Photosensitizers and exposure times to light showed tissue compatibility in isogenic mice / Fotossensibilizadores e tempo de exposição a luz em relação a compatibilidade de tecidos em ratos isogênicos

Objective: The aim of this study was to evaluate the subcutaneous tissue response after different protocols to photodynamic therapy (PDT). In Phase 1, were tested the diode laser (used for 1min) associated to the photosensitizer phenothiazine chloride solution (PCS) in different concentrations. In Phase 2 ­ the diode laser and LED were tested associated to two different photosensitizers, PCS and Curcumin, in different exposure times of light application. Material and Methods: After 7, 21 and 63-days the animals were euthanized and the subcutaneous tissue processed to histological analysis. Qualitative and semi-quantitative descriptions of the inflammatory process and immunohistochemical technique were performed. The obtained data were analyzed by Kruskal-Wallis and Dunn's post-test (α= 0.5). Results: On Phase 1, the tissue response was very similar among the groups. For the inflammatory infiltrate, PCS with concentration of 10mg/mL exhibited the most intense reaction (p > 0.05). On Phase 2, at 7-days period, the analyzed parameters presented small magnitude and after 21 and 63-days, all the parameters demonstrated tissue compatibility. Conclusion: Both photosensitizers presented proper tissue compatibility regardless the different concentrations used on Phase 1 and different durations of light exposure on Phase 2 (AU)

Objetivo: Este estudo avaliou a resposta do tecido subcutâneo após terapia fotodinâmica, utilizando na Fase 1 - laser diodo por 1min e solução fotossensibilizadora de cloreto de fenotiazina (CF) em diferentes concentrações e Fase 2 - laser diodo e LED e dois fotossensibilizadores, CF e Curcumina, em diferentes tempos de exposição da aplicação de luz. Material e Métodos: Após 7, 21 e 63 dias, foram realizadas descrições qualitativas e semiquantitativas do processo inflamatório e técnica de imunoistoquímica. Os dados foram analisados pelo pós-teste de Kruskal-Wallis e Dunn (α = 0,5). Resultados: Na Fase 1, a resposta do tecido foi muito semelhante. O infiltrado inflamatório, na concentração de 10 mg / mL, exibiu reação mais intensa (p > 0,05). Na Fase 2, aos 7 dias, os parâmetros analisados apresentaram pequena magnitude. Aos 21 e 63 dias, todos os parâmetros demonstraram compatibilidade com o tecido. Conclusão: Ambos os fotossensibilizadores apresentaram compatibilidade de tecido adequada, independentemente das diferentes concentrações utilizadas na Fase 1 e diferentes durações de exposição à luz na Fase 2 (AU)

Immunohistochemical and mRNA expression of RANK, RANKL, OPG, TLR2 and MyD88 during apical periodontitis progression in mice.

OBJECTIVE: To evaluate and correlate, in the same research, the mRNA expression and the staining of RANK, RANKL, OPG, TLR2 and MyD88 by immunohistochemistry in the apical periodontitis (AP) progression in mice. MATERIAL AND METHODS: AP was induced in the lower first molars of thirty-five C57BL/6 mice. They were assigned to four groups according to their euthanasia periods (G0, G7, G21 and G42). The jaws were removed and subjected to histotechnical processing, immunohistochemistry and real-time reverse transcription-PCR (qRT-PCR). Data were analyzed with parametric and nonparametric tests (α=0.05). RESULTS: An increase of positive immunoreactivity for RANK, RANKL, OPG, TLR2 and MyD88 was observed over time (p<0.05). The RANKL expression was different between the groups G0 and G42, G21 and G42 (p=0.006), with G42 presenting the higher expression in both comparations. The OPG expression was statistically different between the groups G0 and G7, G7 and G21 and G7 and G42 (p<0.001), with G7 presenting higher expression in all the time points. The TLR2 expression was different between the groups G0 and G42 (p=0.03), with G42 showing the higher expression. The MyD88 expression presented a statistical significant difference between groups G7, G21 and G42 compared with G0 (p=0.01), with G0 presenting the smallest expression in all the comparisons. The Tnfrsf11/Tnfrsf11b (RANKL/OPG) ratio increased with the AP progression (p=0.002). A moderate positive correlation between MyD88 and RANKL (r=0.42; p=0.03) and between MyD88 and TLR2 (r=0.48; p<0.0001) was observed. CONCLUSION: The expression of the RANK, RANKL, OPG, MyD88 and TLR2 proteins as well as the ratio Tnfrsf11/Tnfrsf11b (RANKL/OPG) increased with AP progression. There was also a moderate positive correlation between the expression Myd88-Tnfrsf11 and Tlr2-Myd88, suggesting the relevance of Tlr2-Myd88 in bone loss due to bacterial infection.

