Angiogenesis

, Volume 10, Issue 3, pp 197–216 | Cite as

Transcriptome analysis of endothelial cell gene expression induced by growth on matrigel matrices: identification and characterization of MAGP-2 and lumican as novel regulators of angiogenesis

  • Allan R. Albig
  • Thessa G. Roy
  • Darryl J. Becenti
  • William P. Schiemann
Original Paper

Abstract

Remodeling of vascular microenvironments during normal and tumor-induced angiogenesis is an important, yet poorly understood mechanism by which endothelial cells (ECs) contribute to the activation or resolution of angiogenesis. We used microarray analyses to monitor changes in the transcriptome of ECs undergoing angiogenesis when cultured onto Matrigel matrices. This strategy identified 308 genes whose expression in ECs was altered at least 3-fold by angiogenesis, of which 63 genes were found to encode for secretory proteins. In vitro assays that modeled key steps in the angiogenic process showed that several identified genes possessed pro- or anti-angiogenic activities (e.g., SMOC-2, secreted modular calcium-binding protein-2; CRELD-2, cysteine-rich with EGF-like domains-1; MAGP-2, microfibril-associated glycoprotein-2; lumican; and ECM-1, extracellular matrix protein-1). In particular, MAGP-2 expression potentiated EC proliferation and p38 MAPK activation stimulated by the pro-angiogenic factors, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF); it also stimulated EC invasion and angiogenic sprouting, and more importantly, promoted the development and infiltration of vessels into Matrigel plugs implanted into genetically normal mice. Conversely, lumican inhibited EC activation of p38 MAPK, as well as their invasion, angiogenic sprouting, and vessel formation in mice. Collectively, our findings provide new insights into how EC stromal remodeling regulates angiogenesis activation and resolution, as well as identify two novel EC-secreted stromal proteins that modulate angiogenesis both in vitro and in vivo.

Keywords

Angiogenesis Lumican MAGP-2 Microarray analyses Microenvironment remodeling Stroma 

Notes

Acknowledgements

Members of the Schiemann Laboratory are thanked for critical reading of the manuscript. This research was supported in part by grants from the National Institutes of Health (CA095519) and the Cancer League of Colorado to W.P.S., and by a fellowship from the National Institutes of Health (CA99321) to A.R.A.

