Journal of Structural Biology 152 (2005) 52–63 www.elsevier.com/locate/yjsbi 1047-8477/$ - see front matter  2005 Elsevier Inc. All rights reserved. doi:10.1016/j.jsb.2005.07.009 Investigating native coronary artery endothelium in situ and in cell culture by scanning force microscopy Tobias Reichlin a , Andreas Wild a , Markus Dürrenberger b , A.U. Daniels c , Ueli Aebi a,¤ , Patrick R. Hunziker d , Martin Stolz a a M.E. Müller Institute for Structural Biology, Biozentrum University of Basel, Switzerland b Center for Microscopy, University of Basel, Switzerland c Laboratory for Orthopedic Biomechanics, Felix Platter Hospital, University of Basel, Switzerland d University Hospital of Basel, Medical Intensive Care Unit, University of Basel, Switzerland Received 15 April 2005; received in revised form 9 July 2005; accepted 14 July 2005 Available online 26 August 2005 Abstract The purpose of our studies is to better understand the morphology and functioning of the arteries and their changes in pathogen- esis. The most frequently used imaging techniques are intravascular ultrasound, magnetic resonance imaging, and optical coherence tomography. These methods do not image cell-level structural details and only provide biomechanical properties indirectly. We pres- ent a new protocol for imaging the endothelial surface and measuring elastic properties of vascular tissue by scanning force micros- copy. Full-thickness sections of native pig coronary arteries were prepared. In addition, cultured human umbilical vein endothelial cells were studied as an in vitro model system and for comparison. We encountered a variety of diYculties mostly due to the softness of vascular tissue which required signiWcant adaptations of standard equipment: (i) a new specimen holder designed to stably immo- bilize the coronary arteries; (ii) a phase-contrast microscope incorporated for assessing the status of the cultured endothelial cells and positioning the scanning force microscope (SFM) tip at a site of interest; and (iii) a continuous exchange of the culture medium at 37 °C to assure viability of the cells in the SFM over extended times. We were thus able to investigate both fresh arterial tissue and living endothelial cells in a near-physiological environment. We present initial SFM images of vascular tissue at a spatial resolution similar to scanning electron microscopy, but which also provide a closer view of the bona Wde structure of native tissue. Novel mor- phological features such as distinct granular particles were observed. Moreover, we report initial measurements of vascular tissue surface stiVness, obtained by indentation-type SFM.  2005 Elsevier Inc. All rights reserved. Keywords: Atherosclerosis; Coronary disease; Vulnerable plaques; Soft biological tissues; Endothelial cells; Indentation-type SFM; AFM 1. Introduction Morphologically, the artery walls consist of three concentric layers called intima (the blood-contacting lumen surface), media and adventitia. In normal arteries, the intima consists chieXy of endothelial cells with mini- mal underlying subendothelial connective tissue. Depending on the local mechanical and metabolic needs, the media is composed predominantly of smooth muscle cells alternating with a varying fraction of elastic Wbers. The adventitia is a layer of investing connective tissue. While the media is responsible for modulation of vessel compliance, the intima forms the interface between the artery wall and the blood stream. Therefore, one of the main functions of the vascular endothelium is to main- tain circulatory function and vessel wall homeostasis. There is strong evidence that signaling between the endothelial cell layer and the supporting tissue is of * Corresponding author. Fax: +41 61 267 2109. E-mail address: Ueli.Aebi@unibas.ch (U. Aebi).