Avaliação do colágeno no osso de ratas diabéticas tipo 1 tratadas com isoflavonas
Data
2022-11-29
Autores
Vieira, Magno César 
Orientadores
Simões, Manuel de Jesus
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
Neste trabalho, o modelo “ratas diabéticas ovariectomizadas” foi utilizado pensando
nas mulheres em menopausa e diabéticas com um tratamento alternativo com
isoflavonas da soja, baseamo-nos em dados que mostram um aumento no número
de mulheres que procuram utilizar-se de terapias hormonais. Objetivo: Analisar a
matriz extracelular no osso de ratas diabéticas tratadas com isoflavonas ou 17β-
estradiol. Métodos: Foram utilizadas 60 ratas (Rattus norvegicus albinus), fêmeas,
adultas, ±3 meses de idade. Os animais foram separados em seis (6) grupos, a
saber: controle Sham (n=10) animais não ovariectomizados na fase de estro;
Sham+DM (n=10) controle Sham ratas diabéticas não ovariectomizadas na fase de
estro; OVX (n=10) controle, ratas ovariectomizadas que receberam veículo
propilenoglicol; OVX+DM (n=10) ratas diabéticas ovariectomizadas que receberam
veículo propilenoglicol (fase de estro); OVX+DM+ISO (n=10) animais diabéticos
ovariectomizados tratados com isoflavonas da soja (150mg/Kg, por gavagem);
OVX+DM+E2 (n=10) animais diabéticos ovariectomizados tratados com estrogênio
(17β-estradiol, 10μg/Kg, por via subcutânea). Para a indução do diabetes Tipo 1, as
ratas receberam uma única injeção intraperitoneal de 60 mg/Kg de estreptozotocina
(STZ, Sigma-Aldrich). O diabetes foi confirmado três dias após a injeção da
estreptozotocina. Todos os animais foram tratados durante 30 dias consecutivos, e
ao final, os animais foram anestesiados e os fêmures removidos e processados para
estudo em H.E. e Picro Sirius Red. Para a análise dos resultados foi utilizado Oneway
ANOVA seguido do teste de Tukey. Foram considerados estatisticamente
significativos experimentos cujo valor de p foi menor que 5% (p ≤ 0,05) para
significância estatística. Os cálculos foram feitos com o programa SPSS versão13
(SPSS, Chicago, IL). Resultados: Os resultados obtidos foram os seguintes: Peso
corporal: não foi diferente entre os grupos não diabéticos Sham e OVX e, os grupos
de ratas diabéticas Sham+DM, OVX+DM, OVX+DM+ISO e OVX+DM+E2 (p>0,05).
No entanto, houve uma diminuição significativa do peso corporal nos grupos de ratas
diabéticas em comparação com os grupos não diabéticas Sham e OVX (p<0,001).
Sensibilidade à Insulina: Os valores mais baixos de insulina tolerância foram
observadas no OVX+DM (2,41±0,95). Os grupos Sham (4,64±0,95) e OVX
(4,57±0,58) apresentaram os maiores valores de sensibilidade à insulina (p<0,001).
x
Os grupos OVX+DM+ISO e OVX+DM+E2 apresentaram valores semelhantes,
inferiores aos Sham e OVX e superior ao grupo OVX+DM. Histomorfometria e
Análise Bioquímica de Glicosaminoglicanos Sulfatados: O volume do osso trabecular
foi maior nos grupos de ratas diabéticas ovariectomizadas OVX+DM+E2
(26,35±2,13) e OVX+DM+ISO (18,36±173), tratadas, respectivamente, com 17β
estradiol e isoflavonas e menor no OVX+DM (8,32±2,52), grupo de ratas diabéticas
ovariectomizadas que receberam veículo propilenoglicol em fase de estro.
Resultados semelhantes foram encontrados em relação à espessura do osso
cortical, que foi maior no OVX+DM+E2 (398,4±1,74) e OVX+DM+ISO (295,6±1,45) e
menor no OVX+DM (142,6±2,74) (p<0,05). Com relação ao sulfato de condroitina foi
encontrado nos ossos de todos os grupos de animais estudados. Picro Sirius Red:
As fibras colágenas de substância óssea trabecular e cortical das epífises dos
fêmures apresentaram maior intensidade de birrefringência nos grupos
(OVX+DM+ISO 330,2±2 e OVX+DM+E2 414,8± 33,7) tratados, respectivamente,
com isoflavonas e 17β estradiol
In this work, the "ovariectomized diabetic rats" model was used thinking of menopausal and diabetic women as an alternative treatment with soy isoflavones, based on data that show an increase in the number of women seeking to use hormone therapies. Objective: To analyze the extracellular matrix in the bone of diabetic rats treated with isoflavones or 17β-estradiol. Methods: 60 adult female rats (Rattus norvegicus albinus), ±3 months old, were used. The animals were separated into six (6) groups, namely: Sham (n=10) control Sham animals not ovariectomized at the estrus phase; Sham+DM (n=10) control Sham diabetic rats not ovariectomized at the estrus phase; OVX (n=10) control, ovariectomized rats that received propylene glycol vehicle; OVX+DM (n=10) diabetic ovariectomized rats that received propylene glycol vehicle (estrus phase); OVX+DM+ISO (n=10) diabetic ovariectomized animals treated with soy isoflavones (150mg/Kg, by gavage); OVX+DM+E2 (n=10) diabetic ovariectomized animals treated with estrogen (17β-estradiol, 10μg/Kg, subcutaneously). For the induction of Type 1 diabetes, rats received a single intraperitoneal injection of 60 mg/Kg streptozotocin (STZ, Sigma-Aldrich). Diabetes was confirmed three days after the streptozotocin injection. All animals were treated for 30 consecutive days, then anesthetized and the femurs were removed and processed for H.E. and Picro Sirius Red. One-way ANOVA was used to analyze the results, followed