Supplementary material

References

  1. 1.
    Folkman J, Shing Y (1992) Angiogenesis. J Biol Chem 267:10931–10934PubMedGoogle Scholar
  2. 2.
    Carmeliet P, Jain RK (2000) Angiogenesis in cancer and other diseases. Nature 407:249–257CrossRefPubMedGoogle Scholar
  3. 3.
    Bergers G, Benjamin LE (2003) Tumorigenesis and the angiogenic switch. Nat Rev Cancer 3:401–410CrossRefPubMedGoogle Scholar
  4. 4.
    Hanahan D, Folkman J (1996) Patterns and emerging mechanisms of the angiogenic switch during tumorigenesis. Cell 86:353–364CrossRefPubMedGoogle Scholar
  5. 5.
    Stupack DG, Cheresh DA (2002) ECM remodeling regulates angiogenesis: endothelial integrins look for new ligands. Sci STKE 2002:PE7PubMedCrossRefGoogle Scholar
  6. 6.
    Liotta LA, Kohn EC (2001) The microenvironment of the tumor–host interface. Nature 411:375–379CrossRefPubMedGoogle Scholar
  7. 7.
    Joyce JA (2005) Therapeutic targeting of the tumor microenvironment. Cancer Cell 7:513–520CrossRefPubMedGoogle Scholar
  8. 8.
    Heissig B, Hattori K, Friedrich M, Rafii S, Werb Z (2003) Angiogenesis: vascular remodeling of the extracellular matrix involves metalloproteinases. Curr Opin Hematol 10:136–141CrossRefPubMedGoogle Scholar
  9. 9.
    Albig AR, Neil JR, Schiemann WP (2006) Fibulins 3 and 5 antagonize tumor angiogenesis in vivo. Cancer Res 66:2621–2629CrossRefPubMedGoogle Scholar
  10. 10.
    Albig AR, Schiemann WP (2004) Fibulin-5 antagonizes vascular endothelial growth factor (VEGF) signaling and angiogenic sprouting by endothelial cells. DNA Cell Biol 23:367–379CrossRefPubMedGoogle Scholar
  11. 11.
    Albig AR, Schiemann WP (2005) Identification and characterization of regulator of G protein signaling 4 (RGS4) as a novel inhibitor of tubulogenesis: RGS4 inhibits mitogen-activated protein kinases and vascular endothelial growth factor signaling. Mol Biol Cell 16:609–625CrossRefPubMedGoogle Scholar
  12. 12.
    Schenk PM, Baumann S, Mattes R, Steinbiss HH (1995) Improved high-level expression system for eukaryotic genes in Escherichia coli using T7 RNA polymerase and rare ArgtRNAs. Biotechniques 19:196–198, 200Google Scholar
  13. 13.
    Iruela-Arispe ML, Carpizo D, Luque A (2003) ADAMTS1: a matrix metalloprotease with angioinhibitory properties. Ann NY Acad Sci 995:183–190PubMedGoogle Scholar
  14. 14.
    Brigstock DR (2002) Regulation of angiogenesis and endothelial cell function by connective tissue growth factor (CTGF) and cysteine-rich 61 (CYR61). Angiogenesis 5:153–165CrossRefPubMedGoogle Scholar
  15. 15.
    Gao CF, Vande Woude GF (2005) HGF/SF-Met signaling in tumor progression. Cell Res 15:49–51CrossRefPubMedGoogle Scholar
  16. 16.
    Armstrong LC, Bornstein P (2003) Thrombospondins 1 and 2 function as inhibitors of angiogenesis. Matrix Biol 22:63–71CrossRefPubMedGoogle Scholar
  17. 17.
    Qi JH, Ebrahem Q, Moore N, Murphy G, Claesson-Welsh L, Bond M, Baker A, Anand-Apte B (2003) A novel function for tissue inhibitor of metalloproteinases-3 (TIMP3): inhibition of angiogenesis by blockage of VEGF binding to VEGF receptor-2. Nat Med 9:407–415CrossRefPubMedGoogle Scholar
  18. 18.
    Carmeliet P (2000) Mechanisms of angiogenesis and arteriogenesis. Nat Med 6:389–395CrossRefPubMedGoogle Scholar
  19. 19.
    Sottile J (2004) Regulation of angiogenesis by extracellular matrix. Biochim Biophys Acta 1654:13–22PubMedGoogle Scholar
  20. 20.
    Pupa SM, Menard S, Forti S, Tagliabue E (2002) New insights into the role of extracellular matrix during tumor onset and progression. J Cell Physiol 192:259–267CrossRefPubMedGoogle Scholar
  21. 21.
    Bissell MJ, Radisky DC, Rizki A, Weaver VM, Petersen OW (2002) The organizing principle: microenvironmental influences in the normal and malignant breast. Differentiation 70:537–546CrossRefPubMedGoogle Scholar
  22. 22.
