ARTICLE |
Correspondence to: Helene Solberg, The Finsen Laboratory, Rigshospitalet, Strandboulevarden 49, DK-2100 Copenhagen Ø, Denmark. E-mail: Helene@finsenlab.dk
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Summary |
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uPAR is a cellular receptor for urokinase plasminogen activator, an enzyme involved in extracellular matrix degradation during processes involving tissue remodeling. We have expressed a recombinant soluble form of murine uPAR and raised rabbit polyclonal antibodies to study the expression of uPAR by immunohistochemistry. The immunohistochemical localization of uPAR was determined in normal mouse organs and in tumors formed by the highly metastatic Lewis lung carcinoma. uPAR immunoreactivity was found in the lungs, kidneys, and spleen, and in endothelial cells in the uterus, urinary bladder, thymus, heart, liver, and testis. No uPAR immunoreactivity was detected in muscle. In general, strong uPAR immunoreactivity was observed in organs undergoing extensive tissue remodeling, as exemplified by trophoblast cells in placenta, and in migrating, but not resting, keratinocytes at the edge of incisional wounds. Staining was not detected in any tissue sections derived from uPAR-deficient mice, thus confirming the specificity of the immunohistochemical staining of uPAR in normal mouse tissues. In Lewis lung carcinoma, uPAR immunoreactivity was found in the tumor cells of the primary tumor and in lung metastases. (J Histochem Cytochem 49:237246, 2001)
Key Words: uPAR, immunohistochemistry, mouse, tissue remodeling, cancer
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Introduction |
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Plasmin is an extracellular serine protease capable of degrading proteins of the extracellular matrix (ECM) and basement membranes. It is synthesized as an inactive proenzyme, plasminogen, which can be converted to plasmin by two plasminogen activators, tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA) (
uPAR has been found on a variety of cell types in vivo, including monocytes and macrophages, granulocytes, keratinocytes, trophoblasts, myofibroblasts (Ploug et al; 1992,
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Materials and Methods |
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Animals and Tissues
Female BALB/c mice or uPAR gene-targeted mice of mixed 129/Black Swiss background, 68 weeks old, were anesthetized with 0.03 ml/10 g of a 1:1 mixture of Dormicum (midazolam 5 mg/ml) and Hypnorm (fluanison 5 mg/ml and fentanyl 0.1 mg/ml) and perfused with cold PBS. Organs for lysate preparations were immediately snap-frozen on dry ice and kept at -80C until used. If the tissue was collected for immunohistochemistry the perfusion of the mice with PBS was followed by perfusion with 4% paraformaldehyde in PBS. Organs were fixed overnight in 4% paraformaldehyde and paraffin embedded. Lewis lung carcinomas were obtained according to previously described methods (
Antibodies
The following antibodies were purchased from DAKO (Glostrup, Denmark): rabbit anti-von Willebrand Factor (A82), rabbit anti-keratin (Z622), biotinylated swine anti-rabbit (E431), peroxidase-conjugated swine anti-rabbit (P217), alkaline phosphatase-conjugated swine anti-rabbit (D306), and alkaline phosphatase-conjugated biotinavidin complexes (AP-ABC, K376). Rat monoclonal antibody to mouse macrophages, clone BM-8, was from BMA Biomedicals (Augst, Switzerland). Rat anti-mouse CD34 (clone MEC 14.7) was obtained from Hycult Biotechnology (Uden, Netherlands). The tyramide signal amplification (TSA) kit was purchased from NEN (Boston, MA).
Tissue Extracts
Tissue and cell extracts were prepared as described (
Chemical Crosslinking Assay and Western Blotting Analysis
Twenty µl detergent-phase extract or cell culture media from transfectants was incubated with either 100 nM mouse ATF (mouse ATF was obtained as described by
Cloning and Transfection
A fragment of the receptor for mouse urokinase plasminogen activator (smuPAR) cDNA sequence was prepared by standard PCR technique using muPAR cDNA as template (
Purification of smuPAR
Conditioned media from CHOsmuPAR cells grown in 10% fetal calf serum in -Minimum Essential Medium (MEM) (Gibco; Life Technologies, Roskilde, Denmark) was collected and 0.1 M Tris, pH 8.1, EDTA, and 1 mM PMSF was added before filtration through a 0.8-µm filter. smuPAR was purified by immunoaffinity chromatography using a monoclonal antibody raised against human uPAR in uPAR-deficient mice (KOR1). This antibody recognizes an epitope that is conserved between mouse and human uPAR (unpublished results). The immunoaffinity purification was followed by further purification by reverse-phase HPLC. The purified smuPAR was analysed by SDS-PAGE and the purity was checked by silver staining of the gel.
