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Os benefícios à saúde relacionados ao consumo moderado de vinho incluem diferentes mecanismos, nos quais estão envolvidos tanto etanol quanto compostos fenólicos que são constituintes do mesmo. Com o objetivo de avaliar variações glicêmicas, ponderais e o depósito de triglicérides, colesterol e glicogênio hepáticos com uso regular de vinho tinto em camundongo ApoE Knockout, foram utilizados 60 camundongos machos adultos ApoE Knockout de peso médio de 30 gramas, distribuídos em três grupos de 20 animais: grupo vinho, grupo etanol e grupo água, os quais receberam 50 mL de vinho e 50 mL água, 6mL de etanol e 94mL de água e somente água respectivamente por quatro meses. Os parâmetros avaliados foram: variações glicêmicas, ponderais, acúmulo de triglicerídeos, colesterol e glicogênio hepáticos. O grupo vinho teve em relação a sua massa corporal uma área sob a curva maior que a dos outros dois grupos, mas com um percentual pequeno de aumento. A concentração do triglicerídeo hepático foi maior no grupo vinho 57% em relação ao grupo etanol, que foi 31,6% menor que o controle (p<0,01%). A concentração do colesterol hepático foi menor no grupo vinho (23,6%), assim como no grupo etanol (24,5%), (p<0,05%). A concentração do glicogênio hepático foi maior no grupo vinho (16%), porém não alcançando significado estatístico. A glicemia em jejum no dia da eutanásia foi maior no grupo etanol em relação aos demais grupos, porém não demonstrou diferença estatisticamente significante. Na análise histológica não foi observada diferença significativa entre os grupos, embora o peso médio em gramas nas gorduras, retroperitoneal e subcutâneas tenha sido aproximadamente duas vezes maior no grupo vinho. Concluiu-se que neste estudo o uso regular e crônico de vinho tinto aumentou triglicerídeo hepático, porém o álcool diminui o colesterol hepático. O aumento do triglicerídeo pode ser devido ao alto valor calórico do vinho ou alguma propriedade lipogênica desconhecida que levou ao aumento importante das gorduras retroperitoneais e subcutâneas em camundongos ApoE Knockout. / The benefits to health related to regular consume of red wine includes different mechanisms in which are involved both ethanol and fenolics compounds of the wine. With the objective to evaluate glycemia, lipid profile and weight variations with regular use of red wine by ApoE Knockout mices, sixty adults ApoE Knockout mices weighing around 30g were distributed into 3 groups of 20 animals each: 1.Wine that received 50mL of wine plus 50mL of water, 2. Ethanol and Water groups, 6mL of ethanol plus 94mL of water and just water respectively for 4 months. We evaluate glycemia, weight variations and liver glycogen, triglycerides and cholesterol. The wine group had in relation to its mass body an area under the curve larger than the other two groups, but with a small percentage of increase. The concentration of liver triglycerides was higher in the wine 57% compared to ethanol group, which was 31.6% lower than the control (p<0.01%). The concentration of liver cholesterol was lower in wine (23.6%) and in ethanol group (24.5%) (p<0.05%). The liver glycogen concentration was higher in the wine (16%), although not reaching statistical significance. The fasting glicemia on the day of euthanasia was higher in the ethanol group compared to other groups, but not statistically significant difference. In histological analysis was not significantly different between groups, although the average weight in grams fat, retroperitoneal and subcutaneous was approximately two times higher in the wine group. It was concluded that in this study the regular and chronic use of red wine increased liver triglyceride, however alcohol decreases liver cholesterol. The increase of the triglyceride may be due to the high caloric value of wine or some lipogenic unknown property that led to an important increase in retroperitoneal and subcutaneous fat tissue in ApoE Knockout mice.

