Dissertations / Theses on the topic 'ACTN1'

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
OBJETIVO: Estudar as associações entre os polimorfismos genéticos ECA I/D, ACTN3 R577X e PON1 C(-107)T com diabetes e hipertensão em pacientes atendidos pelo Sistema Único de Saúde. MÉTODOS: Foram coletados dados bioquímicos dos prontuários dos pacientes crônicos de três Unidades Básicas de Saúde da Família localizadas na cidade de Rio Grande. Foram aplicados questionários de consumo alimentar, nível de atividade física e socioeconômico. Foi realizada a avaliação nutricional dos pacientes e, em seguida, a coleta de saliva para a extração do DNA genômico para análise dos polimorfismos genéticos. Os polimorfismos foram analisados por PCR simples, multiplex e por polimorfismo de tamanho de fragmento de restrição. RESULTADOS: O grupo caso obteve maiores valores de peso, IMC, circunferência da cintura, percentual de gordura, pressão arterial, colesterol total, triglicerídeos e glicemia em comparação ao grupo controle. Foram encontradas associações do genótipo XX do polimorfismo R577X da ACTN3 com valores mais elevados de glicemia e triglicerídeos. O genótipo TT do polimorfismo C(-107)T da PON1 também obteve níveis maiores de triglicerídeos comparado aos outros 2 genótipos. CONCLUSÃO: O genótipo XX da ACTN3 foi fortemente relacionado com níveis baixos de HDL e altos valores de triglicerídeos, colesterol total e glicemia. Ainda encontramos altos valores de triglicerídeos e LDL no genótipo TT e menores níveis de colesterol total ligados ao genótipo CC da PON1.
OBJECTIVE: To study associations between polymorphisms in the angiotensin converting enzyme (ACE I/D), actinin 3 (ACTN3 R577X) and paraoxonase 1 (PON1 C-(107)T with chronic diseases (diabetes and hypertension) in patients attended by Sistema Único de Saúde (SUS). METHODS: Biochemical data were collected from the charts in the Basic Health Units of the Family located in the same region of the city of Rio Grande. Charts about food consumption, physical activity level and socioeconomic were applied. Nutritional data were evaluated, genomic DNA was extracted from collected saliva samples and used for analysis of genetic polymorphism. RESULTS: The case group had higher values of weight, BMI, waist circumference, body fat percentage, blood pressure, cholesterol, triglycerides and blood glucose in comparison with control group. Associations were found of XX genotype of the polymorphism R577X ACTN3 with higher values of blood glucose, and triglycerides levels. The TT genotype of the C(-107)T PON1 also had higher levels of triglycerides compared to other 2 genotypes. CONCLUSION: The genotype XX of ACTN3 was strong related with low HDL levels and high triglycerides, total cholesterol and glucose levels. Still, we realized high triglycerides and LDL leves in TT genotype of PON1 and lower leves of total cholesterol linked to CC genotype of PON1.

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The search for better sports performance has encouraged studies on the interaction between environmental and genetic factors. In this perspective, while environmental factors stimulate morphofunctional adaptations, genetic polymorphisms modulate the genes responsible for these adaptations. Therefore, the identification of the candidate genes and their respective polymorphisms with potential to influence the phenotypes related to this performance have been the target of the researchers of the area. Among the promising polymorphisms are the I/D of the angiotensin converting enzyme (ACE) gene and the α-actinin-3 (ACTN3) gene R577X. In the field of sports genetics, swimming has been studied, but research on the younger athletes is rare. The objective of this study was to analyze the association between ACE and ACTN3 polymorphisms in sports performance indicators in 120 brazilian swimmers (75 Boys and 45 Girls), aged 15 to 17 years (16.76 ± 0.6 years ), affiliated to the Brazilian Confederation of Aquatic Sports. 102 non-athletes of the same age group (16.51 ± 0.95 years) residing in the Federal District were part of the control group (56 Boys and 46 Girls). These were subdivided by official swimming tests (short: ≤200m vs long: ≥400m), related to the phenotypes of sports performance (strength vs. power) and by the competitive level (elite vs. sub-elite). The elite status (international experiences) and the technical index (TI) were adopted as indicators of performance. It was also evaluated the total genothype score (TGS) associated to the strength / power phenotypes. The technique of scraping buccal mucosa epithelial cells with the aid of a specific swab was used to collect the samples. In the athletes was collected during the XXIV Brazilian Junior Swimming Championship and among the students, in the intervals of the Physical Education classes. The genotyping of the polymorphisms was performed through the polymerase chain reaction technique obeying standardized and scientifically validated protocols. All the volunteers signed the agreement with prior consent of those responsible. Chi-square and Mann-Whitney tests were used when the variables were not normally distributed. Pearson's correlation and t-test for independent samples were used for the parametric data. The groups (athletes and non-athletes) demonstrated Hardy-Weinberg equilibrium for the genotypic and allelic distribution of polymorphisms. Sub-elite athletes (≤200m and ≥ 400m) presented allelic and genotype frequencies in both polymorphisms very close to those observed for the control group. The elite group of athletes was formed by specialists in short competitions (≤200m). Significant primacy of the DD genotype of ACE was observed for elite athletes. The D allele and DD genotype were also predominant among athletes of the same phenotypic (strength/power) group identified in the upper quartile (Q3) of TI, with significant differences especially in favor of elite athletes. Analysis of the ACTN3 polymorphism revealed that the R allele was predominant in all groups, except for the elite group, which had a frequency of the heterozygote RX genotype significantly higher. The best TI’s were verified among the athletes (≤200m) genotyped for RX and RR, with supremacy for the homozygote among elite athletes. In the joint evaluation of the two polymorphisms, the elite group presented significant genotypic supremacy of DD + RX addition compared to the other groups that presented higher occurrence of DD + RR homozygotes. The TGS analysis showed that athletes with better genotype profiles (score ≥75) also had the best TI’s. The results of the study suggest that the elite status and the best TI’s verified among juvenile athletes were influenced positively by the genotype associations typically expected.
A busca pelo melhor desempenho esportivo tem incentivado estudos sobre a interação entre os fatores ambientais e genéticos. Nesta perspectiva, enquanto os fatores ambientais estimulam as adaptações morfofuncionais os polimorfismos genéticos modulam os genes responsáveis por estas adaptações. Por isso, a identificação dos genes candidatos e seus respectivos polimorfismos com potencial de influenciar os fenótipos relacionados a este desempenho têm sido alvo dos pesquisadores da área. Dentre os polimorfismos promissores destacam-se o I/D do gene da enzima conversora da angiotensina (ECA) e o R577X do gene da α-actinina-3 (ACTN3). Na área da genética do esporte a natação tem sido estudada, mas são raras as pesquisas dedicadas aos atletas mais jovens, notadamente na população brasileira. O objetivo do estudo foi analisar a associação entre os polimorfismos da ECA e da ACTN3 aos indicadores de desempenho esportivo em 120 atletas da natação brasileira (75 Rapazes e 45 Moças), na faixa etária dos 15 aos 17 anos (16,76 ± 0,6 anos), filiados à Confederação Brasileira de Desportos Aquáticos (6,46 ± 2,13 anos). 102 jovens escolares (56 Rapazes e 46 Moças) não-atletas da mesma faixa etária (16,51 ± 0,95 anos) residentes no Distrito Federal fizeram parte do grupo controle. Estes foram subdivididos pelas provas oficiais da natação (curtas: ≤200m vs longas: ≥400m), relacionadas tipicamente aos fenótipos opostos do desempenho atlético (respectivamente: força/potência vs resistência) e pelo nível competitivo (elite vs sub-elite). O status de elite (experiências internacionais) e o índice técnico (IT) foram adotados como indicadores de desempenho. Foi avaliado também o score total dos genótipos (TGS) associados aos fenótipos da força/potência. O material genético dos atletas foi coletado durante o XXIV Campeonato Brasileiro Juvenil de Natação e entre os escolares, nos intervalos das aulas de Educação Física. A técnica de raspagem das células epiteliais da mucosa bucal com o auxílio de swab específico foi utilizada para a coleta das amostras. A genotipagem dos polimorfismos foi realizada através da técnica de reação em cadeia da polimerase obedecendo protocolos padronizados e cientificamente validados. Todos os voluntários assinaram o termo de assentimento com prévia anuência dos responsáveis. Os testes do Qui-Quadrado e Mann-Whitney foram utilizados quando as variáveis não apresentavam distribuição normal. A correlação de Pearson e o teste t para amostras independentes foram utilizados para os dados paramétricos. Os grupos (atletas e não-atletas) demonstraram equilíbrio Hardy-Weinberg para a distribuição genotípica e alélica dos polimorfismos. Os atletas sub-elite (≤200m e ≥400m) apresentaram frequências alélicas e genotípicas em ambos os polimorfismos muito próximas às verificadas para o grupo controle. O grupo de atletas de elite foi formado por especialistas em provas curtas (≤200m). Observou-se supremacia significativa do genótipo DD da ECA para os atletas de elite. O alelo D e o genótipo DD foram predominantes também entre os atletas do mesmo grupo fenotípico (força/potência) identificados no quartil superior (Q3) do IT, com diferenças significativas especialmente em prol dos atletas de elite. A análise do polimorfismo da ACTN3 revelou que o alelo R foi predominante em todos os grupos, exceto para o grupo de elite, os quais apresentaram frequência do genótipo heterozigoto RX significativamente superior. Os melhores IT foram verificados entre os atletas (≤200m) genotipados para RX e RR, com supremacia para o homozigoto entre os atletas de elite. Na avaliação conjunta dos dois polimorfismos o grupo de elite apresentou significativa supremacia genotípica da adição DD+RX frente aos demais grupos que apresentaram maior ocorrência dos homozigotos DD+RR. A análise do TGS demonstrou que os atletas com melhores perfis genofenotípicos (score ≥75) apresentaram também os melhores IT. Os resultados do estudo sugerem que o status de elite e os melhores IT verificados entre atletas juvenis sofreram influência positiva das associações genofenotípicas tipicamente esperada.

