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An Innovative Triple Immunogold Labeling Method to Investigate the Hemopoietic Stem Cell Niche In Situ

Published online by Cambridge University Press:  16 September 2009

Sarah L. Ellis ,
Brenda Williams ,
Stephen Asquith ,
Ivan Bertoncello  and
Susan K. Nilsson
Sarah L. Ellis
Affiliation:
Microscopy Core Facility, Peter MacCallum Cancer Centre, East Melbourne, Victoria 3002, Australia Australian Stem Cell Centre, Clayton, Victoria 3168, Australia Department of Pathology, University of Melbourne, Parkville, Victoria 3052, Australia
Brenda Williams
Affiliation:
Australian Stem Cell Centre, Clayton, Victoria 3168, Australia
Stephen Asquith
Affiliation:
Microscopy Core Facility, Peter MacCallum Cancer Centre, East Melbourne, Victoria 3002, Australia
Ivan Bertoncello
Affiliation:
Australian Stem Cell Centre, Clayton, Victoria 3168, Australia Department of Pathology, University of Melbourne, Parkville, Victoria 3052, Australia
Susan K. Nilsson*
Affiliation:
Australian Stem Cell Centre, Clayton, Victoria 3168, Australia Department of Pathology, University of Melbourne, Parkville, Victoria 3052, Australia
*
Corresponding author. E-mail: Susie.nilsson@csiro.au
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Abstract

The ultrastructural study of rare cells within their niche in situ is very difficult. We have developed a method for locating individual transplanted cells and simultaneously identifying and analyzing the molecules and cellular phenotypes surrounding them in situ using transmission electron microscopy. This innovative method involves triple immunogold labeling combined with serial ultrathin sectioning. We demonstrate the validity of this approach by examining the niche of individual transplanted cells from a population highly enriched for hemopoietic stem cells and the ultrastructural expression of two key stem cell regulatory molecules, hyaluronic acid and osteopontin. In addition, we describe the phenotypes of the surrounding cells.

Keywords

hemopoietic stem cellhemopoietic progenitor cellhomingnicheimmunogoldhyaluronic acidosteopontin
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 15 , Issue 5 , October 2009 , pp. 403 - 414
Copyright
Copyright © Microscopy Society of America 2009

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References

REFERENCES

Arai, F., Hirao, A., Ohmura, M., Sato, H., Matsuoka, S., Takubo, K., Ito, K., Koh, G.Y. & Suda, T. (2004). Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche. Cell 118(2), 149–161.CrossRefGoogle ScholarPubMed
Arnold, J., Ellis, S., Radley, J. & Williams, N. (1991). Compensatory mechanisms in platelet production: The response of Sl/Sld mice to 5-fluorouracil. Exp Hematol 19(1), 24–28.Google Scholar
Avigdor, A., Goichberg, P., Shivtiel, S., Dar, A., Peled, A., Samira, S., Kollet, O., Hershkoviz, R., Alon, R., Hardan, I., Ben-Hur, H., Naor, D., Nagler, A. & Lapidot, T. (2004). CD44 and hyaluronic acid cooperate with SDF-1 in the trafficking of human CD34+ stem/progenitor cells to bone marrow. Blood 103(8), 2981–2989.Google Scholar
Baschong, W. & Stierhof, Y.-D. (1998). Preparation, use, and enlargement of ultrasmall gold particles in immunoelectron microscopy. Microsc Res Tech 42(1), 66–79.3.0.CO;2-P>CrossRefGoogle ScholarPubMed
Bendayan, M. (1984). Protein A-gold electron microscopic immunocytochemistry: Methods, applications, and limitations. J Electron Microsc Tech 1, 243–270.Google Scholar
Bendayan, M., Nanci, A. & Kan, F. (1987). Effect of tissue processing on colloidal gold cytochemistry. J Histochem Cytochem 35, 983–996.CrossRefGoogle ScholarPubMed
