Higher-order genome organization is important for the regulation of gene expression by bringing different cis-regulatory elements and promoters in proximity. The establishment and maintenance of long-range chromatin interactions occur in response to cellular and environmental cues with the binding of transcription factors and chromatin modifiers. Understanding the organization of the nucleus in differentiation and cancer has been a long standing challenge and is still not well-understood. In this thesis, I explore the dynamic changes in the higher-order chromatin structure in bone differentiation and breast cancer. First, we show dynamic chromatin contact between a distal regulatory element and the promoter of Runx2 gene, which encodes the Runtrelated transcription factor 2 (RUNX2) that is essential for bone development. Next, via using a genome-wide approach, we show that breast cancer cells have altered long-range chromatin contacts among small, gene-rich chromosomes and at telomeres when compared with mammary epithelial cells. Furthermore, we assess the changes in nuclear structure and gene expression of breast cancer cells following Runt-related transcription factor 1 (RUNX1) deficiency, an event frequently observed in breast cancer. Finally, I present the role of the central ATPase subunit of the SWI/SNF complex, SMARCA4 (BRG1), in mediating nuclear structure and gene expression. Taken together, the research presented in this thesis reveals novel insight and paradigm for the dynamic changes in disease and differentiation, as well as uncovers previously unidentified roles for two chromatin regulatory proteins, RUNX1 and SMARCA4.

Submitted by Ana Lúcia Torres (bfmhuap@gmail.com) on 2017-09-28T12:43:48Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇÃO MONIQUE BABINSKI_Pós Defesa 2016 SEM APENDICES (1).pdf: 1899605 bytes, checksum: 5667910f275aadea7cc77dcc4a1ea0c1 (MD5) / Approved for entry into archive by Ana Lúcia Torres (bfmhuap@gmail.com) on 2017-09-28T12:44:09Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇÃO MONIQUE BABINSKI_Pós Defesa 2016 SEM APENDICES (1).pdf: 1899605 bytes, checksum: 5667910f275aadea7cc77dcc4a1ea0c1 (MD5) / Made available in DSpace on 2017-09-28T12:44:09Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇÃO MONIQUE BABINSKI_Pós Defesa 2016 SEM APENDICES (1).pdf: 1899605 bytes, checksum: 5667910f275aadea7cc77dcc4a1ea0c1 (MD5) Previous issue date: 2016 / Complexo Hospitalar de Niterói / O objetivo deste estudo foi avaliar os efeitos da restrição proteica e calórica materna durante a lactação, analisando se haveria recuperação, o conhecido “catch-up” das dimensões morfológicas femorais após a restauração da dieta nutricional. Ao nascimento da ninhada, as progenitoras foram divididas nos seguintes grupos: Controle (C) com dieta a 23% de proteína ad libitum; Restrição Proteico-Calórica (RPC) com dieta a 8% de proteína e Restrição Calórica (RC) com dieta a 23% de proteína, em quantidades restritas de acordo com a ingestão do grupo RPC. Após o desmame, todos os filhotes foram separados das mães e receberam livre acesso a uma dieta padrão de laboratório contendo 23% de proteína, até 180 dias, quando os ratos foram eutanasiados. O fêmur foi excisado e as dimensões femorais macroscópicas foram medidas usando pontos anatômicos pré-estabelecidos. Após a morfometria macroscópica, os fêmures foram fixados em formol 10% (24h) e descalcificados por EDTA. Foram realizadas secções transversais de 10μm de espessura no terço médio diafisário dos espécimes, corados com HE e levados ao microscópio para análise qualitativa e histomorfometria da cortical óssea. Os resultados morfométricos macroscópicos mostraram que a maioria das medidas nos grupos de RC e RPC foram significativamente menores do que no grupo controle. As maiores reduções ocorrendo no grupo de RPC e com várias anormalidades histoarquiteturais, eg., diminuição da área cortical e aumento da área medular. Cortical com lacunas aumentadas significativamente nos grupos RC e RPC. Nossos resultados mostram que a restrição proteico-calórica durante a lactação leva a uma recuperação incompleta na idade adulta. O fêmur mostrou redução significativa na maior parte dos parâmetros macro e microscópicos nos dois grupos tratados, em particular o grupo RPC, quando comparado com o grupo C / The aim of this study was to evaluate the effects of maternal protein and energy restriction during lactation, analyzing on morphological dimensions whether there was catch-up growth through normative nutrition restored, as well as on mechanical axis of femur of the offspring at adulthood. At parturition, Wistar rat dams were randomly assigned to the following groups: 1) control group (C) - free access to a standard laboratory diet containing 23 % protein, 2) protein-energy restricted group (PER) - free access to an isoenergetic, protein-restricted diet containing 8 % protein, and 3) energy-restricted group (ER) – fed with restricted amounts of a standard laboratory diet. At weaning, all pups