Hostname: page-component-cd9895bd7-jkksz Total loading time: 0 Render date: 2024-12-23T06:16:19.290Z Has data issue: false hasContentIssue false
  • English
  • Français

Imaging of Vascular Smooth Muscle Cells with Soft X-Ray Spectromicroscopy

Published online by Cambridge University Press:  09 November 2011

Julia Sedlmair ,
Sophie-Charlotte Gleber ,
Semra Öztürk Mert ,
Michael Bertilson ,
Olov von Hofsten ,
Jürgen Thieme  and
Thomas Pfohl
Julia Sedlmair*
Affiliation:
Institute for X-Ray Physics, Georg-August-University Göttingen, Friedrich-Hund-Pl. 1, D-37077 Göttingen, Germany
Sophie-Charlotte Gleber
Affiliation:
Argonne National Laboratory, APS, 9700 S. Cass Avenue, Building 401, Argonne, IL 60439-4837, USA
Semra Öztürk Mert
Affiliation:
Max-Planck-Institute for Dynamics and Self-Organization, Bunsenstr. 10, D-37073, Göttingen, Germany
Michael Bertilson
Affiliation:
Biomedical and X-Ray Physics, Department of Applied Physics, Royal Institute of Technology, AlbaNova, SE-106 91 Stockholm, Sweden
Olov von Hofsten
Affiliation:
Biomedical and X-Ray Physics, Department of Applied Physics, Royal Institute of Technology, AlbaNova, SE-106 91 Stockholm, Sweden
Jürgen Thieme
Affiliation:
Brookhaven National Laboratory, NSLS II, Building 817, Upton, NY 11973, USA
Thomas Pfohl
Affiliation:
Max-Planck-Institute for Dynamics and Self-Organization, Bunsenstr. 10, D-37073, Göttingen, Germany Department of Chemistry, University of Basel, Klingelbergstr. 80, CH-4056, Basel, Switzerland
*
Corresponding author. E-mail: [email protected]
Get access

Abstract

Using X-ray microscopy and spectromicroscopy, vascular smooth muscle cells (VSMCs) were imaged, prepared without using additional embedding material or staining, but by applying simple, noncryo fixation techniques. The cells were imaged with a compact source transmission X-ray microscope and a scanning transmission X-ray microscope (STXM). With the STXM, spectromicroscopy was performed at the C K-edge and the Ca LIII,II-edges. VSMCs were chosen because of their high amount of actin stress fibers, so that the actin cytoskeleton should be visible. Other parts of the cell, such as the nucleus and organelles, were also identified from the micrographs. Both in the spectra and the images, the effects of the different preparation procedures were observable. Furthermore, Ca hotspots were detected and their density is determined.

Keywords

X-ray microscopyvascular smooth muscle cellsspectromicroscopyNEXAFScalciumradiation damage
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 17 , Issue 6 , December 2011 , pp. 991 - 1001
Copyright
Copyright © Microscopy Society of America 2011

