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Production of two novel monoclonal antibodies that distinguish mouse lymphatic and blood vascular endothelial cells

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Abstract

We produced two novel rat monoclonal antibodies (LA102 and LA5) to identify mouse lymphatic vessels and blood vessels, respectively. We characterized the two antibodies as to the morphological and functional specificities of endothelial cells of both types of vessels. The antibodies were produced by a rapid differential immunization of DA rats with collagenase- and neuraminidase-treated mouse lymphangioma tissues. LA102 specifically reacted with mouse lymphatic vessels except the thoracic duct and the marginal sinus of lymph nodes, but not with any blood vessels. In contrast, LA5 reacted with most mouse blood vessels with a few exceptions, but not with lymphatics. LA102 recognized a protein of 25–27 kDa, whereas LA5 recognized a molecule of 12–13 kDa. Neither antibody recognized any currently identified lymphatic or vascular endothelial cell antigens. Immunoelectron microscopy revealed that the antigens recognized by LA102 and LA5 were localized on both luminal and abluminal endothelial cell membranes of each vessel type. Interestingly, LA102 immunoreactivity was strongly expressed on pinocytic or transport vesicle membrane in the cytoplasm of lymphatic endothelium. Besides endothelial cells, both antibodies also recognized some types of lymphoid cells. Since, the LA102 antigen molecule is expressed on some lymphoid cells, it may play important roles in the migration of lymphoid cells and in some transport mechanisms through lymphatic endothelial cells.

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References

  • Anderson CR, Ponce AM, Price RJ (2004) Absence of OX-43 antigen expression in invasive capillary sprouts: identification of a capillary sprout-specific endothelial phenotype. Am J Physiol Heart Circ Physiol 286:H346–H353

    Article  PubMed  CAS  Google Scholar 

  • Baluk P, Tammela T, Ator E, Lyubynska N, Achen MG, Hicklin DJ, Jeltsch M, Petrova TV, Pytowski B, Stacker SA, Yla-Herttuala S, Jackson DG, Alitalo K, McDonald DM (2005) Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation. J Clin Invest 115:247–257

    Article  PubMed  CAS  Google Scholar 

  • Banerji S, Ni J, Wang SX, Clasper S, Su J, Tammi R, Jones M, Jackson DG (1999) LYVE-1, a new homologue of the CD44 glycoprotein, is a lymph-specific receptor for hyaluronan. J Cell Biol 144:789–801

    Article  PubMed  CAS  Google Scholar 

  • Baumhueter S, Singer MS, Henzel W, Hemmerich S, Renz M, Rosen SD, Lasky LA (1993) Binding of L-selectin to the vascular sialomucin CD34. Science 262:436–438

    Article  CAS  Google Scholar 

  • Breiteneder-Geleff S, Soleiman A, Kowalski H, Horvat R, Amann G, Kriehuber E, Diem K, Weninger W, Tschachler E, Alitalo K, Kerjaschki D (1999) Angiosarcomas express mixed endothelial phenotypes of blood and lymphatic capillaries: podoplanin as a specific marker for lymphatic endothelium. Am J Pathol 154:385–394

    PubMed  CAS  Google Scholar 

  • DeLisser HM, Newman PJ, Albelda SM (1993) Platelet endothelial cell adhesion molecule (CD31). Curr Top Microbiol Immunol 184:37–45

    PubMed  CAS  Google Scholar 

  • Ezaki T, Baluk P, Thurston G, La Barbara A, Woo C, McDonald DM (2001) Time course of endothelial cell proliferation and microvascular remodeling in chronic inflammation. Am J Pathol 158:2043–2055

    PubMed  CAS  Google Scholar 

  • Ezaki T, Kuwahara K, Morikawa S, Matsuno K, Sakaguchi N (2004) Characterization of adjuvant-induced rat lymphangiomas as a model to study the lymph drainage from abdominal cavity. Jpn J Lymphol 27:1–10

    Google Scholar 

  • Ezaki T, Matsuno K, Fujii H, Hayashi N, Miyakawa K, Ohmori J, Kotani M (1990) A new approach for identification of rat lymphatic capillaries using a monoclonal antibody. Arch Histol Cytol 53(Suppl):77–86

    PubMed  Google Scholar 

  • Ezaki T, Matsuno K, Kotani M (1991) Thymic nurse cells (TNC) in spontaneous thymoma BUF/Mna rats as a model to study their roles in T-cell development. Immunology 73:151–158

