Effect of in vitro passage of healthy human gingival fibroblasts on cellular morphology and cytokine expression.

Cytokines have been implicated in the regulation of antibody and inflammatory responses, but their role in periodontal diseases has not been elucidated. In the present study, cytokine production by human gingival fibroblasts (HGF) following in vitro passage was assessed in order to determine the basal levels of cytokine message and protein and to determine if the cellular morphology and the profile of cytokines produced differed with passage. The HGF cell line F-CL was established by explantation from non-inflamed gingival tissue, and cells from passages 1-10 were studied. The number of cells was determined in confluent cultures and cell morphology was examined by light microscopy. Fibroblasts from confluent cultures were examined for cytokine mRNA by reverse transcription-polymerase chain reaction and culture supernatants were assessed for cytokines by enzyme-linked immunosorbent assay. The morphology of F-CL fibroblasts in passages 1-4 was normal, while fibroblasts in passages 5-10 were larger. In general, the number of cells decreased from early to late passage. Fibroblasts from passages 1-10 contained message for interleukin-1 beta, -6 and -8, but not for interleukin-1 alpha or tumour necrosis factor-alpha. Interleukin-6 was detected in culture supernatants of F-CL fibroblasts by the enzyme immunoassay and its level decreased with increasing passage.