Specific Polyclonal Rabbit Anti-mouse uPAR Antibodies
Preimmune serum was collected from four individual rabbits. Rabbits were injected SC with 20 µg smuPAR in Freund's complete adjuvant in PBS on Day 0, followed by SC injection of 20 µg smuPAR in Freund's incomplete adjuvant in PBS on Day 18 and every 28 days thereafter. Blood was drawn on Day 28 and every 28 days thereafter. The serum was checked for antibodies specific for mouse uPAR by ELISA. Briefly, ELISA plates were coated overnight with 10 ng/well of purified soluble mouse uPAR. After blocking, the diluted rabbit serum was added to the plates. Bound IgG was detected using peroxidase-coupled swine anti-rabbit antibodies. Color was developed using O-phenylenediamine (OPD) and the optical density measured at 490 nm. The rabbit anti smuPAR IgG was purified using protein GSepharose and the concentration of purified IgG was determined using a protein assay from BioRad (Copenhagen, Denmark).
Immunohistochemistry
Paraffin sections were deparaffinized in xylene, digested with 0.03% trypsin for 10 min at 37C, blocked in 5% swine serum in TBS with 0.25% bovine serum albumin (BSA) (TBSBSA), and incubated with 10 µg/ml polyclonal rabbit IgG in TBSBSA raised against soluble mouse uPAR. Then biotinylated swine anti-rabbit was applied, followed by APABC according to the manufacturer's instructions. Slides were developed for 1030 min. with Fast Red and counterstained with Mayer's hematoxylin. For signal amplification with biotinyl tyramide (
As negative controls, we used preimmune rabbit IgG, rabbit anti-mouse uPAR IgG preincubated with 10-fold molar excess of smuPAR, and tissues from uPAR-deficient mice.
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Results |
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Organ distribution of mouse uPAR
To determine the level of uPAR expression in mice under normal physiological conditions, extracts were prepared from various tissues of BALB/C mice. Detergent-phase extracts were analyzed by a chemical crosslinking method based on the ability of EDC/NHS to covalently crosslink 125I-labeled human ATF to mouse uPAR (
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Generation of Polyclonal Rabbit Antibodies to Mouse uPAR
To study the histological localization of uPAR in the mouse, polyclonal antibodies against mouse uPAR were generated by immunization of rabbits using recombinant mouse uPAR as antigen. Soluble mouse uPAR was produced by transfection of CHO cells with a construct encoding a soluble form of mouse uPAR. Purification of smuPAR from conditioned media was done by immunoaffinity chromatography, using a monoclonal antibody (KOR 1) that binds to an epitope conserved between mouse and human uPAR. The immunoaffinity purification was followed by a further purification by reverse-phase HPLC. The purity and electrophoretic mobility of the mouse uPAR antigen were monitored by SDS-PAGE and silver staining (Fig 2, Lane 1). To test whether the recombinant smuPAR was folded correctly, its ligand-binding properties were examined by surface plasmon resonance using an optical biosensor (Biacore 2000), as described previously for the human protein (
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The purified smuPAR preparation was then used as antigen for immunization of four rabbits and serum was tested for specific IgG by ELISA. Specificity of the purified IgG from these rabbits was subsequently examined by Western blotting analysis of detergent phase extracts of Lewis lung carcinoma (Fig 2, Lane 2), spleen (Fig 2, Lanes 3 and 4), and lung (Fig 2, Lanes 5 and 6) from wild-type (Fig 2, Lanes 3 and 5) or uPAR-deficient (Fig 2, Lanes 4 and 6) mice. A single band corresponding to the migration of purified smuPAR was seen in extacts from Lewis lung carcinoma and wild-type spleen and lung, whereas no bands were observed in extracts prepared from uPAR-deficient mice. No bands were obtained when preimmune IgG from the same rabbit was used (data not shown).
