation or neoplasia.
The American Journal of Pathology, 2013
Alveolar type II epithelial cell (ATII) apoptosis and proliferation of mesenchymal cells are the hallmarks of idiopathic pulmonary fibrosis, a devastating disease of unknown cause characterized by alveolar epithelial injury and progressive fibrosis. We used a mouse model of bleomycin (BLM)einduced lung injury to understand the involvement of p53-mediated changes in urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) levels in the regulation of alveolar epithelial injury. We found marked induction of p53 in ATII cells from mice exposed to BLM. Transgenic mice expressing transcriptionally inactive dominant negative p53 in ATII cells showed augmented apoptosis, whereas those deficient in p53 resisted BLM-induced ATII cell apoptosis. Inhibition of p53 transcription failed to suppress PAI-1 or induce uPA mRNA in BLM-treated ATII cells. ATII cells from mice with BLM injury showed augmented binding of p53 to uPA, uPA receptor (uPAR), and PAI-1 mRNA. p53-binding sequences from uPA, uPAR, and PAI-1 mRNA 3 0 untranslated regions neither interfered with p53 DNA binding activity nor p53-mediated promoter transactivation. However, increased expression of p53binding sequences from uPA, uPAR, and PAI-1 mRNA 3 0 untranslated regions in ATII cells suppressed PAI-1 and induced uPA after BLM treatment, leading to inhibition of ATII cell apoptosis and pulmonary fibrosis. Our findings indicate that disruption of p53efibrinolytic system cross talk may serve as a novel intervention strategy to prevent lung injury and pulmonary fibrosis. (Am J Pathol 2013, 183: 131e143; http://dx.Idiopathic pulmonary fibrosis is a progressive and fatal lung disease that is refractory to current therapy. A better understanding of the underlying mechanisms is necessary for development of novel treatments. Dysregulated fibrinolysis and induction of p53 are often associated with lung injury and precede development of pulmonary fibrosis. 1 These changes occur in a mouse model of bleomycin (BLM)einduced lung injury and accelerated pulmonary fibrosis. 1 p53 Expression increases substantially in type II alveolar epithelial (ATII) cells after BLM-or cigarette smokeeinduced lung injury, 1e3 in association with induction of plasminogen activator inhibitor-1 (PAI-1) and suppression of urokinase-type plasminogen activator (uPA) and uPA receptor (uPAR) expression.
Urokinase Expression by Tumor Suppressor Protein p53
American Journal of Respiratory Cell and Molecular Biology, 2008
Lung carcinoma (H1299) cells deficient in p53 (p53 2/2) express large amounts of urokinase-type plasminogen activator (uPA) protein and uPA mRNA, and exhibit slower degradation of uPA mRNA than that of p53-expressing nonmalignant Beas2B human airway epithelial cells. Expression of p53 protein in H1299 cells, upon transfection with p53 cDNA, suppressed basal as well as uPA-induced expression of uPA protein in both conditioned media and cell lysates, and decreased the level of steady-state uPA mRNA primarily due to increased uPA mRNA turnover. Inhibition of p53 expression by RNA silencing (SiRNA) in Beas2B cells enhanced basal and uPA-mediated uPA protein and mRNA expression with stabilization of uPA mRNA. Purified p53 binds to the uPA mRNA 39 untranslated region (UTR) in a sequence-specific manner and endogenous uPA mRNA associates with p53 protein isolated from Beas2B cytosolic extracts. p53 binds to a 35-nucleotide uPA 39UTR sequence and insertion of this sequence into b-globin mRNA accelerates degradation of otherwise stable b-globin mRNA. These observations confirm a new role for p53 as a uPA mRNA binding protein that down-regulates uPA mRNA stability and decreases cellular uPA expression.
