title: Three-Dimensional Coculture Model to Analyze the Cross Talk Between Endothelial and Smooth Muscle Cells.
abstract: The response of blood vessels to physiological and pathological stimuli partly depends on the cross talk between endothelial cells (EC) lining the luminal side and smooth muscle cells (SMC) building the inner part of the vascular wall. Thus, the in vitro analysis of the pathophysiology of blood vessels requires coculture systems of EC and SMC. We have developed and validated a modified three-dimensional sandwich coculture (3D SW-CC) of EC and SMC using open μ-Slides with a thin glass bottom allowing direct imaging. The culture dish comprises an intermediate plate to minimize the meniscus resulting in homogenous cell distribution. Human umbilical artery SMC were sandwiched between coatings of rat tail collagen I. Following SMC quiescence, human umbilical vein EC were seeded on top of SMC and cultivated until confluence. By day 7, EC had formed a confluent monolayer and continuous vascular endothelial (VE)-cadherin-positive cell/cell contacts. Below, spindle-shaped SMC had formed parallel bundles and showed increased calponin expression compared to day 1. EC and SMC were interspaced by a matrix consisting of laminin, collagen IV, and perlecan. Basal messenger RNA (mRNA) expression levels of E-selectin, angiopoietin-1, calponin, and intercellular adhesion molecule 1 (ICAM-1) of the 3D SW-CC was comparable to that of a freshly isolated mouse inferior vena cava. Addition of tumor necrosis factor alpha (TNF α) to the 3D SW-CC induced E-selectin and ICAM-1 mRNA and protein induction, comparable to the EC and SMC monolayers. In contrast, the addition of activated platelets induced a significantly delayed but more pronounced activation in the 3D SW-CC compared to EC and SMC monolayers. Thus, this 3D SW-CC permits analyzing the cross talk between EC and SMC that mediate cellular quiescence as well as the response to complex activation signals.
authors.affiliation: 1 Skin and Endothelium Research Division (SERD), Department of Dermatology, Medical University of Vienna , Vienna, Austria .
authors.forename: Minu Karthika
authors.initials: MK
authors.lastname: Ganesan
authors.affiliation: 1 Skin and Endothelium Research Division (SERD), Department of Dermatology, Medical University of Vienna , Vienna, Austria .
authors.forename: Richard
authors.initials: R
authors.lastname: Finsterwalder
authors.affiliation: 1 Skin and Endothelium Research Division (SERD), Department of Dermatology, Medical University of Vienna , Vienna, Austria .
authors.forename: Heide
authors.initials: H
authors.lastname: Leb
authors.affiliation: 2 Department of Vascular Biology and Thrombosis Research, Medical University of Vienna , Vienna, Austria .
authors.forename: Ulrike
authors.initials: U
authors.lastname: Resch
authors.affiliation: 1 Skin and Endothelium Research Division (SERD), Department of Dermatology, Medical University of Vienna , Vienna, Austria .
authors.forename: Karin
authors.initials: K
authors.lastname: Neumüller
authors.affiliation: 2 Department of Vascular Biology and Thrombosis Research, Medical University of Vienna , Vienna, Austria .
authors.forename: Rainer
authors.initials: R
authors.lastname: de Martin
authors.affiliation: 1 Skin and Endothelium Research Division (SERD), Department of Dermatology, Medical University of Vienna , Vienna, Austria .
authors.forename: Peter
authors.initials: P
authors.lastname: Petzelbauer
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D002450:Cell Communication; D002454:Cell Differentiation; D002455:Cell Division; D002478:Cells, Cultured; D018920:Coculture Techniques; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D008954:Models, Biological; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D014469:Umbilical Arteries / Q000166:cytology / Q000378:metabolism*; D014471:Umbilical Veins / Q000166:cytology / Q000378:metabolism*
chemical_list: D015415:Biomarkers
keywords: 3D model; coculture; cross talk; endothelial cells; extracellular matrix; platelets; smooth muscle cells