Histological Localization of uPAR in Organs From Healthy Mice
Immunohistochemistry was used to map the histological localization of uPAR protein under normal and pathological conditions in the mouse. For all organs examined, tissues from at least two animals were investigated, giving similar results. A tissue section from each organ was stained with polyclonal rabbit antibodies against cytokeratin to ensure that proteins were intact after fixation and embedding of the tissue, as judged by positive staining of keratin in epithelial cells. Equivalent tissue from mice rendered uPAR-deficient by homologous recombination (
Immunohistochemistry of lung tissue showed uPAR staining in the alveoli and pulmonary vessels, whereas the bronchial epithelium was negative (Fig 3A). No staining was seen with either preimmune rabbit IgG on wild-type lung tissue (Fig 3B) or the polyclonal immune IgG on equivalent lung tissue obtained from uPAR-deficient mice (Fig 3C). By evaluation of adjacent sections of lung tissue stained with antibodies against von Willebrand factor, cytokeratin, and macrophage clone BM-8, it was evident that endothelial cells of the alveoli express uPAR immunoreactivity, whereas we were unable to clearly identify immunostaining of pneumocytes and alveolar macrophages (data not shown).
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In the kidney, the highest intensity of uPAR immunoreactivity was observed in glomeruli and in vessels in the medulla. Vessels in the cortex showed weaker uPAR staining (Fig 3D). Immunohistochemical staining with an antibody against CD34, an endothelial cell marker, showed a staining pattern indistinguishable from that obtained with uPAR-specific antibodies, suggesting that all endothelial cells of the kidney express uPAR. No uPAR immunoreactivity was seen in kidneys from uPAR-deficient mice (Fig 3E).
In the spleen, uPAR immunoreactivity was seen in the perifollicular zone, an area that contains a high number of leukocytes with a cell type composition similar to that found in peripheral blood (Fig 3F). In the liver, uPAR immunoreactivity was detected only in the arteries and arterioles. No other cells were positive, including veins, sinosoids, and Kuppfer cells (data not shown). In the heart, uPAR was detected in the endothelial cells of arteries, whereas capillaries showed little or no immunoreactivity. CD34 immunostaining demonstrated the presence of multiple capillaries that were generally devoid of uPAR immunoreactivity (data not shown).
As a common denominator, pronounced uPAR immunoreactivity was consistently observed in normal mouse organs undergoing a defined tissue remodeling process. First, sections of 16-hr-old mouse skin wounds showed strong uPAR reactivity in the moving keratinocytes at the edge of the wound, whereas the non-migrating keratinocytes were negative. uPAR immunoreactivity was also seen in granulocytes infiltrating the area just below the wound crush. Endothelial cells in the wound area and the nearby normal skin stained positive for uPAR protein (Fig 4A). Second, during late pregnancy, spongiotrophoblast cells in the placenta showed distinct uPAR immunoreactivity localized to the plasma membrane. Endothelial cells in the placenta stained positive for uPAR (Fig 4B), as did the decidual cells bordering the uterine epithelium (data not shown). Third, in post-lactational involuting mammary glands, weak to moderate staining could be detected in the regressing glandular tissue (Fig 4C). Strong uPAR immunoreactivity was also seen in granulocytes localized in the lymph nodes of the mammary gland and in cells, probably macrophages, present in the subcapsular sinuses of the lymph nodes (data not shown). For all three tissues undergoing tissue remodeling, no uPAR immunoreactivity was seen when equivalent tissues obtained from uPAR-deficient mice were stained with the same rabbit anti-mouse uPAR IgG (Fig 4D4F).
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Lewis Lung Carcinoma
Primary tumors of Lewis lung carcinoma showed clear surface-localized uPAR immunoreactivity (Fig 5A). There was pronounced heterogeneity in the staining of parts of individual tumors: some parts stained strongly, whereas other parts were completely devoid of uPAR immunoreactivity. In addition, vessels and neutrophils infiltrating the necrotic foci of the tumor showed uPAR immunoreactivity (Fig 5A). Metastases in the lung also showed uPAR immunoreactivity (data not shown). As control for the staining specificity, rabbit anti-mouse uPAR IgG was incubated with excess antigen before immunostaining. After antigen absorption of the antibody (Fig 5B) and with preimmune IgG (data not shown), no staining of the tumor was seen.
