Role of Candida species from HIV infected children in enamel caries lesions: an in vitro study

Objectives This study analyzed the capacity of Candida spp. from dental biofilm of HIV infected (HIV+) children to demineralize primary molar enamel in vitro by Transversal Microhardness (TMH), Polarized Light Microscopy (PLM) and the quantity of calcium ions (Ca2+) released from the enamel. Material and Methods Candida spp. samples were isolated from the supragingival biofilm of HIV+ children. A hundred and forty (140) enamel blocks were randomly assigned to six groups: biofilm formed by C. albicans (Group 1); mixed biofilm formed by C. albicans and C. tropicalis (Group 2); mixed biofilm formed by C. albicans and C. parapsilosis (Group 3); mixed biofilm formed by C. albicans, C. parapsilosis and C. glabrata (Group 4); biofilm formed by C. albicans ATCC (Group 5) and medium without Candida (Group 6). Enamel blocks from each group were removed on days 3, 5, 8 and 15 after biofilm formation to evaluate the TMH and images of enamel were analyzed by PLM. The quantity of Ca2+ released, from Groups 1 and 6, was determined using an Atomic Absorption Spectrophotometer. The SPSS program was used for statistical analysis and the significance level was 5%. Results TMH showed a gradual reduction in enamel hardness (

TiF4 and NaF varnishes as anti-erosive agents on enamel and dentin erosion progression in vitro

Objective This study assessed the effect of fluoride varnishes on the progression of tooth erosion in vitro. Material and Methods: Forty-eight enamel and 60 root dentin samples were previously demineralized (0.1% citric acid, pH 2.5, 30 min), leading to a baseline and erosive wear of 12.9 and 11.4 µm, respectively. The samples were randomly treated (6 h) with a 4% TiF4 varnish (2.45%F-, pH 1.0), a 5.42% NaF varnish (2.45%F-, pH 5.0), a placebo varnish and no varnish (control). The samples were then subjected to erosive pH cycles (4x90 s/day in 0.1% citric acid, intercalated with artificial saliva) for 5 days. The increment of the erosive tooth wear was calculated. In the case of dentin, this final measurement was done with and without the demineralized organic matrix (DOM). Enamel and dentin data were analyzed using ANOVA/Tukey’s and Kruskal-Wallis/Dunn tests, respectively (

The effect of mouthwashes containing biguanides on the progression of erosion in dentin

Background\ud Dental erosion is caused by frequent exposure to acids without the involvement of microorganism. This study analyzed the effect of biguanides (polyhexamethylene biguanide – PHMB and chlorhexidine – CHX) on dentin erosion due to their possible influence on the enzymatic degradation of the demineralized organic matrix.\ud \ud \ud Method\ud Sixty bovine dentin specimens were prepared. On both sides of their surface, nail varnish was applied to maintain the reference surfaces for the determination of dentin loss. Samples were cyclically de- and remineralized for 6 days. Demineralization was performed with a 0.87 M citric acid solution (6×5 min daily). Thereafter, samples were treated with distilled water (negative control), 0.12% CHX (positive control), 0.07% PHMB, Sanifill Perio Premium™ (0.07% PHMB plus 0.05% NaF), or F solution (0.05% NaF) for 1 min and then subjected to enzymatic challenge for 10 min using a bacterial collagenase (Clostridium hystoliticum, 100 μg/ml). Dentin loss was assessed using profilometry (μm) daily. Data were analyzed using 2-way repeated measures-ANOVA and Bonferroni’s test (p < 0.05).\ud \ud \ud Results\ud Dentin loss progressed significantly for all groups during the 6 days. After the 3rd day, Sanifill Premium™, CHX, and PHMB significantly reduced dentin erosion compared to control. On the 6th day, the lowest mean (±SD) dentin loss was observed for Sanifill Perio Premium™ (94.4 ± 3.9 μm). PHMB and CHX led to intermediate dentin loss (129.9 ± 41.2 and 135.3 ± 33.5 μm, respectively) that was significantly lower than those found for negative control (168.2 ± 6.2 μm). F (157.4 ± 6.1 μm) did not significantly differ from negative control.\ud \ud \ud Conclusions\ud Sanifill Perio Premium™ mouthwash has a good potential to reduce dentin loss, which might be associated with the presence of PHMB.PIBIT/CNPq/USP-2010.1.6720.25.

