Plasminogen activator system, vascular endothelial growth factor, and colorectal cancer progression.

AIMS: To plasminogen activator system (PAS) consists of the plasminogen activators (urokinase (uPA) and tissue-type (tPA) plasminogen activators), the uPA receptor (uPAR), and the plasminogen activator inhibitors (PAI-1 and PAI-2). Plasminogen activators activate plasminogen to plasmin, which can break down extracellular matrix (ECM) components. Vascular endothelial growth factor (VEGF) is a mitogen for endothelial cells and is involved in angiogenesis. VEGF has been shown to upregulate uPA and this may facilitate tumour angiogenesis further. METHODS: PAS components and VEGF were determined by enzyme linked immunosorbent assay (ELISA) in paired colorectal tumour and normal tissue (n = 50) and correlated with pathological staging. RESULTS: uPA, uPAR, PAI-1, and VEGF values were significantly higher in tumour tissue (for example, tumour uPA: median, 2.3 (range, 0.1-6.7) ng/mg protein v normal uPA: median, 0.2 (range, 0-2.6) ng/mg protein). tPA was significantly higher in normal mucosa and there was no difference in PAI-2. uPA, uPAR, PAI-1, and VEGF values significantly correlated with each other and with Dukes's staging (uPA in adenomas: median, 0.42 (range, 0.1-1.2) ng/mg protein; upA in Dukes's B tumours: median, 2.1 (range, 0.4-4.3) ng/mg protein; and uPA in Dukes's D tumours: median, 4.0 (range, 3.7-4.2) ng/mg protein) and lymphatic invasion. In addition PAI-1 also correlated with tumour size and differentiation. CONCLUSION: The involvement of the PAS and VEGF in colorectal cancer appears to be complex. uPA, uPAR, PAI-1, and VEGF were upregulated in tumour tissue and this correlated with Dukes's staging and lymphatic invasion.     

Differential localization and expression of urokinase plasminogen activator (uPA), its receptor (uPAR), and its inhibitor (PAI-1) mRNA and protein in endometrial tissue during the menstrual cycle

Normal endometrium is a highly dynamic tissue, which responds to ovarian steroids with cyclic proliferation, differentiation (secretion), and degradation (menstruation). The urokinase plasminogen activator (uPA)-dependent proteolytic cascade as well as ligand activation of the uPA receptor (uPAR) is critically involved in physiological as well as pathophysiological aspects of tissue expansion and remodelling. Cyclic variation and distribution of uPA, uPAR and plasminogen activator inhibitor 1 (PAI-1) mRNA were examined by in situ hybridization, real-time PCR and northern blot in normal endometrium. Their corresponding proteins were localized with immunohistochemistry. uPA mRNA is exclusively expressed by stromal cells, whereas uPA protein is present in both epithelial and stromal cells. Immunostaining for uPA protein is reduced or undetectable at midcycle, thus coinciding with peak concentration of uPA in the uterine fluid. uPAR mRNA is expressed by epithelial cells in the proliferative phase and by stromal cells in the secretory phase. However, epithelial cells stain for uPAR protein throughout the cycle, suggesting that uPAR may detach from stromal cells and then bind to epithelial cells in the secretory phase. PAI-1 mRNA is located in vessel walls. The late secretory phase has greatly increased expression of all three mRNA and their proteins, mainly in pre-decidual cells in the superficial stroma. Discordant localization of the mRNA and proteins suggest that uPA is produced by stromal cells, released and bound to epithelial cells in both the proliferative and secretory phases, whereas uPAR is released from the stroma and bound to epithelial cells in the secretory phase. Also, the present data together with earlier reports suggest that uPA is released from the epithelial cells to the uterine fluid.     