Burry, R. (1995). Pre-embedding immunocytochemistry with silver-enhanced small gold particles. In Immunogold-Silver Staining: Principles, Methods, and Applications, Hayat, M. (Ed.), pp. 217–230. Boca Raton, FL: CRC Press.Google Scholar
Burry, R.W., Vandre, D.D. & Hayes, D.M. (1992). Silver enhancement of gold antibody probes in pre-embedding electron microscopic immunocytochemistry. J Histochem Cytochem 40(12), 1849–1856.Google Scholar
Calvi, L.M., Adams, G.B., Weibrecht, K.W., Weber, J.M., Olson, D.P., Knight, M.C., Martin, R.P., Schipani, E., Divieti, P., Bringhurst, F.R., Milner, L.A., Kronenberg, H.M. & Scadden, D.T. (2003). Osteoblastic cells regulate the haematopoietic stem cell niche. Nature 425, 841–846.CrossRefGoogle ScholarPubMed
Celso, C.L., Fleming, H.E., Wu, J.W., Zhao, C.X., Miake-Lye, S., Fujisaki, J., Cote, D., Rowe, D.W., Lin, C.P. & Scadden, D.T. (2009). Live-animal tracking of individual haematopoietic stem/progenitor cells in their niche. Nature 457, 92–96.CrossRefGoogle ScholarPubMed
Chan, J., Aoki, C. & Pickel, V.M. (1990). Optimization of differential immunogold-silver and peroxidase labeling with maintenance of ultrastructure in brain sections before plastic embedding. J Neurosci Meth 33(2–3), 113–127.Google Scholar
Danscher, G. (1981). Histochemical demonstration of heavy metals. A revised version of the sulphide silver method suitable for both light and electron microscopy. Histochem 71, 1–16.CrossRefGoogle Scholar
Danscher, G. (1983). A silver method for counterstaining plastic embedded tissue. Stain Technol 58, 365–372.Google Scholar
Danscher, G. & Norgaad, J. (1983). Light microscopic visualization of colloidal gold on resin-embedded tissue. J Histochem Cytochem 31, 1394–1398.CrossRefGoogle ScholarPubMed
Griffiths, G. & Hoppeler, H. (1986). Quantitation in immunocytochemistry: Correlation of immunogold labelling to absolute number of membrane antigens. J Histochem Cytochem 34, 1389–1398.Google Scholar
Gu, J., Choi, T.-S., Whittlesey, M., Slap, S. & Anderson, V. (1995). Development of microwave immunohistochemistry. Cell Vis 2, 257–259.Google Scholar
Hacker, G., Grimelius, L., Danscher, G., Bernatzky, G., Muss, W., Adam, H. & Thurner, J. (1988). Silver acetate autometallography: An alternative enhancement technique for immunogold-silver staining (IGSS) and silver amplification of gold, silver, mercury and zinc in tissues. J Histotechnol 11, 213–221.Google Scholar
Hainfeld, J. (1987). A small gold-conjugate antibody label: Improved resolution for electron microscopy. Science 263, 450–453.CrossRefGoogle Scholar
Hainfeld, J. (1988). Gold cluster-labeled antibodies. Nature 333, 281–282.Google Scholar
Hainfeld, J. & Furuya, F. (1992). A 1.4-nm gold cluster covalently attached to antibodies improves immunolabeling. J Histochem Cytochem 40, 177–184.CrossRefGoogle ScholarPubMed
Haylock, D.N., Williams, B., Johnston, H.M., Liu, M.C.P., Rutherford, K.E., Whitty, G.A., Simmons, P.J., Bertoncello, I. & Nilsson, S.K. (2007). Hemopoietic stem cells with higher hemopoietic potential reside at the bone marrow endosteum. Stem Cells 25(4), 1062–1069.CrossRefGoogle ScholarPubMed
Hendrikx, P.J., Martens, A.C.M., Hagenbeek, A., Keij, J.F. & Visser, J.W.M. (1996). Homing of fluorescently labeled murine hematopoietic stem cells. Exp Hematol 24, 129–140.Google Scholar
Holgate, C.S., Jackson, P., Cowen, P.N. & Bird, C.C. (1983). Immunogold-silver staining: New method of immunostaining with enhanced sensitivity. J Histochem Cytochem 31(7), 938–944.CrossRefGoogle ScholarPubMed
Junqueira, L.C. & Carneiro, J. (2005). Blood cells. In Basic Histology: Text and Atlas, Foltin, J., Lebowitz, H. & Boyle, P.J. (Eds.), pp. 223–237. New York: McGraw-Hill.Google Scholar