Os benefícios à saúde relacionados ao consumo moderado de vinho incluem diferentes mecanismos, nos quais estão envolvidos tanto etanol quanto compostos fenólicos que são constituintes do mesmo. Com o objetivo de avaliar variações glicêmicas, ponderais e o depósito de triglicérides, colesterol e glicogênio hepáticos com uso regular de vinho tinto em camundongo ApoE Knockout, foram utilizados 60 camundongos machos adultos ApoE Knockout de peso médio de 30 gramas, distribuídos em três grupos de 20 animais: grupo vinho, grupo etanol e grupo água, os quais receberam 50 mL de vinho e 50 mL água, 6mL de etanol e 94mL de água e somente água respectivamente por quatro meses. Os parâmetros avaliados foram: variações glicêmicas, ponderais, acúmulo de triglicerídeos, colesterol e glicogênio hepáticos. O grupo vinho teve em relação a sua massa corporal uma área sob a curva maior que a dos outros dois grupos, mas com um percentual pequeno de aumento. A concentração do triglicerídeo hepático foi maior no grupo vinho 57% em relação ao grupo etanol, que foi 31,6% menor que o controle (p<0,01%). A concentração do colesterol hepático foi menor no grupo vinho (23,6%), assim como no grupo etanol (24,5%), (p<0,05%). A concentração do glicogênio hepático foi maior no grupo vinho (16%), porém não alcançando significado estatístico. A glicemia em jejum no dia da eutanásia foi maior no grupo etanol em relação aos demais grupos, porém não demonstrou diferença estatisticamente significante. Na análise histológica não foi observada diferença significativa entre os grupos, embora o peso médio em gramas nas gorduras, retroperitoneal e subcutâneas tenha sido aproximadamente duas vezes maior no grupo vinho. Concluiu-se que neste estudo o uso regular e crônico de vinho tinto aumentou triglicerídeo hepático, porém o álcool diminui o colesterol hepático. O aumento do triglicerídeo pode ser devido ao alto valor calórico do vinho ou alguma propriedade lipogênica desconhecida que levou ao aumento importante das gorduras retroperitoneais e subcutâneas em camundongos ApoE Knockout. / The benefits to health related to regular consume of red wine includes different mechanisms in which are involved both ethanol and fenolics compounds of the wine. With the objective to evaluate glycemia, lipid profile and weight variations with regular use of red wine by ApoE Knockout mices, sixty adults ApoE Knockout mices weighing around 30g were distributed into 3 groups of 20 animals each: 1.Wine that received 50mL of wine plus 50mL of water, 2. Ethanol and Water groups, 6mL of ethanol plus 94mL of water and just water respectively for 4 months. We evaluate glycemia, weight variations and liver glycogen, triglycerides and cholesterol. The wine group had in relation to its mass body an area under the curve larger than the other two groups, but with a small percentage of increase. The concentration of liver triglycerides was higher in the wine 57% compared to ethanol group, which was 31.6% lower than the control (p<0.01%). The concentration of liver cholesterol was lower in wine (23.6%) and in ethanol group (24.5%) (p<0.05%). The liver glycogen concentration was higher in the wine (16%), although not reaching statistical significance. The fasting glicemia on the day of euthanasia was higher in the ethanol group compared to other groups, but not statistically significant difference. In histological analysis was not significantly different between groups, although the average weight in grams fat, retroperitoneal and subcutaneous was approximately two times higher in the wine group. It was concluded that in this study the regular and chronic use of red wine increased liver triglyceride, however alcohol decreases liver cholesterol. The increase of the triglyceride may be due to the high caloric value of wine or some lipogenic unknown property that led to an important increase in retroperitoneal and subcutaneous fat tissue in ApoE Knockout mice.

