Filamentous Actin Disruption and Diminished Inositol Phosphate Response in Gingival Fibroblasts Caused by Treponema denticola

Abstract

Previous reports have shown that Treponema denticolacauses rearrangement of filamentous actin (F-actin) in human gingival fibroblasts (HGF). The purpose of this investigation was to determine the effect of T. denticola on the generation of inositol phosphates (IPs) in relation to a time course for F-actin disruption in HGF. Cultured HGF were exposed to washed cells of T. denticola ATCC 35405 for 140 min. Changes in the fluorescence intensity of rhodamine-phalloidin-labeled F-actin in serial optical sections of single HGF were quantified by confocal microscopy image analysis. The percentage of cells with stress fiber disruption was also determined by fluorescence microscopy. Challenge with T. denticola caused a significant reduction in F-actin within the first hour, especially at the expense of F-actin in the ventral third of the cells, and a significant increase in the percentage of HGF with altered stress fiber patterns. Significant concentration-dependent disruption of stress fibers was also caused by HGF exposure to a Triton X-100 extract of T. denticola outer membrane (OM). IPs were measured by a radiotracer assay based on the incorporation ofmyo-[3H]inositol into IPs in HGF incubated with LiCl to inhibit endogenous phosphatases. HGF challenge with several strains of T. denticola and the OM extract ofT. denticola ATCC 35405 resulted in a diminished accumulation of radiolabeled IPs relative to both 15 and 1% fetal bovine serum, which served as strongly positive and background control agonists, respectively. The significantly diminished IP response toT. denticola ATCC 35405 occurred within 60 min, concomitant with significant reduction of total F-actin and disruption of stress fibers. Pretreatment with the proteinase inhibitor phenylmethylsulfonyl fluoride, which had previously been found to block T. denticola’s degradation of endogenous fibronectin and detachment of HGF from the extracellular matrix, had little effect on F-actin stress fiber disruption and the IP response. Therefore, in addition to its major surface chymotrypsin-like properties, T. denticola expresses cytopathogenic activities that diminish the generation of IPs during the time course associated with significant cytoskeletal disruption in fibroblasts.

Spirochetes at the forefront of periodontal infections

“…Exposure to T. denticola induces cytoskeletal reorganization and coincident cell shrinkage, membrane blebbing, and loss of volume regulation in host cells (6,27,174,175,182). The major outer sheath protein antigen (Msp) of T. denticola is one of the bettercharacterized virulence determinants of oral treponemes that may mediate some of these effects.…”

The major outer sheath protein of Treponema denticola inhibits the binding step of collagen phagocytosis in fibroblasts

“…Treponema denticola type strain ATCC 35405 was grown in modified NOS medium (Dawson and Ellen, 1990). This strain induces actin filament reorganization in human gingival fibroblasts (HGFs) (Yang et al, 1998). Enriched Msp complex was prepared as described previously (Egli et al, 1993) with modifications (Wang et al, 2001).…”

Polymerase chain reaction detection of Treponema denticola in endodontic infections within root canals

“…1996, Fenno & McBride 1998. Adhesion to host cells causes filamentous actin disruption and diminished inositol phosphate response (Yang et al . 1998).…”