Statins and oral biofilm: Simvastatin as a promising drug to control periodontal dysbiosis.

OBJECTIVES: This study evaluated antimicrobial activity of atorvastatin, pravastatin, rosuvastatin, and simvastatin against oral bacteria, and the interaction of simvastatin with standard antimicrobials (amoxicillin and metronidazole). METHODS: Minimal inhibitory concentration assays were performed with Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, Actinomyces odontolyticus, Streptococcus oralis, Streptococcus mitis, Streptococcus salivarius, Streptococcus sanguinis, and Streptococcus gordonii; checkerboard microdilution assays between simvastatin and standard antimicrobials; monospecies and multispecies biofilms. RESULTS: Simvastatin showed the best antimicrobial activity against most species (MIC range from 3.12 to 25 µg/ml), highlighting the sensitivity of P. gingivalis. In the checkerboard assay, synergistic interaction was found between simvastatin and amoxicillin against S. oralis and S. sanguinis. P. gingivalis biofilm was inhibited by simvastatin at 10 and 50× Minimal inhibitory concentration, with similar effects to metronidazole. For multispecies biofilm, SMV reduced the biofilm metabolic activity (79%) and total counts (87%), comparable to amoxicillin. Simvastatin also reduced bacterial counts of Veilonnella parvula, P. gingivalis, Streptococcus mutans, Actinomyces naeslundii, P. intermedia, and Capnocytophaga ochracea in the multispecies biofilm. CONCLUSIONS: Simvastatin showed antimicrobial and antibiofilm activity against oral bacteria and may contribute to the control of dysbiosis, and may be considered in clinical studies as an adjuvant in the treatment of periodontitis.

Quantification and gene expression of Lactobacillus casei group species associated with dentinal lesions in early childhood caries.

BACKGROUND: Considering that the Lactobacillus casei group is strongly associated with caries progression, the use of lactobacilli as probiotics must be balanced due to their possible involvement in dental caries. OBJECTIVE: This study aimed to detect and quantify L. paracasei, L. rhamnosus, and L. casei group species in the active and arrested dentinal lesions of preschoolers. It also aimed to determine the expression profiles of lactobacilli genes related to adhesion, extracellular polymeric substance regulation, and pyruvate oxidation. METHODS: Total ribonucleic acid (RNA) was extracted from dentinal lesion samples (25 active, 13 arrested) of children between 2 and 5 years of age. The samples were converted to complementary deoxyribonucleic acid (cDNA), and quantitative polymerase chain reaction (qPCR) analyses were performed to quantify and determine the relative abundance (measured by percentage of total bacteria) of L. paracasei, L. rhamnosus, and L. casei group species. The expression profiles of L. paracasei/casei genes (spaC and spxB) and L. rhamnosus genes (spaE and wzb) were assessed. The Student t-test and the Mann-Whitney U test were used for comparisons. RESULTS: The L. casei group species were found to be part of the viable microbial community in dentinal caries. L. paracasei (p = 0.001), L. rhamnosus (p = 0.022), and L. casei (p = 0.004) group species were abundant in the active dentinal lesions compared to the arrested dentinal lesions. Only the wzb gene (p = 0.006) exhibited a statistically significant difference between the active and arrested lesions in terms of its expression profile; it was expressed to a higher extent in the active dentinal lesions. CONCLUSIONS: The L. casei group species presented in large numbers in the active dentinal caries lesions, indicating that these microorganisms are related to caries activity, and the wzb gene may play an important role in caries progression.

Physical-Mechanical Properties of Bulk Fill Composites Submitted to Biodegradation by Streptococcus mutans.