The objective was to determine the frequency of ACTN3 in soccer players of different categories and their relationship to performance, hormonal responses and indicators of muscle damage after the game. We evaluated 367 male players under- 15 category, sub-17, U-20 and professional and amateur players (N. 14) and a control group (N. 100). All subjects were genotyped for the polymorphism ACTN3- RR, RX and XX. Were applied tests of aerobic capacity, squat jump (SJ), ountermovement jump (CMJ) and sprint test of 30 m (V30) subdivided into 10 m (V10) and 20 m (V20). Blood collections were performed in the pre, post, 2 and 4 h after the match and were determined the concentrations of interleukin-6 (IL-6), cortisol, testosterone, ratio testosterone/cortisol (T/C), creatinekinase (CK), muscle alphaactin and between the groups RR/RX and XX. The frequency of ACTN3 in players was not different from control group. However it was noted that the group of professional players had a lower frequency of XX and an increased frequency RX (p<0.05) compared to younger players. Concentrations of CK were higher after the game for both groupsand remained high until the time 4 h (p <0.05) but not different. We identified increased IL-6 post game for the XX group (p <0.05) returning to normal at the time 2 h. There was an increase of testosterone after the game in the group RR/RX that lasted until 4 am (p<0.05). Being that this same pattern was not observed for group XX. In the moments Post, 2 and 4 h testosterone concentrations were higher in RR/RX compared to the group XX. The group RR/RX showed increased levels of cortisol Post the game. In both groups, cortisol concentrations were lower at times 2 and 4 h compared with the pre (p<0.05), indicating a decrease of concentration. We conclude that the frequency of genotype XX and RX decreasesand increases respectively in older categories in football, indicating a natural selection of individuals of character stronger and mixed in this sport. The RR genotype and allele R is present in more soccer playerswith superior physical performance in tests of strength and speed, while the reverse was not proved in relation to resistance. XX individuals have higher responses of IL-6 to a football game and individuals RR/RX have higher values of markers of CK Postgame.
O objetivo foi determinar a frequência do ACTN3 em jogadores de futebol de diferentes categorias e a sua relação com o desempenho, respostas hormonais e de indicadores de dano muscular pós jogo. Foram avaliados 367 jogadores masculinos das categorias sub-15, sub-17, sub-20 e profissionais, e jogadores amadores (N. 14) e um grupo controle (N. 100). Todos os indivíduos foram genotipados para o polimorfismo ACTN3- RR, RX e XX. Foram realizados testes de capacidade aeróbia, salto agachado (SJ), com contra-movimento (CMJ) e teste de velocidade de 30m (V30) subdivido em 10m (V10) e 20 (V20). Foram realizadas coletas sanguíneas nos momentos pré, pós, 2 e 4h de jogo e foram determinadas as concentrações de Interleucina-6 (IL-6), Cortisol, testosterona, relação testosterona/cortisol (T/C), creatina-quinase (CK) e alfa-actina muscular entre os grupos RR/RX e XX. A frequência do ACTN3 em jogadores não foi diferente do grupo controle. No entanto observou-se que o grupo de jogadores profissionais apresentou uma menor frequência do XX e aumento do RX (p<0,05) em relação aos jogadores mais jovens. Não foi encontrada relação da frequência genotípica com o desempenho em testes físicos. As concentrações de CK pós jogo foram maiores para ambos os grupo e se mantiveram altas até o momento 4 h (p<0,05), mas não diferiram entre si. Identificou-se aumento da IL-6 pós jogo para o grupo XX (p<0,05) retornando aos valores normais no momento 2 h. Houve aumento da testosterona pós jogo no grupo RR/RX que se manteve até 4h (p<0,05). Sendo que esse mesmo comportamento não foi observado para o grupo XX. Nos momentos pós, 2 e 4 h as concentrações de testosterona foram maiores no grupo RR/RX em comparação ao grupo XX. O grupo RR/RX apresentou aumento das concentrações de cortisol pós jogo. Em ambos os grupos a concentração de cortisol foi menor nos momentos 2 e 4 h em comparação com a fase pré (p<0,05), indicando uma diminuição desta concentração. Conclui-se que a frequência do genótipo XX e o RX diminui e aumenta respectivamente em categorias mais velhas no futebol, indicando uma seleção natural de indivíduos de caráter mais forte e misto neste esporte. Indivíduos XX, apresentam maiores respostas de IL-6 a um jogo de futebol e indivíduos RR/RX apresentam maiores valores de marcadores de CK pós jogo.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Introdução: Alguns estudos longitudinais têm mostrado que a força pode influenciar a economia de corrida (EC). Entretanto, outros modelos [p.ex., dano muscular (DM) induzido por corridas em declive (CrED)] mostram que a magnitude e a cinética de recuperação da EC e da força máxima medida em exercícios de cadeia cinética aberta [pico de torque isométrico de extensão do joelho (PTI)] podem ser diferentes. Fatores como o tipo de exercício usado para medir a força e a forma pela qual a força se manifesta (máxima e explosiva), podem explicar, pelo menos em parte, estes dados antagônicos. O objetivo geral deste estudo foi utilizar o modelo de DM induzido pela CrED para analisar a relação entre força máxima e explosiva e a EC. Método: Para isso, foram empregadas três intervenções para modular o DM induzido pela CrED e verificar seus efeitos sobre a força e a EC. Participaram do estudo 85 sujeitos ativos (22,3 ± 2,4 anos, 78 ± 9,4 kg, 176,9 ± 5 cm) que foram aleatoriamente separados em grupo controle (CON), placebo (PLA), suplementação (SUP), isométrico (ISO) e combinado (COMB). CON foi dividido em indivíduos com presença e ausência da α-actinina-3, resultante de polimorfismo do gene ACTN3. Todos os grupos realizaram uma CrED (-15%) por 30 min a 70% VO2max. SUP ingeriu um suplemento rico em compostos flavonoides antes e após a CrED, PLA ingeriu um placebo nos mesmos períodos, ISO realizou 10 contrações isométricas máximas (CIM) dois dias antes da CrED e COMB ingeriu o suplemento e realizou 10 CIM antes da CrED. Foram avaliados marcadores indiretos de DM (força, dor, amplitude de movimento e circunferência da coxa, e atividade sérica de CK) e EC antes e imediatamente, 24, 48, 72 e 96 horas após a CrED. Alterações ao longo do tempo e entre grupos foram analisadas por meio de ANOVA de dois caminhos com post-hoc de Bonferroni. Resultados: Os resultados obtidos demonstraram comprometimento significante da EC após a CrED, assim como alterações significantes em marcadores de DM. Foi identificado um efeito de aceleração da recuperação nas alterações no DM e EC para os grupos SUP e COMB (que apresentaram retorno do VO2 e PTI a níveis basais no dia seguinte e dois dias após a CrED, respectivamente) em relação ao grupo CON. Os grupos que consumiram o suco antioxidante também apresentaram atenuação significante da dor muscular após a CrED. Entretanto, a diferença de cinética de recuperação entre força e EC se manteve mesmo frente aos tratamentos. O grupo ISO apresentou aceleração da recuperação do PTI, mas não da EC, assim como atenuação da dor muscular em relação ao grupo CON. Alterações na altura de saltos foram significantes, sendo que os valores retornaram a níveis basais antes dos valores de PTI. Em relação aos polimorfismos do gene ACTN3, não houve influência do genótipo nas alterações na EC. Entretanto, como esperado, houve maior perda de força para indivíduos RR (que sintetizam a α-actinina-3). Conclusão: Em conjunto, os resultados obtidos sugerem que há, sim, uma relação entre aspectos neuromusculares e a EC. Entretanto, outros fatores, que não a capacidade de produção de força isoladamente, parecem contribuir para essa relação. Ademais, as estratégias de atenuação do DM investigadas no presente estudo parecem ser eficazes na aceleração da recuperação da EC e da força muscular e o polimorfismo do gene ACTN3 parece ser um dos fatores que explicam a susceptibilidade ao DM.
Introduction: Longitudinal studies have been showing that strength production capacity might influence running economy (RE). However, other study models [e.g., downhill running (DhR) induced muscle damage (MD)] show different magnitudes of change and recovery kinetics between strength measured with open kinetic chain exercises [knee extension isometric peak torque (IPT)] and RE might differ. Factors such as the type of exercised used to measures strength and its form of manifestation (maximal or explosive) might, at least in part, explain these differences. The aim of the present study was to analyze the association between strength production capacity and RE using an DhR induced MD model. Methods: To do so, three different prophylactic strategies were adopted to modulate MD induced by DhR and also analyze their effects on strength production capacity and RE. Eighty five active subjects (22.3 ± 2.4 years, 78 ± 9.4 kg, 176.9 ± 5 cm) participated in the study and were randomly allocated to control (CON), placebo (PLA), supplementation (SUP), isometric (ISO), and combined (COMB) groups. Participants in CON were separated based on their ACTN3 polymorphisms. All groups ran downhill (-15%) for 30 min at 70%VO2max. SUP ingested a flavonoid-rich supplement before and following DhR, PLA ingested an isocaloric placebo at the same time points, ISO performed 10 maximal isometric contractinons (MIC) two days prior to the DhR, and COMB ingested the supplement and performed the MIC before DhR. MD symptoms (strength, soreness, knee joint range of motion, thigh circumference, and serum CK activity) and RE were assessed before, immediately after, and 1-4 days following DhR. Changes over time and between groups were analyzed with two-way ANOVA followed by Bonferroni’s post hoc tests. Results: Results showed that RE was significantly compromised following DhR and MD markers were also affected. Faster recovery of RE and MD markers was identified for the SUP and COMB groups (which reached full recovery of VO2 and IPT 1 and 3 days following the DhR, respectively), as compared to CON. Both groups that consumed the antioxidant supplement also presented significant attenuation of muscle soreness following DhR. However, the difference in the recovery kinetics of strength and RE was maintained with this treatment. ISO presented faster recovery of IPT, but not RE, and attenuation and faster recovery of soreness, as compared to CON. Significant changes in jump height were found for all groups, with faster recovery when compared to IPT. No influence of the ACTN3 gene polymorphism was found for changes in RE. However, as expected, RR individuals presented significantly greater strength loss following DhR. Conclusion: When put together, our results suggest that there is, indeed, an association between neuromuscular aspects and RE. However, factors other than strength production per se seem to contribute to this association. Moreover, the prevention/recovery strategies investigated in the present study seem to be effective to promote faster recovery of RE and strength following DhR. Also, ACTN3 gene polymorphism seems to be one of the factors contributing to susceptibility to MD, but not changes in RE.
FAPESP: 2013/23585-4

