Atividade antimicrobiana, antibiofilme e citotóxica de soluções de nanopartículas de prata e farnesol para desinfecção de canais radiculares
Carregando...
Data
2016-09-19
Autores
Chávez-Andrade, Gisselle Moraima [UNESP]
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
Nanopartículas de prata (NPsAg) apresentam ação bactericida e biocompatibilidade, mostrando potencial para aplicações na Odontologia. Farnesol (FAR) é um álcool encontrado em óleos essencias de frutas cítricas e apresenta atividade antibacteriana/antibiofilme. O objetivo deste estudo foi avaliar a atividade antimicrobiana, antibiofilme e a citotoxicidade de soluções de NPsAg e FAR, para serem utilizadas na desinfecção de canais radiculares. O estudo foi dividido em duas publicações. Na Publicação 1, a atividade antimicrobiana foi avaliada sobre Enterococcus faecalis, Candida albicans ou Pseudomonas aeruginosa, por meio da determinação da concentração inibitória mínima (CIM) e microbicida mínima (CMM) pelo método de microdiluição e coloração com resazurina. A avaliação da atividade sobre a biomassa dos biofilmes foi realizada por meio do ensaio de cristal violeta. A capacidade anti-adesão de biofilme foi avaliada em blocos de dentina radicular bovina após tratamento da dentina com as soluções por 3 min. Foram realizadas análises em microscopia eletrônica de varredura (MEV) e contagem de UFC mL-1 log 10. Os dados foram submetidos à análise estatística (α=0,05). No teste antibacteriano sobre células planctônicas de E. faecalis, os valores de CIM/CMM das soluções de NPsAg e FAR foram de 42,5/50μM e 0,85/1,0%, respectivamente. Para C. albicans, os valores de CIM/CMM de NPsAg e FAR foram de 27,5/37,5μM e 1,75/2,5%, respectivamente. Para P. aeruginosa, os valores de CIM/CMM de NPsAg e FAR foram de 32,5/32,5μM e 2,5/2,75%, respectivamente. Ambas as soluções mostraram atividade sobre a biomassa dos biofilmes, porém foram menos efetivas do que NaOCl. Na avaliação da capacidade anti-adesão de biofilme, NPsAg e FAR mostraram menor quantidade de biofilme aderido ao substrato quando comparados ao controle (SS-salina), porém sem diferença entre as duas soluções (p>0,05), exceto sobre C. albicans. Na contagem de UFC mL-1, pode-se observar que não houve diferença entre as soluções de NPsAg e FAR (p>0,05), sendo similares ao controle (SS), para as 3 cepas avaliadas. Na Publicação 2, foi avaliada a citotoxicidade e genotoxicidade das soluções em cultura de fibroblastos L929, por meio dos ensaios de MTT e Cometa, respectivamente. Para avaliar ex vivo a efetividade antibacteriana, foram realizados diferentes protocolos de irrigação na etapa final do preparo biomecânico em dentes humanos extraídos contaminados com E. faecalis por 21 dias. Após 3 coletas microbiológicas (inicial, pós-irrigação e final), foi realizada a contagem de UFC mL-1 em cada grupo. Os dados obtidos foram analisados estatísticamente (α=0,05). No ensaio MTT, NPsAg e FAR foram menos citotóxicos do que NaOCl. No ensaio Cometa, ambas as soluções mostraram menor % de DNA na cauda, em comparação ao controle positivo (H2O2). No ensaio antibacteriano, todas os protocolos de irrigação foram mais efetivos que o C+ (sem irrigação), com exceção de SS/FAR. NaOCl e sua combinação com NPsAg e FAR levaram a completa eliminação bacteriana (p>0,05). Em comparação à coleta inicial, tanto a coleta pós-irrigação quanto final, apresentaram redução microbiana significativa (p<0,05). NPsAg e FAR mostraram baixa citotoxicidade e genotoxicidade, contribuem com a redução de E. faecalis e apresentam potencial para uso em protocolos de irrigação final do SCR.