The aim of this study was to evaluate the Streptococcus mutans biofilm influence on the roughness (Ra), gloss (GU), surface hardness (KHN) and flexural strength (FS) of high viscosity bulk fill composites. Filtek Bulk Fill (FBF), Tetric N Ceram Bulk Fill (TNC), X-tra fil Bulk Fill (XF) and Filtek Z350 (FZ) were used. Ten discs of each composite were prepared for Ra, KHN and GU and 20 bars for the FS. After 24 h, specimens were polished and initial analyzes performed. Samples were sterilized and subjected to biodegradation for 7 days and final analyzes performed. Representative samples of each group were evaluated in Scanning Electron Microscope. Data were submitted to ANOVA two factors and Tukey test. XF presented the highest values (p<0.05) of Ra before and after biodegradation (0.1251; 0.3100), and FZ (0.1443) the lowest after biodegradation (p<0.05). The highest GU values (p<0.05) were observed for FZ (71.7; 62) and FBF (69.0; 64.6), and the lowest (p<0.05) for TNC (61.4; 53.3) and XF (58.5; 53.5), both before and after biodegradation. For KHN the highest values were obtained by XF (151.7; 106), and the (p< 0.05) lowest values for TNC (62.2; 51.8), both before and after biodegradation. The highest values (p<0.05) of FS were observed for FZ (127.6) and the lowest (p<0.05) for TNC (86.9); after biodegradation, XF (117.7) presented the highest (p<0.05) values compared to TNC and FZ." In conclusion, biodegradation increased Ra and decreased GU and KHN for all. Concerning FS, degradation provided a significant decreased value only for FZ.

Combined Effectiveness of ß-Cyclodextrin Nanoparticles in Photodynamic Antimicrobial Chemotherapy on In Vitro Oral Biofilms.

Objective: This study aimed to investigate if ß-cyclodextrin nanoparticles potentiate the photodynamic antimicrobial chemotherapy (PACT) effects in single and microcosm oral biofilms using methylene blue (MB) and a red laser. Background data: Studies of PACT have demonstrated promising effects; however, the association of nanoparticles with photosensitizers could enhance the antimicrobial result. Materials and methods: Biofilms were grown on enamel blocks either with Streptococcus mutans or in a microcosm model (salivary microorganisms) supplemented with sucrose. PACT using 50 µM MB associated or not with 32 µM encapsulated ß-cyclodextrin with MB for 5 min, followed by irradiation with red laser (λ = 660 nm, 320 J/cm2), was conducted and the counts of viable microorganisms in proper selective media were determined. Data were analyzed by one-factor ANOVA, followed by Tukey's test, or Kruskal-Wallis test, followed by Dunn's post hoc test, all with a significance level of 5%. Results: In the single-species biofilm model, a significant reduction in S. mutans counts was found for all groups when light was present. In the microcosm biofilm model, no significant difference was found among the groups for total streptococci, but a significant reduction of S. mutans was observed for the PACT group of encapsulated ß-cyclodextrin+MB. However, no statistically significant difference was observed among the PACT groups. Conclusions: PACT with ß-cyclodextrin mediated with MB associated with a red laser reduced S. mutans in microcosm biofilms. However, the presence of ß-cyclodextrin nanoparticles did not potentiate the PACT effects in single or microcosm oral biofilms.

Trans,trans-farnesol, an antimicrobial natural compound, improves glass ionomer cement properties.

A series of experiments were conducted to characterize a novel restorative material. We explored the effect on biological, physical and chemical properties of glass ionomer cement (GIC) adding-the naturally occurring tt-farnesol (900 mM). Two groups were accomplished for all assays: GIC+tt-farnesol and GIC (control). Biological assays: 1) agar diffusion against some cariogenic bacteria; 2) S. mutans biofilm formation and confocal laser scanning microscopy-CLSM. 3) gtfB, gtfC, gtfD, gbpB, vicR, and covR expression; 4) MTT and microscopic morphology. Physical properties assays: 1) roughness; 2) hardness; 3) compressive strength and 4) diametral tensile strength. Chemical assay: Raman spectroscopy. The adding of tt-farnesol to GIC led to larger zones of inhibition (p<0.05), biofilms with a short-term reduction in bacterial viability but similar biomass (p>0.05). Polysaccharides levels increased over time, similarly over groups (p>0.05). Viable and non-viable S. mutans were seen on the specimens' surface by CLSM but their virulence was not modulated by tt-farnesol. The tt-farnesol increased the HaCaT cell viability without impact on compressive and diametral tensile strength and roughness although the hardness was positively affected (p<0.05). Raman confirmed the presence of tt-farnesol. The incorporation of tt-farnesol into GIC inhibited the growth of cariogenic bacteria but had a little effect on the composition, structure and physiology of the biofilm matrices. Also, the tt-farnesol increased the hardness and the biocompatibility of the GIC, not influencing negatively other physical properties of the restorative material.

Effect of iodonium salt and chitosan on the physical and antibacterial properties of experimental infiltrants.