    Davis GE, Senger DR (2005) Endothelial extracellular matrix: biosynthesis, remodeling, and functions during vascular morphogenesis and neovessel stabilization. Circ Res 97:1093–1107CrossRefPubMedGoogle Scholar
  23. 23.
    Kahn J, Mehraban F, Ingle G, Xin X, Bryant JE, Vehar G, Schoenfeld J, Grimaldi CJ, Peale F, Draksharapu A, Lewin DA, Gerritsen ME (2000) Gene expression profiling in an in vitro model of angiogenesis. Am J Pathol 156:1887–1900PubMedGoogle Scholar
  24. 24.
    Bell SE, Mavila A, Salazar R, Bayless KJ, Kanagala S, Maxwell SA, Davis GE (2001) Differential gene expression during capillary morphogenesis in 3D collagen matrices: regulated expression of genes involved in basement membrane matrix assembly, cell cycle progression, cellular differentiation and G-protein signaling. J Cell Sci 114:2755–2773PubMedGoogle Scholar
  25. 25.
    Aitkenhead M, Wang SJ, Nakatsu MN, Mestas J, Heard C, Hughes CC (2002) Identification of endothelial cell genes expressed in an in vitro model of angiogenesis: induction of ESM-1, βig-h3, and NrCAM. Microvasc Res 63:159–171CrossRefPubMedGoogle Scholar
  26. 26.
    Sulochana KN, Fan H, Jois S, Subramanian V, Sun F, Kini RM, Ge R (2005) Peptides derived from human decorin leucine-rich repeat 5 inhibit angiogenesis. J Biol Chem 280:27935–27948CrossRefPubMedGoogle Scholar
  27. 27.
    Davies Cde L, Melder RJ, Munn LL, Mouta-Carreira C, Jain RK, Boucher Y (2001) Decorin inhibits endothelial migration and tube-like structure formation: role of thrombospondin-1. Microvasc Res 62:26–42CrossRefPubMedGoogle Scholar
  28. 28.
    Kao WW, Funderburgh JL, Xia Y, Liu CY, Conrad GW (2006) Focus on molecules: lumican. Exp Eye Res 82:3–4CrossRefPubMedGoogle Scholar
  29. 29.
    Chakravarti S, Magnuson T, Lass JH, Jepsen KJ, LaMantia C, Carroll H (1998) Lumican regulates collagen fibril assembly: skin fragility and corneal opacity in the absence of lumican. J Cell Biol 141:1277–1286CrossRefPubMedGoogle Scholar
  30. 30.
    Vij N, Roberts L, Joyce S, Chakravarti S (2004) Lumican suppresses cell proliferation and aids Fas-Fas ligand mediated apoptosis: implications in the cornea. Exp Eye Res 78:957–971CrossRefPubMedGoogle Scholar
  31. 31.
    Vij N, Roberts L, Joyce S, Chakravarti S (2005) Lumican regulates corneal inflammatory responses by modulating Fas-Fas ligand signaling. Invest Ophthalmol Vis Sci 46:88–95CrossRefPubMedGoogle Scholar
  32. 32.
    Ping Lu Y, Ishiwata T, Asano G (2002) Lumican expression in alpha cells of islets in pancreas and pancreatic cancer cells. J Pathol 196:324–330CrossRefPubMedGoogle Scholar
  33. 33.
    Leygue E, Snell L, Dotzlaw H, Hole K, Hiller-Hitchcock T, Roughley PJ, Watson PH, Murphy LC (1998) Expression of lumican in human breast carcinoma. Cancer Res 58:1348–1352PubMedGoogle Scholar
  34. 34.
    Naito Z, Ishiwata T, Kurban G, Teduka K, Kawamoto Y, Kawahara K, Sugisaki Y (2002) Expression and accumulation of lumican protein in uterine cervical cancer cells at the periphery of cancer nests. Int J Oncol 20:943–948PubMedGoogle Scholar
  35. 35.
    Lu YP, Ishiwata T, Kawahara K, Watanabe M, Naito Z, Moriyama Y, Sugisaki Y, Asano G (2002) Expression of lumican in human colorectal cancer cells. Pathol Int 52:519–526CrossRefPubMedGoogle Scholar
  36. 36.
    Vuillermoz B, Khoruzhenko A, D’Onofrio MF, Ramont L, Venteo L, Perreau C, Antonicelli F, Maquart FX, Wegrowski Y (2004) The small leucine-rich proteoglycan lumican inhibits melanoma progression. Exp Cell Res 296:294–306CrossRefPubMedGoogle Scholar
  37. 37.
    Dhanasekaran SM, Barrette TR, Ghosh D, Shah R, Varambally S, Kurachi K, Pienta KJ, Rubin MA, Chinnaiyan AM (2001) Delineation of prognostic biomarkers in prostate cancer. Nature 412:822–826CrossRefPubMedGoogle Scholar
  38. 38.
    Lapointe J, Li C, Higgins JP, van de Rijn M, Bair E, Montgomery K, Ferrari M, Egevad L, Rayford W, Bergerheim U, Ekman P, DeMarzo AM, Tibshirani R, Botstein D, Brown PO, Brooks JD, Pollack JR (2004) Gene expression profiling identifies clinically relevant subtypes of prostate cancer. Proc Natl Acad Sci USA 101:811–816CrossRefPubMedGoogle Scholar
