Culture of primary human gingival fibroblasts on biodegradable membranes

Abstract

Repair and regeneration of periodontal tissues by tissue engineering is dependent on the use of biodegradable polymer scaffolds which serve as a carrier for cells or bioactive substances. There is a need to understand how a specific biomaterial may influence gene expression. The aim of this investigation was to develop and to optimize an in vitro technique for the adhearance and proliferation of primary human gingiva cells on implantable and biodegradable matrices. Square pieces of Bio-Gide ® matrix (BG) and slices of Ethisorb ® tamponade (ET) were coated with poly- l-lactide. The stability of coated and uncoated scaffolds was investigated by incubation in standard culture medium. Various concentrations of the cells were seeded onto coated and uncoated polymer matrices in tissue culture dishes without shaking (“static seeding”) or continuous shaking (“agitated seeding”). Cultures were grown for 4 week and were then evaluated by light and scanning electron microscopy. After a culture period of 10 d, BG-carriers showed a delicate consistency which made histological processing difficult. Cells were grown only sparsely in coated and non-coated BG-scaffolds. Contrary, ET-specimens were stable during a 4 week culture period. After “static seeding” a significantly higher number of cells resulted in comparison to those in “agitated” cultures. The cells were evenly distributed throughout the ET-carriers and produced extracellular matrix compounds as well. Furthermore, the examination with RT-PCR (reverse transcription-polymerase chain reaction) revealed that the cells synthesized and secreted type I collagen, and expressed genes implicated in transducing bone morphogenetic protein (BMP) signals. Messenger RNAs for BMP-2, -4, -7, the BMP type I receptors Act R-I (alk 2, activin-like kinase receptor), BMPR-IA (alk 3), -IB (alk 6), and the type II receptor BMPR-II were detected. These data reveal that static seeding favors the adherence and proliferation of primary gingival cells on polyglactin matrices. This system may serve as a valuable tool for periodontal tissue engineering.