Le déficit familial de LCAT (FLD) est une maladie caractérisée par un défaut de l’activité de l’enzyme lecithin:cholesterol acyltransferase (LCAT). Ce défaut résulte en une concentration plasmatique de C-HDL extrêmement basse, des opacités cornéennes prématurées, la présence d’anémie, de protéinurie et d’insuffisance rénale. Nous avons identifié les premiers patients canadiens-français atteints de déficit familial de LCAT. Deux frères, présentant les signes classiques de FLD étaient homozygotes pour une nouvelle mutation du gène de la LCAT: la mutation c.102delG. Cette mutation se traduit au niveau protéique par un changement du cadre de lecture au niveau du codon His35 et l’insertion d’un codon stop en position 61 entraînant une abolition de l’activité LCAT in vitro et in vivo. La présence de cette mutation cause une réduction importante du C-HDL chez les hétérozygotes (22%) et les homozygotes (88%) ainsi qu’une baisse du C-LDL chez les hétérozygotes (35%) et les homozygotes (58%). De plus, le profil lipidique différait de manière importante entre les deux frères atteints de FLD qui présentaient des génotypes APOE différents. Nous suggérons que APOE est un gène qui modifie le phénotype du FLD et pourrait expliquer l’hétérogénéité des profils lipidiques chez les patients atteints de FLD. Nos résultats suggèrent également que l’association du génotype LCAT-/- a un allèle APOE ε2 est un nouveau mécanisme conduisant à la dysbétalipoproteinemie. Finalement nous avons montré des différences importantes dans les sous-populations des HDL chez les deux sujets atteints de FLD. Le porteur de l’allèle APOE ε2 présentait une proportion beaucoup plus importante de HDL immatures (preβ discoïdaux) par rapport a son frère (77.9% vs. 31.0%). / Familial LCAT deficiency (FLD) is a disease characterized by a defect in the enzyme lecithin:cholesterol acyltransferase (LCAT) resulting in low HDL-C, premature corneal opacities, anaemia as well as proteinuria and renal failure. We have identified the first French Canadian kindred with familial LCAT deficiency. Two brothers, presenting classical signs of FLD, were shown to be homozygous for a novel LCAT mutation. This c.102delG mutation occurs at the codon for His35 and causes a frameshift that stops transcription at codon 61 abolishing LCAT enzymatic activity both in vivo and in vitro. It has a dramatic effect on the lipoprotein profile, with an important reduction of HDL-C in both heterozygotes (22%) and homozygotes (88%) and a significant decrease in LDL-C in heterozygotes (35%) as well as homozygotes (58%). Furthermore, the lipoprotein profile differs markedly between the two affected brothers who had different APOE genotypes. We propose that APOE could be an important modifier gene explaining heterogeneity in lipoprotein profiles observed among FLD patients. Our results suggest that a LCAT-/- genotype associated with an APOE ε2 allele could be a novel mechanism leading to dysbetalipoproteinemia. Finally we have identified major differences in the HDL sub-populations of both subjects affected by FLD. The carrier of the APOE ε2 allele presented a much higher proportion of immature HDL particles (discoid preβ) compared to his brother (77.9% vs. 31.0%).

Le déficit familial de LCAT (FLD) est une maladie caractérisée par un défaut de l’activité de l’enzyme lecithin:cholesterol acyltransferase (LCAT). Ce défaut résulte en une concentration plasmatique de C-HDL extrêmement basse, des opacités cornéennes prématurées, la présence d’anémie, de protéinurie et d’insuffisance rénale. Nous avons identifié les premiers patients canadiens-français atteints de déficit familial de LCAT. Deux frères, présentant les signes classiques de FLD étaient homozygotes pour une nouvelle mutation du gène de la LCAT: la mutation c.102delG. Cette mutation se traduit au niveau protéique par un changement du cadre de lecture au niveau du codon His35 et l’insertion d’un codon stop en position 61 entraînant une abolition de l’activité LCAT in vitro et in vivo. La présence de cette mutation cause une réduction importante du C-HDL chez les hétérozygotes (22%) et les homozygotes (88%) ainsi qu’une baisse du C-LDL chez les hétérozygotes (35%) et les homozygotes (58%). De plus, le profil lipidique différait de manière importante entre les deux frères atteints de FLD qui présentaient des génotypes APOE différents. Nous suggérons que APOE est un gène qui modifie le phénotype du FLD et pourrait expliquer l’hétérogénéité des profils lipidiques chez les patients atteints de FLD. Nos résultats suggèrent également que l’association du génotype LCAT-/- a un allèle APOE ε2 est un nouveau mécanisme conduisant à la dysbétalipoproteinemie. Finalement nous avons montré des différences importantes dans les sous-populations des HDL chez les deux sujets atteints de FLD. Le porteur de l’allèle APOE ε2 présentait une proportion beaucoup plus importante de HDL immatures (preβ discoïdaux) par rapport a son frère (77.9% vs. 31.0%). / Familial LCAT deficiency (FLD) is a disease characterized by a defect in the enzyme lecithin:cholesterol acyltransferase (LCAT) resulting in low HDL-C, premature corneal opacities, anaemia as well as proteinuria and renal failure. We have identified the first French Canadian kindred with familial LCAT deficiency. Two brothers, presenting classical signs of FLD, were shown to be homozygous for a novel LCAT mutation. This c.102delG mutation occurs at the codon for His35 and causes a frameshift that stops transcription at codon 61 abolishing LCAT enzymatic activity both in vivo and in vitro. It has a dramatic effect on the lipoprotein profile, with an important reduction of HDL-C in both heterozygotes (22%) and homozygotes (88%) and a significant decrease in LDL-C in heterozygotes (35%) as well as homozygotes (58%). Furthermore, the lipoprotein profile differs markedly between the two affected brothers who had different APOE genotypes. We propose that APOE could be an important modifier gene explaining heterogeneity in lipoprotein profiles observed among FLD patients. Our results suggest that a LCAT-/- genotype associated with an APOE ε2 allele could be a novel mechanism leading to dysbetalipoproteinemia. Finally we have identified major differences in the HDL sub-populations of both subjects affected by FLD. The carrier of the APOE ε2 allele presented a much higher proportion of immature HDL particles (discoid preβ) compared to his brother (77.9% vs. 31.0%).

