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No. of bitstreams: 1 NAT?LIA_PREDELLA_CIGNACHI_TES.pdf: 11102267 bytes, checksum: 121fbf4d2b9ae1c4a36e21d23ab3140d (MD5) Previous issue date: 2018-01-25 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / This thesis encompasses two parts: firstly, we compared the bone healing in female and male mice, after induction of type 1 diabetes (T1D); secondly, the bone regeneration was evaluated in a menopause model induced by bilateral ovariectomy (OVX), with or without T1D induction. For the part I, the animals (female and male) were initially assigned into two groups, namely control or T1D (elicited by streptozotocin; STZ). In the part II, the females were divided into four experimental groups: sham-operated or OVX, with or without STZ T1D induction. After T1D induction, a monocortical femoral defect was created. In either parts of the present study, we evaluated the effects of supplementation with vitamin D3 and/or insulin. In the second part, the effects of estrogen replacement were also analyzed. Following 21 days of bone defect creation, the animals were euthanized; the femurs and blood were collected for posterior analysis. Both T1D females and males presented a reduction in body weight gain, associated with hyperglycemia. There were no changes in the serum levels of the pro-inflammatory cytokines [interleukin-1? (IL-1?), tumor necrosis factor (TNF) and interferon-? (IFN-?)] in all the evaluated groups. T1D mice of both sexes presented a delayed bone regeneration, according to the histological and micro-CT assessment. The supplementation with vitamin D3 restored the bone healing in female and male T1D mice, reaching values close to controls. The insulin therapy improved the bone remodeling in T1D mice of both sexes, but the effects of this hormone were superior in males. The evaluation of osteoclast activity did not reveal significant differences among the experimental groups. Real time PCR revealed slight differences in the mRNA expression of two transcription factors related to osteoblast differentiation, namely runx2 and osterix, as measured in the area into the bone defect. A higher upregulation of both factors was seen in T1D males treated with vitamin D3. Conversely, vitamin D3-treated T1D females displayed an upregulation of insulin-like growth factor 1 (IGF-1), further indication sex-related differences for the treatments. Besides the experimental protocols described for the 12 part I of this thesis, in the part II, we also evaluated some behavioral locomotor parameters and serum levels of calcium and alkaline phosphatase. OVX animals presented increased body weight gain, accompanied by uterus atrophy. Otherwise, T1D induction elicited a reduction of body weight gain, which was more pronounced in OVX-T1D animals. Serum levels of alkaline phosphatase were divergent in the non-diabetic and T1D OVX animals. Calcium or cytokine levels were similar in all the experimental groups. The sham-operated T1D, the non-diabetic OVX and the OVX-T1D groups presented a delayed bone regeneration, as indicated by histological and micro-CT analysis. Estrogen replacement improved the bone healing in all OVX groups. There was a trend toward an upregulation of IGF-1 mRNA in non-diabetic OVX animals, which was not mirrored in OVX-T1D mice. Locomotor parameters remained unaltered, except by a general reduction of rearing numbers in T1D animals. / A presente tese est? dividida em duas partes: na parte I, comparou-se a regenera??o ?ssea em camundongos f?meas e machos, com diabetes do tipo 1 (T1D). Na parte II, foi avaliada a regenera??o ?ssea no modelo de menopausa experimental induzido por ovariectomia (OVX), com ou sem T1D. Na parte I, os animais (machos e f?meas) foram divididos em dois grandes grupos: controle e T1D (induzido por estreptozotocina; STZ). Na parte II, os animais foram divididos em quatro grandes grupos: f?meas falso-operadas e OVX, com ou sem T1D. Ap?s a indu??o do T1D, foi criado um defeito ?sseo monocortical no f?mur. Nas duas partes do trabalho, foram avaliados os efeitos da suplementa??o com vitamina D3 e/ou insulina. Na segunda parte, tamb?m se avaliou o efeito da reposi??o hormonal com estradiol. Decorridos 21 dias do procedimento da cria??o do defeito, os animais foram eutanasiados; o f?mur e o sangue foram coletados para an?lises posteriores. Tanto as f?meas, quanto os machos T1D, apresentaram uma redu??o do ganho de peso corporal, associado ? hiperglicemia. N?o houve altera??es nos n?veis s?ricos das citocinas pr?-inflamat?rias interleucina-1? (IL-1?), fator de necrose tumoral (TNF) ou interferon-? (IFN?). Os animais T1D, de ambos os sexos, apresentaram um comprometimento na regenera??o ?ssea, como demonstrado pelas an?lises histol?gicas e de micro-CT. A suplementa??o com vitamina D3 reestabeleceu a regenera??o ?ssea em f?meas e machos T1D, apresentando valores pr?ximos aos encontrados nos animais do grupo controle. A terapia com insulina melhorou a remodela??o ?ssea nas f?meas e machos T1D; por?m, os efeitos foram mais pronunciados nos machos. A avalia??o da atividade osteocl?stica n?o revelou diferen?as significativas entre os grupos experimentais, ap?s a indu??o de T1D, em machos ou f?meas. Os resultados do PCR em tempo real para runx2 e osterix (dois fatores de transcri??o relacionados aos osteoblastos), no tecido ?sseo, n?o demonstraram nenhuma diferen?a significativa, exceto por um aumento nos n?veis de RNAm dos dois fatores nos camundongos machos T1D, que receberam suplementa??o com vitamina 10 D3. Por outro lado, f?meas T1D que receberam vitamina D3 apresentaram um aumento na express?o dos n?veis de RNAm para o fator de crescimento semelhante ? insulina do tipo 1 (IGF-1), quando comparado com os machos que tiveram o mesmo tratamento, sugerindo assim, diferen?as relacionadas ao sexo. Al?m das an?lises j? mencionadas anteriormente, na parte II da tese, par?metros comportamentais e n?veis s?ricos de c?lcio e fosfatase alcalina tamb?m foram analisados. Os resultados da segunda parte do trabalho demonstraram que os animais submetidos a OVX tiveram um aumento do peso corporal, com atrofia uterina. Em contrapartida, quando foi induzido T1D, houve uma diminui??o do peso corporal mais acentuada no grupo OVX que no grupo falso-operado. Os animais falso-operados e OVX T1D apresentaram hiperglicemia, confirmando o desenvolvimento do diabetes. Os n?veis s?ricos de fosfatase alcalina foram divergentes entre os grupos n?o-diab?ticos OVX e OVX T1D. N?o houve varia??es dos n?veis de c?lcio. Os animais falso-operados T1D, n?o diab?ticos OVX e OVX T1D apresentaram preju?zos similares na regenera??o ?ssea, como observado pelas an?lises histol?gicas e imagens de micro-CT. A reposi??o com estradiol melhorou a cicatriza??o ?ssea nos animais n?o-diab?ticos OVX e OVX T1D. Houve uma tend?ncia ao aumento nos n?veis de RNAm de IGF-1 no grupo OVX, o que n?o foi observado quando da associa??o de T1D e menopausa. N?o foram observadas diferen?as na atividade locomotora dos animais com T1D e/ou OVX, a n?o ser por uma diminui??o no n?mero de rearings no grupo falso-operado T1D, independente do tratamento com vitamina D3, insulina e estradiol, de forma isolada ou em associa??o.