Kacena, M.A., Gundberg, C.M. & Horowitz, M.C. (2006). A reciprocal regulatory interaction between megakaryocytes, bone cells, and hematopoietic stem cells. Bone 39(5), 978–984.Google Scholar
Kiel, M.J., Yilmaz, O.H., Iwashita, T., Yilmaz, O.H., Terhorst, C. & Morrison, S.J. (2005). SLAM family receptors distinguish hematopoietic stem and progenitor cells and reveal endothelial niches for stem cells. Cell 121(7), 1109–1121.Google Scholar
Kissa, K., Murayama, E., Zapata, A., Cortes, A., Perret, E., Machu, C. & Herbomel, P. (2008). Live imaging of emerging hematopoietic stem cells and early thymus colonization. Blood 111(3), 1147–1156.Google Scholar
Lemoli, R.M. & D'Addio, A. (2008). Hematopoietic stem cell mobilization. Haematologica 93(3), 321–324.Google Scholar
Leunissen, J.L.M. & DeMey, J.R. (1989). Preparation of gold probes. In Immuno-Gold Labeling in Cell Biology, Verkleji, A.J. & Leunissen, J.L.M. (Eds.), pp. 3–16. Boca Raton, FL: CRL Press.Google Scholar
Lucocq, J. (1992). Quantitation of gold labeling and estimation of labeling efficiency with a stereological counting method. J Histochem Cytochem 40, 1929–1936.CrossRefGoogle ScholarPubMed
Mayack, S.R. & Wagers, A.J. (2008). Osteolineage niche cells initiate hematopoietic stem cell mobilization. Blood 112(3), 519–531.Google Scholar
McKee, M.D., Glimcher, M.J. & Nanci, A. (1992). High-resolution immunolocalization of osteopontin and osteocalcin in bone and cartilage during endochondral ossification in the chicken tibia. Anat Rec 234, 479–492.CrossRefGoogle ScholarPubMed
McKee, M.D. & Nanci, A. (1995). Post-embedding colloidal-gold immunocytochemistry of non-collagenous extracellular matrix proteins in mineralized tissues. Microsc Res Technol 31, 44–62.CrossRefGoogle Scholar
McKee, M. & Nanci, A. (1996). Osteopontin: An interfacial extracellular matrix protein in mineralized tissues. Conn Tissue Res 35, 197–205.CrossRefGoogle ScholarPubMed
Newman, G.R. & Hobot, J.A. (1987). Modern acrylics for post-embedding immunostaining techniques. J Histochem Cytochem 35(9), 971–981.Google Scholar
Nilsson, S.K., Dooner, M.S. & Quesenberry, P.J. (1997a). Synchronized cell-cycle induction of engrafting long-term repopulating stem cells. Blood 90(11), 4646–4650.CrossRefGoogle ScholarPubMed
Nilsson, S.K., Dooner, M.S., Tiarks, C.Y., Weier, H.-U.G. & Quesenberry, P.J. (1997b). Potential and distribution of transplanted hematopoietic stem cells in a nonablated mouse model. Blood 89(11), 4013–4020.Google Scholar
Nilsson, S.K., Haylock, D.N., Johnston, H.M., Occhiodoro, T., Brown, T.J. & Simmons, P.J. (2003). Hyaluronan is synthesized by primitive hemopoietic cells, participates in their lodgment at the endosteum following transplantation, and is involved in the regulation of their proliferation and differentiation in vitro. Blood 101(3), 856–862.Google Scholar
Nilsson, S.K., Hulspas, R., Weier, H.-U.G. & Quesenberry, P.J. (1996). In situ detection of individual transplanted bone marrow cells using FISH on sections of paraffin-embedded whole murine femurs. J Histochem Cytochem 44(9), 1069–1074.Google Scholar
Nilsson, S.K., Johnston, H.M. & Coverdale, J.A. (2001). Spatial localization of transplanted hemopoietic stem cells: inferences for the localization of stem cell niches. Blood 97, 2293–2299.Google Scholar
Nilsson, S.K., Johnston, H.M., Whitty, G.A., Williams, B., Webb, R.J., Denhardt, D.T., Bertoncello, I., Bendall, L.J., Simmons, P.J. & Haylock, D.N. (2005). Osteopontin, a key component of the hematopoietic stem cell niche and regulator of primitive hematopoietic progenitor cells. Blood 106(4), 1232–1239.Google Scholar
