Osteogenic Differentiation of Human Gingival Fibroblasts

Objectives: Periodontitis results in bone loss and eventual tooth loss. The current available options for bone regeneration include guided tissue regeneration and the use of bioactive agents. These methods can effectively regenerate bone in small periodontal defects. However, larger periodontal defects are still challenging to treat. Hence, the use of human gingival fibroblasts (HGFs) as a cell-based therapy has been suggested as an alternative treatment option. HGFs are readily available with minimal donor site morbidity and may be ideal for tissue engineering efforts in regenerating lost alveolar bone. We hypothesized that osteogenic induction of HGFs would promote bone regeneration. Therefore, the aim of this study was to investigate the extent of osteogenic differentiation of HGFs in vitro. Our long-term aim is to determine the osteogenic potential for HGFs to develop a cell-based therapy for bone regeneration in periodontitis patients.

Methods: HGF strains isolated from six different donors were exposed to osteogenic medium for four weeks (basic medium, 50mg/mL ascorbic acid, 100nM dexamethasone, 100nM vitamin D3, and 10mM beta-glycerophosphate). Osteogenesis was evaluated based on Von Kossa staining, scanning electron microscope (SEM) and energy-dispersive X-ray (EDX) after four weeks.

Results: Based on Von Kossa staining, cultures were stratified into categories based on the magnitude of calcification: no calcification (0%), poor calcification (<10%), moderate calcification (10-40%), and significant calcification (>50%). There was no calcification in the controls cultured in non-osteogenic medium. Four cell strains showed significant calcification, one moderate calcification and one showed no calcification with osteogenic treatment. Calcified cultures showed diffuse or patterned calcifications. SEM and EDX mineral distribution confirmed the presence of calcium (Ca) and phosphate (P) in the calcifying nodules with Ca/P ratio ranging between 2 and 2.5.

Conclusions: Under appropriate osteogenic conditioning, HGFs may provide a source for cell-based therapy for bone regeneration in periodontitis patients.