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Discussion |
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uPAR has been implicated in ECM degradation under normal physiological processes that involve tissue remodeling and during cancer cell invasion and metastasis. Mice rendered uPAR-deficient by gene targeting (
Immunoreactivity for uPAR was detected throughout the alveoli of the lungs. This finding is consistent with the relatively high level of uPAR protein detected in extracts of lung tissue by chemical crosslinking. Because of the rich capillary network, it was difficult to rigorously conclude that pneumocytes and macrophages are actually responsible for some of the uPAR staining. Immunohistochemical double labeling is required to clarify a possible contribution from such cells. In situ hybridizations on lung tissue have thus far detected uPAR mRNA only in a few scattered cells bordering the alveolar space, possibly representing alveolar macrophages (
Crosslinking analysis revealed appreciable amounts of functional uPAR protein in the kidneys, an organ that has previously been shown to contain high amounts of uPA (
Peripheral blood leukocytes express uPAR, and it has previously been reported that the cell surface expression of uPAR rapidly increases due to a translocation of a certain subset of granules after leukocyte activation (
The confinement of uPAR immunoreactivity to the migrating keratinocytes of incisional skin wounds is in complete accordance with in situ hybridization for uPAR mRNA (
Involuting mammary glands reveal uPAR immunoreactivity in the regressing glandular tissue, which also expresses uPA (
During late pregnancy, uPAR immunoreactivity was localized to spongiotrophoblast cells of the placenta, as well as in the decidual cells bordering the uterine epithelium. The decidual cells have previously been shown also to possess uPA reactivity (
uPAR and uPA have previously been detected only in activated endothelial cells in vivo during neovascularization or inflammation (
The highly metastatic Lewis lung carcinoma showed distinct uPAR staining localized to the surface of the tumor cells in both primary tumor and metastases of the lung. However, the staining in the primary tumor was very heterogeneous, showing some areas with pronounced uPAR immunoreactivity whereas others were totally devoid of staining. Although all Lewis lung cells in culture probably express uPAR, the lack of expression in some of the tumor cells in vivo is probably not the result of downregulation. Rather, the expression of uPAR at the invasive front is likely the result of upregulation of uPAR induced by potential signal(s) from the stroma. By analogy, only cancer cells located at the invasive front in human colon cancer express uPAR, whereas many colon cancer cell lines uniformly express uPAR in vitro (
The generation of uPAR-deficient mice (
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Acknowledgments |
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Supported by the Danish Cancer Society and the Danish Research Council.
We thank Dr Thomas Bugge for the uPAR-deficient mice. We are grateful to Anette Bartels, Pia Gottrup Knudsen, and Helle Malmstedt for excellent technical assistance.
Received for publication September 22, 2000; accepted October 4, 2000.
