CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / O objetivo deste estudo foi investigar os nÃveis de RNA mensageiro (RNAm) para ativina-A em folÃculos prÃ-antrais e antrais caprinos e os efeitos do hormÃnio folÃculo estimulante (FSH), ativina-A e fator de crescimento e diferenciaÃÃo - 9 (GDF-9) sobre o crescimento e a expressÃo de RNAm para ativina-A, GDF-9, proteÃna morfogenÃtica Ãssea (BMP) -2, -4, -6, -7, -15 e receptor de FSH (R-FSH) em folÃculos prÃ-antrais caprinos cultivados in vitro. Inicialmente, folÃculos primordiais, primÃrios e secundÃrios, bem como complexos cumulus-oÃcito (COCs) e cÃlulas da granulosa mural/teca de pequenos e grandes folÃculos antrais foram isoladas mecanicamente a partir de ovÃrios de cabra e a expressÃo de RNAm para ativina-A foi avaliada por PCR em tempo real. Para os estudos in vitro, folÃculos secundÃrios caprinos foram isolados e cultivados por 6 dias na presenÃa de FSH sozinho (50 ng/mL) ou em combinaÃÃo com ativina-A (100 ng/mL) ou GDF-9 (200 ng/mL). Isoladamente ou em combinaÃÃo com FSH, a influÃncia de ativina-A na expressÃo de ativina-A e R-FSH, e o efeito do GDF-9 sobre os nÃveis de RNAm para GDF-9, R-FSH e BMP -2, -4 , -6, -7 e -15 em folÃculos secundÃrios apÃs 6 dias de cultivo foram testados. Para isso, apÃs a extraÃÃo do RNA total e sÃntese do cDNA, os nÃveis de RNAm para ativina-A, GDF-9, R-FSH e BMP -2, -4, -6, -7 e -15 foram quantificados por PCR em tempo real. Os resultados mostraram que folÃculos secundÃrios apresentavam nÃveis menores de RNAm para ativina-A comparado aos folÃculos primÃrios (p<0,05). NÃo houve diferenÃa nos nÃveis de RNAm para ativina-A entre folÃculos primordial e primÃrio ou secundÃrio (p>0,05). Aliado a isso, nÃo houve diferenÃa entre COCs de pequenos e grandes folÃculos antrais (p>0,05). AlÃm disso, as cÃlulas da granulosa e da teca de grandes folÃculos antrais apresentaram maiores nÃveis de RNAm para ativina-A do que de pequenos folÃculos antrais (p<0,05). Quando comparamos a expressÃo do RNAm para ativina-A entre COCs de pequenos e grandes folÃculos antrais e suas cÃlulas da granulosa e da teca, nenhuma diferenÃa nos nÃveis de expressÃo foi observada (p>0,05). ApÃs o cultivo in vitro de folÃculos secundÃrios, a presenÃa de FSH sozinho ou associado com ativina-A ou GDF-9 aumentou a sobrevivÃncia, crescimento folicular e formaÃÃo de antro (p<0,05). O FSH tambÃm aumentou o RNAm para ativina-A, enquanto os folÃculos cultivados com ativina-A apresentaram nÃveis aumentados de RNAm para R-FSH (p<0,05). AlÃm disso, o GDF-9 diminuiu a expressÃo de RNAm para BMP -2 e -15 (p<0,05), mas nÃo teve efeito sobre a expressÃo de BMP -4, -6 e -7 (p>0,05). Em conclusÃo, durante a transiÃÃo de folÃculo primÃrio para secundÃrio hà uma diminuiÃÃo do RNAm para ativina-A, enquanto ocorre um aumento da expressÃo deste fator durante o crescimento de folÃculos antrais. FSH, ativina-A e GDF-9 estimulam o crescimento de folÃculos secundÃrios caprinos apÃs 6 dias de cultivo. ApÃs cultivo folicular, FSH controla a expressÃo de ativina-A e a expressÃo do R-FSH à regulada por ativina-A. Ainda, GDF-9 reduz a expressÃo do RNAm para BMP -2 e -15. / The aim of this study was to investigate the levels of messenger RNA (mRNA) for activin-A on goat preantral and antral follicles and the effects of follicle stimulating hormone (FSH), activin-A and growth and differentiation factor - 9 (GDF-9) on growth and mRNA expression for activin-A, GDF-9, bone morphogenetic protein (BMP) -2, -4, -6, -7, -15 and FSH receptor (FSH-R) in goat preantral follicles cultured in vitro. Initially, primordial, primary and secondary follicles, as well as cumulus-oocyte complexes (COCs) and mural granulosa/theca cells of small and large antral follicles were isolated mechanically from goat ovaries and the expression of mRNA for activin-A was