were separated of dams and received free access to a standard laboratory diet containing 23% protein until 180 days, when the rats were anesthetized and sacrificed. The dimensions of excised pup femur were measured directly using pre-established anatomical points. After macroscopic morphometry, the femurs were fixed in 10% formalin (24h) and decalcified by EDTA. Cross sections of 10μm thickness were made in the middle third (diaphyseal) of the specimens stained with HE and taken to the microscope for qualitative analysis and histomorphometry of cortical bone. Morphometric analysis of the femur (macroscopic) showed that most of the measurements in the ER and PER groups were significantly lower than in the control group, with the greatest reductions occurring in the PER group and several histoarchitectural abnormalities, e.g., decrease in cortical area and increased medullary area. Cortical bone with gaps increased significantly in ER and PER groups. Our results show that protein and energy restriction during lactation leads to an incomplete catch-up growth in adulthood. The femur showed significant reduction in most of the parameters in the two treated groups, particularly the PER group, when compared to the control group

<p>Hormon rasta ima ključnu ulogu u mnogim fiziolo&scaron;kim procesima, anabolički efekti, stimulisanje rasta dugih kostiju, regulacija transkripcije gena u ciljnim ćelijama su uglavnom posredovani preko mitogenog polipeptida, insulinu sličan faktor rasta tip 1 (insulin like growth factor 1-IGF-1). Hormon rasta indukuje proizvodnju IGF-1 u jetri, koji reaguje sa receptorima ciljnih organa indukujući rast, odnosno IGF-1 posreduje svim stimulativnim dejstvima hormona rasta na kost, hrskavicu, rast mi&scaron;ić a i na metabolizam masti i ugljenih hidrata. U proceni redovnosti, bezbednosti i efikasnosti terapije hormonom rasta koristi se merenje koncentracije IGF-1 u serumu. Istraživanje je urađeno kao retrospektivno-prospektivna studija, a obuhvatilo je 80 pacijenata na terapiji hormonom rasta koja se kontroli&scaron;u i leče na Odeljenju za endokrinologiju, dijabetes i bolesti metabolizma Instituta za zdravstvenu za&scaron;titu dece i omladine Vojvodine u Novom Sadu. Istraživani uzorak je obuhvatio 80 pacijenata, od kojih 35 dece sa nedostatkom hormona rasta, 24 dece rođene male za gestacionu dob i 21 devojčicu sa Tarnerovim sindromom. Svi ispitanici su praćeni od početka primene hormona rasta i tokom prve dve godine terapije hormonom rasta. U ovom istraživanju su praćeni auksolo&scaron;ki i laboratorijski parametri u cilju ispitivanja odgovora na terapiju hormonom rasta. Praćene su bazalne vrednosti IGF-1 i promene nivoa IGF-1 u serumu tokom terapije hormonom rasta i kori&scaron;ćene da bi se ispitao odgovor na terapiju hormonom rasta, praćenjem brzine rasta, promena skora standardnih devijacija - SSD za telesnu visinu i ko&scaron;tanog sazrevanja. Ciljevi istraživanja su bili da se utvrdi povezanost vrednosti insulinu sličnog faktora rasta tip 1, brzine rasta i ko&scaron;tanog sazrevanja tokom terapije hormonom rasta. Takođe je poređena brzina rasta dece sa deficitom hormona rasta, devojčica sa T arnerovim sindromom i dece rođene male za gestaciono doba na terapiji hormonom rasta. U istraživanom uzorku, dvogodi&scaron;njim praćenjem terapije hormonom rasta je postignut dobar odgovor na terapiju, među decom sa nedostatkom hormona rasta je 71,5% postiglo normalnu telesnu visinu (&plusmn;2 SSDTV) posle dve godine terapije hormonom rasta, 79,2% dece rođene male za gestacionu dob i 42,9% devojčica sa Tarnerovim sindromom. Značajna zastupljenost dece prepubertetskog uzrasta na početku terapije hormonom rasta, među decom sa nedostatkom hormona rasta 77,2%, među decom rođenom malom za gestacionu dob 79,1% i među devojčicama sa Tarnerovim sindromom 90,5% &scaron;to je značajno uticalo na uspe&scaron;nost terapije. Tokom terapije hormonom rasta je utvrđeno povećanje brzine rasta i SSD TV kod sve tri grupe ispitanika. U sve tri grupe ispitanika je tokom terapije hormonom rasta utvrđen porast nivoa IGF-1 seruma i SSDIGF-1 i ubrzanje ko&scaron;tanog sazrevanja tokom terapije hormonom rasta. Za prvih &scaron;est meseci terapije nema statistički značajnih razlika među grupama u brzini rasta (p&gt;0,05), dok je za period prve i druge godine terapije hormonom rasta utvrđeno da postoji statistički značajna razlika među grupama (p&lt;0,05), da je brzina rasta kod devojčica za Tarnerovim sindromom statistički značajno manja i od brzine rasta kod dece sa nedostatkom hormona rasta (p &lt;0,05), i od brzine rasta kod dece rođene male za gestacionu dob (p&lt;0,05). Među decom sa nedostatkom hormona rasta i dece rođene male za gestacionu dob nema statistički značajne razlike u brzini rasta (p&gt;0,5). U ovom istraživanju je praćenjem auskolo&scaron;kih i laboratrijskih parametara tokom dvogodi&scaron;nje primene hormona rasta, konstruisano vi&scaron;e matematičkih modela za predviđanje odgovora na terapiju hormona rasta koji su statistički veoma značajani sa visokim koeficijentom vi&scaron;estruke linearne korelacije. U ovom istraživanju nije dobijena statistički značajna korelacija izmedju nivoa promene IGF-1 i brzine rasta za ceo uzorak, kao ni za decu sa nedostatkom hormona rasta, decu rođenu malu za gestacionu dob i devojčice za Tarnerovim sindromom. Nije dobijena statistički značajna korelacija izmedju nivoa promene IGF-1 i ubrzanja ko&scaron;tanog sazrevanja za ceo uzorak i za tri grupe pacijenata.