Resinous infiltrants are indicated in the treatment of incipient carious lesions, and further development of these materials may contribute to greater control of these lesions. The aim of this study was to analyze the physical and antibacterial properties of experimental infiltrants containing iodonium salt and chitosan. Nine experimental infiltrants were formulated by varying the concentration of the diphenyliodonium salt (DPI) at 0, 0.5 and 1 mol%; and chitosan at 0, 0.12 and 0.25 g%. The infiltrants contained the monomeric base of triethylene glycol dimethacrylate and bisphenol-A dimethacrylate ethoxylate in a 75 and 25% proportion by weight, respectively; 0.5 mol% camphorquinone and 1 mol% ethyl 4-dimethylaminobenzoate. The degree of conversion was evaluated using Fourier transformer infrared spectroscopy, and the flexural strength and elastic modulus using the three-point bending test. Sorption and solubility in water, and antibacterial analysis (minimum inhibitory concentration and minimum bactericidal concentration) were also analyzed. Data was analyzed statistically by two-way ANOVA and Tukey's test (p<0.05), with the exception of the antibacterial test, which was evaluated by visual inspection. In general, the infiltrant group containing 0.5% DPI and 0.12% chitosan showed high values of degree of conversion, higher values of elastic modulus and flexural strength, and lower sorption values in relation to the other groups. Antibacterial activity was observed in all the groups with DPI, regardless of the concentration of chitosan. The addition of DPI and chitosan to experimental infiltrants represents a valid option for producing infiltrants with desirable physical and antibacterial characteristics.

Extraction and purification of RNA from human carious dentine: an approach to enable bacterial gene expression studies / Extração e purificação de RNA proveniente de dentina humana cariada: uma abordagem para viabilizar estudos de expressão gênica

Background: RNA isolation from bacteria within dentine caries lesions could be difficult due to reduced amount of collectable biomass and high mRNA instability. Attempting to overcome this challenge we describe one protocol developed to extract and purify total RNA from dentine lesions. Objective: customize a bacterial RNA extraction and purification method from human carious dentine. Methods: quantity and purity of extracted RNA were measured with a microvolume UV-VIS spectrophotometer, RNA integrity was assessed by standard denaturing agarose gel electrophoresis and images were captured under ultraviolet light with camera and analyzed. DNase treatment removed genomic DNA and an additional step of purification was carried out in silica spin column. Results: final yield (ng/µl) was 67.01 ± 22.33, absorbance ratio A260/A280 = 2.0 ± 0.07 and RNA integrity were obtained. The purified samples were reversely transcribed and the expression of atpD and fabM gene from Streptococcus mutans analyzed by quantitative real-time PCR. Conclusion: the extraction methodology developed produced high-quality RNA from dentine microbiota for transcriptional analysis.

Introdução: o isolamento de RNA de bactérias dentro de lesões de dentina cariada pode ser difícil devido à quantidade reduzida de biomassa e alta instabilidade de mRNA. Na tentativa de superar esse desafio, descrevemos um protocolo desenvolvido para extrair e purificar o RNA total das lesões dentinárias. Objetivo: personalizar um método de extração e purificação de RNA bacteriano a partir da dentina cariada humana. Métodos: a quantidade e a pureza do RNA extraído foram medidas com um espectrofotômetro UV-VIS de microvolume, a integridade do RNA foi avaliada por eletroforese em gel de agarose desnaturante padrão e as imagens foram capturadas sob luz ultravioleta e analisadas. O tratamento com DNase removeu o DNA genômico e uma etapa adicional de purificação foi realizada em coluna de spin de sílica. Resultados: o rendimento final (ng / µl) foi de 67,01 ± 22,33, a razão de absorbância A260 / A280 = 2,0 ± 0,07 e a integridade do RNA foram obtidas. As amostras purificadas foram transcritas reversamente e a expressão do gene atpD e fabM de Streptococcus mutans analisadas por PCR quantitativo em tempo real (RT-qPCR). Conclusão: a metodologia de extração desenvolvida produziu RNA de alta qualidade da microbiota dentinária para análises transcricionais.

