Hindawi Publishing Corporation Journal of Biomedicine and Biotechnology Volume 2011, Article ID 528276, 8 pages doi:10.1155/2011/528276 Research Article Colocalization of Serum Amyloid A with Microtubules in Human Coronary Artery Endothelial Cells Katja Lakota, 1 Nataˇ sa Resnik, 2 Katjuˇ sa Mrak-Poljˇ sak, 1 Sneˇ zna Sodin- ˇ Semrl, 1 and Peter Veraniˇ c 2 1 Department of Rheumatology, University Medical Centre Ljubljana, Ljubljana, Slovenia 2 Institute of Cell Biology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia Correspondence should be addressed to Peter Veraniˇ c, peter.veranic@mf.uni-lj.si Received 2 June 2011; Accepted 6 July 2011 Academic Editor: Beric Henderson Copyright © 2011 Katja Lakota et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Serum amyloid A (SAA) acts as a major acute phase protein and represents a sensitive and accurate marker of inflammation. Besides its hepatic origin, as the main source of serum SAA, this protein is also produced extrahepatically. The mRNA levels of SAA become significantly elevated following proinflammatory stimuli, as well as, are induced through their own positive feedback in human primary coronary artery endothelial cells. However, the intracellular functions of SAA are so far unknown. Colocalization of SAA with cytoskeletal filaments has previously been proposed, so we analyzed the colocalization of SAA with all three cytoskeletal elements: actin filaments, vimentin filaments, and microtubules. Immunofluorescent double-labeling analyses confirmed by PLA method revealed a strict colocalization of SAA with microtubules and a very infrequent attachment to vimentin while the distribution of actin filaments appeared clearly separated from SAA staining. Also, no significant colocalization was found between SAA and endomembranes labeled with the fluorescent lipid stain DiO 6 . However, SAA appears to be located also unbound in the cytosol, as well as inside the nucleus and within nanotubes extending from the cells or bridging neighboring cells. These different locations of SAA in endothelial cells strongly indicate multiple potential functions of this protein. 1. Introduction The acute phase response represents an evolutionarily con- served mechanism of inflammatory events designed to rapidly react to infections, wounds, and injuries. It can lead to a dramatic increase (up to 1000 fold) in the levels of acute phase proteins (APPs) in the circulation and, ultimately, brings about resolution of the inflammatory reaction [1, 2]. Serum amyloid A (SAA) one of the major APPs in humans is mainly produced by the liver, although extrahepatic synthesis is also prevalent [3]. SAA originates from an evolutionarily conserved multi- gene family [4] ranging from invertebrates (with a wound- healing role in sea cucumbers [5]), vertebrates, to humans, where it represents an accurate and sensitive marker of infla- mmation [2]. Human SAA1 and SAA2 are the inducible iso- types (addressed jointly as SAA1/2, with over 95% sequence identity); SAA3, was thought in the past to be a pseudogene, not expressed in humans; however, recently there has been a report of its mammary-associated expression found in milk [6]. SAA4 was found to be the constitutively expressed iso- type [4]. There have been three acute phase SAA isotypes reported in the mouse SAA1, SAA2, and SAA3, with SAA3 being the primarily extrahepatic isoform [7]. SAA is a small protein (104 amino acids in length), 11.7 kDa in size, lipo- philic, and poorly soluble in aqueous solution, originally described as a component of normal serum [8]. SAA frag- ments were found in amyloidosis, and the accumulation of these fibrils can lead to organ failure and ultimately death [9]. Murine and human SAA have been shown to form hexamers in solution, which can lead to the formation of membrane channels that could be involved in important pathological roles [10, 11]. SAA1/2 has a variety of multiple functions in humans, among them it acts as a cytokine and chemokine, induces matrix metalloproteinases, interferes with platelet functions, replaces Apo-A1 in high density lipoprotein particles in the circulation during inflammation, binds cholesterol and