</p> / <p>Growth hormone plays a key role in many physiological processes. The anabolic effects, the stimulation of growth of the long bones and the regulation of gene transcription in the target cells are mediated mainly via mitogenic polypeptide and insulin-like growth factor type 1 (insulin like growth factor 1-IGF-1). Growth hormone induces the production of IGF-1 in the liver, which interacts with receptors of the target organs inducing growth, that is, IGF-1 mediates all the stimulating effects of growth hormone on bone, cartilage, muscle growth and the metabolism of fats and carbohydrates. In assessing the regularity, safety and efficacy of growth hormone therapy, measuring the concentration of IGF-1 in serum is used. The survey was conducted as a retrospective-prospective study and involved 80 patients treated with growth hormone, monitored and treated at the Department of Endocrinology, Diabetes and Metabolic Diseases, at the Institute for Health Protection of Children and Youth of Vojvodina in Novi Sad. Investigated sample included 80 patients, of whom 35 children have growth hormone deficiency, 24 children were born small for gestational age and 21 girls with Turner syndrome. All the patients were monitored from the beginning of the administration of growth hormone and during the first two years of growth hormone therapy. In this study, auxological and laboratory parameters were monitored for the purpose of examining the response to treatment of growth hormone. The basal values of IGF-1 and changes in IGF-1 levels in serum, along with monitoring the rate of growth velocity and recent changes in standard deviation - SSD for body height and bone maturation, were monitored during growth hormone therapy and used for the evaluation of the response to growth hormone therapy. The objectives of the study were to determine the correlation of insulin-like growth factor type 1 values, the growth velocity and maturation of bone during growth hormone therapy. Also, the growth velocity in children with growth hormone deficiency was compared with the growth velocity in girls with Turner syndrome and in children born small for gestational age while treated with growth hormone. Two-year monitoring of growth hormone therapy in the study sample has show n good response to therapy. 71.5% of children with growth hormone deficiency, 79.2% of children born small for gestational age, and 42.9% of girls with Turner syndrome achieved normal body height (&plusmn; 2 SSDTV) after two years of growth hormone therapy. There was a significant share of children at prepubertal age at the beginning of growth hormone therapy: 77.2% of children with growth hormone deficiency, 79.1% of children born small for gestational age and 90.5% of girls with Turner syndrome, which significantly influenced the success of the therapy. During the growth hormone therapy there was an increase of growth velocity and SSD TV in all three groups of children. An increase in levels of IGF-1 serum and SSDIGF-1 and acceleration of bone maturation were determined in all three groups of patients during growth hormone therapy. For the first six months of therapy there was no statistically significant difference between groups in growth velocity (p&gt; 0.05), while the period of the first and second year of growth hormone therapy showed a statistically significant difference between groups (p &lt;0.05). The growth velocity in girls with Turner syndrome was significantly lower than the growth velocity in children with growth hormone deficiency (p &lt;0.05) and in children born small for gestational age (p &lt;0.05). Between children with growth hormone deficiency and children born small for gestational age there was no statistically significant difference in growth velocity (p&gt; 0.5). By monitoring auxological and laboratory parameters during the two years of application of growth hormone, several highly statistically significant mathematical models for predicting the response to treatment of growth hormone were constructed in this study with a high coefficient of multiple linear correlation. In this study, there was no statistically significant correlation between the level of change in IGF-1 and growth velocity for the entire sample, as well as for children with growth hormone deficiency, children born small for gestational age and girls for Turner syndrome. There was no statistically significant correlation between the level of change in IGF-1 and acceleration of bone maturation for the entire sample and for the three groups of patients.</p>