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O desenvolvimento de mudanças neuropatológicas características da doença de Alzheimer (DA) nos portadores de síndrome de Down (SD) a partir dos 40 anos de idade e a alta taxa de indivíduos com SD em famílias nas quais a DA está presente sugerem uma susceptibilidade comum entre as duas doenças. Além disso, mulheres jovens (<35 anos na concepção) que tiveram filhos com a SD têm um risco cinco vezes maior de desenvolverem DA do que a população em geral. Esta alta suscetibilidade compartilhada poderia envolver um processo de envelhecimento acelerado, que levaria à SD em filhos de mães jovens e a um risco aumentado de demência nestas mães. Até o momento, os genes APOE, PSEN1 e TNF-&#945;, envolvidos direta ou indiretamente na deposição cerebral de beta amilóide e reconhecidos dentre os determinantes genéticos para a DA, parecem também intervir na patogênese da demência na SD. Neste estudo, analisamos a freqüência dos alelos &#949;2, &#949;3 e &#949;4 do gene APOE e dos polimorfismos -48C>T no gene PSEN1 e -850C>T no gene TNF-&#945; em mães de portadores da SD em comparação com mães de indivíduos cromossomicamente normais. Para isto, amostras de DNA foram extraídas a partir do sangue periférico de 114 mães de portadores da trissomia livre do cromossomo 21 com idade inferior a 35 anos no momento da concepção e de 110 mães controle. Para a triagem dos polimorfismos dos alelos do gene APOE e dos polimorfismos PSEN1 -48C>T e TNF-&#945; -850C>T foi realizada a técnica de PCR seguida da digestão dos produtos amplificados com as endonucleases de restrição HhaI, HgaI e HincII, respectivamente. Os produtos digeridos foram analisados através de eletroforese em géis de poliacrilamida corados por prata. A análise estatística através do teste exato de Fisher revelou que não houve diferenças significativas entre as distribuições genotípicas observadas nas amostras caso e controle tanto para os alelos do gene APOE, quanto para os polimorfismos PSEN1-48C>T e TNF-&#945; -850C>T (c2 =4,26; g.l. = 4; P =0,37; c2 =0,22; g.l. = 2; P =0,89; c2 = 4,24; g.l. = 2; P = 0,12, respectivamente). Diferenças significativas também não foram observadas tanto na análise da interação gênica multiplicativa entre os alelos dos genes APOE, PSEN1 e TNF-&#945; como na interação entre estes alelos e polimorfismos participantes do metabolismo do folato (MTHFR 677C>T, MTHFR 1298A>C, MTRR 66A>G, MTR 2756A>G, RFC-1 80G>A, TC 776C>G e CBS 844ins68). No Brasil, este é o primeiro relato da investigação destes genes entre as mães de indivíduos com SD. Nossos resultados sugerem que os alelos APOE e4, PSEN1-48 C e TNF-&#945; -850T não são fatores de risco independentes para a DA nestas mães. Entretanto, em virtude da complexidade tanto da SD como da DA, a análise de variantes em outros genes é de fundamental importância para melhor elucidar o aparecimento insidioso de DA em mães jovens que tiveram filhos com SD. / The development of neuropathological changes of Alzheimer's disease (AD) in patients with Down syndrome (DS) from 40 years old and the high rate of DS births in families of DA individuals suggest a common susceptibility between the two diseases. Besides a five-fold higher risk for AD has been reported in young mothers of DS individuals. This shared genetic susceptibility involving DS and AD could be due to an accelerated ageing process, leading to the birth of a DS child in the offspring of young mothers and to an increased risk of dementia among them. Between the genetic risk factors for AD, APOE, PSEN1 and TNF-&#945; genes are directly or indirectly involved in cerebral amyloid deposition and seem to play a role in the pathogenesis of dementia in DS. In this study, we analyzed the frequency of APOE alleles &#949;2, &#949;3 e &#949;4 and the polymorphisms -48C>T in the PSEN1 gene and -850C>T in the TNF-&#945; gene in DS young mothers in comparison to mothers who