Proteomic analysis of secretory and maturation-stage enamel matrix in mice susceptible or resistant to dental fluorosis, chronically exposed to fluoride in the drinking water

Análise proteômica da matriz do esmalte nos estágios de secreção e maturação em camundongos susceptíveis ou resistentes à fluorose dentária, expostos cronicamente ao fluoreto através da água de beber. Os mecanismos pelos quais a ingestão excessiva de fluoreto (F) durante a amelogênese levam à fluorose dentária ainda não são precisamente conhecidos. Tem sido demonstrado que determinadas linhagens de camundongos são mais susceptíveis que outras à fluorose dentária, o que faz destas linhagens o modelo ideal para se estudarem os fenômenos moleculares envolvidos nesta patologia. No presente estudo, foi empregada uma abordagem proteômica para avaliar alterações na expressão de proteínas da matriz do esmalte dentário nos estágios de secreção e maturação, em duas linhagens de camundongos com diferentes susceptibilidades à fluorose (A/J, susceptível e 129P3/J, resistente). Camundongos de ambos os gêneros, representantes das linhagens 129P3/J (n=200) e A/J (n=200) foram distribuídos em dois grupos para cada linhagem, que receberam ração com baixa concentração de F e água de beber contendo 0 (controle) ou 50 mg/L F por 6 semanas. A concentração de F foi analisada no plasma e no esmalte dos incisivos. Para a análise proteômica, foi realizada a raspagem da matriz do esmalte dos incisivos, nos estágios de secreção e maturação. Para a extração das proteínas do esmalte, foram pesados 15 mg de pó da matriz do esmalte, aos quais foi adicionado 1 mL de tampão de lise contendo uréia 7 M, tiouréia 2 M, CHAPS 4 %, DTT 1 %, anfólitos carreadores 0,5 % pH 3-10 e um coquetel de inibidores de proteases. As proteínas do esmalte extraídas para cada grupo foram separadas pela técnica de eletroforese bidimensional e posteriormente submetidas a LC-ESI-MS/MS. A concentração média (±DP) de F encontrada no plasma dos camundongos foi de 0,023±0,010 e 0,019±0,007 mg/L para os animais do grupo controle, das linhagens A/J e 129P3/J, respectivamente, e de 0,151±0,043 e de 0,252±0,060 mg/L de F para os animais das linhagens A/J e 129P3/J, respectivamente, tratados com água contendo 50 mg/L de F. A ANOVA a 2 critérios revelou que houve diferença significativa entre os tratamentos (F= 658,0, p<0,0001), mas não entre as linhagens (F=3,3 p=0,075), tendo havido interação significativa entre estes critérios (F=16,50, p=0,0002). Já a concentração média (±DP) de F incorporado no esmalte dos incisivos dos camundongos foi 471,4±215,8 mg/kg e 151,7±58,8 mg/kg para os animais do grupo controle das linhagens 129P3/J e A/J, respectivamente, e 2711,2±1019,2 mg/kg, 1756,9±921,6 mg/kg para os animais das linhagens 129P3/J e A/J, respectivamente, tratados com água contendo 50 mg/L de F. A ANOVA a 2 critérios encontrou diferença significativa entre as linhagens (F=11,36, p=0,0016) e entre os tratamentos (F=103,50, p<0,0001), sem interação significativa entre ambos (F=2,82, p=0,1004). Os resultados proteômicos mostraram redução na abundância de proteínas no estágio de maturação, quando comparado com o de secreção. Foi observado que o tratamento com F aumentou consideravelmente o número de spots proteicos detectados em ambos os estágios, sendo que este aumento foi