Phenotypic analysis of bovine chondrocytes cultured in 3D collagen sponges: effect of serum substitutes
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Springer 2005
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Phenotypic analysis of bovine chondrocytes cultured in 3D collagen sponges: effect of serum substitutes Karen E. Yates1,2, Florin Allemann1,2 and Julie Glowacki1,2,* 1
Department of Orthopedic Surgery, Brigham and Women’s Hospital, Harvard Medical School, 75 Francis Street, Boston, MA 02115, USA; 2Skeletal Biology Research Center, Massachusetts General Hospital, Harvard School of Dental Medicine, Boston MA, USA; *Author for correspondence (e-mail: jglowacki @rics.bwh.harvard.edu; phone: +1-617-732-5397; fax:+1-617-732-6937) Received 24 April 2004; accepted in revised form 28 October 2004
Key words: 3D scaffolds, Chondrocytes, ITS, Nutridoma, Serum substitutes, Tissue engineering
Abstract Repair of damaged cartilage usually requires replacement tissue or substitute material. Tissue engineering is a promising means to produce replacement cartilage from autologous or allogeneic cell sources. Scaffolds provide a three-dimensional (3D) structure that is essential for chondrocyte function and synthesis of cartilage-specific matrix proteins (collagen type II, aggrecan) and sulfated proteoglycans. In this study, we assessed porous, 3D collagen sponges for in vitro engineering of cartilage in both standard and serum-free culture conditions. Bovine articular chondrocytes (bACs) cultured in 3D sponges accumulated and maintained cartilage matrix over 4 weeks, as assessed by quantitative measures of matrix content, synthesis, and gene expression. Chondrogenesis by bACs cultured with Nutridoma as a serum replacement was equivalent or better than control cultures in serum. In contrast, chondrogenesis in insulin-transferrinselenium (ITS+3) serum replacement cultures was poor, apparently due to decreased cell survival. These data indicate that porous 3D collagen sponges maintain chondrocyte viability, shape, and synthetic activity by providing an environment favorable for high-density chondrogenesis. With quantitative assays for cartilage-specific gene expression and biochemical measures of chondrogenesis in these studies, we conclude that the collagen sponges have potential as a scaffold for cartilage tissue engineering.
Abbreviations: 2D – two-dimensional; 3D – three-dimensional; AGG – aggrecan; bACs – bovine articular chondrocytes; COL I – type I collagen; COL II – type II collagen; FBS – fetal bovine serum; G3PDH – glyceraldehyde-3-phosphate dehydrogenase; HEPES – 4-(2-hydroxyethyl)piperazine-1-ethansulfonic acid; ITS – insulin-transferrin-sodium selenite; Nut – nutridoma; OD – optical density; RT-PCR – reverse transcriptase-polymerase chain reaction; s-GAG – sulfated glycosaminoglycan
Introduction Cartilage has a poor capacity for healing and regeneration, and therefore most defects require replacement tissue or substitute material. In
mosaicplasty, cores of healthy tissues from nonload-bearing surfaces of the joint are transplanted into the defect (Hangody and Fules 2003). In another procedure, cells are isolated from healthy cartilage, expanded in vitro, and transplanted as a
46 suspension into the defec
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