Cytokines have been implicated in regulating antibody and inflammatory responses; however, their role in periodontal disease has not been elucidated. In the present study, we have assessed cytokine production by human gingival fibroblast (HGF) following in vitro passage, in order to determine if the morphology and profile of cytokines produced differed based on the number of passages. HGF cell lines were established by the explant method using non-inflamed gingival tissue and cell from passage 1-10 were studies. The number of cells were determined in confluent cultures and cell morphology was examined by light microscopy. Fibroblasts from confluent cultures were examined for cytokine mRNA by RT-PCR; culture supernatants (CS) were assessed for cytokines by ELISA. Our results showed that cells from passage 1- 4 had normal fibroblast morphology, while cells in passage 5-10 were larger in size. In general the number of cells decreased from early to late passage. Fibroblasts from passages 1-10 contained message for IL-6, but not for IL-1a, IL-1b, and TNF-a, IL-6 was detected in CS and the level decreased with increasing passage. IL-1a was present in CS from passage 2, 4 and 5, while TNF-a was detected in CS from passages 1, 2, 3, 6, 9, and 10. Our results indicated that the regulation of cytokine production, especially IL-6, by fibroblasts changes with in vitro passage.
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