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OBJECTIVES: To answer the questions: (1) Does reducing estrogen levels influence the microbial composition of the oral cavity? (2) Does the presence of periapical lesion (PL) cause changes in the oral microbiota? (3) Since estrogen deficiency alters the oral microbiota, can this be one of the factors that contribute to the increase of the PL? MATERIALS AND METHODS: Thirty-six rats were divided into four groups: sham (control), ovariectomy (OVX), control with PL (Sham + PL), and OVX + PL. After 9 weeks of OVX, the lower first molars were submitted to PL induction. After 21 days, the microbiological collection of the oral cavity was performed, and the animals were euthanized. The contents were evaluated by the checkerboard DNA-DNA hybridization method, to verify the prevalence of 40 bacterial species (divided into 7 microbial complexes). The blocks containing the lower first molars were submitted to histotechnical processing and staining with hematoxylin and eosin (HE), for the measurement of the periapical lesion area. The results were submitted to ANOVA and Kruskal-Wallis tests and Tukey and Dunn post-tests, with a significance level of 5%. RESULTS: In conditions of estrogen deficiency, there was alteration of the oral microbiota. The OVX groups had a higher amount of bacteria compared to the SHAM group in most of the microbial complexes (p < 0.001). The animals in the control group (with or without lesion) did not present a statistically significant difference (p > 0.001) in any of the microbial complexes. The PLs in OVX animals were significantly higher compared to SHAM animals (p < 0.001). CONCLUSIONS: Hypoestrogenicity conditions interfere in the oral microbiota by increasing the amount of bacteria in the saliva and influencing the progression of periapical lesions. CLINICAL RELEVANCE: This inedited study shows that deficiency of estrogen leads to alteration of the oral microbiota.

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OBJECTIVE: The role of oestrogen in craniofacial growth still remains unclear. Therefore, the present study aimed to assess the effect of oestrogen deficiency on maxilla and mandible dimensions. SETTING AND SAMPLE POPULATION: The study was conducted at the Department of Pediatric Dentistry at the School of Dentistry of Ribeirão Preto, University of São Paulo, and used forty female Wistar rats. MATERIAL AND METHODS: Ovariectomy (OVX) and placebo surgery (Sham) were performed when animals were twenty-one days old (prepubertal stage). Dimensions of the maxilla and mandible were assessed by craniometric analysis using radiographs, during and after puberty of the animals (45 and 63 days old, respectively). Quantitative real-time PCR and immunohistochemical analyses were performed to determine the expression and localization, respectively, of oestrogen receptor alpha (ERα) and oestrogen receptor beta (ERß) in different growth sites of the evaluated structures at puberty. The differences between the groups for each outcome were evaluated using the t test with an established alpha error of 5%. RESULTS: There were significant differences between the OVX and Sham groups for horizontal and vertical linear measurements in the maxilla and the mandible at both pubertal and post-pubertal stages (P < .05). The ovariectomized rats showed significantly greater measures for all dimensions assessed. No differences in gene expression of ERα and ERß were identified at the different growth sites between the OVX and Sham groups (P > .05). Immunohistochemical analyses revealed the presence of both oestrogen receptors in osteoblasts and chondrocytes in the midpalatal suture and mandibular condyle, respectively, in the OVX and Sham groups. CONCLUSION: Our results suggest that oestrogen deficiency from the prepubertal stage might increase the growth of the maxilla and mandible in female rats.

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This study evaluated the effect of antimicrobial photodynamic therapy (aPDT) on the endodontic treatment of apical periodontitis (AP). AP was induced in 48 premolars of 6 dogs. After biomechanical preparation, the teeth were divided into 4 groups: Calcium-Hydroxide (CH)/120d and CH/180d: root canals filled with CH-based dressing for 15 days before obturation; aPDT/120d and aPDT/180d: conditioning with phenothiazine photosensitizer (10 mg/mL) for 1 minute and irradiation with diode laser in the same session as obturation. Root filling was performed with AH Plus sealer. After the experimental periods, animals were euthanized and teeth were submitted for histology. HE staining was performed for descriptive analysis of the periapical region, measurement of apical periodontitis and for inflammatory cells, and blood vessels count. Immunohistochemistry was performed for osteopontin (OPN) and alkaline phosphatase (ALP). Data were analyzed statistically by two-way ANOVA and chi-square test (α = 5%). Teeth in Group CH/120d presented only a slightly enlarged periodontal ligament (PL) with advanced repair. Group aPDT/120d presented the PL moderately enlarged, with moderate inflammatory infiltrate and few collagen fibers. The same pattern was observed at 180 days. AP lesions in CH-treated groups were smaller than those in aPDT-treated groups (p < 0.001) with more blood vessels (p < 0.0001), regardless of the evaluation period, without significant differences in the number of inflammatory cells (p > 0.05). CH-treated groups showed significantly more intense immunostaining for ALP and OPN (p < 0.001) in both periods. Although aPDT stimulated angiogenesis and expression of bone formation markers, the two-session endodontic treatment with CH-based dressing promoted better apical periodontitis repair.

