Análise da dinâmica do tecido ósseo mandibular, colo de fêmur e coluna vertebral de ratas com osteoporose induzida tratadas com ácido zoledrônico e administração sistêmica de ozônio
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Data
2021-02-22
Autores
Lima Neto, Tiburtino José [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
Este trabalho teve como objetivo analisar o potencial da terapia com ozônio na dinâmica do tecido ósseo de ratas ovariectomizadas tratadas com ácido zolendrônico. Para tanto, 110 ratas Wistar, fêmeas, 6 meses de idade foram submetidas à ovariectomia bilateral (Ovx). Após três meses, dez animais foram submetidos à eutanásia para caracterização da arquitetura do tecido ósseo em microtomografia (Micro ct), os animais restantes foram divididos em dois grupos, o grupo ZOL e o grupo SAL e foi iniciado o tratamento nos animais do grupo ZOL com ácido zoledrônico (100 µg/Kg/28 dias) e, no grupo SAL foi administrado 0,45 ml de solução salina usando o mesmo protocolo do ZOL. Após três meses desta terapia, 10 animais de cada grupo foram submetidos à eutanásia para caracterização da arquitetura óssea (Micro-ct) e os animais restantes foram novamente divididos aleatoriamente, em que os animais do grupo ZOL foram subdivididos em ZOL (n=20) e ZOL+OZN (n=20) e, os animais dos grupos SAL foram subdivididos em SAL (n=20) e SAL+OZN (n=20). Após este procedimento foi iniciada a ozonioterapia em concentração de 0,7mg/kg a cada dois dias até o final do experimento. Após 30 e 60 dias do início da terapia com ozônio, seis animais de cada grupo foram submetidos à eutanásia para a análise e caracterização estrutural óssea das regiões de mandíbula, cabeça femoral e coluna vertebral. Uma parte das peças correspondentes ao colo femoral foram destinada a testes biomecânicos. As demais amostras coletadas foram descalcificadas e as lâminas histológicas foram coradas em hematoxilina e eosina para a análise histométrica (área de osso neoformado), contagem de células inflamatórias (linfócitos) e contagem de osteócitos. Os órgãos do metabolismo e absorção como mesentério, fígado, rins, pulmão e cérebro foram removidos para a avaliação do infiltrado inflamatório e ou qualquer proliferação celular desordenada. Para os parâmetros quantitativos de porcentagem de tecido ósseo na região de cabeça femoral e mandíbula, o teste ANOVA-2 fatores (grupos vs períodos de análise) foi aplicado e em seguida o pós-teste Tukey, quando p<0,05. Já este parâmetro na coluna vertebral, o teste ANOVA-1 fator (grupos experimentais, somente no período de 30 dias) foi aplicado e pós-teste de Tukey. Em todos os testes foi considerado p<0,05. O tratamento com ácido zoledrônio e ozônio (ZOL+OZN) apresentou maior porcentagem de volume ósseo, maior número de osteócitos e maior número de células inflamatórias na região de cabeça femoral, mandíbula com resultados estatisticamente significantes (p<0,05). Na análise biomecânica do colo femoral, o módulo de elasticidade foi semelhante para os grupos ZOL e ZOL+OZN (p>0,05), em comparação aos grupos SAL e SAL+OZN (p<0,05). Este estudo in vivo mostrou que existe um efeito sinérgico entre o ozônio e o ácido zoledrônico com manutenção da massa óssea e restauração da vitalidade do tecido ósseo em ratas ovariectomizadas. Além disso, que a terapia com ozônio foi segura no modelo experimental proposto nesta investigação.