  39. 39.
    Chan I (2004) The role of extracellular matrix protein 1 in human skin. Clin Exp Dermatol 29:52–56CrossRefPubMedGoogle Scholar
  40. 40.
    Hamada T, McLean WH, Ramsay M, Ashton GH, Nanda A, Jenkins T, Edelstein I, South AP, Bleck O, Wessagowit V, Mallipeddi R, Orchard GE, Wan H, Dopping-Hepenstal PJ, Mellerio JE, Whittock NV, Munro CS, van Steensel MA, Steijlen PM, Ni J, Zhang L, Hashimoto T, Eady RA, McGrath JA (2002) Lipoid proteinosis maps to 1q21 and is caused by mutations in the extracellular matrix protein 1 gene (ECM1). Hum Mol Genet 11:833–840CrossRefPubMedGoogle Scholar
  41. 41.
    Oyama N, Chan I, Neill SM, Hamada T, South AP, Wessagowit V, Wojnarowska F, D’Cruz D, Hughes GJ, Black MM, McGrath JA (2003) Autoantibodies to extracellular matrix protein 1 in lichen sclerosus. Lancet 362:118–123CrossRefPubMedGoogle Scholar
  42. 42.
    Kowalewski C, Kozlowska A, Chan I, Gorska M, Wozniak K, Jablonska S, McGrath JA (2005) Three-dimensional imaging reveals major changes in skin microvasculature in lipoid proteinosis and lichen sclerosus. J Dermatol Sci 38:215–224CrossRefPubMedGoogle Scholar
  43. 43.
    Wang L, Yu J, Ni J, Xu XM, Wang J, Ning H, Pei XF, Chen J, Yang S, Underhill CB, Liu L, Liekens J, Merregaert J, Zhang L (2003) Extracellular matrix protein 1 (ECM1) is over-expressed in malignant epithelial tumors. Cancer Lett 200:57–67CrossRefPubMedGoogle Scholar
  44. 44.
    Kebebew E, Peng M, Reiff E, Duh QY, Clark OH, McMillan A (2005) ECM1 and TMPRSS4 are diagnostic markers of malignant thyroid neoplasms and improve the accuracy of fine needle aspiration biopsy. Ann Surg 242:353–361PubMedGoogle Scholar
  45. 45.
    Han Z, Ni J, Smits P, Underhill CB, Xie B, Chen Y, Liu N, Tylzanowski P, Parmelee D, Feng P, Ding I, Gao F, Gentz R, Huylebroeck D, Merregaert J, Zhang L (2001) Extracellular matrix protein 1 (ECM1) has angiogenic properties and is expressed by breast tumor cells. FASEB J 15:988–994CrossRefPubMedGoogle Scholar
  46. 46.
    Mirancea N, Hausser I, Beck R, Metze D, Fusenig NE, Breitkreutz D (2006) Vascular anomalies in lipoid proteinosis (hyalinosis cutis et mucosae): basement membrane components and ultrastructure. J Dermatol Sci 42:231–239CrossRefPubMedGoogle Scholar
  47. 47.
    Rocnik EF, Liu P, Sato K, Walsh K, Vaziri C (2006) The novel SPARC family member SMOC-2 potentiates angiogenic growth factor activity. J Biol Chem 281:22855–22864CrossRefPubMedGoogle Scholar
  48. 48.
    Vannahme C, Gosling S, Paulsson M, Maurer P, Hartmann U (2003) Characterization of SMOC-2, a modular extracellular calcium-binding protein. Biochem J 373:805–814CrossRefPubMedGoogle Scholar
  49. 49.
    Vannahme C, Smyth N, Miosge N, Gosling S, Frie C, Paulsson M, Maurer P, Hartmann U (2002) Characterization of SMOC-1, a novel modular calcium-binding protein in basement membranes. J Biol Chem 277:37977–37986CrossRefPubMedGoogle Scholar
  50. 50.
    Funk SE, Sage EH (1993) Differential effects of SPARC and cationic SPARC peptides on DNA synthesis by endothelial cells and fibroblasts. J Cell Physiol 154:53–63CrossRefPubMedGoogle Scholar
  51. 51.
    Sage EH, Reed M, Funk SE, Truong T, Steadele M, Puolakkainen P, Maurice DH, Bassuk JA (2003) Cleavage of the matricellular protein SPARC by matrix metalloproteinase 3 produces polypeptides that influence angiogenesis. J Biol Chem 278:37849–37857CrossRefPubMedGoogle Scholar
  52. 52.
    Kupprion C, Motamed K, Sage EH (1998) SPARC (BM-40, osteonectin) inhibits the mitogenic effect of vascular endothelial growth factor on microvascular endothelial cells. J Biol Chem 273:29635–29640CrossRefPubMedGoogle Scholar
  53. 53.
    Jendraschak E, Sage EH (1996) Regulation of angiogenesis by SPARC and angiostatin: implications for tumor cell biology. Semin Cancer Biol 7:139–146CrossRefPubMedGoogle Scholar