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The aim of this study was to evaluate the radiographic periapical repair and the synthesis of inflammatory mediators after endodontic treatment in a single session, using different irrigation protocols, in teeth with apical periodontitis. Experimental apical periodontitis were induced in dog's teeth randomly assigned into 4 groups: G1 - Irrigation by Negative Apical Pressure (n= 20); G2 - Passive Ultrasonic Irrigation (n= 20), G3 - Positive Pressure Irrigation (n= 20); G4 - apical periodontitis without treatment (n= 20). After 180 days, the animals were euthanized, the tissues removed and submitted to histotechnical processing for immunohistochemical analysis of osteopontin (OPN), tumor necrosis factor-a (TNF-a) and interleukin 1-a (IL-1a). Radiographic analysis was performed using the Periapical Index (PAI), obtained prior to and 180 days following endodontic treatment. Data were analyzed using Wilcoxon signed-rank test, Fisher's Exact test or Kruskal-Wallis test and Dunn's post-test (a = 5%). Radiographically, after endodontic treatment, apical periodontitis persisted in 35% of G1 specimens, 40% of G2 and 40% of G3 (p>0.05), although a PAI reduction was observed (p<0.05). By immunohistochemical evaluation, endodontic treatment resulted in lower synthesis of TNF-a and OPN in periapical region, compared to apical periodontitis without treatment (p<0.05). Production of IL-1 was not modulated by endodontic treatment (p>0.05). Periapical healing was observed in approximately 60% of the cases after endodontic treatment performed in a single session with lower synthesis of TNF-a and OPN in the periapical region, regardless of the irrigation protocol used.

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Abstract The aim of this study was to evaluate the radiographic periapical repair and the synthesis of inflammatory mediators after endodontic treatment in a single session, using different irrigation protocols, in teeth with apical periodontitis. Experimental apical periodontitis were induced in dog's teeth randomly assigned into 4 groups: G1 - Irrigation by Negative Apical Pressure (n= 20); G2 - Passive Ultrasonic Irrigation (n= 20), G3 - Positive Pressure Irrigation (n= 20); G4 - apical periodontitis without treatment (n= 20). After 180 days, the animals were euthanized, the tissues removed and submitted to histotechnical processing for immunohistochemical analysis of osteopontin (OPN), tumor necrosis factor-a (TNF-a) and interleukin 1-a (IL-1a). Radiographic analysis was performed using the Periapical Index (PAI), obtained prior to and 180 days following endodontic treatment. Data were analyzed using Wilcoxon signed-rank test, Fisher's Exact test or Kruskal-Wallis test and Dunn's post-test (a = 5%). Radiographically, after endodontic treatment, apical periodontitis persisted in 35% of G1 specimens, 40% of G2 and 40% of G3 (p>0.05), although a PAI reduction was observed (p<0.05). By immunohistochemical evaluation, endodontic treatment resulted in lower synthesis of TNF-a and OPN in periapical region, compared to apical periodontitis without treatment (p<0.05). Production of IL-1 was not modulated by endodontic treatment (p>0.05). Periapical healing was observed in approximately 60% of the cases after endodontic treatment performed in a single session with lower synthesis of TNF-a and OPN in the periapical region, regardless of the irrigation protocol used.