Antimicrobial activity of mouth rinses against bacteria that initially colonizes dental's surface / Atividade antimicrobiana de enxaguatórios bucais sobre bactérias que iniciam a colonização das superfícies dentais

Abstract Introduction Much advertising in mouthwash is conveyed in all media appealing to the anti-plaque effect and rendering a disservice to the community. Mouth rinses are available over-the-count and differ on their compositions and antimicrobial effectiveness. Objective In this study, we evaluated the antimicrobial activity of 35 widely available mouth rinses against bacterial species involved in initiation of dental biofilm - Streptococcus gordonii, Streptococcus mitis, Streptococcus oralis, Streptococcus salivarius, and Streptococcus sanguinis. Material and method The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) of the evaluated mouth rinses were determined according to the Clinical & Laboratory Standards Institute protocols. Data were submitted to Kruskal-Wallis test and Mann-Whitney post hoc (α=0.05). Result About 70% of the mouth rinses achieved high antibacterial activity and 30%, a low antibacterial activity against all the species tested. The most ineffective mouth rinse showed antibacterial activity (MIC) at 1:1 dilution, while the most effective showed activity even at 1:2048 dilution, which may imply prolonged effect in the mouth. About 51% of mouth rinses showed bactericidal activity, and it was verified that cetylpyridinium chloride or chlorhexidine digluconate containing in the formulation were associated with the highest activity. Conclusion Most - but not all - mouth rinses commercially available are effective in inhibiting in vitro initial colonizers of dental surfaces.

Resumo Introdução Muita publicidade sobre enxaguatórios bucais é veiculada em todos os meios de comunicação apelando para o efeito anti-placa e prestando um desserviço à comunidade. Grande quantidade de enxaguatórios bucais está disponível no mercado e estes diferem em suas composições e eficácia antimicrobiana. Objetivo Neste estudo, avaliamos a atividade antimicrobiana de 35 enxaguatórios bucais amplamente disponíveis contra espécies bacterianas envolvidas na iniciação do biofilme dental - Streptococcus gordonii, Streptococcus mitis, Streptococcus oralis, Streptococcus salivarius e Streptococcus sanguinis. Material e método A Concentração Inibitória Mínima (CIM) e a Concentração Bactericida Mínima (CBM) dos enxaguatórios avaliados foram determinadas de acordo com os protocolos do Clinical & Laboratory Standards Institute. Os dados foram submetidos ao teste Kruskal-Wallis e Mann-Whitney post hoc (α=0,05). Resultado Aproximadamente 70% dos enxaguatórios bucais alcançaram alta atividade antibacteriana e 30%, baixa atividade antibacteriana contra todas as espécies testadas. O enxaguatório bucal mais ineficaz mostrou atividade antibacteriana (CIM) na diluição de 1:1, enquanto a mais eficaz mostrou atividade mesmo na diluição de 1:2048, o que pode implicar em efeito prolongado na boca. Cerca de 51% dos enxaguatórios bucais apresentaram atividade bactericida, e verificou-se que formulações contendo cloreto de cetilpiridíneo ou digluconato de clorexidina estavam associados à maior atividade. Conclusão A maior parte - mas não todos - dos enxaguatórios bucais comercialmente disponíveis são eficazes na inibição de colonizadores iniciais de superfícies dentárias in vitro.

Effect of iodonium salt and chitosan on the physical and antibacterial properties of experimental infiltrants

Abstract Resinous infiltrants are indicated in the treatment of incipient carious lesions, and further development of these materials may contribute to greater control of these lesions. The aim of this study was to analyze the physical and antibacterial properties of experimental infiltrants containing iodonium salt and chitosan. Nine experimental infiltrants were formulated by varying the concentration of the diphenyliodonium salt (DPI) at 0, 0.5 and 1 mol%; and chitosan at 0, 0.12 and 0.25 g%. The infiltrants contained the monomeric base of triethylene glycol dimethacrylate and bisphenol-A dimethacrylate ethoxylate in a 75 and 25% proportion by weight, respectively; 0.5 mol% camphorquinone and 1 mol% ethyl 4-dimethylaminobenzoate. The degree of conversion was evaluated using Fourier transformer infrared spectroscopy, and the flexural strength and elastic modulus using the three-point bending test. Sorption and solubility in water, and antibacterial analysis (minimum inhibitory concentration and minimum bactericidal concentration) were also analyzed. Data was analyzed statistically by two-way ANOVA and Tukey's test (p<0.05), with the exception of the antibacterial test, which was evaluated by visual inspection. In general, the infiltrant group containing 0.5% DPI and 0.12% chitosan showed high values of degree of conversion, higher values of elastic modulus and flexural strength, and lower sorption values in relation to the other groups. Antibacterial activity was observed in all the groups with DPI, regardless of the concentration of chitosan. The addition of DPI and chitosan to experimental infiltrants represents a valid option for producing infiltrants with desirable physical and antibacterial characteristics.

