Efeito do PTH tópico incrementado ao Biogran no processo de reparo alveolar em ratos submetidos à orquiectomia: análise imunoistoquímica, por microtomografia computadorizada e microscopia confocal a laser.
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Data
2020-09-17
Autores
Frigério, Paula Buzo [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
Esse estudo teve como objetivo avaliar um enxerto ósseo sintético (BioGran®) em associação ao PTH 1-34 tópico no reparo alveolar de ratos orquiectomizados. 96 ratos foram divididos em dois grupos: SHAM (cirurgia fictícia) e ORQ (orquiectomia bilateral), os quais foram divididos em três subgrupos, CLOT (defeito alveolar sem biomaterial); BG (defeito com BioGran®); BG-PTH (defeito com BioGran® + PTH tópico). A exodontia do incisivo superior direito e inserção do biomaterial com ou sem PTH 1-34 tópico foi realizada 30 dias após a orquiectomia e cirurgia fictícia. A 14 dias após a exodontia, foi administrado o fluorocromo calceína e passados mais 28 dias, foi administrado o fluorocromo vermelho de alizarina. A eutanásia foi realizada a 60 dias após a cirurgia de exodontia, na qual as maxilas contendo os alvéolos em reparação foram removidas para análise imunoistoquímica direcionada para os imunomarcadores: OPG, RANKL, TRAP, OC, Wnt e Bcat; para a microtomografia computadorizada, usou-se os parâmetros BV, BV/TV, Tb.Th, Tb.N, Tb.Sp e Po(Tot); e a análise por microscopia confocal a laser determinou MAR, AON e a histometria e dinâmica óssea. Os dados quantitativos da Micro-CT e da microscopia confocal (MAR, AON e histometria e dinâmica óssea), passaram pela análise de homocedasticidade (teste de Shapiro-Wilk) para distinção dos dados paramétricos e não paramétricos. Os dados paramétricos foram comparados pelo teste ANOVA One-Way e pós-teste Tukey e para os não paramétricos foi utilizado o teste ANOVA Two-Way e pós-teste de Tukey. Adotou-se o nível de significância p < 0,05. Nos resultados da imunoistoquímica, observou-se expressão equilibrada de OPG e RANKL no grupo BG-PTH, tanto no grupo SHAM quanto em ORQ. Para TRAP a expressão foi leve nos grupos SHAM/ORQ BG-PTH e para os grupos SHAM CLOT e ORQ CLOT a expressão foi de leve a moderada. Quanto as proteínas OC, Wnt e Bcat indicaram uma maior marcação após a funcionalização de BioGran® com teriparatida. Para os demais grupos, CLOT e BG, a marcação foi leve a moderada. Na micro-CT, foi obervado diferença estatística para os parâmetros, BV/TV, Tb.Th, Tb.Sp e Po(tot) na comparação dos grupos ORQ/SHAM CLOT vs BG vs BG-PTH. Os resultados apontaram que para o grupo SHAM houve uma perspectiva de melhora quando existe a funcionalização BioGran® + teriparatida e a funcionalização apenas do biomaterial colaborou com a manutenção do alvéolo reparacional dos demais grupos. Na análise de microscopia confocal, para MAR houve diferença estatística entre os grupos ORQ BG e SHAM BG-PTH, o grupo que apresentou maior precipitação de cálcio foi o SHAM BG-PTH. O parâmetro AON mostrou maiores resultados para grupo SHAM CLOT e quanto a histometria e dinâmica óssea que faz a comparação entre os grupos ORQ/SHAM CLOT vs BG vs BG-PTH, os grupos que apresentaram maiores valores para alizarina (osso novo) foram ORQ BG e SHAM BG e para calceína (osso antigo) foi ORQ CLOT. A partir de todos os resultados apresentados, podemos notar que a utilização do BioGran® com ou sem PTH 1-34 tópico promoveu discreta melhora no reparo alveolar de ratos orquiectomizados.