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Bâldea, I., Schimmelpfennig, B., Plaschke, M., Rothe, J., Schirmer, J., Trofimov, A.B. & Fanghänel, T. (2007). C 1s near edge X-ray absorption fine structure (NEXAFS) of substituted benzoic acids—A theoretical and experimental study. J Electron Spectrosc 154, 109–118.CrossRefGoogle Scholar
Beetz, T. (2004). Soft X-ray diffraction imaging with and without lenses and radiation damage studies. PhD thesis. Stony Brook, NY: Stony Brook University.Google Scholar
Beetz, T. & Jacobsen, C. (2003). Soft X-ray radiation-damage studies in PMMA using a cryo-STXM. J Synchrotron Rad 10(3), 280–283.CrossRefGoogle ScholarPubMed
Benzerara, K., Yoon, T., Tyliszczak, T., Constantz, B., Spormann, A. & Brown, G.B. Jr. (2004). Scanning transmission X-ray microscopy study of micro-bial calcification. Geobiol 2(4), 249–259.CrossRefGoogle Scholar
Berglund, M., Rymell, L., Peuker, M., Wilhein, T. & Hertz, H.M. (2000). Compact water-window transmission X-ray microscopy. J Microsc 197(3), 268–273.CrossRefGoogle ScholarPubMed
Bertilson, M.C., von Hofsten, O., Thieme, J., Lindblom, M., Holmberg, A., Takman, P.A.C, Vogt, U. & Hertz, H.M. (2009). First application experiments with the Stockholm compact soft X-ray microscope. J Phys Conf Ser 186, 012025.CrossRefGoogle Scholar
Bertsch, P.M. & Hunter, D.B. (2001). Applications of synchrotron-based X-ray microprobes. Chem Rev 101, 1809–1842.CrossRefGoogle ScholarPubMed
Betzig, E., Patterson, G.H., Sougrat, R., Lindwasser, O.W., Olenych, S., Bonifacino, J.S., Davidson, M.W., Lippincott-Schwartz, J. & Hess, H.F. (2006). Imaging intracellular fluorescent proteins at nanometer resolution. Science 313(1642), 810–813.CrossRefGoogle ScholarPubMed
Buckley, C.J. (1995). Measuring and mapping in mineralized tissues by absorption difference imaging. Rev Sci Instrum 66(2), 1318–1321.CrossRefGoogle Scholar
Chao, W., Kim, J., Rakewa, S., Fischer, P. & Anderson, E.H. (2009). Demonstration of 12 nm resolution fresnel zone plate lens based soft X-ray microscopy. Opt Express 17(20), 17669–17677.CrossRefGoogle ScholarPubMed
Cheng, H. & Lederer, W. (2008). Calcium sparks. Physiol Rev 88(4), 1491–1545.CrossRefGoogle ScholarPubMed
CXRO (2011). X-Ray Interactions with Matter. Berkeley, CA: Center for X-Ray Optics, Lawrence Berkeley National Laboratory (retrieved Jan. 2011).Google Scholar
Egner, A. & Hell, S.W. (2005). Fluorescence microscopy with super-resolved optical sections. Trends Cell Biol 15(4), 207–215.CrossRefGoogle ScholarPubMed
Fleet, M.E. & Liu, X. (2009). Calcium L2, 3-edge XANES of carbonates, carbonate apatite, and oldhamite (CaS). Am Mineral 94(8-9), 1235–1241.CrossRefGoogle Scholar
Gleber, S.-C. (2009). Soft X-ray stereo microscopy for investigation of dynamics and elemental distribution of colloidal systems from the environment. PhD thesis. Gottingen, Germany: Georg-August-University.Google Scholar
Greco, T., Hodara, R., Parastatidis, I., Heijnen, H., Dennehy, M., Liebler, D. & Ischiropoulos, H. (2006). Identification of s-nitrosylation motifs by site-specific mapping of the s-nitrosocysteine proteome in human vascular smooth muscle cells. Proc Nat Acad Sci 103(19), 7420–7425.CrossRefGoogle ScholarPubMed
Guttmann, P., Heim, S., Werner, S., Niemann, B., Follath, R. & Schneider, G. (2007). X-ray microscopy at the new U41-FSGM beam line. BESSY—Annual Report 2007, pp. 297–300.Google Scholar
Hitchcock, A., Morin, C, Zhang, X. & Araki, T. (2005). Soft X-ray spectromicroscopy of biological and synthetic polymer systems. J Electron Spectrosc 144–147, 259–269.CrossRefGoogle Scholar
Hitchcock, A.P., Dynes, J.J., Lawrence, J.R., Obst, M., Swerhone, G.D.W., Korber, D.R. & Leppard, G.G. (2009). Soft X-ray spectromicroscopy of nickel sorption in a natural river biofilm. Geobiol 7(4), 432–453.CrossRefGoogle Scholar
Huang, B., Wang, W., Bates, M. & Zhuang, X. (2008). Three-dimensional super-resolution imaging by stochastic optical reconstruction microscopy. Science 319(5864), 810–813.CrossRefGoogle ScholarPubMed