Okada, S., Nakauchi, H., Nagayoshi, K., Nishikawa, S., Miura, Y. & Suda, T. (1992). In vivo and in vitro stem cell function of c-kit- and Sca-1-positive murine hematopoietic cells. Blood 80(12), 3044–3050.Google Scholar
Oprins, A., Geuze, H. & Slot, J. (1994). Cryosubstitution dehydration of aldehyde-fixed tissue—A favorable approach to quantitative immunocytochemistry. J Histochem Cytochem 42, 497–503.Google Scholar
Papayannopoulou, T. (2004). Current mechanistic scenarios in hematopoietic stem/progenitor cell mobilization. Blood 103(5), 1580–1585.Google Scholar
Pohl, K. & Stierhof, Y.-D. (1998). Action of gold chloride (“gold toning”) on silver-enhanced 1 nm gold markers. Microsc Res Tech 42(1), 59–65.Google Scholar
Robinson, J.M., Takizawa, T. & Vandré, D.D. (2000). Applications of gold cluster compounds in immunocytochemistry and correlative microscopy: Comparison with colloidal gold. J Microsc 199(3), 163–179.Google Scholar
Robinson, J.M. & Vandré, D.D. (1997). Efficient immunocytochemical labeling of leukocyte microtubules with fluoroNanogold: An important tool for correlative microscopy. J Histochem Cytochem 45(5), 631–642.Google Scholar
Roth, J. (1982). The preparation of A-gold complexes with 3 nm and 15 nm gold particles and their use in labelling multiple antigens on ultrathin sections. Histochem 14, 791–801.CrossRefGoogle Scholar
Sacchetti, B., Funari, A., Michienzi, S., Di Cesare, S., Piersanti, S., Saggio, I., Tagliafico, E., Ferrari, S., Robey, P.G., Riminucci, M. & Bianco, P. (2007). Self-renewing osteoprogenitors in bone marrow sinusoids can organize a hematopoietic microenvironment. Cell 131(2), 324–336.Google Scholar
Schofield, R. (1978). The relationship between the spleen colony-forming cell and the haemopoietic stem cell. Blood Cells 4, 7–25.Google ScholarPubMed
Shingu, K., Helfritz, A., Kuhlmann, S., Zielinska-Skowronek, M., Jacobs, R., Schmidt, R. & von Hörsten, S. (2002). Kinetics of the early recruitment of leukocyte subsets at the sites of tumor cells in the lungs: Natural killer (NK) cells rapidly attract monocytes but not lymphocytes in the surveillance of micrometastasis. Int J Cancer 99(1), 74–81.CrossRefGoogle Scholar
Shingu, K., Kruschinski, C., Luhrmann, A., Grote, K., Tschernig, T., von Horsten, S. & Pabst, R. (2003). Intratracheal macrophage-activating lipopeptide-2 reduces metastasis in the rat lung. Am J Respir Cell Mol Biol 28(3), 316–321.CrossRefGoogle ScholarPubMed
Shiozawa, Y., Havens, A.M., Pienta, K.J. & Taichman, R.S. (2008). The bone marrow niche: Habitat to hematopoietic and mesenchymal stem cells, and unwitting host to molecular parasites. Leukemia 22(5), 941–950.Google Scholar
Stier, S., Ko, Y., Forkert, R., Lutz, C., Neuhaus, T., Grunewald, E., Cheng, T., Dombkowski, D., Calvi, L.M., Rittling, S.R. & Scadden, D.T. (2005). Osteopontin is a hematopoietic stem cell niche component that negatively regulates stem cell pool size. J Exp Med 201(11), 1781–1791.Google Scholar
Stierhof, Y.-D., Schwatz, H., Durrenberger, M., Villiger, W. & Kellenberger, E. (1991). Yield of immunolabel compared to resin sections and thawed cryosections. In Colloidal Gold: Principles, Methods, and Applications, Hayat, M. (Ed.), pp. 88–115. San Diego, CA: Academic Press.Google Scholar
Stierhof, Y.-D., Schwarz, H. & Frank, H. (1986). Transverse sectioning of plastic-embedded immunolabeled cryosections: Morphology and permeability to protein A-colloidal gold complexes. Mol Struct Res 97, 187–196.Google Scholar
Sugiyama, T., Kohara, H., Noda, M. & Nagasawa, T. (2006). Maintenance of the hematopoietic stem cell pool by CXCL12-CXCR4 chemokine signaling in bone marrow stromal cell niches. Immunity 25(6), 977–988.Google Scholar