MMP2 and MMP9 are Associated with Apical Periodontitis Progression and Might be Modulated by TLR2 and MyD88

Abstract The aim of this study was to evaluate the expression of MMP2 and MMP9 during apical periodontitis (AP) progression in TLR2 (TLR2 KO) and in MyD88 (MyD88 KO) knockout mice compared to wild type (WT) mice. AP was induced in mandibular first molars of TLR2 KO (n= 18), MyD88 KO (n= 18), and WT mice (n= 18). After 7, 21, and 42 days, the animals were euthanized and the jaws were dissected and subjected to histotechnical processing. Subsequent sections were stained by immunohistochemistry and evaluated for detection of MMP2 and MMP9. Statistical analysis of the semi-quantitative analysis of immunohistochemistry was performed using chi-square test (α = 0.05). In the initial periods of AP progression, an increased expression of MMP9 in the TLR2 KO and MyD88 KO mice was observed. In the final periods of AP progression, a reduction of MMP2 expression and an increase of MMP9 expression in the TLR2 KO mice were observed. MMP2 and MMP9 production was modulated for TLR2 and MyD88 during apical periodontitis progression.

Resumo O objetivo deste estudo foi avaliar a expressão de MMP2 e MMP9 durante a progressão da periodontite apical (AP) em camundongos knockout para TLR2 (TLR2 KO) e MyD88 (MyD88 KO) comparados aos camundongos wild type (WT). A AP foi induzida nos primeiros molares inferiores dos camundongos TLR2 KO (n = 18), MyD88 KO (n = 18) e WT (n = 18). Após 7, 21 e 42 dias, os animais foram eutanaziados e as mandíbulas foram dissecadas e submetidas a processamento histotécnico. As lâminas foram coradas por imuno-histoquímica e analisadas para a detecção de MMP2 e MMP9. A análise estatística semi-quantitativa da imuno-histoquímica foi realizada pelo teste qui-quadrado (α = 0,05). Nos períodos iniciais de progressão AP, foi observada uma expressão aumentada de MMP9 nos camundongos TLR2 KO e MyD88 KO. Nos períodos finais de progressão AP, observou-se uma redução da expressão de MMP2 e um aumento da expressão de MMP9 nos camundongos TLR2 KO. A produção de MMP2 e MMP9 foi modulada por TLR2 e MyD88 durante a progressão da periodontite apical.

MMP2 and MMP9 are Associated with Apical Periodontitis Progression and Might be Modulated by TLR2 and MyD88.

The aim of this study was to evaluate the expression of MMP2 and MMP9 during apical periodontitis (AP) progression in TLR2 (TLR2 KO) and in MyD88 (MyD88 KO) knockout mice compared to wild type (WT) mice. AP was induced in mandibular first molars of TLR2 KO (n= 18), MyD88 KO (n= 18), and WT mice (n= 18). After 7, 21, and 42 days, the animals were euthanized and the jaws were dissected and subjected to histotechnical processing. Subsequent sections were stained by immunohistochemistry and evaluated for detection of MMP2 and MMP9. Statistical analysis of the semi-quantitative analysis of immunohistochemistry was performed using chi-square test (α = 0.05). In the initial periods of AP progression, an increased expression of MMP9 in the TLR2 KO and MyD88 KO mice was observed. In the final periods of AP progression, a reduction of MMP2 expression and an increase of MMP9 expression in the TLR2 KO mice were observed. MMP2 and MMP9 production was modulated for TLR2 and MyD88 during apical periodontitis progression.