Resumo O objetivo deste estudo foi avaliar o reparo periapical e a síntese de mediadores inflamatórios após tratamento endodôntico em dentes de cães com lesão periapical, em sessão única, utilizando diferentes protocolos de irrigação. Lesões periapicais foram induzidas experimentalmente em dentes de cães e aleatoriamente divididas em 4 grupos: G1 - Irrigação por Pressão Apical Negativa (n = 20); G2 - Irrigação Ultrassônica Passiva (n = 20), G3 - Irrigação por Pressão Positiva (n = 20); G4 - Lesão periapical sem tratamento (n = 20). Após 180 dias, os animais foram eutanasiados, as peças removidas e submetidas ao processamento histotécnico para análise imunohistoquímica para osteopontina (OPN), fator de necrose tumoral-a (TNF-a) e interleucina 1-a (IL-1a). A análise radiográfica do reparo das lesões periapicais foi realizada por meio do Índice Periapical, obtido antes e 180 dias após o tratamento endodôntico. Os resultados obtidos foram submetidos à análise estatística por meio dos testes de sinais de Wilcoxon, Exato de Fisher ou Kruskal-Wallis seguido pelo pós-teste de Dunn (a = 5%). O exame radiográfico após o tratamento endodôntico, mostrou a persistência de áreas radiolúcidas periapicais e descontinuidade da lâmina dura em 35% dos espécimes do G1, 40% do G2 e 40% do G3, embora uma redução no PAI tenha sido observada (p<0,05). Pela análise imuno-histoquímica, o tratamento endodôntico resultou na menor síntese de TNF-a e de OPN na região periapical, comparativamente à lesão periapical sem tratamento (p<0,05). A produção de IL-1a não foi modulada pelo tratamento endodôntico (p>0,05). Reparo da lesão periapical foi observado em cerca de 60% dos casos após tratamento endodôntico realizado em sessão única e menor síntese de TNF-a e de OPN na região periapical, independente do protocolo de irrigação utilizado.

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Recent studies suggested that genetics contribute to differences in dental fluorosis (DF) susceptibility among individuals having the same environmental exposure. This study evaluated if MMP2, MMP9 and MMP20 are expressed during enamel development and assessed the association between polymorphisms in these genes with DF. Mice susceptible and resistant to DF were used to evaluate if MMPs were candidate genes for DF. The animals received fluoride and their enamels were used for immunohistochemistry. Additionally, 481 subjects from a city with fluoridation of public water supplies were recruited. Genotyping was performed using real time PCR. Allele/genotype frequencies were compared between groups. MMP2, MMP9 and MMP20 immunostaining was detected in both animal groups. DF was observed in 22.4% of the subjects. A borderline association was observed in MMP2 (rs243865), MMP9 (rs17576) and in MMP20 (rs1784418) (p = 0.06, p = 0.08 and p = 0.06 respectively). Briefly, MMPs were expressed during enamel maturation and genetic polymorphisms were not associated with DF.

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Abstract This study evaluated the effect of antimicrobial photodynamic therapy (aPDT) on the endodontic treatment of apical periodontitis (AP). AP was induced in 48 premolars of 6 dogs. After biomechanical preparation, the teeth were divided into 4 groups: Calcium-Hydroxide (CH)/120d and CH/180d: root canals filled with CH-based dressing for 15 days before obturation; aPDT/120d and aPDT/180d: conditioning with phenothiazine photosensitizer (10 mg/mL) for 1 minute and irradiation with diode laser in the same session as obturation. Root filling was performed with AH Plus sealer. After the experimental periods, animals were euthanized and teeth were submitted for histology. HE staining was performed for descriptive analysis of the periapical region, measurement of apical periodontitis and for inflammatory cells, and blood vessels count. Immunohistochemistry was performed for osteopontin (OPN) and alkaline phosphatase (ALP). Data were analyzed statistically by two-way ANOVA and chi-square test (α = 5%). Teeth in Group CH/120d presented only a slightly enlarged periodontal ligament (PL) with advanced repair. Group aPDT/120d presented the PL moderately enlarged, with moderate inflammatory infiltrate and few collagen fibers. The same pattern was observed at 180 days. AP lesions in CH-treated groups were smaller than those in aPDT-treated groups (p < 0.001) with more blood vessels (p < 0.0001), regardless of the evaluation period, without significant differences in the number of inflammatory cells (p > 0.05). CH-treated groups showed significantly more intense immunostaining for ALP and OPN (p < 0.001) in both periods. Although aPDT stimulated angiogenesis and expression of bone formation markers, the two-session endodontic treatment with CH-based dressing promoted better apical periodontitis repair.