had had chromosomicaly normal children. With this purpose, DNA samples were extracted from 114 mothers of DS individuals (full trisomy 21) with less than 35 years-old at conception, and from 110 control mothers. For the screening of the APOE alleles, PSEN1 -48C>T and TNF-&#945; -850C>T polymorphisms, polymerase chain reaction (PCR) was accomplished, followed by the digestion of the amplification products with the restriction endonucleases HhaI, HgaI and HincII, respectively. The digested products were analysed by polyacrilamide gels electrophoresis followed by silver staining. No significant differences were observed between the genotype distributions observed in control and case samples with respect APOE alleles, PSEN1 -48C>T and TNF-&#945; -850C>T polymorphisms (Fisher exact test: c2 =4,26; g.l. = 4; P =0,37; c2 =0,22; g.l. = 2; P =0,89; c2 = 4,24; g.l. = 2; P = 0,12, respectively). No differences were also observed in the gene multiplicative interaction analysis for APOE, PSEN1 and TNF-&#945; alleles and in the interaction of these alleles with folate pathway polymorphisms (MTHFR 677C>T, MTHFR 1298A>C, MTRR 66A>G, MTR 2756A>G, RFC-1 80G>A, TC 776C>G e CBS 844ins68). This is the first study reporting the investigation of these genes in Brazilian DS mothers. Our findings suggest that APOE &#949;4, PSEN1 -48C and TNF-&#945; -850T alleles are not independent risk factors for AD in young DS mothers. However, considering the complexity of both DS and AD, polymorphisms in other genes should be investigated to evaluate the actual relationship between the genetic risk factors and AD in DS young mothers.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O desenvolvimento de mudanças neuropatológicas características da doença de Alzheimer (DA) nos portadores de síndrome de Down (SD) a partir dos 40 anos de idade e a alta taxa de indivíduos com SD em famílias nas quais a DA está presente sugerem uma susceptibilidade comum entre as duas doenças. Além disso, mulheres jovens (<35 anos na concepção) que tiveram filhos com a SD têm um risco cinco vezes maior de desenvolverem DA do que a população em geral. Esta alta suscetibilidade compartilhada poderia envolver um processo de envelhecimento acelerado, que levaria à SD em filhos de mães jovens e a um risco aumentado de demência nestas mães. Até o momento, os genes APOE, PSEN1 e TNF-&#945;, envolvidos direta ou indiretamente na deposição cerebral de beta amilóide e reconhecidos dentre os determinantes genéticos para a DA, parecem também intervir na patogênese da demência na SD. Neste estudo, analisamos a freqüência dos alelos &#949;2, &#949;3 e &#949;4 do gene APOE e dos polimorfismos -48C>T no gene PSEN1 e -850C>T no gene TNF-&#945; em mães de portadores da SD em comparação com mães de indivíduos cromossomicamente normais. Para isto, amostras de DNA foram extraídas a partir do sangue periférico de 114 mães de portadores da trissomia livre do cromossomo 21 com idade inferior a 35 anos no momento da concepção e de 110 mães controle. Para a triagem dos polimorfismos dos alelos do gene APOE e dos polimorfismos PSEN1 -48C>T e TNF-&#945; -850C>T foi realizada a técnica de PCR seguida da digestão dos produtos amplificados com as endonucleases de restrição HhaI, HgaI e HincII, respectivamente. Os produtos digeridos foram analisados através de eletroforese em géis de poliacrilamida corados por prata. A análise estatística através do teste exato de Fisher revelou que não houve diferenças significativas entre as distribuições genotípicas observadas nas amostras caso e controle tanto para os alelos do gene APOE, quanto para os polimorfismos PSEN1-48C>T e TNF-&#945; -850C>T (c2 =4,26; g.l. = 4; P =0,37; c2 =0,22; g.l. = 2; P =0,89; c2 = 4,24; g.l. = 2; P = 0,12, respectivamente). Diferenças significativas também não foram observadas tanto na análise da interação gênica multiplicativa entre os alelos dos genes APOE, PSEN1 e TNF-&#945; como na interação