Immunohistochemical and mRNA expression of RANK, RANKL, OPG, TLR2 and MyD88 during apical periodontitis progression in mice

Abstract Objective To evaluate and correlate, in the same research, the mRNA expression and the staining of RANK, RANKL, OPG, TLR2 and MyD88 by immunohistochemistry in the apical periodontitis (AP) progression in mice. Material and Methods AP was induced in the lower first molars of thirty-five C57BL/6 mice. They were assigned to four groups according to their euthanasia periods (G0, G7, G21 and G42). The jaws were removed and subjected to histotechnical processing, immunohistochemistry and real-time reverse transcription-PCR (qRT-PCR). Data were analyzed with parametric and nonparametric tests (α=0.05). Results An increase of positive immunoreactivity for RANK, RANKL, OPG, TLR2 and MyD88 was observed over time (p<0.05). The RANKL expression was different between the groups G0 and G42, G21 and G42 (p=0.006), with G42 presenting the higher expression in both comparations. The OPG expression was statistically different between the groups G0 and G7, G7 and G21 and G7 and G42 (p<0.001), with G7 presenting higher expression in all the time points. The TLR2 expression was different between the groups G0 and G42 (p=0.03), with G42 showing the higher expression. The MyD88 expression presented a statistical significant difference between groups G7, G21 and G42 compared with G0 (p=0.01), with G0 presenting the smallest expression in all the comparisons. The Tnfrsf11/Tnfrsf11b (RANKL/OPG) ratio increased with the AP progression (p=0.002). A moderate positive correlation between MyD88 and RANKL (r=0.42; p=0.03) and between MyD88 and TLR2 (r=0.48; p<0.0001) was observed. Conclusion The expression of the RANK, RANKL, OPG, MyD88 and TLR2 proteins as well as the ratio Tnfrsf11/Tnfrsf11b (RANKL/OPG) increased with AP progression. There was also a moderate positive correlation between the expression Myd88-Tnfrsf11 and Tlr2-Myd88, suggesting the relevance of Tlr2-Myd88 in bone loss due to bacterial infection.

Three-Dimensional Micro-Computed Tomography Analyses of Induced Periapical Lesions in Transgenic Mice.

The aim of this study was to evaluate the three-dimensional (3D) parameters given by the micro-computed tomography (µCT) analysis of experimentally induced periapical lesions in wild type (WT) and knockout mice for the interleukin 22 (IL-22 KO). Periapical lesions were induced in the mandibular first molars of wild type and IL-22 KO mice (n = 12 teeth/group). The animals were euthanized after the experimental periods of 7, 21 and 42 days. The mandibles were removed and exposed to µCT scanning. The analyses were performed by the CTAn software for the tree-dimensional parameters: Tissue Volume (TV), Lesion Volume (LV), Tissue Surface (TS), Lesion Surface (LS), Intersection Surface (IS), and Trabecular Pattern factor (Tb.Pf). After that, the tissue was subjected to routine histologic procedures and to immunohistochemistry analysis. Statistical analysis was performed in the GraphPad software. A t-test was used to compare the differences between the groups with significance level of 5%. The evaluation of the 3D parameters showed statistical significant difference between the groups only at the latest period of periapical lesion development (42 days), for the TV, LV, TS, LS and IS parameters. The immunohistochemistry evaluation confirmed the immunostaining for IL-22 only in the WT mice, surrounding the periapical lesion. There were no differences regarding the trabecular alveolar bone (Tb.Pf) that could influence the lesion development. In conclusion, the 3D parameters showed that the absence of IL-22 leads to detectable differences at 42 days of lesion progression, resulting in smaller periapical lesions.