maior para os animais da linhagem A/J, indicando uma tentativa de se combater os efeitos deletérios do F e reforçando a maior susceptibilidade aos seus efeitos desta linhagem. A identificação das proteínas com expressão diferencial revelou que tanto a linhagem quanto o tratamento com F levaram à expressão diferencial de proteínas pertencentes a todas as categorias funcionais.The mechanisms by which excessive ingestion of fluoride (F) during amelogenesis leads to fluorosis are still not precisely known. It has been shown that certain strains of mice are more susceptible to dental fluorosis than others, which turns these strains the ideal model for studying the molecular phenomena involved in this pathology. In the present study, we employed a proteomic approach to identify and evaluate changes in protein expression of secretory and maturation-stage enamel matrix in two strains of mice with different susceptibilities to dental fluorosis (A/J, susceptible and 129P3/J, resistant). Mice of both genders, from 129P3/J (n=200) and A/J (n=200) strains were divided into two groups for each strain. They received a low-F diet and drinking water containing 0 (control) or 50 mg/L F for 6 weeks. The F concentration was analyzed in plasma and enamel incisors. For proteomic analysis, the enamel matrix of secretory and maturation stages (incisors) was scrapped. For the extraction of enamel proteins, 1 ml of lysis buffer containing 7 M urea, 2 M thiourea, 4% CHAPS, 1% DTT, 0.5% ampholytes carriers pH 3-10 and a cocktail of protease inhibitors was added to 15 mg of enamel matrix powder. The enamel proteins extracted for each group were separated by two-dimensional electrophoresis and subsequently subjected to LC-ESIMS/ MS. The mean (± SD) F concentrations found in plasma were 0.023 ± 0.010 and 0.019 ±0.007 mg/L for the control group, A/J and 129P3/J strains, respectively; and 0.151 ± 0.043 and 0.252 ± 0.060 mg/L F for A/J and 129P3/J mice, respectively, treated with water containing 50 mg/L F. Two-way ANOVA revealed significant differences between treatments (F = 658.0, p <0.0001), but not between strains (F = 3.3 p = 0.075), with was significant interaction between these criteria (F = 16.50, p = 0.0002). The mean (± SD) F concentrations in the enamel of the incisors were 471.4 ± 215.8 mg/kg and 151.7 ± 58.8 mg/kg for the control group of 129P3/J and A/J strains, respectively; and 2711.2 ± 1019.2 mg/kg and 1756.9 ± 921.6 mg/kg for 129P3/J and A/J mice, respectively, treated with water containing 50 mg/L F. Two-way ANOVA detected significant differences between the strains (F=11.36, p=0.0016) and between treatments (F=103.50, p <0.0001), without significant interaction between these criteria (F=2.82, p=0.1004). The proteomic results revealed a reduction in the abundance of proteins in the maturation stage, as compared with the secretory stage. Treatment with F greatly increased the number of protein spots detected in both stages. This increase was greater for A/J mice, indicating an attempt to fight the deleterious effects of F and, thus, reinforcing the susceptibility of this strain to the effects of F. The identification of differentially expressed proteins revealed that both the strain and the treatment with F led to differential expression of proteins belonging to all functional categories