Greitumas – vienas iš svarbiausių judamųjų gebėjimų. Jis įvairiose sporto šakose pasireiškia skirtingomis formomis. Greitumo pratybos, tai tokios pratybos, kurios nemažina maksimalaus raumens susitraukimo ir atsipalaidavimo greičio, o jį padidina. Yra žinoma, kad jėgos pratybos yra kenksmingos greitumui. Mūsų nuomone, didžiausia problema sporto moksle - kaip padidinti jėgą nesumažinant raumens susitraukimo ir atsipalaidavimo greičio. Taip pat labai aktualu sužinoti, kaip skirtingų genų variantų žmonės geba didinti greitumą ir jo pasireiškimo formas. Šiuo metu pasirodė daug straipsnių apie ACTN3 R577X polimorfizmą. Yra žinoma, kad α-aktinino-3 baltymo nėra pas 16 % pasaulio žmonių. Įdomu tai, kad mutavusio (X) alelio ir ypač pilno α-aktinino-3 (alelis XX) deficito dažnis yra ženkliai mažesnis tarp sprinto ir galingumo atletus. Todėl mes savo tyrime analizavome greitumo pratybų įtaką didesnio jėgos indėlio reikalaujantiems pratimams ir gautus rezultatus lyginome tarp skirtingų RR ir XX genotipų. Tyrimo tikslas – ištirti maksimalaus greitumo padidėjimo įtaką didesnio jėgos indėlio reikalaujantiems judamiesiems gebėjimams priklausomai nuo RR (gaminasi α-aktininas-3) ir XX (nesigamina α-aktininas-3) genotipo. Tyrimo objektas – greitumo pokytis ir to pokyčio įtaka didesnės jėgos reikalaujantiems judamiesiems gebėjimams RR ir XX genotipo grupėse. Tyrimo uždaviniai: 1. Nustatyti ir įvertinti, kaip po 10 maksimalaus greitumo lavinimo pratybų kito RR ir XX grupių greitumo judamasis... [toliau žr. visą tekstą]
Running speed is one of the most important physical properties. It has various forms. Speed training does not decrease muscle contraction and relaxation speed. It is well known, that strength training decrease running speed results. That’s why we think that maybe the most important problem in sport science is how to increase strength without maximum muscle contraction and relaxation speed decrease. Also it is important to know how people with different genotypes can increase running speed. There are only few articles about ACTN3 R577X polymorphism. A common nonsense mutation (R577X) in the ACTN3, resulting in a premature stop codon and lack of detectable protein in homozygous individuals for the ACTN3 null allele (XX genotype), has been demonstrated in the general human population with 16 % prevalence in Caucasians. Sprint athletes have lower 577XX genotype frequency endurance athletes. That’s why we analyze running speed workouts influence to more strength required physical properties between RR and XX groups. The aim of the study - to investigate running speed increment influence to more strength required physical properties and compare results between RR (with α-actinin-3) and XX (without α-actinin-3) groups. The object of the study - running speed change and this change influence to more strength required physical properties between RR (with α-actinin-3) and XX (without α-actinin-3) groups. Study tasks: 1. To investigate how after maximum speed workouts vary running speed... [to full text]