Huang, X., Nelson, J., Kirz, J., Lima, E., Marchesini, S., Miao, H., Neiman, A., Shapiro, D., Steinbrenner, J., Stewart, A., Turner, J. & Jacobsen, C. (2009). Soft X-ray diffraction microscopy of a frozen hydrated yeast cell. Phys Rev Lett 103(19), 198101.CrossRefGoogle ScholarPubMed
Jacobsen, C, Wirick, S., Flynn, G. & Zimba, C. (2000). Soft X-ray spec-troscopy from image sequences with sub-100 nm spatial resolution. J Microsc 197(2), 173–184.CrossRefGoogle Scholar
Janiak, R., Wilson, S., Montague, S. & Hume, J. (2001). Heterogeneity of calcium stores and elementary release events in canine pulmonary arterial smooth muscle cells. Am J Physiol 280(1), C22–C33.CrossRefGoogle ScholarPubMed
Kirz, J., Jacobsen, C. & Howells, M. (2009). Soft X-ray microscopes and their biological applications. Q Rev Biophys 28(1), 33–130 (reprint of 1995).CrossRefGoogle Scholar
Kolczewski, C., Püttner, R., Martins, M., Schlachter, A.S., Snell, G., Santanna, M.M., Hermann, K. & Kaindl, G. (2006). Spectroscopic analysis of small organic molecules: A comprehensive near-edge X-ray-absorption fine-structure study of C6-ring-containing molecules. J Chem Phys 124(3), 034302.CrossRefGoogle ScholarPubMed
Lange, K. & Gartzke, J. (2006). F-actin-based Ca signaling—A critical comparison with the current concept of Ca signaling. J Cell Physiol 209(2), 270–287.CrossRefGoogle ScholarPubMed
Larabell, C.A. & Le Gros, M.A. (2004). X-ray tomography generates 3-D reconstructions of the yeast, Saccharomyces cerevisiae, at 60 nm resolution. Mol Biol Cell 15(3), 957–962.CrossRefGoogle ScholarPubMed
Lawrence, J., Swerhone, G., Leppard, G., Araki, T., Zhang, X., West, M. & Hitchcock, A.P. (2003). Scanning transmission X-ray, laser scanning, and transmission electron microscopy mapping of the exopolymeric matrix of microbial biofilms. Appl Environ Microbiol 69(9), 5543.CrossRefGoogle ScholarPubMed
Lerotic, M., Jacobsen, C., Schäfer, T. & Vogt, S. (2004). Cluster analysis of soft X-ray spectromicroscopy data. Ultramicroscopy 100, 35–57.CrossRefGoogle ScholarPubMed
McDermott, G., Le Gros, M.A., Knoechel, C.G., Uchida, M. & Larabell, C.A. (2009). Soft X-ray tomography and cryogenic light microscopy: The cool combination in cellular imaging. Trends Cell Biol 19(11), 587–595.CrossRefGoogle ScholarPubMed
McGeown, G. (2010). Gl brown lecture 2010: Seeing is believing: Imaging ca2+-signalling events in living cells. Exp Physiol 95(11), 1049–1060.CrossRefGoogle Scholar
Meyer-Ilse, W., Hamamoto, D., Nair, A., Leliévre, S.A., Denbeaux, G., Johnson, L., Pearson, A.L., Yager, D., Le Gros, M.A. & Larabell, C.A. (2001). High resolution protein localization using soft X-ray microscopy. J Microsc 201(3), 395–403.CrossRefGoogle ScholarPubMed
Mitrea, G., Thieme, J., Guttmann, P., Heim, S. & Gleber, S. (2008). X-ray spectromicroscopy with the scanning transmission X-ray microscope at BESSY II. J Synchrotron Rad 15(1), 26–35.CrossRefGoogle ScholarPubMed
Neu, T.R., Manz, B., Volke, F., Dynes, J.J., Hitchcock, A.P. & Lawrence, J.R. (2010). Advanced imaging techniques for assessment of structure, composition and function in biofilm systems. FEMS Microbiol Ecol 72, 1–21.CrossRefGoogle ScholarPubMed
Norlund, K.L.I., Southam, G., Tyliszcak, T., Hu, Y., Karunkaran, C., Obst, M., Hitchcock, A.P. & Warren, L.A. (2009). Microbial architecture of environmental sulfur processes: A novel syntrophic sulfur-metabolizing consortia. Org Geochem 43(23), 8781–8786.Google ScholarPubMed
Parker, I., Choi, J. & Yao, Y. (1996). Elementary events of insp3-induced ca2+ liberation in xenopus oocytes: Hot spots, puffs and blips. Cell Calcium 20(2), 105–121.CrossRefGoogle ScholarPubMed
Parkinson, D.Y., McDermott, G., Etkin, L.D., Le Gros, M.A. & Larabell, C.A. (2008). Quantitative 3-D imaging of eukaryotic cells using soft X-ray tomography. J Struct Biol 162(3), 380–386.CrossRefGoogle ScholarPubMed
