Characterization of fibroblast morphology on bioactive surfaces using vertical scanning interferometry Christopher M. Revell a , Jeffrey A. Dietrich a , C. Corey Scott a,b , Andreas Luttge c , L. Scott Baggett d , Kyriacos A. Athanasiou a, ⁎ a Department of Bioengineering, Rice University, 6100 Main Street, Keck Hall, Suite 116, Houston, Texas 77005, United States b MSTP, Baylor College of Medicine, One Baylor Plaza, N201, Houston, Texas 77030, United States c Department of Earth Science, Rice University, 6100 Main Street, Keith-Weiss Geology Lab, Room 321, Houston, Texas 77005, United States d Jones Graduate School of Management, Rice University, 6100 Main Street, Jones Graduate School, Room 248, Houston, Texas 77005, United States Received 7 July 2005; received in revised form 19 July 2006; accepted 20 July 2006 Abstract Tissue donor scarcity is a major hindrance to articular cartilage tissue engineering. Previous research shows that dermal fibroblasts express chondrocytic markers after seeding on aggrecan-coated surfaces. Since cell roundness appears to correlate with chondrocytic behavior of dermal fibroblasts, this study quantified roundness by measuring cell height and surface area–volume ratio. In addition to aggrecan as a surface coating, collagen type II and decorin, two other major extracellular matrix components of articular cartilage, were examined. Aggrecan, collagen type II, and decorin were coated onto a glass substrate using three application techniques: static drying, airbrush, and painting. Vertical scanning interferometry (VSI) is a novel technique that allows for the expedient morphological determination of single cells. Interferometry was used for the characterization of protein-coated surfaces in addition to characterizing the morphology of single dermal fibroblasts after 24 h of seeding. Fibroblast height was found to vary from 1.0 to 4.0 μm and protein coating, application technique, and seeding position were significant factors (p b 0.002). The largest cell heights were observed on aggrecan and collagen type II coated surfaces using the air brush and static applications. Additionally, variations were observed for surface area–volume ratio, ranging from 1.75 to 11.94 μm - 1 with decorin resulting in the lowest ratio, followed by collagen type II and aggrecan. This study identifies optimal coating conditions for stimulating morphology in dermal fibroblasts that is characteristic of the chondrocytic phenotype. These conditions can be employed to attempt articular cartilage regeneration and bypass difficulties due to a paucity of donor tissue. © 2006 Elsevier B.V./International Society of Matrix Biology. All rights reserved. Keywords: Vertical scanning interferometry; Cell morphology; Cartilage matrix proteins; Fibroblasts; Surface modification 1. Introduction Shortage of tissue for clinical application of tissue engi- neering of articular cartilage presents a serious problem, as chondrocytes have a low capacity for proliferation and long-term culture causes a loss of functionality (Schnabel et al., 2002). Previous work has shown chondrocytes, in vivo, maintain a spherical morphology. These cells, when moved from the in vivo environment to an in vitro culture environment, begin to change both morphologically as well as phenotypically. It has been shown that chondrocytes, after only one or two passages in vitro, lose all of their phenotypic markers, mainly a severe down- regulation of collagen type II and aggrecan and a severe up- regulation of collagen type I (Darling and Athanasiou, 2005). One approach to address this dedifferentiation and tissue scarcity in the field of cartilage regeneration is focused on initiating phenotypic changes in cells from non-traditional tissues using protein-coated surfaces. Modification of surfaces is a widely employed technique for controlling cell attachment, prolifera- tion, and differentiation (Barbucci et al., 2005; Carlsson et al., 1979; McKeehan and Ham, 1976). One method utilizes protein coatings to modify the surface and investigate the ability of Matrix Biology 25 (2006) 523 – 533 www.elsevier.com/locate/matbio ⁎ Corresponding author. Tel.: +1 713 348 6385; fax: +1 713 348 5877. E-mail address: athanasiou@rice.edu (K.A. Athanasiou). 0945-053X/$ - see front matter © 2006 Elsevier B.V./International Society of Matrix Biology. All rights reserved. doi:10.1016/j.matbio.2006.07.007