Sundström, G., Löfvenberg, E., Hassan, I. & Engström-Laurent, A. (2002). Localisation and distribution of hyaluronan in normal bone marrow matrix: A novel method to evaluate impending fibrosis? Eur J Haematol 68(4), 194–202.CrossRefGoogle ScholarPubMed
Suzuki, N., Ohneda, O., Minegishi, N., Nishikawa, M., Ohta, T., Takahashi, S., Engel, J.D. & Yamamoto, M. (2006). Combinatorial Gata2 and Sca1 expression defines hematopoietic stem cells in the bone marrow niche. Proc Natl Acad Sci 103(7), 2202–2207.CrossRefGoogle ScholarPubMed
Takizawa, T. & Robinson, J. (1994). Use of 1.4-nm immunogold particles for immunocytochemistry on ultra-thin cryosections. J Histochem Cytochem 42, 1615–1623.Google Scholar
Takizawa, T. & Robinson, J.M. (2000). Analysis of antiphotobleaching reagents for use with fluoroNanogold in correlative microscopy. J Histochem Cytochem 48(3), 433–436.Google Scholar
van den Pol, A. (1986). Tyrosine hydroxylase immunoreactive neurons throughout the hypothalamus receive glutamate decarboxylase immunoreactive synapses: A double pre-embedding immunocytochemical study with particulate silver and HRP. J Neurosci 6(3), 877–891.Google Scholar
Vandré, D. & Burry, R. (1992). Immunoelectron microscopic localization of phosphoproteins associated with the mitotic spindle. J Histochem Cytochem 40(12), 1837–1847.CrossRefGoogle ScholarPubMed
Visnjic, D., Kalajzic, Z., Rowe, D.W., Katavic, V., Lorenzo, J. & Aguila, H.L. (2004). Hematopoiesis is severely altered in mice with an induced osteoblast deficiency. Blood 103(9), 3258–3264.Google Scholar
von Horsten, S., Helfritza, A., Kuhlmanna, S., Navea, H., Tschernig, T., Pabst, R., Ben-Eliyahu, S., Meyer, M., Schmidt, R. & Schmitz, C. (2000). Stereological quantification of carboxyfluorescein-labeled rat lung metastasis: A new method for the assessment of natural killer cell activity and tumor adhesion in vivo and in situ. J Immunol Methods 239, 25–34.Google Scholar
Wang, X.-Q., Duan, X.-M., Liu, L.-H., Fang, Y.-Q. & Tan, Y. (2005). Carboxyfluorescein diacetate succinimidyl ester fluorescent dye for cell labeling. Acta Biochimica et Biophysica Sinica 37(6), 379–385.Google Scholar
Weiss, L. (1976). The hematopoietic microenvironment of the bone marrow: An ultrastructural study of the stroma in rats. Anat Rec 186, 161–184.Google Scholar
Weiss, L. & Geduldig, U. (1991). Barrier cells: Stromal regulation of hematopoiesis and blood cell release in normal and stressed murine bone marrow. Blood 78, 975–990.Google Scholar
Wilson, A., Oser, G.M., Jaworski, M., Blanco-Bose, W.E., Laurenti, E., Adolphe, C., Essers, M.A., MacDonald, H.R. & Trumpp, A. (2007). Dormant and self-renewing hematopoietic stem cells and their niches. Ann NY Acad Sci 1106, 64–75.Google Scholar
Xie, Y., Yin, T., Wiegraebe, W., He, X.C., Miller, D., Stark, D., Perko, K., Alexander, R., Schwartz, J., Grindley, J.C., Park, J., Haug, J.S., Wunderlich, J.P., Li, H., Zhang, S., Johnson, T., Feldman, R.A. & Li, L. (2009). Detection of functional haematopoietic stem cell niche using real-time imaging. Nature 457(7225), 97–101.Google Scholar
Young, B., Lowe, J.S., Stevens, A. & Heath, J.W. (2007). Blood. In Wheater's Functional Histology. A Text and Colour Atlas, Ozols, I. (Ed.), pp. 46–64. Philadelphia, PA: Churchill Livingstone Elsevier.Google Scholar
Zhang, J., Niu, C., Ye, L., Huang, H., He, X., Tong, W.G., Ross, J., Haug, J., Johnson, T., Feng, J.Q., Harris, S., Wiedemann, L.M., Mishina, Y. & Li, L. (2003). Identification of the haematopoietic stem cell niche and control of the niche size. Nature 425(6960), 836–841.CrossRefGoogle ScholarPubMed