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Medicina, Programa de Pós-Graduação em Patologia Molecular, 2013.
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O exercício físico, por aumentar o consumo de oxigênio, pode produzir um desequilíbrio entre a geração de espécies reativas de oxigênio (ERO) e a capacidade de defesa antioxidante do organismo, levando ao estresse oxidativo. Esta sobrecarga oxidativa pode acarretar dano celular e graves lesões musculares com consequente processo inflamatório. Neste contexto, muitos atletas consomem suplementos antioxidantes para evitar os danos oxidativos, a inflamação e o consequente comprometimento do desempenho. O pequi (Caryocar brasiliense Camb.), um fruto típico do cerrado brasileiro, contém diversos componentes antioxidantes, importantes para reduzir ERO produzidas durante a atividade física. Visto que nutrientes presentes na alimentação podem interagir com o genoma humano para influenciar a saúde e a doença, e variabilidade genética também pode influenciar a resposta à dieta, estudos que avaliem esta interação podem contribuir para futuras intervenções dietéticas baseadas no conhecimento do requerimento nutricional, do estado nutricional e do genótipo. Alguns polimorfismos já foram descritos na literatura como agentes interferentes do desempenho atlético em determinadas categorias de esportes. Entre eles, várias variantes sob a forma de polimorfismos de nucleotídeo único (SNPs) já foram identificadas, incluindo os SNPs R577X no gene da α-actinina-3 (ACTN-3), T→ G na região promotora do gene da eritropoetina (EPO) e G6002A no receptor de eritropoetina (EPOR). O gene da ACTN-3 codifica a proteína α-actinina-3, componente altamente conservado da maquinaria contrátil das fibras rápidas da musculatura esquelética, e é expresso apenas em miofibras tipo II, que são fibras glicolíticas do músculo esquelético. O polimorfismo da ACTN-3 resulta na substituição de uma arginina (R) por um códon prematuro de parada (X) na posição 577 e cria uma proteína não funcional. A eritropoetina (EPO) é o principal hormônio endógeno regulador da eritropoiese, que permite a sobrevivência, proliferação, diferenciação das células progenitoras e, consequentemente, aumento da oxigenação dos tecidos. Assim, polimorfismos no gene da EPO ou de seu receptor (EPOR) podem afetar o desempenho atlético, por influenciarem a expressão de EPO. O objetivo deste trabalho foi investigar a influência dos SNPs rs1815739 no gene da ACTN-3, rs1617640 na região promotora do gene da EPO, e rs121918116 no gene da EPOR na peroxidação lipídica (Teste Tbars), hemograma completo e dosagens bioquímicas de creatina quinase (CK), aspartato aminotransferase (AST), alanina aminotransferase (ALT) e proteína Creativa (PCR e PCR-us), em uma amostra de corredores de rua do Distrito Federal (N=123), antes e depois da suplementação com 400mg de óleo de pequi em cápsulas ingeridas diariamente por 14 dias consecutivos. As amostras de sangue foram coletadas imediatamente após cada uma das corridas para realizar os testes. O DNA foi obtido por extração da fração leucocitária, e a genotipagem foi realizada por PCR seguida por corte com enzimas de restrição. Os dados estatísticos para as frequências alélicas e genotípicas foram gerados pelo programa Genepopweb®, e os demais dados pelo programa Statistical Package for the Social Sciences (SPSS). Os resultados demonstraram que todos os genótipos estudados encontraram-se em equilíbrio de Hardy- Weinberg (EHW). Apesar dos muitos parâmetros analisados apresentarem diferença significativa entre os sexos, faixas etárias e distâncias percorridas, nenhuma correlação entre esses parâmetros e os polimorfismos analisados foi encontrada. O polimorfismo da EPO influenciou os resultados do eritrograma e do plaquetograma, sugerindo uma vantagem aeróbica para o genótipo TG, e uma desvantagem para o genótipo GG sobre possíveis eventos de complicações microvasculares, enquanto não foi encontrada associação do polimorfismo da ACTN-3 com o desempenho de resistência. Ambos os polimorfismos influenciaram as respostas dos corredores ao óleo de pequi, onde respostas significativas foram observadas para o genótipo selvagem da EPO (TT) nos valores de eritrócitos (p=0,000), hematócrito (HCT, p=0,001), hemoglobina corpuscular média (HCM, p=0,000) e concentração hemoglobínica corpuscular média (CHCM, p=0,001); e para os genótipos TT e TG (p=0,000 para ambos) nos valores da amplitude ou variação da distribuição do tamanho dos eritrócitos (RDW). Diferenças significativas também foram observadas no plaquetograma só para os genótipos TT e TG. O polimorfismo da ACTN-3 influenciou principalmente os valores de AST e CK, onde os heterozigotos RX tiveram uma redução significativa nos valores de AST (p=0,037), e os homozigotos XX, nos valores de CK (p=0,010) após a suplementação o óleo de pequi. Esses resultados de resposta diferenciada de cada polimorfismo à suplementação com cápsulas de pequi demonstram a importância dos efeitos da nutrigenômica no desempenho do atleta. _______________________________________________________________________________________ ABSTRACT
Physical exercise, by increasing oxygen consumption, can produce an imbalance between the generation of reactive oxygen species (ROS) and antioxidant defense capacity of the organism, leading to oxidative stress. This oxidative overload can cause cell damage and severe muscle damage with consequent inflammation. In this context, many athletes consume antioxidant supplements to prevent oxidative damage, inflammation and consequent impairment of performance. Pequi (Caryocar brasiliense Camb.), a typical fruit of the Brazilian Cerrado, contains several antioxidant components that are important to reduce ROS produced during physical activity. Since nutrients present in food can interact with the human genome to influence health and disease, and genetic variability may also influence the response to diet, studies evaluating this interaction may contribute to future dietary interventions based on knowledge of nutritional requirement, nutritional status and genotype. Some polymorphisms have been described in the literature as agents interfering in athletic performance in certain categories of sports. Among them, several variants in the form of single nucleotide polymorphisms (SNPs) have been identified, including the SNPs R577X in the α-actinin-3 gene (ACTN-3), T→G in the promoter region of the erythropoietin (EPO) gene, and G6002A in the erythropoietin receptor (EPOR). The ACTN-3 gene encodes protein α-actinin-3, a highly conserved component of the contractile machinery of the fast skeletal muscle fibers, and is expressed only in type II myofibers, which are glycolytic fibers of skeletal muscle. The ACTN-3 polymorphism results in substitution of an arginine (R) by a premature stop codon (X) at position 577 and creates a non-functional protein. Erythropoietin (EPO) is the main endogenous hormone regulator of erythropoiesis, which allows the survival, proliferation, and differentiation of progenitor cells and, hence, increased oxygenation of the tissues. Thus, polymorphisms in the EPO gene or in its receptor can affect athletic performance by influencing the expression of EPO. The aim of this study was to investigate the influence of the SNPs rs1815739 in the ACTN-3 gene, rs1617640 in the promoter region of the EPO gene, and rs121918116 EPOR gene in the lipid peroxidation (TBARS assay), taking a complete hemogram and biochemical dosages of creatine kinase (CK), aspartate aminotransferase (AST), alanine aminotransferase (ALT) and C-reactive protein (CRP and hs-CRP), in a sample of street runners from the Federal District (N=123), before and after supplementation of 400mg of pequi oil capsules taken daily for 14 consecutive days. Blood samples were taken immediately after racing to perform the tests. DNA was obtained by extraction of leukocyte fraction, and genotyping was performed by PCR followed by cut with restriction enzymes. Statistical data for the allele and genotype frequencies were generated by the program Genepopweb®, and other data by the program Statistical Package for the Social Sciences (SPSS). Results showed that all genotypes were found in Hardy-Weinberg Equilibrium (HWE). Although many examined parameters showed significant differences between sexes, age group and distance covered, no correlation between these parameters and the analyzed polymorphisms was found. EPO polymorphism influenced results of erythrogram and plateletgram, suggesting an aerobic advantage for TG genotype and a disadvantage for GG genotype concerning possible microvascular complication events, while no association was found for ACTN-3 polymorphism with endurance performance. Both polymorphisms influenced runner’s response to pequi oil, where the significant responses were observed for EPO wild type genotype (TT) in the red blood cells, hematocrit, mean corpuscular hemoglobin and mean corpuscular hemoglobin concentration values; and for TT and TG genotypes in the red cell distribution width values. Significant differences were also observed in the plateletgram only for TT and TG genotypes. ACTN-3 mainly influenced AST and CK values, where heterozygous RX had significant decrease in AST values (p=0.037), and homozygous XX in CK values (p=0.010) after pequi oil supplementation. These results of differential response of each polymorphism to the supplementation with pequi capsules demonstrate the importance of the effects of nutrigenomics on athletic performance.