    PubMed  CAS  Google Scholar 

  • Ezaki T, Yao L, Matsuno K (1995) The identification of proliferating cell nuclear antigen (PCNA) on rat tissue cryosections and its application to double immunostaining with other markers. Arch Histol Cytol 58:103–115

    PubMed  CAS  Google Scholar 

  • Feng D, Nagy JA, Pyne K, Dvorak HF, Dvorak AM (2004) Ultrastructural localization of platelet endothelial cell adhesion molecule (PECAM-1, CD31) in vascular endothelium. J Histochem Cytochem 52:87–101

    PubMed  CAS  Google Scholar 

  • Garlanda C, Dejana E (1997) Heterogeneity of endothelial cells. Specific markers. Arterioscler Thromb Vasc Biol 17:1193–1202

    CAS  Google Scholar 

  • Gougos A, Letarte M (1990) Primary structure of endoglin, an RGD-containing glycoprotein of human endothelial cells. J Biol Chem 265:8361–8364

    PubMed  CAS  Google Scholar 

  • Hallmann R, Mayer DN, Berg EL, Broermann R, Butcher EC (1995) Novel mouse endothelial cell surface marker is suppressed during differentiation of the blood brain barrier. Dev Dyn 202:325–332

    PubMed  CAS  Google Scholar 

  • Hirakawa S, Hong YK, Harvey N, Schacht V, Matsuda K, Libermann T, Detmar M (2003) Identification of vascular lineage-specific genes by transcriptional profiling of isolated blood vascular and lymphatic endothelial cells. Am J Pathol 162:575–586

    PubMed  CAS  Google Scholar 

  • Hong YK, Shin JW, Detmar M (2004) Development of the lymphatic vascular system: a mystery unravels. Dev Dyn 231:462–473

    Article  PubMed  CAS  Google Scholar 

  • Jackson DG (2004) Biology of the lymphatic marker LYVE-1 and applications in research into lymphatic trafficking and lymphangiogenesis. APMIS 112:526–538

    Article  PubMed  CAS  Google Scholar 

  • Jackson DG, Prevo R, Clasper S, Banerji S (2001) LYVE-1, the lymphatic system and tumor lymphangiogenesis. Trends Immunol 22:317–321

    Article  PubMed  CAS  Google Scholar 

  • Ji RC (2005) Characteristics of lymphatic endothelial cells in physiological and pathological conditions. Histol Histopathol 20:155–175

    PubMed  CAS  Google Scholar 

  • Kahn HJ, Bailey D, Marks A (2002) Monoclonal antibody D2-40, a new marker of lymphatic endothelium, reacts with Kaposi’s sarcoma and a subset of angiosarcomas. Mod Pathol 15:434–440

    Article  PubMed  Google Scholar 

  • Kaipainen A, Korhonen J, Mustonen T, van Hinsbergh VW, Fang GH, Dumont D, Breitman M, Alitalo K (1995) Expression of the fms-like tyrosine kinase 4 gene becomes restricted to lymphatic endothelium during development. Proc Natl Acad Sci USA 92:3566–3570

    Article  PubMed  CAS  Google Scholar 

  • Karpanen T, Alitalo K (2001) Lymphatic vessels as targets of tumor therapy. J Exp Med 194:F37–F42

    Article  PubMed  CAS  Google Scholar 

  • Kuwahara K, Matsuo T, Nomura J, Igarashi H, Kimoto M, Inui S, Sakaguchi N (1994) Identification of a 52-kDa molecule (p52) coprecipitated with the Ig receptor-related MB-1 protein that is inducibly phosphorylated by the stimulation with phorbol myristate acetate. J Immunol 152:2742–2752

    PubMed  CAS  Google Scholar 

  • Lesley J, Hyman R, Kincade PW (1993) CD44 and its interaction with extracellular matrix. Adv Immunol 54:271–335

    Article  PubMed  CAS  Google Scholar 

  • Mancardi S, Stanta G, Dusetti N, Bestagno M, Jussila L, Zweyer M, Lunazzi G, Dumont D, Alitalo K, Burrone OR (1999) Lymphatic endothelial tumors induced by intraperitoneal injection of incomplete Freund’s adjuvant. Exp Cell Res 246:368–375