by Tukey's test. Experiments whose p value was less than 5% (p ≤ 0.05) for statistical significance were considered statistically significant. Calculations were done with SPSS version13 program (SPSS, Chicago, IL). Results: The results obtained were as follows: Body weight: there was no diference between the nondiabetic groups Sham and OVX and, the diabetic rat groups Sham+DM, OVX+DM, OVX+DM+ISO and OVX+DM+E2 (p>0.05). However, there was a significant decrease in body weight in the diabetic rat groups compared to the non-diabetic Sham+DM groups (p<0.001). Insulin sensitivity: The lowest values of insulin tolerance were observed in OVX+DM (2.41±0.95). Groups Sham (4.64±0.95) and OVX (4.57±0.58) had the highest insulin sensitivity values (p<0.001). The OVX+DM+ISO and OVX+DM+E2 groups showed similar values, lower than Sham and OVX and higher than the OVX+DM group. Histomorphometry and Biochemical Analysis of Sulfated Glycosaminoglycans: The volume of trabecular bone was higher xii in the groups of ovariectomized diabetic rats OVX+DM+E2 (26.35±2.13) and OVX+DM+ISO (18.36±173) treated with 17β estradiol and isoflavones, respectively, and lower in OVX+DM (8.32±2.52), a group of ovariectomized diabetic rats that received propylene glycol vehicle in the estrus phase. Similar results were found regarding cortical bone thickness, which was higher in OVX+DM+E2 (398.4±1.74) and OVX+DM+ISO (295.6±1.45) and lower in OVX+DM (142.6±2.74) (p<0.05). Chondroitin sulfate was found in the bones of all groups of animals studied. Picro Sirius Red: Collagen fibers of trabecular and cortical bone substance of femoral epiphyses showed higher birefringence intensity in the groups (OVX+DM+ISO 330.2±2 and OVX+DM+E2 414.8± 33.7) treated, respectively, with isoflavones and 17β estradiol.
In this work, the "ovariectomized diabetic rats" model was used thinking of menopausal and diabetic women as an alternative treatment with soy isoflavones, based on data that show an increase in the number of women seeking to use hormone therapies. Objective: To analyze the extracellular matrix in the bone of diabetic rats treated with isoflavones or 17β-estradiol. Methods: 60 adult female rats (Rattus norvegicus albinus), ±3 months old, were used. The animals were separated into six (6) groups, namely: Sham (n=10) control Sham animals not ovariectomized at the estrus phase; Sham+DM (n=10) control Sham diabetic rats not ovariectomized at the estrus phase; OVX (n=10) control, ovariectomized rats that received propylene glycol vehicle; OVX+DM (n=10) diabetic ovariectomized rats that received propylene glycol vehicle (estrus phase); OVX+DM+ISO (n=10) diabetic ovariectomized animals treated with soy isoflavones (150mg/Kg, by gavage); OVX+DM+E2 (n=10) diabetic ovariectomized animals treated with estrogen (17β-estradiol, 10μg/Kg, subcutaneously). For the induction of Type 1 diabetes, rats received a single intraperitoneal injection of 60 mg/Kg streptozotocin (STZ, Sigma-Aldrich). Diabetes was confirmed three days after the streptozotocin injection. All animals were treated for 30 consecutive days, then anesthetized and the femurs were removed and processed for H.E. and Picro Sirius Red. One-way ANOVA was used to analyze the results, followed by Tukey's test. Experiments whose p value was less than 5% (p ≤ 0.05) for statistical significance were considered statistically significant. Calculations were done with SPSS version13 program (SPSS, Chicago, IL). Results: The results obtained were as follows: Body weight: there was no diference between the nondiabetic groups Sham and OVX and, the diabetic rat groups Sham+DM, OVX+DM, OVX+DM+ISO and OVX+DM+E2 (p>0.05). However, there was a significant decrease in body weight in the diabetic rat groups compared to the non-diabetic Sham+DM groups (p<0.001). Insulin sensitivity: The lowest values of insulin tolerance were observed in OVX+DM (2.41±0.95). Groups Sham (4.64±0.95) and OVX (4.57±0.58) had the highest insulin sensitivity values (p<0.001). The OVX+DM+ISO and OVX+DM+E2 groups showed similar values, lower than Sham and OVX and higher than the OVX+DM group. Histomorphometry and Biochemical Analysis of Sulfated Glycosaminoglycans: The volume of trabecular bone was higher xii in the groups of ovariectomized diabetic rats OVX+DM+E2 (26.35±2.13) and OVX+DM+ISO (18.36±173) treated with 17β estradiol and isoflavones, respectively, and lower in OVX+DM (8.32±2.52), a group of ovariectomized diabetic rats that received propylene glycol vehicle in the estrus phase. Similar results were found regarding cortical bone thickness, which was higher in OVX+DM+E2 (398.4±1.74) and OVX+DM+ISO (295.6±1.45) and lower in OVX+DM (142.6±2.74) (p<0.05). Chondroitin sulfate was found in the bones of all groups of animals studied. Picro Sirius Red: Collagen fibers of trabecular and cortical bone substance of femoral epiphyses showed higher birefringence intensity in the groups (OVX+DM+ISO 330.2±2 and OVX+DM+E2 414.8± 33.7) treated, respectively, with isoflavones and 17β estradiol.
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Citação
Avaliação do colágeno no osso de ratas diabéticas tipo 1 tratadas com isoflavonas