entre estes alelos e polimorfismos participantes do metabolismo do folato (MTHFR 677C>T, MTHFR 1298A>C, MTRR 66A>G, MTR 2756A>G, RFC-1 80G>A, TC 776C>G e CBS 844ins68). No Brasil, este é o primeiro relato da investigação destes genes entre as mães de indivíduos com SD. Nossos resultados sugerem que os alelos APOE e4, PSEN1-48 C e TNF-&#945; -850T não são fatores de risco independentes para a DA nestas mães. Entretanto, em virtude da complexidade tanto da SD como da DA, a análise de variantes em outros genes é de fundamental importância para melhor elucidar o aparecimento insidioso de DA em mães jovens que tiveram filhos com SD. / The development of neuropathological changes of Alzheimer's disease (AD) in patients with Down syndrome (DS) from 40 years old and the high rate of DS births in families of DA individuals suggest a common susceptibility between the two diseases. Besides a five-fold higher risk for AD has been reported in young mothers of DS individuals. This shared genetic susceptibility involving DS and AD could be due to an accelerated ageing process, leading to the birth of a DS child in the offspring of young mothers and to an increased risk of dementia among them. Between the genetic risk factors for AD, APOE, PSEN1 and TNF-&#945; genes are directly or indirectly involved in cerebral amyloid deposition and seem to play a role in the pathogenesis of dementia in DS. In this study, we analyzed the frequency of APOE alleles &#949;2, &#949;3 e &#949;4 and the polymorphisms -48C>T in the PSEN1 gene and -850C>T in the TNF-&#945; gene in DS young mothers in comparison to mothers who had had chromosomicaly normal children. With this purpose, DNA samples were extracted from 114 mothers of DS individuals (full trisomy 21) with less than 35 years-old at conception, and from 110 control mothers. For the screening of the APOE alleles, PSEN1 -48C>T and TNF-&#945; -850C>T polymorphisms, polymerase chain reaction (PCR) was accomplished, followed by the digestion of the amplification products with the restriction endonucleases HhaI, HgaI and HincII, respectively. The digested products were analysed by polyacrilamide gels electrophoresis followed by silver staining. No significant differences were observed between the genotype distributions observed in control and case samples with respect APOE alleles, PSEN1 -48C>T and TNF-&#945; -850C>T polymorphisms (Fisher exact test: c2 =4,26; g.l. = 4; P =0,37; c2 =0,22; g.l. = 2; P =0,89; c2 = 4,24; g.l. = 2; P = 0,12, respectively). No differences were also observed in the gene multiplicative interaction analysis for APOE, PSEN1 and TNF-&#945; alleles and in the interaction of these alleles with folate pathway polymorphisms (MTHFR 677C>T, MTHFR 1298A>C, MTRR 66A>G, MTR 2756A>G, RFC-1 80G>A, TC 776C>G e CBS 844ins68). This is the first study reporting the investigation of these genes in Brazilian DS mothers. Our findings suggest that APOE &#949;4, PSEN1 -48C and TNF-&#945; -850T alleles are not independent risk factors for AD in young DS mothers. However, considering the complexity of both DS and AD, polymorphisms in other genes should be investigated to evaluate the actual relationship between the genetic risk factors and AD in DS young mothers.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2017-06-14T19:33:16Z No. of bitstreams: 1 alicebelleigolirezende.pdf: 4554252 bytes, checksum: 45624650ab925d7640e01b2b01e2a245 (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2017-06-29T12:16:33Z (GMT) No. of bitstreams: 1 alicebelleigolirezende.pdf: 4554252 bytes, checksum: 45624650ab925d7640e01b2b01e2a245 (MD5) / Made available in DSpace on 2017-06-29T12:16:33Z (GMT). No. of bitstreams: 1 alicebelleigolirezende.pdf: 4554252 bytes, checksum: 45624650ab925d7640e01b2b01e2a245 (MD5) Previous issue date: 2011-08-30 / FAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas Gerais / A aterosclerose é uma doença imune-inflamatória