Influence of donor age and culture conditions on tissue engineering of mucosa autografts.

In oral surgery the transplantation of tissue engineered mucosa is used more frequently. The conventional single cell suspension culture method (SCSM) involves murine feeder cells and foetal calf serum. The explant technique (ET) has been used as alternative culture procedure. Aim was to study the efficacy of the ET and the SCSM without feeder cells to grow primary cultures and to test the effects of donor age, of extracellular matrix proteins (ECMP), and of autogenous serum on cell growth in explant cultures. These factors were assessed in cultures of 58 patients overall. In 48 cultures of 12 patients primary cell growth was compared between the ET and the SCSM. Eighteen of 24 cultures were established with the ET whereas only 3 of 24 were established with the SCSM. To test the influence of donor age on cell multiplication, the proliferation rate (DNA synthesis measured by bromodeoxyuridine uptake) and the overall growth (DNA content) was determined in cultures of five young and five old donors. In cultures from old donors (mean age 56 years) proliferation was lower but more sustained relative to the cultures from the young donors (mean age 25 years). In old donors overall in vitro cell growth was only 2/3 of that in young donors. In cultures of 20 donors the influence on cell adhesion and growth of the ECMP fibronectin and laminin was assessed by planimetry. While ECMP augmented explant adhesion, these substances did not enhance keratinocyte growth significantly. Comparing the influence of autogenous and foetal calf serum on cell growth no differences were observed in all cultures of the six donors. In conclusion, the ET without additional ECMP coating and with autogenous instead of foetal calf serum are now used to culture gingival keratinocytes for tissue engineering mucosa grafts. Consequently xenogenous components are avoided, being a considerable advantage.