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Literature Cited |
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![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
AlmusJacobs F, Varki N, Sawdey MS, Loskutoff DJ (1995) Endotoxin stimulates expression of the murine urokinase receptor gene in vivo. Am J Pathol 147:688-698[Abstract]
Andreasen PA, Kjøller L, Christensen L, Duffy MJ (1997) The urokinase-type plasminogen activator system in cancer metastasis: a review. Int J Cancer 72:1-22[Medline]
Appella E, Robinson EA, Ullrich SJ, Stoppelli MP, Corti A, Cassani G, Blasi F (1987) The receptor-binding site of urokinase. J Biol Chem 261:4437-4440
Bacharach E, Itin A, Keshet E (1992) In vivo patterns of expression of urokinase and its inhibitor PAI-1 suggest a concerted role in regulating physiological angiogenesis. Proc Natl Acad Sci USA 89:10686-10690[Abstract]
Behrendt N, Ploug M, Patthy L, Houen G, Blasi F, Danø K (1991) The ligand binding domain of the cell surface receptor for urokinase-type plasminogen activator. J Biol Chem 266:7842-7847
Behrendt N, Rønne E, Ploug M, Petri T, Løber D, Nielsen LS, Schleuning W-D, Blasi F, Appella E, Danø K (1990) The human receptor for urokinase plasminogen activator. NH2-terminal amino acid sequence and glycosylation variants. J Biol Chem 265:6453-6460
Bobrow MN, Harris TD, Shaughnessy KJ, Litt GJ (1989) Catalyzed reporter deposition, a novel method of signal amplification. Application to immunoassays. J Immunol Methods 125:279-285[Medline]
Bohuslav J, Horejsi V, Hansmann C, Stöckl J, Weidle UH, Majdic O, Bartke I, Knapp W, Stockinger H (1995) Urokinase plasminogen activator receptor, ß2-integrins, and src-kinases within a single receptor complex of human monocytes. J Exp Med 181:1381-1390[Abstract]
Bugge T, Suh TT, Flick MJ, Daugherty CC, Rømer J, Solberg H, Ellis V, Danø K, Degen JL (1995) The receptor for urokinase-type plasminogen activator is not essential for mouse development or fertility. J Biol Chem 270:16886-16894
Crowley CW, Cohen RL, Lucas BK, Liu G, Shumn MA, Levinson AD (1993) Prevention of metastasis by inhibition of the urokinase receptor. Proc Natl Acad Sci USA 90:5021-5025[Abstract]
Danø K, Behrendt N, Brünner N, Ellis V, Ploug M, Pyke C (1994) The urokinase receptor: protein structure and role in plasminogen activation and cancer invasion. Fibrinolysis 8:189-203
Dubuisson L, Monvoisin A, Nielsen BS, Le Bail B, Bioculac-Sage P, Rosenbaum J (2000) Expression and cellular localization of the urokinase-type plasminogen activator and its receptor in hepatocellular carcinomas. J Pathol 190:190-195[Medline]
Ellis V, Ploug M, Plesner T, Danø K (1996) Gene expression and function of the cellular receptor for u-PA (uPAR). In GlassGreenwalt P, ed. Fibrinolysis in Disease: Molecular and Hemovascular Aspects of Fibrinolysis. Boca Raton, FL, CRC Press, 30-42
Ellis V, Scully MF, Kakkar VV (1989) Plasminogen activation initiated by single-chain urokinase-type plasminogen activator. Potentiation by U937 monocytes. J Biol Chem 264:2185-2188
Estreicher A, Wohlwend A, Belin D, Schleuning WD, Vassalli J-D (1989) Characterization of the cellular binding site for the urokinase-type plasminogen activator. J Biol Chem 264:1180-1189
Fazioli F, Resnati M, Sidenius N, Higashimoto Y, Apella E, Blasi F (1997) A urokinase-sensitive region of the human urokinase receptor is responsible for its chemotactic activity. EMBO J 16:7279-7286
Ganesh S, Sier CF, Heerding MM, Griffioen G, Lamers CB, Verspaget HW (1994) Urokinase receptor and colorectal cancer survival. Lancet 344:401-402. [letter][Medline]
Gårdsvoll H, Danø K, Ploug M (1999) Mapping part of the functional epitope for ligand binding on the receptor for urokinase-type plasminogen activator by site-directed mutagenesis. J Biol Chem 274:37995-38003