associada ao acúmulo de lípides na camada íntima das artérias. O baço desempenha uma importante função imunológica, mas a sua participação no desenvolvimento da aterosclerose permanece controversa. Considerando que a incidência de esplenectomias totais ainda é alta e que a doença aterosclerótica é prevalente, é de grande importância que se verifique se o tecido esplênico influencia no processo de aterosclerose. Neste trabalho, o impacto da esplenectomia no desenvolvimento da aterosclerose foi avaliado em camundongos deficientes em apoproteína E (apoE). Os animais foram divididos em dois grupos: grupo controle (CT), composto de camundongos deficientes em apoE submetidos a um procedimento cirúrgico simulado; e grupo esplenectomia total (ET), camundongos deficientes em apoE submetidos à esplenectomia total. Trinta dias após a cirurgia, os animais foram submetidos a uma dieta aterogênica enriquecida com 0,15% de colesterol. Após oito semanas, os animais foram eutanasiados e o sangue, o coração e a aorta foram submetidos à análise. Os níveis séricos de colesterol total e de anticorpos anti-LDL oxidada foram avaliados. As áreas de lesão aterosclerótica na raiz da aorta foram quantificadas por morfometria em cortes histológicos corados com hematoxilina-eosina. As lesões ateroscleróticas nas porções torácica e abdominal das aortas foram determinadas pela porcentagem da superfície luminal corada com Sudan IV em relação à área total do vaso. Os resultados mostraram que os níveis séricos de colesterol total não foram alterados pela esplenectomia e que os títulos de anticorpos IgG anti-LDL oxidada foram semelhantes entre os grupos. No entanto, foi observada uma maior porcentagem de lesões ateroscleróticas nas porções torácica e abdominal das aortas de animais esplenectomizados. Além disso, camundongos esplenectomizados apresentaram lesões ateroscleróticas na raiz da aorta significativamente maiores do que os animais controles. Estes dados indicam, em conjunto, que a esplenectomia está associada ao aumento das lesões ateroscleróticas em camundongos deficientes em apoE submetidos à dieta aterogênica, sugerindo um papel ateroprotetor do baço no modelo estudado. / Atherosclerosis is an inflammatory immune disease associated with lipid accumulation in the intima layer of arteries. The spleen plays an important immune function, but its influence in development of atherosclerosis remains unclear. Evaluation of the role of the spleen in atherosclerosis is justified due to the high frequency of total splenectomies and the high prevalence of atherosclerotic disease. In this work, the effect of splenectomy on the development of atherosclerosis in apolipoprotein E (apoE) deficient mice was investigated. ApoE deficient mice were divided into a sham-operated control group (CT) and a splenectomized group (SP). Thirty days after surgery, animals were fed a high fat western diet (with 0.15% cholesterol). After eight weeks, mice were euthanized and their blood, heart, and aorta were subjected to analysis. Total serum cholesterol and anti-oxidized LDL antibodies were measured. Atherosclerotic lesion areas in the aortic root were stained with hematoxylin-eosin and quantified by morfometry. The atherosclerotic lesions in the thoracic and abdominal portions of aorta were determined by assessing the percentage of the luminal surface area stained by Sudan IV. Levels of total serum cholesterol did not vary significantly after splenectomy. Anti-oxidized LDL IgG antibodies were similar between groups. However, compared to the control group, atherosclerotic area in the thoracic and abdominal portions of aorta were significantly increased in splenectomized mice. These data were confirmed by the larger lesions in the aortic root in splenectomized mice. These data indicate that splenectomy increases atherosclerotic lesions in apoE deficient mice fed an atherogenic diet, suggesting a atheroprotector role of the spleen.