The purpose of this study is to evaluate a synthetic bone graft (BioGran®) in association with topical PTH 1-34 in the alveolar repair of orchiectomized rats. 96 rats were divided into two groups: SHAM (fictitious surgery) and ORQ (bilateral orchiectomy), which were divided into three subgroups, COAG (alveolar defect without biomaterial); BG (defect with BioGran®); BG-PTH (defect with BioGran® + topical PTH). The extraction of the right upper incisor and insertion of the biomaterial with or without topical PTH 1-34 was performed 30 days after the orchiectomy and fictitious surgery. 14 days after tooth extraction, fluorochrome calcein was administered and after another 28 days, alizarin red fluorochrome was administered. Euthanasia was performed 60 days after extraction surgery, in which the jaws containing the alveoli under repair were removed for immunohistochemical analysis directed to the immunostaining: OPG, RANKL, TRAP, OC, Wnt and Bcat; for computed microtomography, the parameters BV, BV / TV, Tb.Th, Tb.N, Tb.Sp and Po (Tot) were used; and analysis by laser confocal microscopy determined MAR, AON and histometry and bone dynamics. Quantitative data from Micro-CT and confocal microscopy (MAR, AON and histometry and bone dynamics) were analyzed by homoscedasticity (Shapiro-Wilk test) to distinguish between parametric and nonparametric data. Parametric data were compared using the ANOVA One-Way test and Tukey post-test and for non-parametric ANOVA Two-Way test and Tukey posttest. The significance level was set at p < 0.05. In the immunohistochemistry results, a balanced expression of OPG and RANKL was observed in the BG-PTH group, in both SHAM and ORQ groups. For TRAP the expression was light in the SHAM/ORQ BGPTH groups and for the SHAM COAG and ORQ COAG groups the expression was light to moderate. As for the OC proteins, Wnt and Bcat indicated a higher marking after the functionalization of BioGran® with teriparatide. For the other groups, COAG and BG, the marking was light to moderate. In the micro-CT, a statistical difference was observed for the parameters, BV / TV, Tb.Th, Tb.Sp and Po (tot) when comparing the ORQ/SHAM COAG vs BG vs BG-PTH groups. The results showed that for the SHAM group there was a prospect of improvement when there is the BioGran® + teriparatide functionalization and the functionalization of only the biomaterial collaborated with the maintenance of the reparational socket of the other groups. In the analysis of confocal microscopy, for MAR there was a statistical difference between the ORQ BG and SHAM BG-PTH groups, the group that showed greater calcium precipitation was the SHAM BG-PTH. The AON parameter showed the highest results for the SHAM COAG group and as for the histometry and bone dynamics that makes the comparison between the ORQ/SHAM COAG groups vs BG vs BG-PTH, the groups that presented higher values for alizarin (new bone) were ORQ BG and SHAM BG and for calcein (old bone) was ORQ COAG. From all the results presented, we realized that the use of BioGran® with or without topical PTH 1-34 promoted a slight improvement in the alveolar repair of orchiectomized rats.
The purpose of this study is to evaluate a synthetic bone graft (BioGran®) in association with topical PTH 1-34 in the alveolar repair of orchiectomized rats. 96 rats were divided into two groups: SHAM (fictitious surgery) and ORQ (bilateral orchiectomy), which were divided into three subgroups, COAG (alveolar defect without biomaterial); BG (defect with BioGran®); BG-PTH (defect with BioGran® + topical PTH). The extraction of the right upper incisor and insertion of the biomaterial with or without topical PTH 1-34 was performed 30 days after the orchiectomy and fictitious surgery. 14 days after tooth extraction, fluorochrome calcein was administered and after another 28 days, alizarin red fluorochrome was administered. Euthanasia was performed 60 days after extraction surgery, in which the jaws containing the alveoli under repair were removed for immunohistochemical analysis directed to the immunostaining: OPG, RANKL, TRAP, OC, Wnt and Bcat; for computed microtomography, the parameters BV, BV / TV, Tb.Th, Tb.N, Tb.Sp and Po (Tot) were used; and analysis by laser confocal microscopy determined MAR, AON and histometry and bone dynamics. Quantitative data from Micro-CT and confocal microscopy (MAR, AON and histometry and bone dynamics) were analyzed by homoscedasticity (Shapiro-Wilk test) to distinguish between parametric and nonparametric data. Parametric data were compared using the ANOVA One-Way test and Tukey post-test and for non-parametric ANOVA Two-Way test and Tukey posttest. The significance level was set at p < 0.05. In the immunohistochemistry results, a balanced expression of OPG and RANKL was observed in the BG-PTH group, in both SHAM and ORQ groups. For TRAP the expression was light in the SHAM/ORQ BGPTH groups and for the SHAM COAG and ORQ COAG groups the expression was light to moderate. As for the OC proteins, Wnt and Bcat indicated a higher marking after the functionalization of BioGran® with teriparatide. For the other groups, COAG and BG, the marking was light to moderate. In the micro-CT, a statistical difference was observed for the parameters, BV / TV, Tb.Th, Tb.Sp and Po (tot) when comparing the ORQ/SHAM COAG vs BG vs BG-PTH groups. The results showed that for the SHAM group there was a prospect of improvement when there is the BioGran® + teriparatide functionalization and the functionalization of only the biomaterial collaborated with the maintenance of the reparational socket of the other groups. In the analysis of confocal microscopy, for MAR there was a statistical difference between the ORQ BG and SHAM BG-PTH groups, the group that showed greater calcium precipitation was the SHAM BG-PTH. The AON parameter showed the highest results for the SHAM COAG group and as for the histometry and bone dynamics that makes the comparison between the ORQ/SHAM COAG groups vs BG vs BG-PTH, the groups that presented higher values for alizarin (new bone) were ORQ BG and SHAM BG and for calcein (old bone) was ORQ COAG. From all the results presented, we realized that the use of BioGran® with or without topical PTH 1-34 promoted a slight improvement in the alveolar repair of orchiectomized rats.
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Palavras-chave
Osteoporose, Regeneração óssea, Hormônio paratireóideo, Parathyroid hormone, Osteoporosis