Quantitative analysis of biofilm bacteria according to different stages of early childhood caries.

OBJECTIVE: Monitoring selected key species related to health or disease may facilitate caries risk assessment and discovery of novel ecological preventive and therapeutic approaches. This study aimed at quantifying Actinomyces naeslundii, Bifidobacterium spp., Lactobacillus acidophilus, Lactobacillus casei group, Streptococcus gordonii, Mitis group and Streptococcus mutans by quantitative polymerase chain reaction (qPCR) in dental biofilm from Brazilian children with different stages of early childhood caries (ECC). DESIGN: Seventy-five preschool children were clinically evaluated by ICDAS criteria and divided into groups: caries-free (CF; n = 20), enamel caries lesions (ECL; n = 17) and dentine caries lesions (DCL; n = 38). Plaque samples from all children were collected for detection and quantification of the selected bacteria. RESULTS: L. acidophilus and L. casei group were absent in almost all plaque samples. No differences in relative proportions of A. naeslundii, Mitis group and S. gordonii were observed in any stage of caries. However, S. mutans and Bifidobacterium spp. were present at higher concentrations in the biofilm of children with DCL (p < 0.001). Multivariate analysis showed that S. mutans and Bifidobacterium spp. were strongly associated with biofilm in children with DCL. CONCLUSION: Differences were observed in the proportion of acidogenic and aciduric bacteria with dental caries progression. The data indicate that S. mutans and Bifidobacterium spp. in dental biofilm may be involved in some progression processes for ECC.

Viability and Osteogenic Differentiation of Human Periodontal Ligament Progenitor Cells Are Maintained After Incubation With Porphyromonas gingivalis Protein Extract.

BACKGROUND: Porphyromonas gingivalis (Pg) is a major periodontal pathogen that contains immunostimulatory components. Periodontal ligament mesenchymal stem cells (PDLMSCs) are responsible for regeneration of the periodontium that is lost due to periodontitis. Pathologic factors within the microenvironment that impair resident PDLMSCs are not well understood. The present study investigates in vitro the effects of Pg protein extract (PgPE) on biologic properties of CD105-enriched PDL progenitor cell populations (PDL-CD105+). METHODS: Five populations of PDL-CD105+ cells were exposed to PgPE and assessed for cell viability, apoptosis, and proinflammatory gene expression (interleukin-1ß [IL-1ß], tumor necrosis factor-alpha [TNF-α], and IL-6) by quantitative reverse transcription polymerase chain reaction, IL-6 immunostaining, activation of IL-6/signal transducer and activator of transcription (STAT) 3 signaling pathway, and osteogenic differentiation potential. RESULTS: PgPE treatment (2 µg/mL) did not affect cell viability or survival but induced a significant increase in IL-1ß, TNF-α, and IL-6 messenger RNA (mRNA) expression and positive staining for IL-6. A total of 29 genes from the IL-6/STAT3 pathway were upregulated on PgPE stimulation. These genes are related to biologic processes involved in the control of cell survival (B-cell lymphoma 2 [BCL2]), cell proliferation (hepatocytehepatocyte growth factor), cytokine-mediated signaling pathway (suppressor of cytokine signaling 3, C-X-C ligand 8 [CXCL8]), and response to stress (CXCL8, mitogen-activated protein kinase 3, BCL2-associated X protein, and BCL2). Additionally, PgPE treatment caused an increase in alkaline phosphatase mRNA expression in PDL-CD105+ cells after 7 days of osteogenic induction, although mineral nodule formation was comparable to the control group. CONCLUSIONS: These results suggest that the inflammatory profile induced by PgPE treatment in PDL-CD105+ cells did not affect cell viability, apoptosis, or osteogenic differentiation, perhaps due to increased expression of genes involved in the control of cell proliferation and protection against cell death.

Molecular detection of bacteria associated to caries activity in dentinal lesions.