Les malformations du cortex cérébral (MDC) représentent une cause importante de handicap et d'épilepsie pharmaco-résistante. Le séquençage à haut débit a permis une amélioration considérable de l'identification des bases moléculaires des MDC non syndromiques. Toutefois, certaines formes, notamment les MDC complexes, demeurent inexpliquées. Mon projet de thèse a pour objectif de progresser dans la compréhension des MDC complexes en utilisant deux modèles : les microlissencéphalies (MLIS) et le syndrome d'Aicardi (AIC), une forme syndromique particulière associant des malformations de l'oeil et du cerveau uniquement rapporté chez les filles. L'étude par séquençage d'exome en trios de 16 familles MLIS m'a permis d'identifier et de caractériser un nouveau gène, WDR81, impliqué dans le cycle cellulaire. Par la même stratégie, j'ai pu identifier un variant homozygote pathogène dans TLE1, un partenaire majeur de FOXG1 dans la balance prolifération/différenciation de progéniteurs neuronaux, dans une famille consanguine de microcéphalie postnatale dont le phénotype est proche du syndrome FOXG1. En parallèle, mes travaux ont permis de préciser les spectres phénotypiques associés à RTTN, EPG5, COL4A1, COL4A2, TBR1, KIF5C, KIF2A et FOXG1. La deuxième partie de mon projet avait pour objet l'identification des bases moléculaires du syndrome d'Aicardi à partir d'une cohorte internationale de 19 patientes. Après avoir exclu un biais d'inactivation du chromosome X et la présence de microremaniements chromosomiques, j'ai réalisé un séquençage d'exome en trio. Aucun variant récurrent n'a été retrouvé dans les séquences codantes. Dans un second temps, j'ai testé une approche combinant les données du séquençage de génome et l'analyse du transcriptome (RNA-Seq) sur fibroblastes, me permettant d'identifier des transcrits dérégulés qui étaient impliqués dans le développement du cerveau et de l'oeil. J'ai comparé les résultats de cette analyse avec ceux de l'analyse du génome dans le but d'identifier des variants dans ces gènes candidats. En conclusion, mon travail de thèse a permis d'améliorer la connaissance des bases moléculaires des MDC complexes et d'ouvrir des perspectives de nouveaux mécanismes tels que ceux engageant les gènes WDR81 et EPG5, et le rôle des endosomes et de l'autophagie dans les MDC, et aussi TLE1 comme nouvelle cause de microcéphalies postnatales. Mes travaux ont également permis de générer une collection de données de séquençage haut débit (WES, WGS et RNA-Seq) qui seront mises en commun dans le cadre d'un consortium international afin de développer des nouvelles stratégies d'analyse en particulier pour les séquences non codantes. Cette approche permettra également d'ouvrir la voie vers la compréhension des mécanismes cellulaires impliqués dans la formation du cerveau et de l' œil
Malformations of cortical development (MCD) are a major cause of intellectual disability and drug-resistant epilepsy. Next Generation Sequencing (NGS) has considerably improved the identification of the molecular basis of non-syndromic MCD. However, certain forms, including complex MCD, remain unexplained. My PhD project aimed to improve the understanding of complex MCD using two disorders: Microlissencephaly (MLIS) and Aicardi Syndrome (AIC), the latter associating brain and eye malformations and only reported in girls. Trio Whole Exome Sequencing (WES) performed in 16 MLIS families allowed me to identify and functionally characterize a new MLIS gene, WDR81, in which mutations lead to cell cycle alteration. Moreover, using the same strategy, I was able to identify a pathogenic homozygous variant in TLE1 in a patient from consanguineous family with a postnatal microcephaly, suggestive of a FOXG1-like presentation. Interestingly, TLE1 is a major partner of FOXG1, a gene involved in maintaining the balance between progenitor proliferation and differentiation. In parallel, my work allowed me to redefine the phenotypic spectrum associated with RTTN, EPG5, COL4A1 and COL4A2, TBR1, KIF5C, KIF2A and FOXG1. The second part of my PhD program was aimed at identifying the genetic basis of AIC in an international cohort of 19 patients. After excluding a skewed X chromosome inactivation and the presence of chromosomal rearrangements, I performed WES in trios. The analysis of the data from WES did not allow me to identify any recurrent variants. I therefore tested a new approach combining Whole Genome Sequencing (WGS) and RNA-Sequencing (RNA-Seq) on fibroblast cells. I identified a number of deregulated transcripts implicated in brain and eye development. I compared the results of this analysis with the WGS analysis in order to find variants in these candidate genes. In conclusion, these studies have improved the knowledge of the molecular basis of complex MCD, such as TLE1 in postnatal microcephaly, and revealed the pathogenic mechanisms such as WDR81 in cell cycle progression and EPG5 in endosomes and autophagy. My work has also generated a collection of NGS data (WES, WGS and RNA-Seq) that will be shared in an international consortium to develop new analytical strategies, in particular for the non-coding DNA regions. This novel strategy provides opportunities to improve understanding of the cellular mechanisms involved in brain and eye development

Activation-tagging is a powerful functional genomics technique used to identify plant genes and their functions. The random introduction of gene enhancers into the plant genome results in the overexpression of nearby genes, thus the gene responsible for a mutant phenotype can be determined based on the location of the enhancers. In a screen for activation-tagged Arabidopsis lines with aberrations in trichome morphology, a mutant (named P330) with unbranched trichomes was identified. In this thesis, the actin- encoding gene ACT11 was found to be upregulated by T-DNA pSKI015 in P330. Additionally, this line also produces a second mutant phenotype, characterized by a significant reduction in seed set. The findings presented in this thesis build upon previous work that has shown ACT11 is strongly expressed in reproductive organs (such as pollen and ovules) and rapidly elongating tissues in Arabidopsis. The role of actin has been characterized in Arabidopsis trichome morphogenesis, however much remains unclear about actin dynamics in sexual reproduction. Our investigation on the effects of increased ACT11 expression adds to our understanding of these processes and also provides the framework for future studies into trichome morphogenesis and sexual reproduction in flowering plants.
Thesis (Master, Biology) -- Queen's University, 2014-06-24 15:14:59.429

O polimorfismo ACTN3 R577X é um dos mais promissores marcadores genéticos associados à performance desportiva, e os estudos existentes com futebolistas apontam para melhores desempenhos de velocidade e força e um menor risco de lesões associados ao alelo R deste polimorfismo. Assim, o polimorfismo ACTN3 R577X, combinado com outros marcadores, poderá ajudar na melhoria da performance desportiva dos futebolistas no futuro. No entanto, são necessários mais estudos que confirmem estes resultados, e as implicações éticas dos testes genéticos no desporto precisam de ser discutidas.
The ACTN3 R577X polymorphism is one of the most promising genetic markers associated with athletic performance, and the current results link the presence of the R allele of this polymorphism in soccer players to better strength and speed performances and a lower injury risk. So, the ACTN3 R577X polymorphism, combined with other markers, could help on the improvement of the players performance levels in the future. However, further confirmatory studies are necessary, and ethical implications of genetic testing in sports need to be addressed.

O polimorfismo ACTN3 R577X é um dos mais promissores marcadores genéticos associados à performance desportiva, e os estudos existentes com futebolistas apontam para melhores desempenhos de velocidade e força e um menor risco de lesões associados ao alelo R deste polimorfismo. Assim, o polimorfismo ACTN3 R577X, combinado com outros marcadores, poderá ajudar na melhoria da performance desportiva dos futebolistas no futuro. No entanto, são necessários mais estudos que confirmem estes resultados, e as implicações éticas dos testes genéticos no desporto precisam de ser discutidas.
The ACTN3 R577X polymorphism is one of the most promising genetic markers associated with athletic performance, and the current results link the presence of the R allele of this polymorphism in soccer players to better strength and speed performances and a lower injury risk. So, the ACTN3 R577X polymorphism, combined with other markers, could help on the improvement of the players performance levels in the future. However, further confirmatory studies are necessary, and ethical implications of genetic testing in sports need to be addressed.

碩士
國立成功大學
環境醫學研究所
101
Lung cancer is the leading cause of cancer related death worldwide. There is a lack of diagnostic tests that specifically address metastasis. In our previous research of comparison of cell secretomes with low and high metastatic abilities, Alpha-actinin-4 (ACTN4) exhibited high expression levels in the cell secretome with high metastatic ability. ACTN4 is one of the members in the ACTN family of binding filaments that preserve cytoskeletal structure and cell morphology. It is also known to have a critical role in transcriptional regulation; this suggests that ACTN4 may have an association with cancer metastasis. In this study, ACTN4 was investigated for its role in lung adenocarcinoma metastasis. Knockdown of ACTN4 expression in lung adenocarcinoma cell lines showed that ACTN4 was a critical mediator in the regulation of carcinoma invasion and cell migration. In the immunofluorescence and confocal microscopy studies, ACTN4 had an indirect effect on the distribution of Actin to plays an important role in the formation of a pseudopod or lamellipodium structure in the cell. Immunohistochemistry was also performed to study the expression levels of ACTN4 in tissues. We found that the ACTN4 expression significantly correlated with the staging of lung cancer; especially in T-staging and N-staging (which are the factors of metastasis progression). The expressed levels of ACTN4 were significantly higher in the tumor tissues than in the normal tissues. Furthermore, the plasma expression levels of ACTN4 in lung adenocarcinoma patients were significantly higher than in the normal controls by enzyme-linked immunosorbent assay. Our results suggest that ACTN4 protein has a significant association with lung cancer metastasis and thus may be a potential biomarker for early detection of lung adenocarcinoma metastasis.