Silver nanoparticles (AgNPs) have potential for applications in dentistry due to bactericidal action and biocompatibility. Farnesol (FAR) is an alcohol found in essential oils of citrus fruits and it has antibacterial/antibiofilm activity. The aim of this study was to evaluate the antimicrobial, antibiofilm activities and the cytotoxicity of AgNPs and FAR solutions used as root canal disinfectant. The study was divided into two publications. In Publication 1, antimicrobial activity was evaluated against Enterococcus faecalis, Candida albicans or Pseudomonas aeruginosa, by determining the minimum inhibitory concentration (MIC) and the minimum microbicidal concentration (MMC) by microdilution method and rezasurin staining. The anti-biofilm activity was performed by crystal violet assay. The anti-biofilm adhesion ability was evaluated using bovine root dentine blocks after treating them with tested solutions for 3 min. Scanning electron microscopy (SEM) analysis and CFU mL-1 log 10 counting were performed. Data were statistically analyzed (α=0.05). The antibacterial test against planktonic cells of E. faecalis showed MIC/MMC values of 42.5/50μM and 0.85/1.0% for AgNPs and FAR, respectively. The values of MIC/MMC of AgNPs and FAR for C. albicans were 27.5/37.5μM and 1.75/2.5% and for P. aeruginosa were 32.5/32.5μM and 2.5/2.75%, respectively. The anti-adhesion biofilm capacity assay showed smaller amount of biofilm adhered to the substrate treated with AgNPs and FAR when compared to control group (saline solution - SS), but there was no statistical difference between the two solutions (p>0.05), except for C. albicans. The log 10 UFC mL-1 counting showed no statistical difference between AgNPs and FAR solutions (p>0.05), which were similar to the control group (SS) for all strains. In publication 2, the cytotoxicity and genotoxicity of the solutions were evaluated in culture cell of fibroblast L929 by MTT and Comet assays, respectively. To evaluate the ex vivo antibacterial effectiveness, different irrigation protocols were performed in the final stage of root canal instrumentation on extracted human teeth infected with E. faecalis for 21 days. The number of log 10 CFU mL-1 in each group was determined in microbiological samples collected at different periods (initial, post-irrigation and final). Data were statistically analyzed (α=0.05). In the MTT assay, AgNPs and FAR were less cytotoxic than NaOCl. In the Comet assay, both solutions showed lower % of DNA in the tail, compared to positive control (H2O2). In the antimicrobial assay, all the irrigating protocols were more effective than C+ (without irrigation), with exception of SS/FAR. NaOCl and their combinations with AgNPs or FAR led to complete bacterial elimination (p>0.05). In comparison with initial sample, both the post-irrigation and final samples showed significant microbial reduction (p<0.05). AgNPs-PVA and FAR showed low cytotoxicity and genotoxicity, contribute to E. faecalis reduction and exhibit potential for use as a final irrigation protocols of the RCS.
Silver nanoparticles (AgNPs) have potential for applications in dentistry due to bactericidal action and biocompatibility. Farnesol (FAR) is an alcohol found in essential oils of citrus fruits and it has antibacterial/antibiofilm activity. The aim of this study was to evaluate the antimicrobial, antibiofilm activities and the cytotoxicity of AgNPs and FAR solutions used as root canal disinfectant. The study was divided into two publications. In Publication 1, antimicrobial activity was evaluated against Enterococcus faecalis, Candida albicans or Pseudomonas aeruginosa, by determining the minimum inhibitory concentration (MIC) and the minimum microbicidal concentration (MMC) by microdilution method and rezasurin staining. The anti-biofilm activity was performed by crystal violet assay. The anti-biofilm adhesion ability was evaluated using bovine root dentine blocks after treating them with tested solutions for 3 min. Scanning electron microscopy (SEM) analysis and CFU mL-1 log 10 counting were performed. Data were statistically analyzed (α=0.05). The antibacterial test against planktonic cells of E. faecalis showed MIC/MMC values of 42.5/50μM and 0.85/1.0% for AgNPs and FAR, respectively. The values of MIC/MMC of AgNPs and FAR for C. albicans were 27.5/37.5μM and 1.75/2.5% and for P. aeruginosa were 32.5/32.5μM and 2.5/2.75%, respectively. The anti-adhesion biofilm capacity assay showed smaller amount of biofilm adhered to the substrate treated with AgNPs and FAR when compared to control group (saline solution - SS), but there was no statistical difference between the two solutions (p>0.05), except for C. albicans. The log 10 UFC mL-1 counting showed no statistical difference between AgNPs and FAR solutions (p>0.05), which were similar to the control group (SS) for all strains. In publication 2, the cytotoxicity and genotoxicity of the solutions were evaluated in culture cell of fibroblast L929 by MTT and Comet assays, respectively. To evaluate the ex vivo antibacterial effectiveness, different irrigation protocols were performed in the final stage of root canal instrumentation on extracted human teeth infected with E. faecalis for 21 days. The number of log 10 CFU mL-1 in each group was determined in microbiological samples collected at different periods (initial, post-irrigation and final). Data were statistically analyzed (α=0.05). In the MTT assay, AgNPs and FAR were less cytotoxic than NaOCl. In the Comet assay, both solutions showed lower % of DNA in the tail, compared to positive control (H2O2). In the antimicrobial assay, all the irrigating protocols were more effective than C+ (without irrigation), with exception of SS/FAR. NaOCl and their combinations with AgNPs or FAR led to complete bacterial elimination (p>0.05). In comparison with initial sample, both the post-irrigation and final samples showed significant microbial reduction (p<0.05). AgNPs-PVA and FAR showed low cytotoxicity and genotoxicity, contribute to E. faecalis reduction and exhibit potential for use as a final irrigation protocols of the RCS.
Descrição
Palavras-chave
Preparo de canal radicular, Produtos com ação antimicrobiana, Técnicas de cultura de células, Root canal preparation, Products with antimicrobial action, Cell culture techniques