Grøndahl-Hansen J, Peters HA, van Putten WLJ, Look MP, Pappot H, Rønne E, Danø K, Kllijn JG, Brünner N, Foekens JA (1995) Prognostic significance of the receptor for urokinase plasminogen activator in breast cancer. Clin Cancer Res 1:1079-1087[Abstract]
Høyer-Hansen G, Rønne E, Solberg H, Behrendt N, Ploug M, Lund LR, Ellis V, Danø K (1992) Urokinase plasminogen activator cleaves its cellular receptor releasing the ligand-binding domain. J Biol Chem 267:18224-18229
Kim J, Kovalski YK, Ossowski L (1998) Requirement for specific proteases in cancer cell intravasation as revealed by a novel semiquantitative PCR-based assay. Cell 94:353-362[Medline]
Kook YH, Adamski J, Zelent A, Ossowski L (1994) The effect of antisense inhibition of urokinase receptor in human squamous cell carcinoma on malignancy. EMBO J 13:3983-3991[Abstract]
Kristensen P, Eriksen J, Blasi F, Danø K (1991a) Two alternatively spliced mouse urokinase receptor mRNAs with different histological localization in the gastrointestinal tract. J Cell Biol 115:1763-1771[Abstract]
Kristensen P, Eriksen J, Danø K (1991b) Localization of urokinase-type plasminogen activator messenger RNA in the normal mouse by in situ hybridization. J Histochem Cytochem 39:341-349[Abstract]
Larsson LI, Skriver L, Nielsen LS, Grøndahl-Hansen J, Kristensen P, Danø K (1984) Distribution of urokinase-type plasminogen activator immunoreactivity in the mouse. J Cell Biol 98:894-903[Abstract]
Lund LR, Bjørn SF, Sternlicht MD, Nielsen BS, Solberg H, Usher PA, Østerby R, Christensen IJ, Stephens RW, Bugge TH, Danø K, Werb Z (2000) Lactational competance and involution of the mammary gland require plasminogen. Development 127:4481-4492
May AE, Kanse SM, Lund LR, Gisler RH, Imhof BA, Preissner KT (1998) Urokinase receptor (CD87) regulates leukocyte recruitment via ß2 integrins in vivo. J Exp Med 188:1029-1037
Min HY, Doyle LV, Vitt CR, Zandonnella CL, StrattonThomas JR, Shuman MA, Rosenberg S (1996) Urokinase receptor antagonists inhibit angiogenesis and primary tumor growth in syngenic mice. Cancer Res 56:2428-2433[Abstract]
Nielsen LS, Kellerman GM, Behrendt N, Picone R, Danø K, Blasi F (1988) A 55,000-60,000 receptor protein for urokinase-type plasminogen activator: identification in human tumor cell lines and partial purification. J Biol Chem 263:2358-2363
Nykjær A, Petersen CM, Christensen EI, Davidsen O, Gliemann J (1990) Urokinase receptors in human monocytes. Biochim Biophys Acta 1052:399-407[Medline]
Nykjær A, Petersen CM, Møller B, Andreasen P, Gliemann J (1992) Identification and characterization of urokinase receptors in natural killer cells and T-cells-derived lymphokine activated killer cells. FEBS Lett 300:13-17[Medline]
Pedersen H, Brünner N, Francis D, Østerlind K, Rønne E, Hansen HH, Danø K, GrøndahlHansen J (1994) Prognostic impact of urokinase, urokinase receptor, and type 1 plasminogen activator inhibitor in squamous and large cell lung cancer tissue. Cancer Res 54:4671-4675[Abstract]
Pepper MS, Sappino A-P, Stöcklin R, Montesano R, Orci L, Vassalli J-D (1993) Upregulation of urokinase receptor expression on migrating endothelial cells. J Cell Biol 122:673-684[Abstract]
Pierleoni C, Samuelsen GB, Græm N, Rønne E, Nielsen BS, Kaufmann P, Castellucci M (1998) Immunohistochemical identification of the receptor for urokinase plasminogen activator associated with fibrin deposition in normal and ectopic human placenta. Placenta 19:501-508[Medline]
Plesner T, Ploug M, Ellis V, Rønne E, HøyerHansen G, Wittrup M, Pedersen TL, Tscherning T, Danø K, Hansen NE (1994a) The receptor for urokinase-type plasminogen activator and urokinase is translocated from two distinct intracellular compartments to the plasma membrane on stimulation of human neutrophils. Blood 83:808-815
Plesner T, Ralfkiær E, Wittrup M, Johnsen H, Pyke C, Pedersen T, Hansen NE, Danø K (1994b) Expression of the receptor for urokinase-type plasminogen activator in normal and neoplastic blood cells and hematopoietic tissue. Am J Clin Pathol 102:835-841[Medline]