碩士
國立臺灣師範大學
體育學系
97
Purpose: To examine the role of ACTN3, ACE and PPARD polymorphism on physical fitness performance in untrained person. Also, to assess the differences in effect of the polymorphism on phenotype associated with different physical performance. Methods: All 411 subjects were of native Taiwan students and aged from 16-18 years old (241 males and 170 females). Height, weight, BMI and body fat were measured. Physical fitness performance were measured in 40m sprint, 30 sec push-up, 60 sec push-up, 30 sec sit-ups, 60 sec sit-ups, handgrip strength, 25m swim, and 1600m run for males, 800m run for females. The genotype of ACTN3, ACE, and PPARD genes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). One-way ANOVA and Pearson’s product-moment correlation were used for statistical analysis. Results: In ACTN3, the female 30 sec sit-ups performance, RR genotype (19.29 ± 4.49 times) is greater than RX genotype (16.88 ± 4.13 times) (p < .05). In 60 sec sit-ups performance, RR genotype (35.42 ± 8.71 times) is also greater than RX genotype (29.91 ± 8.24 times) (p < .05). In ACE, the female handgrip strength performance, DD genotype (27.23± 4.39 kg) is greater than ID genotype (23.58 ± 4.20 kg) (p < .05). Conclusion: The subjects with ACTN3 RR and ACE DD genotype had better performance in 30 sec sit-ups, 60 sec sit-ups, and handgrip strength.

碩士
國立臺灣大學
微生物學研究所
106
Nuclear receptor interaction protein (NRIP) also named DDB1 and CUL4 associated factor 6 (DCAF6) and IQ motif and WD repeats 1 (IQWD1), is composed of seven WD-40 domains and an IQ motif. NRIP acts as a coactivator to enhance the transcriptional activity of androgen receptor (AR), glucocorticoid receptor (GR) and human papillomavirus (HPV) E2 or prevent AR and HPV E2 from proteasomal degradation. The physiological role of NRIP also determined by using of NRIP global and muscle-specific knockout mice. Previous studies have demonstrated that IQ motif of NRIP associates with calmodulin (CaM) in the presence of calcium (Ca2+) to activate calcineurin (CaN) and Ca2+/calmodulin-dependent protein kinase II (CaMKII) that regulates muscle functions and motor performance. Except NRIP role in skeletal muscle function, our unpublished results demonstrated that NRIP muscle-specific conditional knockout (cKO) mice revealed decrease of heart contraction and performed dilated cardiac hypertrophy. Furthermore, the interaction between NRIP and α-actinin-2 (ACTN2), a heart specific α-actinin isoforms, was also determined in vitro and in vivo. NRIP interacts either ACTN2 or CaM through its IQ motif. According to the study of Michael et al., N-methyl-D-aspartate receptor (NMDAR) containing IQ domain can bind to CaM and ACTN2 in the presence of Ca2+ resulting in recruiting CaMKII to NMDAR-CaM complex for kinase signaling. Like NMDAR, clathrin heavy chain (CHC), endothelial nitric oxide synthase (eNOS), L-type calcium channel (LTCC) and adenosine 2A receptor (A2AR), can interact both with CaM and α-actinin (ACTN). Some of these studies found the binding affinity of proteins with CaM and ACTN would be changed in the presence of Ca2+ that influences different biological consequences. Hence, we hypothesized that CaM and ACTN2 could compete for each other for binding with NRIP through NRIP IQ domain and exerted different biological functions. We firstly combined the His-MBP-NRIP, GST-ACTN2 and CaM proteins and performed in vitro His-tag pull down assay. In addition to in vitro experiments, we also expressed NRIP-GFP, ACTN2-V5 and CaM in 293T cells and performed immunoprecipitation experiments in vivo. We found that the combination of NRIP with ACTN2 and CaM did not achieve consistent results in the presence of Ca2+. Therefore we follow researches over the years and review our experiments.

ACTN3 has been labelled as the ‘gene for speed’ due to the increased frequency of the R allele encoding the α-actinin-3 protein in elite sprint athletes compared to the general population. The results of the first study of this thesis demonstrate that elite athletes who express α-actinin-3 (ACTN3 RR genotype) have faster sprint times compared to those who do not express α-actinin-3 (ACTN3 XX genotype). Further analysis indicates that the ACTN3 genotype accounts for 0.92% in sprint speed amongst elite 200-m athletes. In study two, the same quantitative genetic epidemiological design applied to elite endurance athletes, showed no evidence that a trade-off existed. The endurance athletes with the ACTN3 XX genotype were no faster than those who express the α-actinin-3 protein. These results added to literature that it is unlikely the ACTN3 XX genotype to offer an advantage for endurance performance. While ACTN3 genotype does not appear to influence endurance performance in athletes, studies in mice that completely lack the α-actinin-3 protein suggest the ACTN3 genotype influences the adaptive response to endurance exercise. Based on these findings, the aim of study 3 was to investigate if ACTN3 genotype influences exercise-induced changes in the content of genes and proteins associated with mitochondrial biogenesis. At baseline, there was a compensatory greater α-actinin-2 protein content in ACTN3 XX vs ACTN3 RR participants (p=0.018) but there were no differences in the endurance-related phenotypes measured. There was a main effect of genotype (p=0.006), without a significant interaction effect, for RCAN1-4 or significant exercise-induced expression of genes associated with mitochondrial biogenesis. Together, these results suggest that ACTN3 genotype has a small but significant influence on sprint speed amongst elite sprint athletes. However, loss of α-actinin-3 protein is not associated with higher values for endurance-related phenotypes, endurance performance, or a greater adaptive response to a single session of high-intensity endurance exercise.

Dissertação de Mestrado em Ciências do Desporto, Especialização em Avaliação e Prescrição na Atividade Física
Introdução: O decatlo é uma modalidade desportiva composta por dez provas de atletismo, cumpridas durante dois dias. A performance desportiva nesta modalidade exige uma combinação óptima de diferentes capacidades, nomeadamente resistência, força, velocidade e agilidade. As pesquisas científicas no domínio da biotecnologia e da genética têm identificado um número crescente de genes candidatos com efeito significativo na performance desportiva. Salienta-se em vários estudos de associação que o polimorfismo R577X do gene da alpha-actinin-3 (ACTN3) influencia o desempenho desportivo em atletas de diferentes modalidades. A presença desta proteína funcional (α-actinina 3, ACTN3), enquanto componente estrutural da linha Z do músculo esquelético, parece contribuir para as variações de potência muscular e de velocidade efetiva de contração muscular. Contudo, carecem estudos que determinem o efeito deste polimorfismo no desempenho desportivo no decatlo. Objetivo: Verificar se a variação genética da região R577X do locus ACTN3 apresenta relação com o desempenho desportivo na prova do decatlo. Métodos: A amostra foi composta por 31 atletas de decatlo do gênero masculino, com idade de 18 a 50 (anos). O desempenho desportivo mais recente foi registado nas 10 provas que compõem o decatlo, distintas em provas de velocidade, saltos, arremesso e de lançamento. Procedeu-se à extração do ADN dos sujeitos, recorrendo-se à aplicação dos sítios polimórficos do gene ACTN3 através da técnica modificada de reação em cadeira de polimerase (PCR). Os dados foram agrupados e analisados estatisticamente, tendo sido considerado significativo um valor de P ≤ 0.05. Resultados: Foram identificadas correlações fortes e significativas no desempenho desportivo em particular entre os eventos de velocidade, saltos, arremesso e lançamentos. Foram obtidas as seguintes frequências genotípicas do polimorfismo R577X do ACTN3: RR=51.6% e RX=48.4%, RR=0%. Foi identificado um excesso do alelo R (0.76, p<0.01) nestes atletas. Contudo, não foram encontradas diferenças significativas no desempenho desportivo (pontuação) entre os grupos genotípicos (RR vs RX). Conclusão: Os resultados obtidos confirmam a importância do gene ACTN3 como um marcador genético útil na seleção específica em decatletas de elite brasileira, embora pareça limitado o seu efeito isolado no desempenho desportivo nas diferentes provas.
Introduction: The decathlon is a sport consists of ten track and field events, met for two days. The sporting performance in this mode requires an optimal combination of different capacities, including endurance, strength, speed and agility. Scientific research in the field of biotechnology and genetics have identified a growing number of candidate genes with significant effect on sports performance. It is noted in several studies of association that the R577X polymorphism of the alpha-actinin-3 gene (ACTN3) influences sports performance in athletes of different modalities. The presence of the functional protein (α-actinin 3, ACTN3), as a structural component of skeletal muscle Z-line, appears to contribute to the changes of muscle power and the effective speed of muscle contraction. However, require studies to determine the effect of this polymorphism in sports performance in the decathlon. Objective: To determine if the genetic variation of R577X region of ACTN3 locus is correlated with sporting performance in the decathlon event. Methods: The sample consisted of 31 decathlon athletes males, aged 18-50 (years). The latest sports performance was recorded in 10 events that make up the decathlon, different in speed events, jumping, throwing and release. It proceeded to the extraction of DNA from the subject, resorting to the application of the polymorphic sites in the ACTN3 gene by polymerase chair in modified reaction (PCR). Data were grouped and analyzed statistically and were considered significant at P ≤ 0:05. Results: Strong and significant correlations were identified in sports performance in particular between the speed of events, jumping and throwing and throwing. RR = 51.6% and RX = 48.4%, RR = 0%: The following genotype frequencies of the ACTN3 R577X polymorphism were obtained. An allele of excess R been identified (0.76, p <0.01) in these athletes. However, there were no significant differences in sports performance (score) between the genotype groups (RR vs RX). Conclusion: The results confirm the importance of ACTN3 gene as a useful genetic marker in specific selection in decatletas Brazilian elite, although it seems limited its isolated effect on sporting performance in the tests.

