Abstract
Plasminogen activator inhibitor (PAI)‐1, a serine protease inhibitor, inactivates urokinase‐type plasminogen activator (uPA) and regulates degradation of the extracellular matrix; whether it functions for or against tumor progression, however, has been the subject of controversy. To assess the role of PAI‐1 in invasion and proliferation of hepatocellular carcinoma (HCC) cells, HLE cells were transfected with a vector capable of expressing an antisense PAI‐1 transcript. Analysis of seven stably transfected clones (PAI‐1−) showed reductions of 81% in PAI‐1 mRNA by northern blot analysis and 63% in the cellular PAI‐1 antigen level by enzyme‐linked immunosorbent assay(ELISA). There was no change in the levels of secreted PAI‐1 or PAI‐2. The activity of cellular uPA increased by 54%, without change in the protein level or the secreted uPA activity evaluated by ELISA. Morphologically, PAI‐1 antisense induced a spindle shape with narrower cytoplasmic processes in HLE cells. The forced inhibition of PAI‐1 increased the invasion and the growth of PAI‐1− cells by 75% and 82%, respectively. These results suggest that PAI‐1 plays a role in inhibiting invasion and proliferation, and the balance between uPA and PAI‐1 expression is important to assess the invasiveness of HCC cells.
Keywords: Hepatocellular carcinoma, Plasminogen activator inhibitor‐1, Antisense, Invasion, Proliferation
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REFERENCES
- 1).Nakajima , M. and Chop , A. M.Tumor invasion and extracellular matrix degradative enzyme: regulation of activity by organ factors . Semin. Cancer Biol. , 2 , 115 – 127 ( 1991. ). [PubMed] [Google Scholar]
- 2).Tryggvason , K. , Hoyhtya , M. and Salo , T.Proteolytic degradation of extracellular matrix in tumor invasion . Biochim. Biophys. Acta , 907 , 191 – 217 ( 1987. ). [DOI] [PubMed] [Google Scholar]
- 3).Liotta , L. A. , Steeg , P. S. and Stetler‐Stevenson , W. G.Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation . Cell , 64 , 327 – 336 ( 1991. ). [DOI] [PubMed] [Google Scholar]
- 4).Danø , K. , Andreasen , P. A. , Grøndahl‐Hansen , J. , Kristensen , P. , Nielsen , L. S. and Skriver , L.Plasminogen activators, tissue degradation and cancer . Adv. Cancer Res. , 44 , 139 – 266 ( 1985. ). [DOI] [PubMed] [Google Scholar]
- 5).Mars , W. M. , Kim , T. H. , Stolz , D. B. , Liu , M. L. and Michalopoulos , G. K.Presence of urokinase in serum‐free primary rat hepatocyte cultures and its role in activating hepatocyte growth factor . Cancer Res. , 56 , 2837 – 2843 ( 1996. ). [PubMed] [Google Scholar]
- 6).Jeffers , M. , Rong , S. and Woude , G. F. V.Enhanced tumorigenicity and invasion‐metastasis by hepatocyte growth factor/scatter factor‐met signaling in human cells concomitant with induction of the urokinase proteolysis network . Mol. Cell. Biol. , 16 , 1115 – 1125 ( 1996. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7).Andreasen , P. A. , Georg , B. , Lund , L. R. , Riccio , A. and Stacey , S. N.Plasminogen activator inhibitors: hormonally regulated serpins . Mol. Cell. Endocrinol. , 68 , 1 – 19 ( 1990. ). [DOI] [PubMed] [Google Scholar]
- 8).Kruithof , E. K. O.Plasminogen activator inhibitors — are view . Enzyme , 40 , 113 – 121 ( 1988. ). [DOI] [PubMed] [Google Scholar]
- 9).Montogomery , A. M. P. , Declerck , Y. A. , Lamgley , K. E. , Reisfeld , R. A. and Mueller , B. M.Melanoma‐mediated dissolution of extracellular matrix: contribution of urokinase‐dependent and metalloproteinase‐dependent proteolytic pathways . Cancer Res. , 53 , 693 – 700 ( 1993. ). [PubMed] [Google Scholar]
- 10).Laug , W. E. , Cao , X. R. , Yu , Y. B. , Shimada , H. and Kryithof , E. K. O.Inhibition of invasion of HT1080 sarcoma cells expressing recombinant plasminogen activator inhibitor 2 . Cancer Res. , 53 , 6051 – 6057 ( 1993. ). [PubMed] [Google Scholar]
- 11).Grøndahl‐Hansen , J. , Christensen , I. J. , Rosenquist , C. , Brunner , N. , Mouridsen , H. T. , Danø , K. and Blichert‐Toft , M.High levels of urokinase‐type plasminogen activator and its inhibitor PAI‐1 in cytosolic extracts of breast carcinomas are associated with poor prognosis . Cancer Res. , 53 , 2513 – 2521 ( 1993. ). [PubMed] [Google Scholar]