Les maladies cardiovasculaires (MCV) constituent la première cause de mortalité dans le monde. Des efforts significatifs ont été menés pour améliorer la prévention des MCV et ont abouti à une réduction du taux de mortalité. Cependant, un ralentissement considérable du taux de réduction des MCV a été observé à partir de 2011 malgré les nouvelles avancées thérapeutiques. Ces taux alarmants justifient le besoin de découvrir de nouvelles thérapies afin d’améliorer la prévention des MCV dans la population générale. Des taux élevés de cholestérol total et transporté par les lipoprotéines de basse densité (LDL) sont fortement liés aux MCV athérosclérotiques. Ainsi, les thérapies hypolipémiantes sont les thérapies les plus utilisées pour prévenir les MCV. Nos résultats antérieurs ont identifié Golgi glycoprotein 1 (GLG1) comme étant une nouvelle protéine impliquée dans le métabolisme lipidique. Par conséquent, notre étude actuelle vise à démontrer l'effet de GLG1 sur le développement de l'athérosclérose. Pour étudier cet effet, nous avons réduit l'expression de GLG1 (silençage génique ou knockdown) dans le foie de souris hypercholestérolémiques Apoe-/- en utilisant un virus adéno-associé de type 8 (AAV8) véhiculant un petit ARN en épingle à cheveux (shRNA) ciblant GLG1. Deux semaines post-injection, les souris ont été nourries par un régime occidental pendant 12 semaines. L'étude a révélé que le knockdown de GLG1 réduit significativement les taux plasmatiques de cholestérol total, de lipoprotéines de très basse densité (VLDL), de LDL et diminue les lésions athérosclérotiques au niveau du sinus aortique. Cependant, nos résultats ont démontré que le knockdown de GLG1 à l’aide du système AAV8 n'était pas stable pendant toute l'étude, le rendant inefficace au-delà de 2 mois. En résumé, nos résultats montrent que le knockdown de GLG1 réduit les taux de cholestérol plasmatique ainsi que la progression de l'athérosclérose chez les souris Apoe-/-. Cette réduction s'est produite en dépit de la perte progressive du knockdown de GLG1 et est probablement liée à la réduction de la sécrétion d'apolipoprotéines B100. Ces résultats montrent que GLG1 est une cible thérapeutique prometteuse pour abaisser les taux de cholestérol plasmatique, en particulier les VLDL et LDL, et prévenir le développement de l'athérosclérose. / Cardiovascular diseases (CVD) are the leading cause of death worldwide. Significant efforts have been made to prevent CVD and have resulted in a reduced mortality rate. However, a considerable slowdown in the reduction rate of CVD has been observed starting in 2011 despite new therapeutic advances. These alarming rates justify the need to discover new therapies to improve the prevention of CVD in the general population. High levels of total cholesterol and low-density lipoprotein cholesterol (LDL-C) are strongly linked to atherosclerotic cardiovascular diseases. Thus, lipid-lowering therapies are the most used therapies to prevent CVDs. Our previous study identified Golgi glycoprotein 1 (GLG1) as a new protein involved in lipid metabolism. Therefore, our current study aims to demonstrate the effect of GLG1 on the development of atherosclerosis. To study the effect of GLG1 on atherosclerosis, we reduced GLG1 expression in hypercholesterolemic Apoe-/- mice livers using an adeno-associated virus coding for a short hairpin RNA (shRNA) targeting GLG1 then 2 weeks post-injection mice were fed a Western diet for 12 weeks. The study revealed that GLG1 knockdown significantly reduced plasma total cholesterol levels, especially very low-density lipoprotein (VLDL)- and LDL-C, and atherosclerotic lesions in the aortic root. However, our results showed that AAV-mediated GLG1 knockdown was not stable during the entire study, making it ineffective in lowering plasma cholesterol levels beyond 2 months. In summary, our results show that lowering GLG1 expression reduces plasma cholesterol levels as well as atherosclerosis progression in Apoe-/- mice. Remarkably, the reduction in atherosclerosis occurred in spite of the gradual loss of GLG1 AAV-mediated knockdown and is likely related to the reduced apolipoptotein B100 secretion. These findings demonstrate that GLG1 is a promising therapeutic target for lowering plasma cholesterol levels, particularly VLDL- and LDL-C, and preventing atherosclerosis development.