Plasminogen activator inhibitor-1 as a measure of vascular remodelling in breast cancer

The generation of urokinase plasminogen activator (uPA) by tumours is an important pathway for neoplastic cell invasion and metastasis. Indeed in several tumour types, elevated levels of uPA, its receptor (uPAR) or its inhibitor plasminogen activator inhibitor-1 (PAI-1) is associated with a poorer prognosis. Since endothelial cells also use this proteolytic system to remodel the extracellular matrix during angiogenesis and since angiogenesis, as assessed by microvessel density, is also a predictor of patient survival, this study was designed to investigate the relationship between angiogenesis and the urokinase system in breast tumours. The aims were to assess whether the uPA, uPAR and/or PAI-1 correlates with angiogenic activity and could therefore be a useful objective clinical measure of tumour neovascularization; and to clarify whether the poor outcome associated with high levels of the urokinase system is due to its association with angiogenesis. The study also sought to examine the relationship between the uPA system and vessel remodelling using loss of a basement membrane epitope (LH39) normally associated with established capillaries. The cytosolic levels of uPA, PAI-1 and uPAR were therefore measured by enzyme linked immunoabsorbent assay, together with tumour vascularity, in 136 well-characterized invasive breast carcinomas. There were significant relationships between uPA and uPAR (Spearman r=0.37, p<0.0001), uPA and PAI-1 (Spearman r=0.19, p=0.03) and between uPAR and PAI-1 (Spearman r=0.23 p=0.01). A significant correlation was also observed between PAI-1 and vessel remodelling (Spearman r=0.34, p=0.04), patient age (p=0.01), nodal status (p=0.047) and tumour grade (p=0.04), but no association between tumour vascularity and PAI (p=0.96), uPA (p=0.69) or uPAR (p=0.81) was present. No significant association was seen between any of the urokinase variables and expression of the angiogenic factor thymidine phosphorylase. Furthermore, no significant associations were found between any of the studied parameters and overall survival in a univariate analysis of the cancer patients. A multivariate Cox proportional hazard model of overall survival showed that uPA (p=0.15), but not uPAR (p=0.52) or PAI-1 (p=0.61), gave no additional prognostic information. These findings show that uPA may work via an independent pathway to angiogenesis and therefore combined blockade of uPA and angiogenesis may have additional therapeutic benefits. It also shows, as recently demonstrated in animal models, that PAI-1 may be a key regulator of vascular remodelling in human cancer.     

Immunohistochemical expression of uPA, uPAR, and PAI-1 in breast carcinoma. Fibroblastic expression has strong associations with tumor pathology

The urokinase-type plasminogen activator (uPA) system has been implicated in tumor spread. We have used immunohistochemistry to examine three components of this system, ie, uPA, uPA receptor (uPAR), and plasminogen activator inhibitor-1 (PAI-1), in a pilot study on 142 cases of breast carcinoma. We wished to determine whether there were any relationships between expression of the proteins in either tumor cells or fibroblasts and clinical and pathological features. Strong uPA expression in each cell type was significantly related to high tumor grade (P = 0.013 and 0.008, respectively), and was more common in invasive than in in situ carcinomas (P < 0.0001). Fibroblastic expression of uPAR was only related to the presence of invasion (P < 0.0001). Strong PAI-1 expression in both cell types was seen in high-grade tumors (tumor cells, P = 0.012; fibroblasts, P < 0.001), but only fibroblastic expression was related to the presence of invasion (P = 0.042). Fibroblastic expression of both uPA and uPAR were positively correlated with tumor size. Although patients with strong fibroblastic expression of uPA showed a tendency toward a shorter time to relapse, none of the plasminogen activator proteins were significantly associated with relapse-free survival. These results suggest that strong expression of uPA, uPAR, and PAI-1 in fibroblasts rather than in tumor cells have the most impact on the clinical behavior of breast cancer. Larger prospective studies are needed to confirm these findings.     