Tese de Doutoramento em Ciências do Desporto
Os declínios relacionados com o avanço da idade ocorrem de forma mais pronunciada nas características de potência muscular, fator inerente nas limitações funcionais e na qualidade de vida na população idosa. Estudos têm demonstrado uma variação substancial entre indivíduos em resposta ao treino de força, indicando o potencial papel da componente genética na adaptação muscular induzida pelo treino. O propósito desta investigação foi 1) examinar o efeito de 12 semanas de treino de potência com velocidade elevada na contração isométrica (força de preensão manual), força máxima (1RM) dos grupos musculares dos membros superiores e inferiores, potência muscular (velocidade de caminhar, salto com contramovimento e lançamento da bola medicinal) e testes funcionais (levantar e sentar da cadeira e levantar ir e voltar testes) em mulheres idosas; 2) analisar o impacto de 6 semanas de destreino na força máxima (1RM), potência e capacidade funcional (levantar e sentar da cadeira em 30s). Além disso, foi ainda objetivo identificar a magnitude das diferenças entre os sujeitos de acordo com a variação genética dos polimorfismos dos genes ECA (I/D) e ACTN3 (R577X) de forma singular ou combinada, antes e após 12 semanas de treino de potência com velocidade elevada. Para a realização dos nossos objetivos, a seguinte sequência foi utilizada: (i) revisão da literatura existente; (ii) analisar os efeitos do treino de potência com velocidade elevada na capacidade funcional e na performance muscular em mulheres idosas; (iii) examinar a retenção da performance muscular e da capacidade funcional após a cessação de treino; (iv) analisar a associação entre os polimorfismos dos genes ECA (I/D) e ACTN3 (R577X) de forma singular ou combinada, em resposta ao treino, (v) e analisar a influência dos genótipos musculares na funcionalidade da extremidade inferior após o programa de treino. Resultados sugerem: (i) o treino de potência com velocidade elevada é efetivo na melhoria da performance muscular e da capacidade funcional em mulheres idosas; (ii) o destreino parece induzir amplos declínios na força muscular em relação à produção de potência, preservando desta forma a independência física, mediada em parte, pela efetividade do treino com velocidades elevadas; (iii) os genótipos ECA e ACTN3 exercem uma influência significante (singular ou combinada) em fenótipos musculares em mulheres idosas em resposta ao treino de potência com velocidade elevada; (iv) os polimorfismos dos genes ECA (I/D) e ACTN3 (R577X) são candidatos na variação de fenótipos relacionados com a potência muscular em mulheres idosas, no entanto não exercem uma influência significantiva em tarefas de mobilidade.
Age-realted declines are more pronounced in muscle power characteristics, a cornerstone of functional limitations and quality of life in older people. Studies have also shown substantial variation among subjects in response of resistant training, indicating the potential role of genetic component to training-induced muscle adaptation. The purpose of this investigation was twofold 1) examine the effect of 12 weeks high-speed power training on isometric contraction (handgrip strength), maximal strength (1RM) of the arm and leg muscles, power performance (walking velocity, counter movement jump and ball throwing) and functional tasks (sit-to-stand and get-up and go tests) in older women and 2) analyze the impact of 6 weeks of detraining on maximal strength (1RM), power performance and functional task (sit-to-stand test). Secondly, it was intended to identify the magnitude of the differences between the subjects according to the genetic variation of the human ACE I/D and ACTN3 R577X polymorphisms (single or combined) before and subsequent to 12 weeks of high-speed power training. For the accomplishment of these purposes the following sequence was used: (i) reviewing available literature; (ii) analyzing the effects of high-speed power training on functional capacity and muscle performance in older women; (iii) examine muscle performance and functional capacity retention after training cessation; (iv) analyzing the association between ACE I/D and ACTN3 R/X polymorphisms (single or combined) in response to training stimuli, (v) and analyze the influence of muscular genotypes on lower-extremity function after the program training. Results suggest that: (i) high-speed power training is an effective exercise approach leading to large gains in upper and lower extremity muscle performance and function capacity in older women; (ii) detraining may induce larger declines in muscle strength than in power output and preserved physical independence, mediated in part, by the effectiveness of high-speed power training; (iii) ACE and ACTN3 genotypes (single or combined) exert a significant influence specially in muscular power phenotypes of older women in response to high-speed power training; (iv) and ACE I/D and ACTN3 R577X polymorphisms are likely candidates in the modulation of exercise-related power phenotypes in older women but not a significant influence in mobility traits.

Duchenne muscular dystrophy (DMD) is the second most common fatal genetic disease in humans, with an incidence of 1 in 3300 live male births. DMD is characterized by progressive cycles of skeletal muscle necrosis/regeneration triggered by the absence of the protein dystrophin from the inner surface of the sarcolemma. In DMD and dystrophin-negative mdx mice, regenerated skeletal muscle fibres are branched and deterioration of muscle contractile function with age is correlated with an increase in both the number and complexity of branched fibres. In this thesis, I present four papers in support of my hypothesis, that when the number and complexity of branched fibres in dystrophin-negative muscles reaches a critical threshold, termed ‘tipping point’, the branches in and of themselves, mechanically weaken the muscle and are susceptible to rupturing when subjected to high forces such as those experienced during eccentric/lengthening contractions. Methodologically, the papers utilise a combination of isolated muscle function contractile measurements coupled with single fibre imaging and confocal microscopy of cleared whole muscles. All experiments use intact muscles isolated from the dystrophic mdx mouse, double knockout (dk)Actn3KO/mdx (dKO) mouse and littermate controls. In conclusion, I propose a two-phase model to explain the aetiology of DMD. Phase-one involves the absence of dystrophin triggering a pathological increase in [Ca2+]in resulting in skeletal muscle fibre necrosis followed immediately by regeneration. The process proceeds cyclically, increasing the number of abnormally branched regenerated dystrophin-deficient muscle fibres. Once the number and complexity of branched fibres passes a level I term ‘tipping point’, phase-two occurs; now eccentric contractions cause force deficits as a consequence of branches rupturing. In the final stage, phase-two will tend to have a positive feedback component, as breaking branches will no longer support the eccentrically contracting muscle, placing additional stress on the remaining branches during the contraction. It is important to note that depending on the complexity of branching and forces experienced by the muscle, phase-one and phase-two are not mutually exclusive and will occur simultaneously.