Role of Candida species from HIV infected children in enamel caries lesions: an in vitro study

Abstract Objectives This study analyzed the capacity of Candida spp. from dental biofilm of HIV infected (HIV+) children to demineralize primary molar enamel in vitro by Transversal Microhardness (TMH), Polarized Light Microscopy (PLM) and the quantity of calcium ions (Ca2+) released from the enamel. Material and Methods Candida spp. samples were isolated from the supragingival biofilm of HIV+ children. A hundred and forty (140) enamel blocks were randomly assigned to six groups: biofilm formed by C. albicans (Group 1); mixed biofilm formed by C. albicans and C. tropicalis (Group 2); mixed biofilm formed by C. albicans and C. parapsilosis (Group 3); mixed biofilm formed by C. albicans, C. parapsilosis and C. glabrata (Group 4); biofilm formed by C. albicans ATCC (Group 5) and medium without Candida (Group 6). Enamel blocks from each group were removed on days 3, 5, 8 and 15 after biofilm formation to evaluate the TMH and images of enamel were analyzed by PLM. The quantity of Ca2+ released, from Groups 1 and 6, was determined using an Atomic Absorption Spectrophotometer. The SPSS program was used for statistical analysis and the significance level was 5%. Results TMH showed a gradual reduction in enamel hardness (p<0.05) from the 1st to 15th day, but mainly five days after biofilm formation in all groups. The PLM showed superficial lesions indicating an increase in porosity. C. albicans caused the release of Ca2+ into suspension during biofilm formation. Conclusion Candida species from dental biofilm of HIV+ children can cause demineralization of primary enamel in vitro

TiF4 and NaF varnishes as anti-erosive agents on enamel and dentin erosion progression in vitro

Objective This study assessed the effect of fluoride varnishes on the progression of tooth erosion in vitro. Material and Methods: Forty-eight enamel and 60 root dentin samples were previously demineralized (0.1% citric acid, pH 2.5, 30 min), leading to a baseline and erosive wear of 12.9 and 11.4 &#181;m, respectively. The samples were randomly treated (6 h) with a 4% TiF4 varnish (2.45%F-, pH 1.0), a 5.42% NaF varnish (2.45%F-, pH 5.0), a placebo varnish and no varnish (control). The samples were then subjected to erosive pH cycles (4x90 s/day in 0.1% citric acid, intercalated with artificial saliva) for 5 days. The increment of the erosive tooth wear was calculated. In the case of dentin, this final measurement was done with and without the demineralized organic matrix (DOM). Enamel and dentin data were analyzed using ANOVA/Tukey&#8217;s and Kruskal-Wallis/Dunn tests, respectively (p<0.05). Results The TiF4 (mean&#177;s.d: 1.5&#177;1.1 &#181;m) and NaF (2.1&#177;1.7 &#181;m) varnishes significantly reduced enamel wear progression compared to the placebo varnish (3.9&#177;1.1 &#181;m) and control (4.5&#177;0.9 &#181;m). The same differences were found for dentin in the presence and absence of the DOM, respectively: TiF4 (average: 0.97/1.87 &#181;m), NaF (1.03/2.13 &#181;m), placebo varnish (3.53/4.47 &#181;m) and control (3.53/4.36 &#181;m). Conclusion The TiF4 and NaF varnishes were equally effective in reducing the progression of tooth erosion in vitro

Liver proteome of mice with different genetic susceptibilities to the effects of fluoride

A/J and 129P3/J mice strains have been widely studied over the last few years because they respond quite differently to fluoride (F) exposure. 129P3/J mice are remarkably resistant to the development of dental fluorosis, despite excreting less F in urine and having higher circulating F levels. These two strains also present different characteristics regardless of F exposure. Objective In this study, we investigated the differential pattern of protein expression in the liver of these mice to provide insights on why they have different responses to F. Material and Methods Weanling male A/J and 129P3/J mice (n=10 from each strain) were pared and housed in metabolic cages with ad libitum access to low-F food and deionized water for 42 days. Liver proteome profiles were examined using nLC-MS/MS. Protein function was classified by GO biological process (Cluego v2.0.7 + Clupedia v1.0.8) and protein-protein interaction network was constructed (PSICQUIC, Cytoscape). Results Most proteins with fold change were increased in A/J mice. The functional category with the highest percentage of altered genes was oxidation-reduction process (20%). Subnetwork analysis revealed that proteins with fold change interacted with Disks large homolog 4 and Calcium-activated potassium channel subunit alpha-1. A/J mice had an increase in proteins related to energy flux and oxidative stress. Conclusion This could be a possible explanation for the high susceptibility of these mice to the effects of F, since the exposure also induces oxidative stress