    Article  PubMed  CAS  Google Scholar 

  • Mokhtar N, Hsu SM, Lad RP, Haynes BF, Jaffe ES (1984) Thymoma: lymphoid and epithelial components mirror the phenotype of normal thymus. Hum Pathol 15:378–384

    Article  PubMed  CAS  Google Scholar 

  • Morikawa S, Kuwahara K, Sakaguchi N, Matsushima K, Ezaki T (2004) A new marker for lymphatic endothelial cells recognized by a monoclonal antibody. Lymphology 37(Suppl):105–108

    Google Scholar 

  • Mouta Carreira C, Nasser SM, di Tomaso E, Padera TP, Boucher Y, Tomarev SI, Jain RK (2001) LYVE-1 is not restricted to the lymph vessels: expression in normal liver blood sinusoids and down-regulation in human liver cancer and cirrhosis. Cancer Res 61:8079–8084

    PubMed  CAS  Google Scholar 

  • Niemela H, Elima K, Henttinen T, Irjala H, Salmi M, Jalkanen S (2005) Molecular identification of PAL-E, a widely used endothelial-cell marker. Blood 106:3405–3409

    Article  PubMed  CAS  Google Scholar 

  • Partanen TA, Alitalo K, Miettinen M (1999) Lack of lymphatic vascular specificity of vascular endothelial growth factor receptor 3 in 185 vascular tumors. Cancer 86:2406–2412

    Article  PubMed  CAS  Google Scholar 

  • Petrova TV, Karpanen T, Norrmen C, Mellor R, Tamakoshi T, Finegold D, Ferrell R, Kerjaschki D, Mortimer P, Yla-Herttuala S, Miura N, Alitalo K (2004) Defective valves and abnormal mural cell recruitment underlie lymphatic vascular failure in lymphedema distichiasis. Nat Med 10:974–981

    Article  PubMed  CAS  Google Scholar 

  • Prevo R, Banerji S, Ferguson DJ, Clasper S, Jackson DG (2001) Mouse LYVE-1 is an endocytic receptor for hyaluronan in lymphatic endothelium. J Biol Chem 276:19420–19430. Epub 2001 February 20

    Google Scholar 

  • Robinson AP, White TM, Mason DW (1986) MRC OX-43: a monoclonal antibody which reacts with all vascular endothelium in the rat except that of brain capillaries. Immunology 57:231–237

    PubMed  CAS  Google Scholar 

  • Sakaguchi N, Kashiwamura S, Kimoto M, Thalmann P, Melchers F (1988) B lymphocyte lineage-restricted expression of mb-1, a gene with CD3-like structural properties. EMBO J 7:3457–3464

    PubMed  CAS  Google Scholar 

  • Samelson LE, Schwartz RH (1983) T cell clone-specific alloantisera that inhibit or stimulate antigen-induced T cell activation. J Immunol 131:2645–2650

    PubMed  CAS  Google Scholar 

  • Scavelli C, Weber E, Agliano M, Cirulli T, Nico B, Vacca A, Ribatti D (2004) Lymphatics at the crossroads of angiogenesis and lymphangiogenesis. J Anat 204:433–449

    Article  PubMed  CAS  Google Scholar 

  • Schacht V, Dadras SS, Johnson LA, Jackson DG, Hong YK, Detmar M (2005) Up-regulation of the lymphatic marker podoplanin, a mucin-type transmembrane glycoprotein, in human squamous cell carcinomas and germ cell tumors. Am J Pathol 166:913–921

    PubMed  CAS  Google Scholar 

  • Schacht V, Ramirez MI, Hong YK, Hirakawa S, Feng D, Harvey N, Williams M, Dvorak AM, Dvorak HF, Oliver G, Detmar M (2003) T1alpha/podoplanin deficiency disrupts normal lymphatic vasculature formation and causes lymphedema. EMBO J 22:3546–3556

    Article  PubMed  CAS  Google Scholar 

  • Schlingemann RO, Dingjan GM, Emeis JJ, Blok J, Warnaar SO, Ruiter DJ (1985) Monoclonal antibody PAL-E specific for endothelium. Lab Invest 52:71–76

    PubMed  CAS  Google Scholar 

  • Sleeman JP, Krishnan J, Kirkin V, Baumann P (2001) Markers for the lymphatic endothelium: in search of the holy grail. Microsc Res Tech 55:61–69