Research Doctorate - Doctor of Philosophy (PhD)
ACTN4 is a member of the α-actinin family of filamentous actin crosslinking proteins important for regulation of cytoskeletal integrity and cell movement (Honda, 2015; Honda et al., 1998; Hsu and Kao, 2013). Like other family members, ACTN4 contains an N-terminal actin-binding domain (ABD) with two calponin-homology (CH) repeats, a central rod domain (RD) with 4 spectrin-like repeats (SR) that is essential for anti-parallel dimerisation of ACTN4, and a calmodulin (CaM)-like domain (CLD) at the C-terminus (Broderick and Winder, 2002; Djinovic-Carugo et al., 2002; Hsu and Kao, 2013). Although ACTN4 is ubiquitously expressed in normal non-muscle cells (Hsu and Kao, 2013; Oikonomou et al., 2011), its expression is frequently increased in various types of cancer cells including melanoma cells and high ACTN4 expression levels are often correlated with disease progression and poor patient prognosis (Kakuya et al., 2017; Watanabe et al., 2015; Yamamoto et al., 2009). Indeed, there is increasing evidence pointing to a role of ACTN4 in the pathogenesis of cancer (Kakuya et al., 2017; Noro et al., 2013; Watabe et al., 2014; Yamamoto et al., 2009; Yamamoto et al., 2012). This is not only due to its critical involvement in regulation of cancer cell adhesion, invasion, and metastasis (Gao et al., 2015; Honda et al., 1998; Shao et al., 2014), but also closely associated with its role in regulation of signalling pathways through its interaction with a large array of proteins (Agarwal et al., 2013; Aksenova et al., 2013; Khurana et al., 2011). For example, ACTN4 targets the p65/RelA subunit of NF-κB to the nucleus in breast cancer cells upon stimulation with tumour necrosis factor α (TNF-α) or epithelial growth factor (EGF), where it functions as a co-factor for transactivation of nuclear factor κB (NF-κB) target genes (Aksenova et al., 2013; Babakov et al., 2008). Moreover, ACTN4 interacts with Akt1 and promotes its phosphorylation (activation) thus leading to enhanced cell survival and proliferation (Ding et al., 2006). Although the activity of ACTN4 is known to be modulated by multiple mechanisms such as processing by calpain protease, phosphorylation by protein kinases, and binding to phosphatidylinositol intermediates (Carragher et al., 2001; Shao et al., 2010; Sjoblom et al., 2008), how ACTN4 expression is regulated remains to be defined. RIPK1 is a Ser/Thr protein kinase that plays an important role in cell survival and death signal transduction and is a critical determinant of cell fate in response to cellular stress (Christofferson et al., 2012; Festjens et al., 2007; Jin et al., 2016; Liu et al., 2015; Luan et al., 2015; Wang et al., 2008), in particular, in response to activation of death receptors such as TNF receptor 1 (TNFR1). Upon TNFR1 stimulation, prosurvival complex I is formed via recruiting RIPK1 and other proteins involving cellular inhibitor of apoptosis proteins (cIAP1/2), TNFR-associated death domain (TRADD), and TNFR-associated factors (TRAF2/5) (Ofengeim and Yuan, 2013; Wang et al., 2008). This results in stabilisation of RIPK1 through K63-linked polyubiquitination by cIAPs (Bertrand et al., 2008; Liu et al., 2015). K63-linked polyubiquitin chains serve as substrates for binding of the transforming growth factor-β (TGF-β)-activated kinase 1 (TAK1)/TAK1-binding proteins 2 and 3 (TAB2/3) complex and NF-κB essential modulator (NEMO), leading to activation of NF-κB (Blackwell et al., 2013; Liu et al., 2015; Ofengeim and Yuan, 2013). When cIAPs are blocked, RIPK1 is deubiquitinated and its function is switched to that of promoting apoptosis, or alternatively, necroptosis in certain types of cells (Moquin et al., 2013; Wang et al., 2008). RIPK1 is often upregulated and promotes cell proliferation via activation of NF-κB in melanoma (Liu et al., 2015). Stabilisation of RIPK1 by cIAPs is critical for its increased expression (Bertrand et al., 2008; Liu et al., 2015). Here we demonstrate that ACTN4 is necessary for cIAP-mediated stabilisation of RIPK1 through acting as a scaffold to enable the physical association between cIAP1 and RIPK1, and thus plays a critical role in activation of NF-κB and promotion of melanoma cell proliferation. Moreover, we show that NF-κB signalling is responsible for ACTN4 transcriptional upregulation in melanoma cells.

Nephrotic syndrome (NS) is characterized by proteinuria, hypalbuminemia and edemas. It occurs during first and second glomerulopathies. This disease can be divided into two groups: primary (idiopathic) and secondary. The heredity of the familial nephrotic syndrome is autosomal dominant and autosomal recessive. There are four most important genes that condition the formation of hereditary nephrotic syndrome in adult patienst. These genes are ACTN4, CD2AP, NPHS2 and TRPC6. The gene ACTN4, which encodes protein α-actinin 4, is responsible for the autosomal dominant form of focal segmental glomerulosclerosis (FSGS). FSGS is included in first glomerulopathies. α-Actinin 4 was also researched for some types of carcinomas. There was performed the mutational analysis of the gene ACTN4 on the set of 48 patients with nephrotic syndrome in this diploma thesis. High resolution melting (HRM) analysis and sequencing selected samples were used during this mutation detection. During this process many published and unpublished SNPs and one unpublished candidate mutation that could have causal associations with FSGS were found.

Nephrotic syndrome is characterized by proteinuria, hypoproteinemia, edemas and hyperlipidemia. It occurs in primary (e.g. focal segmental glomerulosclerosis, FSGS or minimal change disease, MCD) and in secondary glomerulopathies (e.g. kidney amyloidosis). In primary forms, great attention is paid to the potential genetic background of the disease and due to new molecular genetic methods genes, whose mutations cause different nephropathies (e.g. ACTN4 or INF2) were identified. The aims of presented doctoral thesis were following. Firstly, to continue the mutational analysis of ACTN4 that was described in the author's diploma thesis in other glomerulopathies. Secondly, to implement the mutational analysis of INF2 and subsequently analyse this gene in patients with FSGS/MCD and in patients from special group characterized by positive family history for end stage renal disease (ESRD) in combination with advanced chronic kidney disease (CKD) or already developed ESRD at the time of diagnosis. Thirdly, mutational analysis of NPHS2 and TRPC6 (methods implemented in laboratory earlier) in selected patients from the special group. Finally, expression analyses of genes important for podocyte function or connected with human immune system. This part also verifies the applicability of NPHS2/SYNPO expression...

Workflow technology has long been employed for the modeling, validation and execution of business processes. A workflow is a formal description of a business process in which single atomic work units (tasks), organized in a partial order, are assigned to processing entities (agents) in order to achieve some business goal(s). Workflows can also employ workflow paths (projections with respect to a total truth value assignment to the Boolean variables associated to the conditional split connectors) in order (not) to execute a subset of tasks. A workflow management system coordinates the execution of tasks that are part of workflow instances such that all relevant constraints are eventually satisfied. Temporal workflows specify business processes subject to temporal constraints such as controllable or uncontrollable durations, delays and deadlines. The choice of a workflow path may be controllable or not, considered either in isolation or in combination with uncontrollable durations. Access controlled workflows specify workflows in which users are authorized for task executions and authorization constraints say which users remain authorized to execute which tasks depending on who did what. Access controlled workflows may consider workflow paths too other than the uncertain availability of resources (users, throughout this thesis). When either a task duration or the choice of the workflow path to take or the availability of a user is out of control, we need to verify that the workflow can be executed by verifying all constraints for any possible combination of behaviors arising from the uncontrollable parts. Indeed, users might be absent before starting the execution (static resiliency), they can also become so during execution (decremental resiliency) or they can come and go throughout the execution (dynamic resiliency). Temporal access controlled workflows merge the two previous formalisms by considering several kinds of uncontrollable parts simultaneously. Authorization constraints may be extended to support conditional and temporal features. A few years ago some proposals addressed the temporal controllability of workflows by encoding them into temporal networks to exploit "off-the-shelf" controllability checking algorithms available for them. However, those proposals fail to address temporal controllability where the controllable and uncontrollable choices of workflow paths may mutually influence one another. Furthermore, to the best of my knowledge, controllability of access controlled workflows subject to uncontrollable workflow paths and algorithms to validate and execute dynamically resilient workflows remain unexplored. To overcome these limitations, this thesis goes for exact algorithms by addressing temporal and resource controllability of workflows under uncertainty. I provide several new classes of (temporal) constraint networks and corresponding algorithms to check their controllability. After that, I encode workflows into these new formalisms. I also provide an encoding into instantaneous timed games to model static, decremental and dynamic resiliency and synthesize memoryless execution strategies. I developed a few tools with which I carried out some initial experimental evaluations.