evaluated by real-time PCR. For in vitro studies, goat secondary follicles were isolated and cultured for 6 days in the presence of FSH alone (50 ng/mL) or in combination with activin-A (100 ng/mL) or GDF-9 (200 ng/mL). Alone or in combination with FSH, the influence of activin-A on expression of activin-A and FSH-R, and the effect of GDF-9 on the levels of mRNA for GDF-9, FSH-R and BMP -2, -4, -6, -7 and -15 in secondary follicles after 6 days of culture were tested. For this, after extraction of total RNA and cDNA synthesis, the levels of mRNA for activin-A, GDF-9, FSH-R and BMP -2, -4, -6, -7 and -15 were quantified by real time PCR. The results showed that secondary follicles had lower levels of mRNA for activin-A, than compared to primary follicles (p<0.05). There was no difference in levels of mRNA for activin-A between primordial and primary or secondary (p>0.05). Allied to this, there was no difference between COCs of small and large antral follicles (p>0.05). Moreover, granulosa and theca cells of large antral follicles showed higher levels of mRNA for activin-A than in small antral follicles (p<0.05). When comparing mRNA expression for activin-A between COCs from small and large antral follicles and their granulosa and theca cells, no difference in expression levels was observed (p>0.05). After in vitro culture of secondary follicles, the presence of FSH either alone or associated with activin-A or GDF-9 increased survival, follicular growth and antrum formation (p<0.05). The FSH increased mRNA for activin-A, while the follicles cultured with activin-A showed increased levels of mRNA for FSH-R (p<0.05). In addition, GDF-9 decreased mRNA expression for BMP -2 and -15 (p<0.05), but had no effect on expression of BMP -4, -6, and -7 (p>0.05). In conclusion, during the transition from primary to secondary follicles there is a reduction of mRNA for activin-A, whereas there is an increased expression of this factor during the growth of antral follicles. FSH, activin-A and GDF-9 stimulate growth of goat secondary follicles after 6-day culture. After follicle culture FSH controls the expression of activin-A, and the expression of FSH-R is regulated by activin-A. Furthermore, GDF-9 reduces the mRNA expression for BMP-2 and -15.

Adipositas und Typ-2-Diabetes sind häufige Erkrankungen des Stoffwechsels. Zur Basistherapie der Adipositas und des Typ-2-Diabetes gehören eine gesunde Ernährungs- weise und die Erhöhung der körperlichen Aktivität unter anderem mit dem Ziel der Gewichtsreduktion. Vermehrte Bewegung führt neben der Verbesserung der körperlichen Leistungsfähigkeit zur Fettmassenreduktion, Verbesserungen der Hyperglykämie, Lipo- proteinstoffwechsels und des Adipokinprofils. Bone morphogenetic proteins (BMPs) werden im Fettgewebe produziert und spielen eine wichtige Rolle in der Adipogenese und Transdifferenzierung von Adipozyten. Während ein Zusammenhang zwischen der BMP-7-Serumkonzentration und Adipositas vor kurzem belegt wurde, ist bisher nicht bekannt, ob weitere BMPs wie BMP-2 und -4 mit Adipositas und Typ-2-Diabetes assoziiert sind. Ziel dieser Arbeit war es deshalb zu untersuchen, ob die BMP-2 und -4 Serumkonzentrationen im Zusammenhang mit Körpergewicht, Fett- verteilung und Parametern des Glukosestoffwechsels bei Patienten mit Adipositas und Typ-2-Diabetes (n=213) stehen. Im Rahmen von drei Interventionsstudien wurde der Einfluss einer hypokalorischen Ernährungsweise über sechs Monate (n=19), eines 45,3 ± 7,4 kg Gewichtsverlustes ein Jahr nach bariatrischer Chirurgie (n=32) sowie eines zwölf- wöchigen Trainingsprogramms (n=60) auf die BMP-2- und -4-Serumkonzentrationen untersucht. Zusätzlich wurde die BMP-2-und -4-mRNA-Expression in humanen omentalen und subkutanen Fettgewebsproben von 161 Patienten charakterisiert. Die BMP-2- und -4-Serumkonzentrationen und die BMP-2- und -4-mRNA-Expression im viszeralen Fettgewebe korrelieren signifikant mit dem BMI und dem Körperfettgehalt. Zirkulierende BMP-4-Spiegel sind geschlechtsabhängig und bei Patienten mit T2D signifikant niedriger als bei gesunden Kontrollpatienten. Sowohl eine moderate Gewichts- reduktion durch kalorienreduzierte Ernährung als auch ein Gewichtsverlust von 45,3 ± 7,4 kg nach bariatrischer Chirurgie führen zu einer signifikanten Reduktion der zirkulierenden BMP-2- und -4-Spiegel. Das zwölfwöchige Trainingsprogramm führte lediglich zu einer signifikanten Reduktion der BMP-2-Serumkonzentration und zu signifikanten Ver- besserung der Leistungsfähigkeit, von Parametern des Glukosestoffwechsels und der Serumkonzentrationen von Adiponektin und Interleukin-6. Zusammengefasst zeigen die Daten, dass erhöhte Serumkonzentrationen von BMP-2 und 4 mit Adipositas assoziiert sind und durch Gewichtsreduktion und Erhöhung der körperlichen Aktivität verringert werden können. Die BMP-2- und -4-mRNA-Expression im viszeralen Fettgewebe kann zu erhöhten Serumkonzentrationen dieser Adipokine bei viszeraler Fettverteilung beitragen.

As BMPs (Bone Morphogenetic Proteins) são membros da superfamília de proteínas TGF-&#946; (Transforming Growth Factor &#946; ), regulam o crescimento e diferenciação de vários tipos celulares em diversos tecidos, e algumas delas desempenham um papel crítico na diferenciação de células de origem mesenquimal em osteoblastos. Particularmente, rhBMP2 e rhBMP7, promovem osteoindução tanto \"in vitro\" como \"in vivo,\" sendo, ambas as proteínas utilizadas terapeuticamente em Ortopedia/Odontologia para reparo ósseo. A expressão diferencial de genes durante a osteodiferenciação de células C2C12 induzida por rhBMP2 e rhBMP7, foi analisada através de microarranjos de DNA, selecionando 31 genes, dos quais 24 foram validados por qPCR, 13 dos quais são relacionados à transcrição, quatro associados a algumas vias de sinalização celular e sete associados à matriz extracelular. Análise funcional destes genes permitirá conhecer, com maiores detalhes, os eventos moleculares que ocorrem durante a diferenciação osteoblástica de células C2C12 induzida por rhBMPs. Em paralelo, foi perseguida a super-expressão de rhBMP2 e rhBMP7 em células HEK293T, demonstrando-se a atividade de rhBMP7, induzindo osteodiferenciação \"in vitro\" e formação de osso \"in vivo\", demonstrando a viabilidade do objetivo de se produzir estas proteínas para futura aplicação como biofármacos no Brasil. / The BMPs (Bone Morphogenetic Proteins) are members of the TGF-&#946; (Transforming Growth Factor &#946;) superfamily of proteins, regulate growth and differentiation of various cell types in various tissues, and some play a critical role in differentiation of mesenchymal cells into osteoblasts. Particularly, rhBMP2 and rhBMP7, promote osteoinduction \"in vitro\" and \"in vivo\" and both proteins are used therapeutically in Orthopedics and Dentistry. The differential expression of genes during osteodifferentiation induced by rhBMP2 and rhBMP7 in C2C12 cells was analyzed through DNA microarrays, allowing the selection of 31 genes, of which 24 were validated by qPCR, 13 of which are related to transcription, four associated with cell signaling pathways and seven are associated with the extracellular matrix. Subsequent functional analysis of these genes should reveal more details on the molecular events which take place during C2C12 cells osteoblastic differentiation induced by rhBMPs In paralel, rhBMPs 2 and 7 were overexpressed in HEK293T cells and BMP7 activity to induce osteodifferentiation \"in vitro\" and bone formation \"in vivo\" was demonstrated, reinforcing the viability of our objective to produce these proteins for future application as biopharmaceuticals in Brazil.