Spitz naevi may express components of the plasminogen activation system

The plasminogen activation (PA) system is involved in the process of invasion and metastasis. Its major components are urokinase (uPA) and tissue-type plasminogen activator (tPA), plasminogen activation inhibitor type 1 and 2 (PAI-1 and PAI-2) and a receptor for urokinase (uPAR). In this study, the expression of plasminogen activation components in Spitz naevi was compared with that in common and dysplastic naevi on the one hand and primary cutaneous melanomas on the other. Spitz naevi had melanocytic positivity for uPA in 0% (0/36), tPA in 30% (6/20), PAI-1 in 10% (3/35), PAI-2 in 40% (8/21) and uPAR in 60% (13/21) of cases. This far exceeded the expression found in common (n = 25) and dysplastic (n = 15) naevi, which only showed melanocytic positivity for PAI-2 (20% and 15% respectively) and in one dysplastic naevus also for uPAR. This was much (for most components significantly) less than the proportion of primary melanomas with tumour cell positivity, which was 30% (11/38) for uPA, 80% (19/24) for tPA, 75% (28/38) for PAI-1, 80% (19/24) for PAI-2 and 80% (19/24) for uPAR. The main findings of this study are that Spitz naevi, firstly, may express plasminogen activator (tPA), inhibitors and the receptor of the PA system, but in a much smaller proportion than cutaneous melanomas; and secondly, do not express urokinase, whereas some of the melanomas do. uPA positivity may therefore be suggestive of melanoma. However, overlapping staining results imply that the PA system has limited value in the differential diagnosis between Spitz naevus and primary melanoma. As serine protease components are expressed, Spitz naevi may use this proteolytic machinery to accomplish matrix degradation, although in a more restricted, possibly transient manner than melanomas.     

Expression of urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, and plasminogen activator inhibitor-1 and -2 in hepatocellular carcinoma

It has become more and more clear in recent decades that the plasminogen activation system, which includes urokinase-type plasminogen activator (uPA), urokinase-type plasminogen activator receptor (uPAR), plasminogen activator inhibitor (PAI)-1 and PAI-2, plays a very important role in the aggressiveness of cancer. Using immunohistochemistry and enzyme-linked immunosorbent assay (ELISA), the expression of these four components of the uPA system was analyzed in 19 cases of hepatocellular carcinoma (HCC) and 18 cases of the adjacent non-cancer tissues which all had chronic active hepatitis with liver fibrosis or liver cirrhosis. Four cases of normal liver tissues, as controls for immunohistochemical stains, were obtained from the hepatectomized liver of patients with metastatic cancer in the liver. The positive rates of uPA, uPAR, PAI-1 and PAI-2 for immunohistochemical stains in cancer tissues were 78.9, 68.4, 57.9 and 31.6%, respectively. Positive signals were mainly distributed in the cytoplasm of the cancer and in stromal cells. Moreover, the strong stains were chiefly located in the invasive front of the cancer cells. No specific stain was detected in four cases of normal liver tissues. In ELISA, there were significant differences between cancer and non-cancer tissues in concentration of uPA, uPAR and PAI-1 (P < 0.0003, 0.0024 and 0.01, respectively), but there was no significant difference in that of PAI-2 (P = 0.37). These results suggest that uPA, uPAR and PAI-1 are related to invasion of HCC.     

Vitamin D3 modulation of plasminogen activator inhibitor type-1 in human breast carcinomas under organ culture

Urokinase plasminogen activator (uPA), its cell-bound receptor (uPAR) and its main inhibitor plasminogen activator type 1 (PAI-1) are present primarily in stromal cells in invasive breast carcinoma. The purpose of this study was to investigate the regulation by 1,25 dihydroxyvitamin-D3 (VD3) of these invasion-associated markers expressed in breast cancer tumors under organ culture, which preserves the interacting network of tumor and stromal cells. Breast carcinoma slices (30 cases), obtained using the Krumdieck tissue slicer, cultured for 48 h in the presence or absence of 100 nM vitamin D3, were embedded in formalin-fixed paraffin. uPA, uPAR, PAI-1 and VD3 receptor (VDR) were analyzed by immunohistochemistry, and their expression, detected in tumor cells and fibroblasts of the specimens, was not statistically changed by culture conditions. The proportion of cases expressing uPA, uPAR and PAI-1 was not affected by VD3 in epithelial cells, but the fraction of cases displaying strong PAI-1 reactivity in fibroblasts was reduced (P=0.016) compared with control slices. Fibroblasts isolated from invasive ductal carcinomas and from normal breast tissues expressed higher VDR mRNA levels than epithelial cells. In cultured tumor fibroblasts, PAI-1 immunostaining and mRNA levels were reduced by VD3-limiting fibroblast contribution to invasion.     

