Prostaglandin E₂ secretion from gingival fibroblasts treated with interleukin‐1β: effects of lipid extracts from Porphyromonas gingivalis or calculus

Abstract

Complex lipids of Porphyromonas gingivalis have been identified in lipid extracts from calculus-contaminated root surfaces and in diseased gingival tissues. However, little is known about the biological effects of these complex lipids on host cells. The purpose of this study was to evaluate the effects of P. gingivalis or calculus lipids on prostaglandin secretion from gingival fibroblasts. Lipids were extracted from paired subgingival plaque and teeth samples, and calculus-contaminated root surfaces before and after scaling and root planing, in order to determine the relevant levels of lipid extracts for the treatment of gingival fibroblasts in culture. Primary cultures of gingival fibroblasts were exposed to lipid extracts from either P. gingivalis or calculus/teeth for a period of 7 days. Control and lipid-treated cultures were exposed to human recombinant interleukin-1beta for 48 h and prostaglandin secretion from interleukin-1beta-treated fibroblasts was compared with control and lipid-treated fibroblasts without interleukin-1beta treatment. These experiments demonstrated that P. gingivalis lipids or calculus-tooth lipids potentiate interleukin-1beta-mediated prostaglandin secretory responses from gingival fibroblasts. Additionally, P. gingivalis or calculus-tooth lipid extracts were readily taken up by gingival fibroblasts as measured by bacterial fatty acid recovery in lipid extracts of cultured fibroblasts. These results indicate that bacterial lipid penetration into gingival tissues in combination with a chronic inflammatory response may substantially potentiate prostaglandin secretion from gingival fibroblasts, thereby promoting tissue destructive processes associated with adult periodontitis.

Structures and biological activities of novel phosphatidylethanolamine lipids of Porphyromonas gingivalis

“…The cultures were then incubated for an additional 24 h at 378C in a humidified 5% CO 2 incubator, after which the medium samples were harvested and frozen. Prostaglandin F 2a (PGF 2a ), 6-keto PGF 1a , and PGE 2 were quantified in medium samples using stable isotope dilution GC-MS analysis, as described previously (1,9). Each histogram bar depicts the mean 6 SD for two trials.…”

“…Effects of P. gingivalis phosphatidylethanolamine lipids on interleukin-1 (IL-1)-mediated prostaglandin secretion from gingival fibroblasts. P. gingivalis phosphatidylethanolamine lipids (10 mg in 20 ml of 100% ethanol) or dimyristoyl phosphatidylethanolamine lipid standard (Avanti Polar Lipids, Inc.) were dispersed into 35 mm plastic culture dishes and air-dried, as described previously (1,9). Gingival fibroblasts were grown, harvested, and treated with lipids as described (1,9).…”

“…P. gingivalis phosphatidylethanolamine lipids (10 mg in 20 ml of 100% ethanol) or dimyristoyl phosphatidylethanolamine lipid standard (Avanti Polar Lipids, Inc.) were dispersed into 35 mm plastic culture dishes and air-dried, as described previously (1,9). Gingival fibroblasts were grown, harvested, and treated with lipids as described (1,9). Approximately 2 3 10 6 cells were deposited into each culture well suspended in Minimum Essential Medium supplemented with 10% fetal bovine serum and 100 U/ml penicillin G, 100 mg/ml streptomycin, and 0.25 mg/ml amphotericin B.…”

“…Several plausible mechanisms could account for periodontal tissue contamination with lipids of P. gingivalis, including direct adhesion between bacteria and host epithelial cells in the gingival sulcus surrounding each tooth (20)(21)(22)(23), direct contact between host tissues and lipid-contaminated diseased teeth, or invasion of periodontal tissues by P. gingivalis (24)(25)(26)(27)(28)(29)(30)(31). Because complex lipids of P. gingivalis are most prevalent on periodontally diseased tooth roots (3,9), this study exposed gingival fibroblasts to lipid films that recreate the approximate lipid levels observed on calculus-contaminated tooth roots. A previous report indirectly estimated P. gingivalis lipid levels on calculus-contaminated tooth roots by quantifying 3-OH iso C 17:0 on root sections of known surface area (9).…”

“…Quantification of prostaglandins in culture medium samples PGF 2a , 6-keto PGF 1a , and PGE 2 were quantified in medium samples using a modification of the method of Luderer, Riley, and Demers (8), as described previously (1,9). Each medium sample was supplemented with 100 ng of D 4 -PGF 2a , D 4 -6-keto PGF 1a , and D 4 -PGE 2 and the pH adjusted to pH 3.5 with concentrated formic acid.…”