Expansion of chondrocytes in a three-dimensional matrix for tracheal tissue engineering.
BACKGROUND
The generation of autologous tracheal implants by tissue-engineering techniques is a promising concept for otherwise untreatable patients. A functional cartilaginous backbone represents a prerequisite for any bioartificial tracheal graft. The aim of this study was to define suitable cell types and culture conditions for the generation of tracheal cartilage.


METHODS
We obtained tracheal, costal, and auricular cartilage from porcine donor animals (n = 10). The chondrocytes were cultured two-dimensionally in cell flasks or mixed with a liquid collagen solution forming a three-dimensional culture system. Labeling with carboxy fluorescein diacetate succinimidyl ester (CFDA SE) and biochemical reduction of formazan served to determine cell viability and proliferation. The extracellular matrix produced by the chondrocytes was characterized by Western blot.


RESULTS
The CFDA SE labeling proved viability and the MTT assays documented a proliferation of the chondrocytes over time in vitro. While the chondrocytes in the three-dimensional cell culture system produced hyaline cartilage composed of collagen II, the two-dimensional culture conditions resulted in nonspecific collagen synthesis.


CONCLUSIONS
Chondrocytes grown in a three-dimensional matrix can effectively proliferate and produce cartilage and are viable for more than 2 weeks. Costal chondrocytes are suitable for tracheal cartilage tissue engineering.
Department of Thoracic and Vascular Surgery, Heidehaus Hospital, Am Leineufer 70, D-30149 Hannover, Germany. twalles@yahoo.com
Thorsten
T
Walles
Bettina
B
Giere
Paolo
P
Macchiarini
Heike
H
Mertsching
D000818:Animals; D015153:Blotting, Western; D002356:Cartilage / Q000166:cytology; D002455:Cell Division; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D024043:Collagen Type II / Q000096:biosynthesis; D003470:Culture Media; D004425:Ear Cartilage / Q000166:cytology; D005109:Extracellular Matrix / Q000737:chemistry; D005452:Fluoresceins / Q000032:analysis; D005456:Fluorescent Dyes / Q000032:analysis; D007583:Jejunum / Q000737:chemistry; D009928:Organ Specificity; D012272:Ribs; D034421:Sus scrofa; D023822:Tissue Engineering / Q000295:instrumentation* / Q000379:methods; D014132:Trachea / Q000166:cytology
D024043:Collagen Type II; D003470:Culture Media; D005452:Fluoresceins; D005456:Fluorescent Dyes; C084577:5-carboxyfluorescein diacetate