Promigratory Effect of Plasminogen Activator Inhibitor-1 on Invasive Breast Cancer Cell Populations

The urokinase-type plasminogen activator (uPA) system is a dynamic complex in which the membrane receptor uPAR binds uPA that binds the plasminogen activator inhibitor (PAI)-1 localized in the extracellular matrix, resulting in endocytosis of the whole complex by the low-density lipoprotein receptor-related protein (LRP). High expression of PAI-1 is paradoxically associated with marked tumor spreading and poor prognosis. We previously reported a nonproteolytic role of the [uPAR:uPA:PAI-1:LRP] complex operative in cell migration. Here we explored whether matrix PAI-1 could be used as a migration support by human breast cancer cells. We showed that the uPA system and LRP are localized at filopodia of invasive cells, and that formation/internalization of the [uPAR:uPA:PAI-1:LRP] complex is required for attachment and migration of cancer cells on plastic and on a PAI-1 coat. PAI-1 increased both filopodia formation and migration of cancer cells suggesting a chemokine-like activity. Migration velocity, expression of the uPA system, use of the [uPAR:uPA:PAI-1:LRP] complex to migrate, and promigratory effects of PAI-1 paralleled cancer cell invasiveness. Phenotyping and functional analysis of invasive cancer cell subclones indicated that different cell subpopulations may use different strategies to migrate depending on both the environment and their expression of the uPA system, some of them taking advantage of abundant available PAI-1.     

In situ localization of mRNA for the fibrinolytic factors uPA, PAI-1 and uPAR in endometriotic and endometrial tissue

Endometriotic tissue grows invasively. The plasminogen-activating system is suggested to participate in degradation of extracellular matrix (ECM) and modulation of cell adhesion and migration. We have previously demonstrated elevated levels of the fibrinolytic factors urokinase plasminogen activator (uPA) and plasminogen activator inhibitor (PAI-1) in endometriotic tissue and endometrium from women with endometriosis. The aim of the present study was to localize the uPA, PAI-1 and urokinase plasminogen activator receptor (uPAR) mRNA in endometriotic tissue and in endometrium both from women with and without endometriosis. With in situ hybridization, we found that uPA mRNA seems to be up-regulated in endometriotic glands and endometrial stroma as well as PAI-1 mRNA in endometriotic and endometrial stroma from women with endometriosis. uPAR mRNA likewise appears to be up-regulated in both glands and stroma in endometriotic tissue and in endometrial glands from patients compared to endometrial glands and stroma from healthy women. These differences might be important for menstrual shedding and adherence of endometrial fragments to peritoneal lining in women developing endometriosis and for the invasive growth of endometriotic tissue.     

The plasminogen activator inhibitor "paradox" in cancer

Proteolysis in general and specifically the plasminogen activating system regulated by urokinase (uPA) its specific receptor, the GPI membrane anchored urokinase receptor (uPAR) and the specific plasminogen activator inhibitor 1 (PAI-1) plays a major role in tumorigenesis, tumor progression, tumor invasion and metastasis formation. This is exemplified by a body of published work showing a positive correlation between the expression of uPA or uPAR in several tumors and their malignancy. It is generally assumed that such a "pro-malignant" effect of the uPA-uPAR system is mediated by increased local proteolysis thus favoring tumor invasion, by a pro-angiogenic effect of this system and also by uPA-uPAR signaling towards the tumor thereby shifting the tumor phenotype to a more "malignant" one. However, when tumor patients are analyzed for long term survival, those with high levels of the inhibitor of the system, PAI-1 have a much worse prognosis than those with lower PAI-1 levels. This indicates that increased overall proteolysis alone cannot be made responsible for the adverse effects of the plasminogen activating system in tumors. Moreover, it becomes increasingly evident that components of the fibrinolytic system secreted by the tumor cells themselves are not solely responsible for a correlation between the plasminogen activating system and tumor malignancy; components of the plasminogen activating system secreted by stroma cells or cells of the immune system such as macrophages contribute also to the impact of fibrinolysis on malignancy. This review summarizes the evidence for the role of plasminogen activator inhibitor-1 in mediating the malignant phenotype and possible mechanism thereby trying to explain the "PAI-1 paradox in cancer" on a molecular level.     