- 12).Nekarda , H. , Schmitt , M. , Ulm , K. , Wenninger , A. , Vogelsang , H. , Becker , K. , Roder , J. D. , Fink , U. and Siewert , J. R.Prognostic impact of urokinase‐type plasminogen activator and its inhibitor PAI‐1 in completely resected gastric cancer . Cancer Res. , 54 , 2900 – 2907 ( 1994. ). [PubMed] [Google Scholar]
- 13).Pedersen , H. , Brunner , N. , Francis , D. , Osterlind , K. , Rønne , E. , Hansen , H. H. , Danø , K. and Grøndahl‐Hansen , J.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 ( 1994. ). [PubMed] [Google Scholar]
- 14).Deng , G. , Curriden , S. A. , Wang , S. , Rosenberg , S. and Loskutoff , D. J.Is plasminogen activator inhibitor‐1 the molecular switch that governs urokinase receptor‐mediated cell adhesion and release ? J. Cell Biol. , 134 , 1563 – 1571 ( 1996. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15).Waltz , D. A. , Natkin , L. R. , Fujita , R. M. , Wei , Y. and Chapman , H. A.Plasmin and plasminogen activator inhibitor type 1 promote cellular motility by regulating the interaction between the urokinase receptor and vitronectin . J. Clin. Invest. , 100 , 58 – 67 ( 1997. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16).Soff , G. A. , Sanderowitz , J. , Gately , S. , Verrusio , E. , Weiss , I. , Brem , S. and Kwaan , H. C.Expression of plasminogen activator inhibitor type1 by human prostate carcinoma cells inhibits primary tumor growth . J. Clin. Invest. , 96 , 2593 – 2600 ( 1995. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17).Alizadeh , H. , Ma , D. , Berman , M. , Bellinham , D. , Comerford , S. A. , Gething , M. J. H. , Sambrook , J. F. and Niederkorn , J. Y.Tissue‐type plasminogen activator induced invasion and metastasis of murine melanomas . Curr. Eye Res. , 14 , 449 – 458 ( 1995. ). [DOI] [PubMed] [Google Scholar]
- 18).Morita , Y. , Hayashi , Y. , Wang , Y. , Kanamaru , T. , Suzuki , S. , Kawasaki , K. , Ohta , K. , Yamamoto , M. , Saitoh , Y. , Itoh , H. and Doe , W. F.Expression of urokinase‐type plasminogen activator receptor in hepatocellular carcinoma . Hepatology , 25 , 856 – 861 ( 1997. ). [DOI] [PubMed] [Google Scholar]
- 19).Lee , F. , Mulligan , R. , Berg , P. and Ringold , G.Glucocorticoids regulate expression of dihydrofolate reductase cDNA in mouse mammary tumor virus chimaeric plasmids . Nature , 294 , 228 – 232 ( 1981. ). [DOI] [PubMed] [Google Scholar]
- 20).Cajot , J. F. , Bamat , J. , Bergonzelli , G. E. , Kruithof , E. K.O. , Medcalf , R. L. , Testuz , J. and Sordat , B.Plasminogen activator inhibitor type 1 is a potent natural inhibitor of extracellular matrix degradation by fibrosarcoma and colon carcinoma cell . Proc. Natl. Acad. Sci. USA , 87 , 6939 – 6943 ( 1990. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21).Ponton , A. , Coulombe , B. and Skuo , D.Decreased expression of tissue inhibitor of metalloproteinases in metastatic tumor cells leading to increased levels of collagenase activity . Cancer Res. , 51 , 2138 – 2143 ( 1991. ). [PubMed] [Google Scholar]
- 22).Pappot , H. , Gardsvoll , H. , Romer , J. , Pedersen , A. N. , Grøndahl‐Hansen , J. , Pyke , C. and Brunner , N.Plasminogen activator inhibitor type 1 in cancer: therapeutic and prognostic implications . Biol. Chem. Hoppe Seyler , 376 , 259 – 267 ( 1995. ). [DOI] [PubMed] [Google Scholar]
- 23).Mimuro , J. , Schleef , R. and Loskutoff , D. J.Extracellular matrix of cultured bovine aortic endothelial cells contains functionally active type 1 plasminogen activator inhibitor . Blood , 70 , 721 – 728 ( 1987. ). [PubMed] [Google Scholar]
- 24).Jankun , J. , Keck , R. W. , Skrzypczak‐Jankun , E. and Swiercz , R.Inhibitors of urokinase reduce size of prostate cancer xenografts in severe combined immunodeficient mice . Cancer Res. , 57 , 559 – 563 ( 1997. ). [PubMed] [Google Scholar]
- 25).Billstrom , A. , Hartley‐Asp , B. , Lancander , I. , Batra , S. and Astedt , B.The urokinase inhibitor p‐aminobenzamidine inhibits growth of a human prostate tumor in SCID mice . Int. J. Cancer , 61 , 542 – 547 ( 1995. ). [DOI] [PubMed] [Google Scholar]
- 26).Rifkin , D. B. , Gleizes , P. E. , Harpel , J. , Nunes , I. , Mungen , J. , Mazzieri , R. and Noguera , I.Plasminogen/plasminogen activator and growth factor activation . Ciba Found Symp. , 212 , 105 – 115 ( 1997. ). [DOI] [PubMed] [Google Scholar]