This work aimed to analyze the potential of ozone therapy in the bone tissue dynamics of ovariectomized rats treated with zoledronic acid. For this purpose, 110 Wistar rats, schoolchildren, 6 months old, underwent bilateral ovariectomy (Ovx). After three months, ten animals were euthanized to characterize bone tissue architecture in microtomography (Micro-ct). The remaining animals were divided into two groups: the ZOL group, the SAL group, and treatment was started on the animals in the group. ZOL with zoledronic acid (100 μg / Kg / 28 days) and no SAL group was administered 0.45 ml of saline using the same protocol as ZOL. After three months of this therapy, 10 animals from each group were euthanized to characterize the bone architecture (Micro-ct), and the remaining animals were again randomly divided, in which the animals in the ZOL group were subdivided into ZOL (n = 20) and ZOL + OZN (n = 20) and, the animals in the SAL groups were subdivided into SAL (n = 20) and SAL + OZN (n = 20). After this procedure, ozone therapy was obtained at a concentration of 0.7 mg/kg every two days until the end of the experiment. After 30 and 60 days from the beginning of ozone therapy, six animals from each group were found to be euthanized for analysis and structural bone characterization of the mandible, femoral head, and spine regions. A part of the pieces corresponding to the cervix was destined for biomechanical tests. The others collected were decalcified, and histological slides were stained in hematoxylin and eosin for histometric analysis (area of newly formed bone), inflammatory cell count (lymphocytes), and osteocyte count. The metabolism and absorption organs such as mesentery, liver, kidneys, lung, and brain were removed to evaluate inflammatory infiltrate and or any disorderly cell proliferation. For the quantitative parameters of the percentage of bone tissue in the femoral head and mandible region, the ANOVA-2 factor test (groups vs analysis periods) was provided and then the Tukey post-test, when p <0.05. Already this parameter in the spine, the ANOVA-1 factor test (experimental groups, only in the period of 30 days) was applied and Tukey's post-test. In all tests, p <0.05 was considered. Treatment with zoledronic acid and ozone (ZOL + OZN) showed a higher percentage of bone volume, a greater number of osteocytes, and a greater number of inflammatory cells in the femoral head and mandible region with statistically significant results (p <0.05). In the biomechanical analysis of the femoral neck, the modulus of elasticity was similar for the ZOL and ZOL + OZN groups (p> 0.05), compared to the SAL and SAL + OZN groups (p <0.05). This in vivo study revealed a synergistic effect between ozone and zoledronic acid with the maintenance of bone mass and restoration of bone tissue vitality in ovariectomized rats. Furthermore, that ozone therapy was safe in the experimental model proposed in this investigation.
This work aimed to analyze the potential of ozone therapy in the bone tissue dynamics of ovariectomized rats treated with zoledronic acid. For this purpose, 110 Wistar rats, schoolchildren, 6 months old, underwent bilateral ovariectomy (Ovx). After three months, ten animals were euthanized to characterize bone tissue architecture in microtomography (Micro-ct). The remaining animals were divided into two groups: the ZOL group, the SAL group, and treatment was started on the animals in the group. ZOL with zoledronic acid (100 μg / Kg / 28 days) and no SAL group was administered 0.45 ml of saline using the same protocol as ZOL. After three months of this therapy, 10 animals from each group were euthanized to characterize the bone architecture (Micro-ct), and the remaining animals were again randomly divided, in which the animals in the ZOL group were subdivided into ZOL (n = 20) and ZOL + OZN (n = 20) and, the animals in the SAL groups were subdivided into SAL (n = 20) and SAL + OZN (n = 20). After this procedure, ozone therapy was obtained at a concentration of 0.7 mg/kg every two days until the end of the experiment. After 30 and 60 days from the beginning of ozone therapy, six animals from each group were found to be euthanized for analysis and structural bone characterization of the mandible, femoral head, and spine regions. A part of the pieces corresponding to the cervix was destined for biomechanical tests. The others collected were decalcified, and histological slides were stained in hematoxylin and eosin for histometric analysis (area of newly formed bone), inflammatory cell count (lymphocytes), and osteocyte count. The metabolism and absorption organs such as mesentery, liver, kidneys, lung, and brain were removed to evaluate inflammatory infiltrate and or any disorderly cell proliferation. For the quantitative parameters of the percentage of bone tissue in the femoral head and mandible region, the ANOVA-2 factor test (groups vs analysis periods) was provided and then the Tukey post-test, when p <0.05. Already this parameter in the spine, the ANOVA-1 factor test (experimental groups, only in the period of 30 days) was applied and Tukey's post-test. In all tests, p <0.05 was considered. Treatment with zoledronic acid and ozone (ZOL + OZN) showed a higher percentage of bone volume, a greater number of osteocytes, and a greater number of inflammatory cells in the femoral head and mandible region with statistically significant results (p <0.05). In the biomechanical analysis of the femoral neck, the modulus of elasticity was similar for the ZOL and ZOL + OZN groups (p> 0.05), compared to the SAL and SAL + OZN groups (p <0.05). This in vivo study revealed a synergistic effect between ozone and zoledronic acid with the maintenance of bone mass and restoration of bone tissue vitality in ovariectomized rats. Furthermore, that ozone therapy was safe in the experimental model proposed in this investigation.
Descrição
Palavras-chave
Osteoporose, Regeneração óssea, Ozônio, Osteoporosis