大鼠肝纤维化过程中4种肝脏细胞对纤溶系统的调节

目的:探讨在大鼠肝纤维化发展过程中,肝细胞、肝星状细胞(HSCs)、Kupffer细胞和内皮细胞(ECs)对纤溶系统的调节作用。方法:采用1周2次皮下注射50%CCl4、持续12周的方法制备大鼠慢性肝纤维化模型;将肝纤维化不同时期的4种肝脏细胞进行分离;采用Northern印迹法及Western印迹法分别测定大鼠各类肝脏细胞中尿激酶型纤溶酶原激活物(uPA)、以及它的抑制物(PAI-1)和受体(uPAR)的mRNA和蛋白表达。结果:Northern印迹法及Western印迹法分析均显示,在大鼠肝纤维化过程中,HSCs和ECs均为PAI-1、uPAR的主要产生细胞,相比之下,HSCs产生更多。结论:在大鼠肝纤维化发生发展过程中,HSCs和ECs对纤溶系统成分的产生起着重要的调节作用,而HSCs则是PAI-1、uPAR最主要的产生细胞。     

uPAR与肿瘤的研究新进展

尿激酶型纤溶酶原激活物受体(urokinase plasminogen activator receptor,uPAR)是一个糖基磷脂酰肌醇锚定的蛋白质,高亲和力结合并激活尿激酶型纤溶酶原激活物(urokinase plasminogen activator,uPA),由此调节细胞表面蛋白水解活性。uPAR在几乎所有人类肿瘤中高表达,其高表达与增强肿瘤增殖、迁移、侵袭有关。本文对uPAR增强肿瘤趋向性、诱导上皮-间质转变与促进胞葬作用等新功能以及靶向uPAR治疗进展进行综述。     

Dysregulation of matrix metalloproteinases and their tissue inhibitors is related to abnormality of left ventricular geometry and function in streptozotocin-induced diabetic minipigs

This study aimed to characterize matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in relation to changes in left ventricle (LV) geometry and function in a porcine model with streptozotocin (STZ)-induced diabetes. In 15 Chinese Guizhou minipigs with STZ-induced diabetes (diabetic group) and 15 age-matched normal controls (control group), Doppler tissue imaging was performed at 6 months of diabetes. Serum MMP-2, -9, TIMP-1, -4 and B-type natriuretic peptide (BNP) were determined. Expression of MMPs, TIMPs, urokinase type-plasminogen activator (uPA), its receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1) in aortic intima and LV myocardium was evaluated, with gelatinolytic activities of tissue MMP-2, -9 accessed by zymography. Left ventricle end-diastolic septum thickness (P < 0.05) and mass (P < 0.05) were increased, whereas peak systolic mitral annulus velocity (Sm, P < 0.001), LV systolic (P = 0.01) and diastolic strain (P < 0.001) were significantly decreased in diabetic group than in controls. Diabetic group showed higher expression of TIMP-1, -4 in aortic intima and LV myocardium (P < 0.01 or P < 0.05), with increased collagen content and elevated serum BNP level (P = 0.004) and lower gelatinolytic activities of tissue MMP-2, -9 (all P < 0.05). Semi-quantitative RT-PCR of those diabetic tissues revealed elevated mRNA levels of major TIMPs, uPA, uPAR and PAI-1. Reduction of serum MMP-2 and -9 levels was observed in diabetic group vs. control group (both P < 0.05). This study features elevated levels of TIMP-1, -4, uPA, uPAR and PAI-1, and decreased activities of MMP-2, -9 in aorta and myocardium in STZ-induced diabetic minipigs, indicating that MMP-TIMP dysregulation is associated with LV hypertrophy, cardiac dysfunction and increased cardiovascular fibrosis in diabetes.     