Resch, G.P., Goldie, K.N., Krebs, A., Hoenger, A. & Small, J.V. (2002). Visualisation of the actin cytoskeleton by cryo-electron microscopy. J Cell Sci 115(9), 1877–1882.CrossRefGoogle ScholarPubMed
Rust, M.J., Bates, M. & Zhuang, X. (2006). Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM). Nat Methods 3(10), 793–796.CrossRefGoogle ScholarPubMed
Schneider, G., Guttmann, P., Heim, S., Rehbein, S., Mueller, F., Nagashima, K., Heymann, J.B., Müller, W.G. & McNally, J. (2010). Three-dimensional cellular ultrastructure resolved by X-ray microscopy. Nat Methods 7(12), 985–987.CrossRefGoogle ScholarPubMed
Small, J.V., Auinger, S., Nemethova, M., Koestler, S., Goldie, K.N., Hoenger, A. & Resch, G.P. (2008). Unravelling the structure of the lamellipodium. J Microsc 231(3), 479–485.CrossRefGoogle ScholarPubMed
Smith, A.P., Urquhart, S.G., Winesett, D.A., Mitchell, G. & Ade, H. (2001). Use of near edge X-ray absorption fine structure spectromicroscopy to characterize multicomponent polymeric systems. Appl Spectrosc 55(12), 1676–1681.CrossRefGoogle Scholar
Solomon, D., Lehmann, J., Kinyangi, J., Liang, B., Heymann, K., Dathe, L., Hanley, K., Wirick, S. & Jacobsen, C. (2009). Carbon (1s) NEXAFS spectroscopy of biogeochemically relevant reference organic compounds. Soil Sci Soc Am J 73(6), 1817.CrossRefGoogle Scholar
Song, C., Jiang, H., Mancuso, A., Amirbekian, B., Peng, L., Sun, R., Shah, S.S., Zhou, Z.H., Ishikawa, T. & Miao, J. (2008). Quantitative imaging of single, unstained viruses with coherent X rays. Phys Rev Lett 101(8), 158101.CrossRefGoogle ScholarPubMed
Stöhr, J. (1992). NEXAFS Spectroscopy. Berlin, Germany: Springer.CrossRefGoogle Scholar
Studer, D., Humbel, B. & Chiquet, M. (2008). Electron microscopy of high pressure frozen samples: Bridging the gap between cellular ultrastructure and atomic resolution. Histochem Cell Biol 130(5), 877–889.CrossRefGoogle Scholar
Takman, P.A.C., Stollberg, H., Johansson, G., Holmberg, A., Lindblom, M. & Hertz, H.M. (2007). High-resolution compact X-ray microscopy. J Microsc 226(2), 175–181.CrossRefGoogle ScholarPubMed
Thieme, J., Sedlmair, J., Gleber, S.-C, Prietzel, J., Coates, J., Eusterhues, K., Abbt-Braun, G. & Salome, M. (2010). X-ray spectromicroscopy in soil and environmental sciences. J Synchrotron Rad 17(2), 149–157.CrossRefGoogle ScholarPubMed
Thompson, A.C, Attwood, D.T., Gullikson, E.M., Howells, M.R., Kim, K.J., Kirz, J., Kortright, J.B., Lindau, I., Pianatta, P. & Robinson, A.L. (2001). X-ray Data Booklet, 2nd ed., Thompson, A.C. & Vaughan, D. (Eds.). Berkeley, CA: Lawrence Berkeley National Laboratory.Google Scholar
Thyberg, J. (2000). Differences in caveolae dynamics in vascular smooth muscle cells of different phenotypes. Lab Invest 80(6), 915–929.CrossRefGoogle ScholarPubMed
Wang, J., Morin, C., Li, L., Hitchcock, A.P., Scholl, A. & Doran, A. (2009). Radiation damage in soft X-ray microscopy. J Electron Spectrosc 170(1-3), 25–36.CrossRefGoogle Scholar
Wiesemann, U. (2003). The scanning transmission X-ray microscope at BESSY II. PhD thesis. Göttingen, Germany: Institute for X-Ray Physics, University of Göttingen.Google Scholar
Wiesemann, U., Thieme, J., Früke, R. & Guttmann, P. (2001). Construction of a scanning transmission X-ray microscope at the undulator U-41 at BESSY II. Nucl Instrum Methods A 467–468, 861–863.CrossRefGoogle Scholar
Willig, K., Rizzoli, S., Westphal, V., Jahn, R. & Hell, S. (2006). STED microscopy reveals that synaptotagmin remains clustered after synaptic vesicle exocytosis. Nature 440, 935–939.CrossRefGoogle ScholarPubMed
Yamamoto, A., Masaki, R., Guttmann, P., Schmahl, G. & Kihara, H. (1998). Studies on intracellular structures of COS cells by X-ray microscopy. J Synchrotron Rad 5(3), 1105–1107.CrossRefGoogle ScholarPubMed
Zenisek, D., Davila, V., Wan, L. & Almers, W. (2003). Imaging calcium entry sites and ribbon structures in two presynaptic cells. J Neurosc 23(7), 2538–2548.CrossRefGoogle ScholarPubMed