Oral bacteria can have effects on fibroblast behavior and motility which are likely mediated by cytoskeletal alterations. Objectives: To examine the effects of Actinobacillus actinomycetemcomitans (Aa) on cytoskeletal organization and in vitro wound healing of human gingival fibroblasts (HGF).
Methods: Sonicated extracts were obtained of Aa (strains 1169 and Y4) after culture under both aerobic and anaerobic conditions in BHI medium. Primary cultures of HGF were then exposed to DMEM alone (control) or a 1:5 dilution of bacterial extracts for 45 minutes. Cytoskeletal organization was examined by immunofluorescent labeling of F-actin, vinculin, and tubulin. Cytoskeletal changes were correlated with previous work in which wounds were created in confluent monolayers of HGF, and repopulation of the wound space was assessed by recording the number of cells and distance traveled from the wound edge.
Results: All forms of Aa extracts impaired HGF wound healing. Aerobic Aa (1169) reduced mean distance migrated from 4.85 to 2.55mm and cells/field from 182.5 to 52.5, whereas anaerobic Aa (1169) reduced distance migrated from 4.34 to 1.8mm and cells/field from 167 to 14.93. Aerobic Aa (Y4) reduced distance migrated from 6.38mm to 1.7mm and cells/field from 374.43 to 77.07, and with anaerobic Aa (Y4) distance migrated was 1.45mm and cells/field was 43.1. The HGF cytoskeleton was characterized by well organized actin stress fibers, a normal microtubular network, and punctate vinculin labeling. However there were no observable differences in cytoskeletal organization in response to any of the bacterial extracts.
Conclusions: HGF wound healing was significantly inhibited by Aa (1169 and Y4) under both aerobic and anaerobic culture conditions. However, cytoskeletal organization was not visibly altered. Further studies are needed to understand how oral bacteria influence fibroblast behavior during wound healing. Supported by NIH/NCRR P20RR018741 and the J. Dean Robertson Society.