Markers of the uPA system and common prognostic factors in breast cancer

The urokinase plasminogen activator (uPA) system includes uPA and plasminogen activator inhibitor types 1 (PAI-1) and 2 that mainly act by regulating extracellular matrix degradation, and it is involved in tumor progression. The -675 4G/5G polymorphism of the PAI-1 gene regulates PAI-1 activity in serum. We aimed at studying the -675 4G/5G polymorphism of the PAI-1 gene and uPA, PAI-1, and cyclooxygenase-2 (COX-2) immunohistochemical expression in a series of breast cancer cases. Homozygosity for the 4G allele of the PAI-1 gene was associated with node-positive breast cancer ( P = .02). We showed a direct correlation between uPA and estrogen receptor expression ( P = .03); negative uPA expression was associated with negative hormonal expression, high tumor grade, and high proliferation index ( P < .05). A direct correlation was seen between uPA and PAI-1, uPA and COX-2, and PAI-1 and COX-2 expression ( P < .05). Interaction between uPA and COX-2 systems in breast cancer deserves further study.     

Lipoprotein receptor-related protein 1 regulates collagen 1 expression, proteolysis, and migration in human pleural mesothelial cells

The low-density lipoprotein receptor-related protein 1 (LRP-1) binds and can internalize a diverse group of ligands, including members of the fibrinolytic pathway, urokinase plasminogen activator (uPA), and its receptor, uPAR. In this study, we characterized the role of LRP-1 in uPAR processing, collagen synthesis, proteolysis, and migration in pleural mesothelial cells (PMCs). When PMCs were treated with the proinflammatory cytokines TNF-α and IL-1β, LRP-1 significantly decreased at the mRNA and protein levels (70 and 90%, respectively; P < 0.05). Consequently, uPA-mediated uPAR internalization was reduced by 80% in the presence of TNF-α or IL-1β (P < 0.05). In parallel studies, LRP-1 neutralization with receptor-associated protein (RAP) significantly reduced uPA-dependent uPAR internalization and increased uPAR stability in PMCs. LRP-1-deficient cells demonstrated increased uPAR t(1/2) versus LRP-1-expressing PMCs. uPA enzymatic activity was also increased in LRP-1-deficient and neutralized cells, and RAP potentiated uPA-dependent migration in PMCs. Collagen expression in PMCs was also induced by uPA, and the effect was potentiated in RAP-treated cells. These studies indicate that TNF-α and IL-1β regulate LRP-1 in PMCs and that LRP-1 thereby contributes to a range of pathophysiologically relevant responses of these cells.     

Expression of urokinase-type plasminogen activator,its receptor,and its inhibitor in gastric adenocarcinoma tissues.

The plasminogen and plasmin system, which is mainly regulated by urokinase-type plasminogen activator (uPA), its receptor (uPAR) and its inhibitor (PAI-1), is generally believed to play a role in cancer invasion and metastasis. This study was conducted to investigate the role of uPA, uPAR and PAI-1 in the invasion and metastasis of gastric adenocarcinoma. The expression of mRNAs for uPA and PAI-1 was determined by Northern blot analysis in nine primary gastric cancer tissues, nine paired metastatic lymph nodes and normal gastric mucosa. The mRNA of uPA was not or faintly detected in normal mucosa, while the expression was increased in both primary gastric cancer tissues and metastatic lymph nodes to a similar degree. The mRNA expression for PAI-1 in the gastric cancer tissues was not different from that in the paired metastatic lymph nodes and normal mucosae. uPAR was determined by immunohistochemical staining, demonstrating that five (56%) and six (67%) out of nine primary gastric cancer tissues and nine paired metastatic lymph nodes were positive, respectively and the intensity was stronger in metastatic lymph nodes. The results support the concept that most gastric cancer cells may have an innately moderate level of uPA and uPAR, and that increase of uPAR expression can be considered to be closely associated with cancer invasion and metastasis.     