Ploug M (1998) Identification of specific sites involved in ligand binding by photoaffinity labeling of the receptor for urokinase plasminogen activator. Residues located at equivalent positions in uPAR domains I and III participate in the assembly of a composite ligand-binding site. Biochemistry 37:16494-16505[Medline]
Ploug M, Ellis V, Danø K (1994) Ligand interaction between urokinase-type plasminogen activator and its receptor probed with 8-anilino-1-naphthalenesulfonate. Evidence for a hydrophobic binding site exposed only in the intact receptor. Biochemistry 30:8991-8997
Ploug M, Plesner T, Rønne E, Ellis V, HøyerHansen G, Hansen NE, Danø K (1992) The receptor for urokinase-type plasminogen activator is deficient on peripheral blood leukocytes in patients with paroxysmal nocturnal hemoglobinuria. Blood 79:1447-1455[Abstract]
Ploug M, Rønne E, Beherendt N, Jensen AL, Blasi F, Danø K (1991) Cellular receptor for urokinase plasminogen activator: carboxyl-terminal processing and membrane anchoring by glycosyl-phosphatidylinositol. J Biol Chem 266:1926-1933
Pyke C, Kristensen P, Ralfkiær E, GrøndahlHansen J, Eriksen J, Blasi F, Danø K (1991) Urokinase-type plasminogen activator is expressed in stromal cells and its receptor in cancer cells at invasive foci in human colon adenocarcinomas. Am J Pathol 138:1059-1067[Abstract]
Pyke C, Ralfkiær E, Rønne E, Høyer-Hansen G, Kirkeby L, Danø K (1994) Immunohistochemical detection of the receptor for urokinase plasminogen activator in human colon cancer. Histopathology 24:131-138[Medline]
Resnati M, Guttinger M, Valcamonica S, Sidenius N, Blasi F, Fazioli F (1996) Proteolytic cleavage of the urokinase receptor substitutes for the agonist-induced chemotactic effect. EMBO J 15:1572-1582[Abstract]
Roldan AL, Cubellis MV, Mascuri MT, Behrendt N, Lund LR, Danø K, Apella E, Blasi F (1990) Cloning and expression of the receptor for human urokinase-plasminogen activator, a central molecule in cell-surface plasmin-dependent proteolysis. EMBO J 9:467-474[Abstract]
Rømer J, Lund LR, Eriksen J, Ralfkiær E, Zeheb R, Gelehrter TD, Danø K, Kristensen P (1991) Differential expression of the receptor of urokinase-type plasminogen activator and its type-1 inhibitor during healing of mouse skin wounds. J Invest Dermatol 97:803-811[Abstract]
Rømer J, Pyke C, Lund LR, Eriksen J, Kristensen P, Rønne E, HøyerHansen G, Danø K, Brünner N (1994) Expression of uPA and receptor by both neoplastic and stromal cells during xenograft invasion. Int J Cancer 57:553-560[Medline]
Skriver L, Larsson LI, Kielberg V, Nielsen LS, Andresen PB, Kristensen P, Danø K (1984) Immunocytochemical localization of urokinase-type plasminogen activator in Lewis lung carcinoma. J Cell Biol 99:753-758
Solberg H, Løber D, Eriksen J, Ploug M, Rønne E, Behrendt N, Danø K, HøyerHansen G (1992) Identification and characterization of the murine cell surface receptor for the urokinase type plasminogen activator. Eur J Biochem 205:451-458[Abstract]
Solberg H, Rømer J, Brünner N, Holm A, Sidenius N, Danø K, Høyer-Hansen G (1994) A cleaved form of the receptor for urokinase-type plasminogen activator in invasive transplanted human and murine tumors. Int J Cancer 58:877-881[Medline]
Teesalu T, Blasi F, Talarico D (1998) Expression and function of the urokinase-type plasminogen activator during mouse hemochorial placental development. Dev Dyn 213:27-38[Medline]
Tressler RJ, Pitot PA, StrattonThomas JR, Forrest LD, Zhuo S, Drummond RJ, Fong S, Doyle MV, Doyle LV, Min HY, Rosenberg S (1999) Urokinase receptor antagonists discovery and application to in vivo models of tumor growth. APMIS 107:168-173[Medline]
Xue W, Kindzelskii AL, Todd RF, Petty HR (1994) Physical association of complement receptor type 3 and urokinase-type plasminogen activator receptor in neutrophil membranes. J Immunol 152:4630-4640