As BMPs (Bone Morphogenetic Proteins) são membros da superfamília de proteínas TGF-&#946; (Transforming Growth Factor &#946; ), regulam o crescimento e diferenciação de vários tipos celulares em diversos tecidos, e algumas delas desempenham um papel crítico na diferenciação de células de origem mesenquimal em osteoblastos. Particularmente, rhBMP2 e rhBMP7, promovem osteoindução tanto \"in vitro\" como \"in vivo,\" sendo, ambas as proteínas utilizadas terapeuticamente em Ortopedia/Odontologia para reparo ósseo. A expressão diferencial de genes durante a osteodiferenciação de células C2C12 induzida por rhBMP2 e rhBMP7, foi analisada através de microarranjos de DNA, selecionando 31 genes, dos quais 24 foram validados por qPCR, 13 dos quais são relacionados à transcrição, quatro associados a algumas vias de sinalização celular e sete associados à matriz extracelular. Análise funcional destes genes permitirá conhecer, com maiores detalhes, os eventos moleculares que ocorrem durante a diferenciação osteoblástica de células C2C12 induzida por rhBMPs. Em paralelo, foi perseguida a super-expressão de rhBMP2 e rhBMP7 em células HEK293T, demonstrando-se a atividade de rhBMP7, induzindo osteodiferenciação \"in vitro\" e formação de osso \"in vivo\", demonstrando a viabilidade do objetivo de se produzir estas proteínas para futura aplicação como biofármacos no Brasil. / The BMPs (Bone Morphogenetic Proteins) are members of the TGF-&#946; (Transforming Growth Factor &#946;) superfamily of proteins, regulate growth and differentiation of various cell types in various tissues, and some play a critical role in differentiation of mesenchymal cells into osteoblasts. Particularly, rhBMP2 and rhBMP7, promote osteoinduction \"in vitro\" and \"in vivo\" and both proteins are used therapeutically in Orthopedics and Dentistry. The differential expression of genes during osteodifferentiation induced by rhBMP2 and rhBMP7 in C2C12 cells was analyzed through DNA microarrays, allowing the selection of 31 genes, of which 24 were validated by qPCR, 13 of which are related to transcription, four associated with cell signaling pathways and seven are associated with the extracellular matrix. Subsequent functional analysis of these genes should reveal more details on the molecular events which take place during C2C12 cells osteoblastic differentiation induced by rhBMPs In paralel, rhBMPs 2 and 7 were overexpressed in HEK293T cells and BMP7 activity to induce osteodifferentiation \"in vitro\" and bone formation \"in vivo\" was demonstrated, reinforcing the viability of our objective to produce these proteins for future application as biopharmaceuticals in Brazil.