Discrimination of different forms of the murine urokinase plasminogen activator receptor on the cell surface using monoclonal antibodies

The urokinase plasminogen activator receptor (uPAR) is a versatile three-domain GPI-anchored protein, which binds urokinase plasminogen activator (uPA) and thereby focalises plasminogen activation on the cell surface. Generation of a proteolytic potential is essential in both normal physiological and pathological extracellular tissue remodelling processes. uPA can also cleave uPAR, resulting in liberation of the amino-terminal domain I, which encompasses binding sites for both uPA and the adhesion molecule, vitronectin. In order to localise the different uPAR forms on the plasma membrane of murine monocyte macrophage-like P388D.1 cells, we have now generated and characterised two high-affinity murine mAbs, mR3 and mR4, raised against murine uPAR. mR3 was found to recognise an epitope located in domain I of uPAR. Surface plasmon resonance analyses and cell binding studies revealed that this mAb was able to bind preformed complexes of murine pro-uPA and murine uPAR. In contrast, mR4 recognises domains II-III in uPAR and does not bind preformed pro-uPA-uPAR complexes in similar analyses. Immunofluorescence microscopy of P388D.1 cells revealed that mR3 stained the cells equally well in the presence or absence of saturation with the amino-terminal fragment of uPA, ATF. However, the signal intensity obtained using another uPAR domain I specific mAb, mR1, was significantly reduced upon ATF saturation. Furthermore, when adding ATF, mR4 selectively stained the cleaved receptor. Applying these newly generated mAbs, we additionally demonstrated that cleaved and intact uPAR was evenly distributed on the surface of these cells.     

Expression and activation of proteases in co-cultures

The present study concerned the expression and activation of metalloproteinase-2 (MMP-2), metalloproteinase-9 (MMP-9) and the urokinase plasminogen activator/urokinase plasminogen activator receptor (uPA/uPAR) system in co-cultures of human colon carcinoma cell spheroids (HT29, LS180, SW948) with human normal colon epithelium (CCD 841 CoTr), myofibroblasts (CCD-18Co) and endothelial cells (HUVEC). Additionally, the influence of monensin on the production and function of the proteases was tested.Tumor cells expressed small amounts of MMP-2, MMP-9 and uPA. Normal cells generally produced proportionally higher concentrations of these proteases (especially MMP-2, compared with significantly smaller yields of MMP-9 and significantly lower amounts of uPAR than tumors. In co-cultures of tumor spheroids with normal cell monolayers, the concentration of the proteases was equal to the sum of the enzymes produced in monocultures of both types of cells. The highest activity of uPA, measured as the reduction of the chromogenic substrate (S-2444), was detected in supernatants and lysates of endothelial cells. Interestingly, in normal cells, the higher expression of proteases, mainly uPA, measured as the level of protein concentration, was closely linked with their lower activity and inversely, in tumor cells, the low level of the expression of the enzymes correlated with their high enzymatic activity. In zymography analysis, mainly pro-MMPs were detected both in culture supernatants and cell lysates. The highest amounts of active forms of the MMPs were detected in tumor spheroids co-cultured with endothelial cells. Monensin inhibited MMPs and uPA secretion but significantly increased uPAR release, mainly from normal cells.In conclusion, during direct interactions of tumor cells with normal cells, MMPs and the uPA/uPAR system play an important role in the degradation of ECM and tumor development, but as we found, there is a reverse relationship between the concentration and the enzymatic activity of MMPs and uPA/uPAR in co-culture models.     

uPA系统与神经胶质瘤局部侵袭

uPA系统包括尿激酶型纤溶酶原激活物（urokinase plasminogen activator, uPA）、 尿激酶型纤溶酶原激活物受体（urokinase plasminogen activation receptor, uPAR）和纤溶酶原激活物抑制剂(plasminogen activator inhibitor, PAI),它们参与了多种人类恶性肿瘤的局部侵袭和转移,是目前肿瘤治疗重要的分子靶点。uPA能激活纤溶酶原降解细胞外基质与基底膜,uPAR则能显著提升uPA的激活纤溶酶原的功能,两者结合能够增强肿瘤的侵袭性与转移能力。PAI主要通过促进血管内皮生长因子（vascular endothelial growth factor,VEGF）的表达,促进新生血管的形成,从而促进肿瘤的局部侵袭,但PAI又可通过抑制uPA-uPAR复合体的生物学活性来抑制肿瘤的局部侵袭。神经胶质瘤是最常见的颅内肿瘤,很少转移到颅外,局部侵袭是其预后不良的主要原因。近年来发现uPA系统的表达水平与神经胶质瘤的恶性程度呈正相关。本文综述了uPA系统对神经胶质瘤局部侵袭的影响及其在治疗中的意义。