Strain-dependent activation of monocytes and inflammatory macrophages by lipopolysaccharide of Porphyromonas gingivalis - PubMed (original) (raw)

Comparative Study

Strain-dependent activation of monocytes and inflammatory macrophages by lipopolysaccharide of Porphyromonas gingivalis

L Shapira et al. Infect Immun. 1998 Jun.

Abstract

Porphyromonas gingivalis is one of the pathogens associated with periodontal diseases, and its lipopolysaccharide (LPS) has been suggested as a possible virulence factor, acting by stimulation of host cells to secrete proinflammatory mediators. However, recent studies have shown that P. gingivalis LPS inhibited some components of the inflammatory response. The present study was designed to test the hypothesis that there are strain-dependent variations in the ability of P. gingivalis LPS to elicit the host inflammatory response. By using LPS preparations from two strains of P. gingivalis, W50 and A7346, the responses of mouse macrophages and human monocytes were evaluated by measuring the secretion of nitric oxide (NO) and tumor necrosis factor alpha (TNF-alpha). Both direct and indirect (priming) effects were investigated. LPS from Salmonella typhosa was used as a reference LPS. P. gingivalis A7436 LPS induced lower secreted levels of NO from the tested cells than S. typhosa LPS but induced similar levels of TNF-alpha. In contrast, LPS from P. gingivalis W50 did not induce NO or TNF-alpha secretion. Preincubation of macrophages with LPS from S. typhosa or P. gingivalis A7436 prior to stimulation with S. typhosa LPS upregulated NO secretion and downregulated TNF-alpha secretion, while preincubation with P. gingivalis W50 LPS enhanced both TNF-alpha and NO secretory responses. These results demonstrate that LPSs derived from different strains of P. gingivalis vary in their biological activities in vitro. The findings may have an impact on our understanding of the range of P. gingivalis virulence in vivo.

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Figures

FIG. 1

SDS-PAGE analysis of LPSs from P. gingivalis A7436 (lane a), P. gingivalis W50 (lane b), S. typhosa (lane c), and E. coli 055:B5 (lane d). Positions of molecular mass standards (in kilodaltons) are indicated on the left.

FIG. 2

Nitrite levels in culture supernatants of macrophages stimulated with LPS isolated from P. gingivalis A7436 or W50 or S. typhosa (S.t.). Thioglycolate-elicited macrophages were cultured with 1 μg of LPS per ml for 24 or 48 h. Culture supernatants were harvested, and nitrite levels were determined by using Gries reagent. Data is presented as means ± standard deviations for four replicates.

FIG. 3

Dose-dependent release of nitrite by LPS-stimulated macrophages. Thioglycolate-elicited macrophages were cultured with increasing doses of LPS, isolated from P. gingivalis A7436 or W50 or S. typhosa (S.t.), for 24 h. Culture supernatants were harvested, and nitrite levels were determined by using Gries reagent. Data is presented as means ± standard deviations for four replicates.

FIG. 4

Nitrite secretion by macrophages stimulated with S. typhosa (S.t.) LPS (1 μg/ml). Thioglycolate-elicited macrophages were cultured with medium alone, S. typhosa LPS, S. typhosa LPS plus P. gingivalis A7436 LPS, or S. typhosa LPS plus P. gingivalis W50 LPS for 24 h. Culture supernatants were harvested, and nitrite levels were determined by using Gries reagent. Data is presented as means ± standard deviations for four replicates.

FIG. 5

Activation of NOS by LPS of S. typhosa (S.t.), P. gingivalis (Pg) A7436, or P. gingivalis W50. Macrophages were cultured in the presence or absence (Non) of LPS (1 μg/ml) for 18 h. Cells were harvested, and cell lysates were analyzed for NOS activity as described in Materials and Methods. Results are for pooled samples from six different wells.

FIG. 6

Dose-dependent secretion of TNF-α by LPS-stimulated macrophages. Macrophages were cultured with increasing doses of S. typhosa (S.t.), P. gingivalis A7436, or P. gingivalis W50 LPS for 24 h. Culture supernatants were harvested, and TNF-α levels were determined by ELISA. Data is presented as means ± standard deviations for four replicates.

FIG. 7

Secretion of TNF-α by LPS-stimulated human monocytes. Human monocytes were cultured with S. typhosa (S.t.), E. coli, P. gingivalis (P.g.) A7436, or P. gingivalis W50 LPS (1 μg/ml) for 24 h. Culture supernatants were harvested, and TNF-α levels were determined by ELISA. Data is presented as means ± standard deviations for four replicates.

FIG. 8

Effect of pretreatment of macrophages with different LPSs on their LPS-induced nitrite production. Macrophages were preexposed to increasing doses of S. typhosa (S.t.), P. gingivalis A7436, or P. gingivalis W50 LPS for 24 h. Cultures were washed, and fresh S. typhosa LPS (1 μg/ml) was added. After an additional 24 h, culture supernatants were harvested and nitrite levels were determined by using Gries reagent. Data is presented as means ± standard deviations for four replicates.

FIG. 9

Effect of pretreatment of macrophages with different LPS preparations on their LPS-induced TNF-α production. Macrophages were preexposed to increasing doses of S. typhosa (S.t.), P. gingivalis A7436, or P. gingivalis W50 LPS for 24 h. Culture wells were washed, and fresh S. typhosa LPS (1 μg/ml) was added. After an additional 24 h, culture supernatants were harvested and TNF-α levels were determined by ELISA. Data is presented as means ± standard deviations for four replicates.

FIG. 10

Effect of pretreatment of human monocytes with different LPS preparations on their LPS-induced TNF-α production. Human monocytes were preexposed to medium alone, S. typhosa (S.t.), P. gingivalis (P.g.) A7436, or P. gingivalis W50 LPS (1 μg/ml) for 24 h. Cultures were washed, and fresh S. typhosa LPS (1 μg/ml) was added. After an additional 24 h, culture supernatants were harvested and TNF-α levels were determined by ELISA. Data is presented as means ± standard deviations for four replicates.

References

1. Aleo J J. Stimulation of macromolecular synthesis by endotoxin-treated 3T6 fibroblasts. Experimentia. 1980;36:546–547. - PubMed
1. Aleo J J, DeRenzis F A, Farber P A, Varboncoeur A P. The presence and the biologic activity of cementum bound endotoxin. J Periodontol. 1974;45:672–675. - PubMed
1. Amar S, Van Dyke T E, Eugster H P, Schultze N, Koebel P, Bluethmann H. Tumor necrosis factor (TNF)-induced cutaneous necrosis is mediated by TNF receptor 1. J Inflammation. 1996;47:180–189. - PubMed
1. Bramanti T E, Wong G G, Weinstraub S T, Holt S C. Chemical characterization and biologic properties of lipopolysaccharides from Bacteroides gingivalis strains W50, W83 and ATCC 33277. Oral Microbiol Immunol. 1989;4:183–192. - PubMed
1. Champagne C M, Holt S C, Van Dyke T E, Gordon B J, Shapira L. Lipopolysaccharide isolated from Porphyromonas gingivalis grown in hemin-limited conditions has a reduced capacity for human neutrophil priming. Oral Microbiol Immunol. 1996;5:319–325. - PubMed

Strain-dependent activation of monocytes and inflammatory macrophages by lipopolysaccharide of Porphyromonas gingivalis - PubMed (original) (raw)