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The aim of the present study was to evaluate, in vitro, the effect of different pre-irradiation times of the photosensitizer in photodynamic therapy in biofilms formed by Streptococcus mutans and Candida albicans, through the evaluation of the microbial load. The factors under study were as follows: times of pre-irradiation of the photosensitizer in three levels (1, 2, or 5 min). For the control of the cariogenic dental biofilm with antimicrobial photodynamic therapy (aPDT), methylene blue (0.01%) was used in association with the diode laser (InGaAlP) with a wavelength of 660 nm. Chlorhexidine digluconate (0.12% CHX) and saline were used as positive and negative controls, respectively. The study design was carried out in complete and randomized blocks. The sample consisted of 15 S. mutans biofilms cultures, randomly divided into five groups and 15 C. albicans cultures, also divided into five groups. The experiment was performed in triplicate (n = 3) and the response variables were obtained through quantitative analysis of bacterial viability, expressed in colony-forming units (CFU) per square millimeter of the specimen area. The data were analyzed with the aid of the ANOVA one-way test and Tukey's post-test. All analyses were performed using the Graph Pad Prism 4.0 program, with a significance level of 5%. For the S. mutans group, only the saline solution presented a statistically significant difference when compared to the other treatments (p < 0.05), that is, the treatment with aPDT, irrespective of the irradiation time applied, was similar to the treatment with CHX and both were more effective in reducing cariogenic biofilm compared to saline. For the group of C. albicans, there was no statistical difference between the groups (p > 0.05). Therefore, it can be concluded that the treatment with aPDT reduced the number of CFUs of S. mutans in a similar way to CHX, independently of the pre-irradiation time applied. No effect of this therapy or of the different pre-irradiation times on the C. albicans biofilm could be observed. In this way, the pre-irradiation time of 1 min can be used to reduce the microbial load of S. mutans.

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AIM: To compare the apical negative pressure irrigation (ANP) with conventional irrigation in the teeth of immature dogs with apical periodontitis. METHODS: Fifty-two immature pre-molar root canals were randomly assigned into 4 groups: ANP (n=15); conventional irrigation (n=17); healthy teeth (control) (n = 10); and teeth with untreated apical periodontitis (control) (n=10). After induction of apical periodontitis, teeth were instrumented using EndoVac® (apical negative pressure irrigation) or conventional irrigation. The animals were euthanized after 90 days. The sections were stained by HE and analyzed under conventional and fluorescence microscopy. TRAP histoenzymology was also performed. Statistical analyses were performed with the significance level set at 5%. RESULTS: There was difference in the histopathological parameters between ANP and conventional groups (p<0.05). The ANP group showed a predominance of low magnitude inflammatory infiltrate, a smaller periodontal ligament, and lower mineralized tissue resorption. There were no differences in the periapical lesion extensions between the ANP and conventional groups (p>0.05). However, a lower number of osteoclasts was observed in the ANP group (p<0.05). CONCLUSION: The EndoVac® irrigation system presented better biological results and more advanced repair process in immature teeth with apical periodontitis than the conventional irrigation system, confirming the hypothesis.

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Abstract Aim: To compare the apical negative pressure irrigation (ANP) with conventional irrigation in the teeth of immature dogs with apical periodontitis. Methods: Fifty-two immature pre-molar root canals were randomly assigned into 4 groups: ANP (n=15); conventional irrigation (n=17); healthy teeth (control) (n = 10); and teeth with untreated apical periodontitis (control) (n=10). After induction of apical periodontitis, teeth were instrumented using EndoVac® (apical negative pressure irrigation) or conventional irrigation. The animals were euthanized after 90 days. The sections were stained by HE and analyzed under conventional and fluorescence microscopy. TRAP histoenzymology was also performed. Statistical analyses were performed with the significance level set at 5%. Results: There was difference in the histopathological parameters between ANP and conventional groups (p<0.05). The ANP group showed a predominance of low magnitude inflammatory infiltrate, a smaller periodontal ligament, and lower mineralized tissue resorption. There were no differences in the periapical lesion extensions between the ANP and conventional groups (p>0.05). However, a lower number of osteoclasts was observed in the ANP group (p<0.05). Conclusion: The EndoVac® irrigation system presented better biological results and more advanced repair process in immature teeth with apical periodontitis than the conventional irrigation system, confirming the hypothesis.

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