    Article  PubMed  CAS  Google Scholar 

  • Stan RV (2005) Structure of caveolae. Biochim Biophys Acta 1746:334–348

    Article  PubMed  CAS  Google Scholar 

  • Swartz MA, Skobe M (2001) Lymphatic function, lymphangiogenesis, and cancer metastasis. Microsc Res Tech 55:92–99

    Article  PubMed  CAS  Google Scholar 

  • Tammela T, Petrova TV, Alitalo K (2005) Molecular lymphangiogenesis: new players. Trends Cell Biol 15:434–441

    Article  PubMed  CAS  Google Scholar 

  • Thompson LF, Ruedi JM, Glass A, Low MG, Lucas AH (1989) Antibodies to 5’-nucleotidase (CD73), a glycosyl-phosphatidylinositol-anchored protein, cause human peripheral blood T cells to proliferate. J Immunol 143:1815–1821

    PubMed  CAS  Google Scholar 

  • Thurston G, Baluk P, Hirata A, McDonald DM (1996) Permeability-related changes revealed at endothelial cell borders in inflamed venules by lectin binding. Am J Physiol 271:H2547–H2562

    PubMed  CAS  Google Scholar 

  • Toshimori K, Araki S, Oura C (1988) Masking of sperm maturation antigen by sialic acid in the epidedymis of the mouse. Histochemistry 90:195–200

    Article  PubMed  CAS  Google Scholar 

  • Wigle JT, Harvey N, Detmar M, Lagutina I, Grosveld G, Gunn MD, Jackson DG, Oliver G (2002) An essential role for Prox1 in the induction of the lymphatic endothelial cell phenotype. EMBO J 21:1505–1513

    Article  PubMed  CAS  Google Scholar 

  • Wigle JT, Oliver G (1999) Prox1 function is required for the development of the murine lymphatic system. Cell 98:769–778

    Article  PubMed  CAS  Google Scholar 

  • Wilting J, Tomarev SI, Christ B, Schweigerer L (2003) Lymphangioblasts in embryonic lymphangiogenesis. Lymphat Res Biol 1:33–40

    Article  PubMed  Google Scholar 

  • Xu B, deWaal RM, Mor-Vaknin N, Hibbard C, Markovitz DM, Kahn ML (2004a) The endothelial cell-specific antibody PAL-E identifies a secreted form of vimentin in the blood vasculature. Mol Cell Biol 24:9198–9206

    Article  CAS  Google Scholar 

  • Xu H, Edwards JR, Espinosa O, Banerji S, Jackson DG, Athanasou NA (2004b) Expression of a lymphatic endothelial cell marker in benign and malignant vascular tumors. Hum Pathol 35:857–861

    Article  CAS  Google Scholar 

  • Zawieja D (2005) Lymphatic biology and the microcirculation: past, present, and future. Microcirculation 12:141–150

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors are grateful to Dr. P Baluk (Cardiovascular Research Institute, UCSF, San Francisco, USA) for his advice and critical review of the manuscript; to Mrs. Y. Yamazaki, Ms. K. Nakata, Ms. H. Kasahara, and Ms. K. Moriya for their devoted technical help. This work was mainly supported by a Grant-in-Aid for Scientific Research (A)(2) (No.14207001) from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.

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Authors and Affiliations

  1. Department of Anatomy and Developmental Biology, Tokyo Women’s Medical University, Tokyo, Japan

    Taichi Ezaki, Shunichi Morikawa & Kazuhiko Shimizu

  2. Department of Immunology, Faculty of Medical and Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan

    Kazuhiko Kuwahara & Nobuo Sakaguchi

  3. Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan

    Kouji Matsushima

  4. Department of Anatomy, Dokkyo University School of Medicine, Tochigi, Japan

    Kenjiro Matsuno

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  1. Taichi Ezaki
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  2. Kazuhiko Kuwahara
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  4. Kazuhiko Shimizu
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  5. Nobuo Sakaguchi
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Correspondence to Taichi Ezaki.

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Ezaki, T., Kuwahara, K., Morikawa, S. et al. Production of two novel monoclonal antibodies that distinguish mouse lymphatic and blood vascular endothelial cells. Anat Embryol 211, 379–393 (2006). https://doi.org/10.1007/s00429-006-0091-3

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