OBJECTIVES: This study aimed at identifying and quantifying Actinomyces naeslundii, Bifidobacterium spp., Streptococcus mitis group, Lactobacillus acidophilus, Lactobacillus casei group, Streptococcus gordonii, and Streptococcus mutans in active and inactive carious dentine lesions of children with early childhood caries by using quantitative polymerase chain reaction. MATERIAL AND METHODS: Fifty-six dentin lesion samples, classified as active (n = 39) or inactive (n = 17), were collected from children aged from 2 to 5 years old. Dentinal-cavitated lesions were evaluated by Nyvad criteria for the assessment of caries lesion activity. RESULTS: Relative quantification revealed that Bifidobacterium spp. and the L. casei group were significantly more abundant in active dentin lesions (p < 0.05). Concentrations of A. naeslundii, S. mitis group, and S. gordonii were not significantly different when comparing dentin lesion activity. The relative proportion of S. mutans was significantly greater in inactive than in active lesions (p < 0.05). Bifidobacterium spp. and L. casei group demonstrated a positive correlation (p = 0.001) in active lesions. The positive detection of L. acidophilus (odds ratio = 15.1) and S. gordonii (odds ratio = 7.7) was significantly associated to the active lesions. CONCLUSIONS: The data indicate that higher detection levels of Bifidobacterium spp. and the L. casei group may be linked to dentin lesion activity. Additionally, the presence of L. acidophilus and S. gordonii was associated with lesion activity. CLINICAL RELEVANCE: Considering that information about the oral microbiota related to dentin caries activity status is relevant, this study provides insights to better understand the differences in the microbiotas between active and arrested dentin cavities.

Dentine bond strength and antimicrobial activity evaluation of adhesive systems.

OBJECTIVES: This study evaluated the dentine bond strength (BS) and the antibacterial activity (AA) of six adhesives against strict anaerobic and facultative bacteria. METHODS: Three adhesives containing antibacterial components (Gluma 2Bond (glutaraldehyde)/G2B, Clearfil SE Protect (MDPB)/CSP and Peak Universal Bond (PUB)/chlorhexidine) and the same adhesive versions without antibacterial agents (Gluma Comfort Bond/GCB, Clearfil SE Bond/CSB and Peak LC Bond/PLB) were tested. The AA of adhesives and control groups was evaluated by direct contact method against four strict anaerobic and four facultative bacteria. After incubation, according to the appropriate periods of time for each microorganism, the time to kill microorganisms was measured. For BS, the adhesives were applied according to manufacturers' recommendations and teeth restored with composite. Teeth (n=10) were sectioned to obtain bonded beams specimens, which were tested after artificial saliva storage for one week and one year. BS data were analyzed using two-way ANOVA and Tukey test. RESULTS: Saliva storage for one year reduces the BS only for GCB. In general G2B and GCB required at least 24h for killing microorganisms. PUB and PLB killed only strict anaerobic microorganisms after 24h. For CSP the average time to eliminate the Streptococcus mutans and strict anaerobic oral pathogens was 30 min. CSB showed no AA against facultative bacteria, but had AA against some strict anaerobic microorganisms. CONCLUSIONS: Storage time had no effect on the BS for most of the adhesives. The time required to kill bacteria depended on the type of adhesive and never was less than 10 min. CLINICAL SIGNIFICANCE: Most of the adhesives showed stable bond strength after one year and the Clearfil SE Protect may be a good alternative in restorative procedures performed on dentine, considering its adequate bond strength and better antibacterial activity.

Evaluation of the effects of transient or continuous PTH administration to odontoblast-like cells.

Parathyroid hormone participates in the metabolism of mineralized tissue. Its role in the formation of dentine is, as yet, incompletely understood. In the present study we analyzed the effect of transient (1 and 24-h/cycle) or continuous hPTH (1-34) treatment in odontoblast-like cells (MDPC-23) to the following parameters: mineral deposition detected by alizarin red, mRNA expression of the type I collagen (COL1), alkaline phosphatase (ALP), biglycan (BGN), matrix metalloproteinase 2 (MMP-2) and dentine sialophosphoprotein (DSPP) quantified by qRT-PCR. MMP-2 and ALP activities were quantified by zymography and colorimetric assay, respectively. The results showed that compared to Control group: intermittent PTH administration (1 and 24-h/cycle) decreased the mineral deposition and ALP activity. DSPP gene expression was not detected in both control and PTH treated cells. The PTH administration for 24-h/cycle increased the ALP, BGN and COL1 mRNA expression and continuous PTH treatment increased BGN and COL1 mRNA expression. Zymography assays showed that compared to Control group: PTH treatment for 1-h/cycle increased the total MMP-2 secretion and the continuous treatment decreased the secreted levels of MMP-2 active-form. Taken together, the results shown that PTH may regulate the odontoblast-like cells-induced secretion, and potentially this hormone can affect in vivo odontoblasts functions.