Biological Chemistry, 2002
domain, kringle(s), a serine protease domain and also a finger domain for t-PA] (Ny et al., 1984; Danø et al., 1994). As serine proteases, PAs are involved in many physiological and pathological processes, including embryogenesis, tissue repair, chemotaxis, fibrinolysis, angiogenesis and tumor invasion. These processes are complex and may involve different types of events, including proteolysis, migration and/or proliferation of a variety of cell types. t-PA is thought to be mainly involved in fibrinolysis, whereas u-PA in association with its receptor (u-PAR) is thought to play a role in the tissue processes of localized and directional proteolysis (Pollanen et al., 1991; Johnsen et al., 1998). u-PA and t-PA control a variety of proteolytic processes: a cascade of events mediated by the activation of plasminogen (PG) in plasmin, which in turn may activate the procollagenases in collagenases (Johnsen et al., 1998); the biological activation of HGF, from the single-to the two-chain form (Mars et al., 1993); and the direct degradation of plasma and cellular FN (pFN, cFN) at specific sites (Gold et al., 1992; Marchina and Barlati, 1996). PAs are involved in many processes. In particular, the following has been demonstrated: u-PA stimulates the migration and proliferation of a variety of tumor and normal cells (Kirchheimer et al., 1989; Blasi, 1997; Chapman, 1997); t-PA is involved in the mitogenesis of cultured human skin fibroblasts (De Petro et al., 1994) and of human aortic smooth muscle cells (Herbert et al., 1994); u-PA, in association with its receptor, and independently of its serine protease domain, may transduce intracellular signals (via PTK, CK2…), which are involved in cell migration and proliferation (Dumler et al., 1999). For the role of PAs in the process of tumor growth and invasion, evidence is emerging that they may play a distinct role, at least in certain types of solid tumors. Indeed, the high levels of u-PA, and not of t-PA, are unfavorable prognostic factors for cancer patients with several types of tumors (Schmitt et al., 2000), whereas the high levels of t-PA detected in certain malignant conditions are associated with a better prognosis (Yamashita et al., 1993; Ferrier et al., 2000). According to our previous data, in human hepatocellular carcinoma the high levels of u-PA mRNA, and not of t-PA mRNA, detected in the tumor biopsies were inversely related to patient survival (De Petro et al., 1998). In breast carcinoma and in primary melanoma of the limb, the high levels of t-PA were correlated with a better prognosis (
Anticancer research, 2013
To investigate possible differences in the effects of soluble factors from oral squamous cell carcinoma (SCC) cells (UT-SCC-87) and normal oral keratinocytes (NOK) on fibroblast expression of genes involved in tumor stroma turnover. Transwell co-cultures with fibroblasts in collagen gels, and SCC cells or NOK in inserts were carried out. Fibroblast gene expression was measured with real-time polymerase chain reaction (PCR). The expression of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) was up-regulated in co-cultures with SCC cells but not with NOK. In contrast, both SCC cells and NOK regulated matrix metalloproteinase-1 (MMP1) and -3, and tissue inhibitor of metalloproteinases-2 (TIMP2) and -3 to a similar extent, while MMP2 and TIMP1 were largely unaffected. Interleukin 1 alpha (IL1α) up-regulated both MMP1 and MMP3 and down-regulated PAI-1, TIMP2 and -3. SCC and NOK regulate fibroblast expression of genes involved in tumor stroma turnov...
1997
p53, a tumor suppressor and a transcription factor, has been shown to transcriptionally activate the expression of a number of important genes involved in the regulation of cell growth, DNA damage, angiogenesis, and apoptosis. In a computer search for other potential p53 target genes, we identified a perfect p53 binding site in the promoter of the human type IV collagenase (also called 72-kDa gelatinase or matrix metalloproteinase 2 [MMP-2]) gene. This p53 binding site was found to specifically bind to p53 protein in a gel shift assay. Transcription assays with luciferase reporters driven by the promoter or enhancer of the type IV collagenase gene revealed that (i) activation of the promoter activity is p53 binding site dependent in p53positive cells but not in p53-negative cells and (ii) wild-type p53, but not p53 mutants commonly found in human cancers, transactivates luciferase expression driven by the type IV collagenase promoter as well as by a p53 site-containing enhancer element in the promoter. Significantly, expression of the endogenous type IV collagenase is also under the control of p53. Treatment of U2-OS cells, a wild-type p53-containing osteogenic sarcoma line, with a common p53 inducer, etoposide, induced p53 DNA binding and transactivation activities in a time-dependent manner. Induction of type IV collagenase expression followed the p53 activation pattern. No induction of type IV collagenase expression can be detected under the same experimental conditions in p53-negative Saos-2 cells. All these in vitro and in vivo assays strongly suggest that the type IV collagenase gene is a p53 target gene and that its expression is subject to p53 regulation. Our finding links p53 to a member of the MMP genes, a family of genes implicated in trophoblast implantation, wound healing, angiogenesis, arthritis, and tumor cell invasion. p53 may regulate these processes by upregulating expression of type IV collagenase.
Gene Regulation by p53 in Human Cancer System
Asian Pacific Journal of Cancer Biology
TP53 proto-oncogene constitutes tumor induction in more than 50% of human cancers as it is mutated frequently in a wide range of cell lines. The transcription of TP53 is postulated to be autoregulated via either binding with TBP and CBF or via direct interaction of p53 protein with TP53 promoter, though further investigation is needed to acknowledge it. Alteration in pathways, regulated through wild type, by mutant p53 (Mutp53) give rise to immortality through interaction with other transcription factors or inducing receptor tyrosine kinases and other signal components. The missense mutation is more frequent constituting more than 60% among all mainly because of the high rate of G>A or C>T transitions in TP53, giving rise to mutation hotspots in R248, R273, etc. In addition to the loss of function, mutations in the TP53 gene also confers oncogenic functions that are not found in wild type p53, referred to as Gain of Function (GOF). GOF mutp53 has been found to promote metastas...