  54. 54.
    Gibson MA, Finnis ML, Kumaratilake JS, Cleary EG (1998) Microfibril-associated glycoprotein-2 (MAGP-2) is specifically associated with fibrillin-containing microfibrils but exhibits more restricted patterns of tissue localization and developmental expression than its structural relative MAGP-1. J Histochem Cytochem 46:871–886PubMedGoogle Scholar
  55. 55.
    Gibson MA, Leavesley DI, Ashman LK (1999) Microfibril-associated glycoprotein-2 specifically interacts with a range of bovine and human cell types via αVβ3 integrin. J Biol Chem 274:13060–13065CrossRefPubMedGoogle Scholar
  56. 56.
    Lemaire R, Bayle J, Mecham RP, Lafyatis R (2007) Microfibril-associated MAGP-2 stimulates elastic fiber assembly. J Biol Chem PMID 282:800–808CrossRefGoogle Scholar
  57. 57.
    Lemaire R, Farina G, Kissin E, Shipley JM, Bona C, Korn JH, Lafyatis R (2004) Mutant fibrillin 1 from tight skin mice increases extracellular matrix incorporation of microfibril-associated glycoprotein 2 and type I collagen. Arthritis Rheum 50:915–926CrossRefPubMedGoogle Scholar
  58. 58.
    Lemaire R, Korn JH, Shipley JM, Lafyatis R (2005) Increased expression of type I collagen induced by microfibril-associated glycoprotein 2: novel mechanistic insights into the molecular basis of dermal fibrosis in scleroderma. Arthritis Rheum 52:1812–1823CrossRefPubMedGoogle Scholar
  59. 59.
    Bodolay E, Koch AE, Kim J, Szegedi G, Szekanecz Z (2002) Angiogenesis and chemokines in rheumatoid arthritis and other systemic inflammatory rheumatic diseases. J Cell Mol Med 6:357–376CrossRefPubMedGoogle Scholar
  60. 60.
    Graham JD, Yager ML, Hill HD, Byth K, O’Neill GM, Clarke CL (2005) Altered progesterone receptor isoform expression remodels progestin responsiveness of breast cancer cells. Mol Endocrinol 19:2713–2735CrossRefPubMedGoogle Scholar
  61. 61.
    Bild AH, Yao G, Chang JT, Wang Q, Potti A, Chasse D, Joshi MB, Harpole D, Lancaster JM, Berchuck A, Olson JA Jr, Marks JR, Dressman HK, West M, Nevins JR (2006) Oncogenic pathway signatures in human cancers as a guide to targeted therapies. Nature 439:353–357CrossRefPubMedGoogle Scholar
  62. 62.
    Creighton C, Kuick R, Misek DE, Rickman DS, Brichory FM, Rouillard JM, Omenn GS, Hanash S (2003) Profiling of pathway-specific changes in gene expression following growth of human cancer cell lines transplanted into mice. Genome Biol 4:R46CrossRefPubMedGoogle Scholar
  63. 63.
    Iacobuzio-Donahue CA, Ashfaq R, Maitra A, Adsay NV, Shen-Ong GL, Berg K, Hollingsworth MA, Cameron JL, Yeo CJ, Kern SE, Goggins M, Hruban RH (2003) Highly expressed genes in pancreatic ductal adenocarcinomas: a comprehensive characterization and comparison of the transcription profiles obtained from three major technologies. Cancer Res 63:8614–8622PubMedGoogle Scholar
  64. 64.
    Miyamoto A, Lau R, Hein PW, Shipley JM, Weinmaster G (2006) Microfibrillar proteins MAGP-1 and MAGP-2 induce Notch1extracellular domain dissociation and receptor activation. J Biol Chem 281:10089–10097CrossRefPubMedGoogle Scholar
  65. 65.
    Nehring LC, Miyamoto A, Hein PW, Weinmaster G, Shipley JM (2005) The extracellular matrix protein MAGP-2 interacts with Jagged1 and induces its shedding from the cell surface. J Biol Chem 280:20349–20355CrossRefPubMedGoogle Scholar
  66. 66.
    Leong KG, Karsan A (2006) Recent insights into the role of Notch signaling in tumorigenesis. Blood 107:2223–2233CrossRefPubMedGoogle Scholar
  67. 67.
    Alva JA, Iruela-Arispe ML (2004) Notch signaling in vascular morphogenesis. Curr Opin Hematol 11:278–283CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science + Business Media B.V. 2007

Authors and Affiliations

  • Allan R. Albig
    • 1
  • Thessa G. Roy
    • 1
  • Darryl J. Becenti
    • 1
  • William P. Schiemann
    • 1
  1. 1.Department of PharmacologyUniversity of Colorado Health Sciences CenterAuroraUSA

Personalised recommendations