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Published on: April 18, 2019
Antimicrobial and immunomodulatory in vitro profile of double antibiotic paste
Maurício Gonçalves da Costa Sousa1, Patrícia Diniz Xavier2, Ana Paula de Castro Cantuária3
1Pós-Graduação em Ciências Genômicas e Biotecnologia, Centro de Análises Proteômicas e Bioquímicas, Universidade Católica de Brasília, Brasília, Brazil.
Aim:
To evaluate the antimicrobial and immunomodulatory activity of double antibiotic paste (DAP) in an in vitro infection model.
Methodology:
The minimum inhibitory and bactericidal concentrations (MIC and MBC) and the antibiofilm activities (TTC assay) of DAP and its components (ciprofloxacin and metronidazole) were evaluated against Staphylococcus aureus and Enterococcus faecalis compared with triple antibiotic paste (TAP). The cellular viability of RAW 264.7 macrophages (24 and 72 h) and L929 fibroblasts (48 and 72 h) was evaluated by MTT. Furthermore, the production of TNF-α, IL-12, IL-6, IL-1α, IL-10 and NO (on RAW 264.7), besides IL-6, TGF-β and NO (on L929), stimulated with DAP in baseline and associated with heat-killed microbial-antigen conditions was measured by ELISA and Griess reaction. Data were analysed using the one-way ANOVA test with Bonferroni's corrections.
Results:
The MBC of pharmacopoeia DAP was similar to TAP for E. faecalis (0.25 μg. mL-1 ) and lower for S. aureus (DAP 1 μg. mL-1 and TAP 2 μg. mL-1 ; p < .001). Ciprofloxacin was the most effective antibiofilm drug from the pastes (35% of reduction for E. faecalis and S. aureus; p < .0001), and both pastes had a similar antibiofilm eradication against both biofilm species (29% and 35% for S. aureus and 76% and 85% for E. faecalis; p < .0001). DAP was cytotoxic against the tested cells. DAP significantly upregulated IL-1α (p < .001), IL-6 (p < .0001), TNF-α (p < .01) and IL-12 (p < .05; in the absence of antigens) and significantly reduced IL-6 (p < .0001; in the presence of HK-S. aureus) and IL-10 (p < .05; in the presence of both antigens) on macrophages. Furthermore, DAP upregulated IL-6 (p < .001) and NO (p < .05; in the absence of antigens), IL-6 (p < .001; in the presence of HK-S. aureus) and reduced NO (p < .001; in the presence of HK-S. aureus).
Conclusions:
Double antibiotic paste and TAP had similar antimicrobial activity against S. aureus and E. faecalis. DAP upregulated pro-inflammatory cytokines mainly in the absence of antigens and had pro- and anti-inflammatory activity in RAW 264.7 macrophages and L929 fibroblasts in the presence of antigens involved in pulp infections.
Insights
Double antibiotic paste (DAP) exhibits antimicrobial efficacy comparable to triple antibiotic paste (TAP) against common root canal bacteria. DAP demonstrates immunomodulatory effects, influencing cytokine production in immune cells during infection.
Area of Science:
- Endodontics
- Microbiology
- Immunology
Background:
- Double antibiotic paste (DAP) is utilized in endodontic treatments.
- Understanding its antimicrobial and immunomodulatory effects is crucial for optimizing treatment outcomes.
Purpose of the Study:
- To evaluate the in vitro antimicrobial and immunomodulatory activity of double antibiotic paste (DAP).
- To compare DAP with triple antibiotic paste (TAP) against Staphylococcus aureus and Enterococcus faecalis.
Main Methods:
- Minimum inhibitory and bactericidal concentrations (MIC/MBC) and antibiofilm activity assays were performed.
- Cellular viability of macrophages and fibroblasts was assessed using MTT assay.
- Cytokine and nitric oxide production was measured via ELISA and Griess reaction.
Main Results:
- DAP demonstrated comparable antimicrobial activity to TAP against E. faecalis and S. aureus.
- Ciprofloxacin within the pastes showed significant antibiofilm efficacy.
- DAP induced significant upregulation of pro-inflammatory cytokines (IL-1α, IL-6, TNF-α, IL-12) in macrophages, with varied effects in the presence of antigens.
Conclusions:
- DAP possesses similar antimicrobial activity to TAP against key endodontic pathogens.
- DAP exhibits complex immunomodulatory effects, acting as both a pro- and anti-inflammatory agent on immune cells in the context of pulp infection antigens.
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