<p>Proper morphogenesis is essential for both form and function of mammalian craniofacial and neural tube development. Craniofacial deformities and neural tube defects are highly prevalent human birth defects. Although studies concerning craniofacial and neural tube development have revealed important genetic and/or environmental factors, understanding the mechanisms underlying proper development and the defects remain incomplete. </p><p>Among many genes that were cloned as the gastrula organizer genes in 1990s, Nog, a secreted BMP antagonist, is expressed in the relevant domains during craniofacial and neural tube development. Previous studies show that Nog null embryos exhibit fully penetrant spina bifida (open spine) and to the lesser extent exencephaly (open brain). Moreover, Nog null mice display deformities in skeletal structures including defects in craniofacial skeleton. As such, Nog is essential for proper neural tube and craniofacial development. However, it is still not clear that which domain(s) of Nog are responsible for proper craniofacial development or neural tube closure. In addition, it is also an important question when, and in what capacity Nog is necessary during development of craniofacial and neural tube.</p> / Dissertation

Doctor of Philosophy / Department of Biological & Agricultural Engineering / Kyle Douglas-Mankin / Soil erosion is one of the most important of today’s global environmental problems. Over the past few decades, soil conservation practices were implemented to reduce soil erosion in the United States. However, excessive sediment still remains among the most prevalent water quality problems. Agricultural fields and in particular ephemeral gullies (EGs) are considered to be a major contributor of sediment. The overall goal of this study was to improve modeling utility to identify and quantify sources of sediment. Specific objectives were: (1) to develop and demonstrate a method of field-scale targeting using Soil and Water Assessment Tool (SWAT) and to use this method as a targeted, flexible approach to pay explicitly for sediment-yield reductions; (2) to evaluate topographic index models (Slope Area [SA], Compound Topographic Index [CTI], Slope Area Power [SAP] and Wetness Topographic Index [WTI]) and a physical-based model (Overland Flow Turbulent [OFT]) in predicting spatial EG location and lengths. Black Kettle Creek watershed was the focus of an innovative project to pay for modeled field sediment reductions. An Arc-Geographical Information System (GIS) tool bar was developed that post processed SWAT hydrologic response unit output to field boundaries and prepared maps of high-priority fields by sediment, total nitrogen, and total phosphorus and was demonstrated to be useful for field-scale targeting. Calibrated SWAT model was used to establish baseline sediment yields. Various Best Management Practices (BMPs) were simulated and payments to implement each BMP for a given field were calculated. This study helped to guide determination of appropriate farmer support payments and quantified the important influence of BMP type and site-specific conditions for use in targeting conservation practice funding to achieve maximum soil-loss reductions per dollar spent. Extreme care should be used in selecting the source of spatial model input data when using SWAT for field-level targeting. Automated geospatial models were developed in a GIS environment to spatially locate and derive length of EGs using topographic index and physical based models. EG predictions were better for the SA model among the four topographic index models tested. Individual calibration of topographic index model threshold for each application site was needed. An OFT model (physical based model), which utilized topography, precipitation, soil, landuse/landcover and SWAT-based runoff estimates, did not need individual site calibration, and may have broader applicability than empirical based models.

A total maximum daily load (TMDL) assessment in the Upper North Bosque River (UNBR) has mandated reductions of soluble reactive phosphorus (SRP). The large concentrations of dairies in the UNBR watershed have been identified as a source of the SRP. Agricultural best management practices (BMPs) can be used to reduce in-stream loads of manure nutrients from confined dairy feeding operations (CAFOs). A new BMP utilizes turfgrass sod to export composted dairy manure nutrients out of the impaired watershed in a sustainable manner. Previous plot-scale experiments have showed that 46 to 77% of applied phosphorus (P) and 36 to 47% of applied nitrogen (N) were removed in a single sod harvest. Two, 1.4 ha turfgrass fields were instrumented to measure runoff flow and sediment and nutrient transport. One turfgrass field was topdressed with composted dairy manure and fertilizer N and the other with fertilizer N only. A total of 3.5% of the applied manure P and 3.1% of applied manure N were lost in the surface runoff over a 1.5 year period. The runoff data from the experimental fields were used to calibrate and validate Soil and Water Assessment Tool (SWAT) model simulations of flow, sediment, organic, and mineral nutrients. The Nash-Sutcliffe model fit statistic was greater than 0.6 for flow, sediment, and nutrients during the calibration period and greater than 0.3 during the validation period. Research results indicated that turfgrass sod can be used to export composted dairy manure out of impaired watersheds to improve water and soil quality.

Concentrated animal feeding operations (CAFOs) have caused water quality concerns in many rural watersheds, sometimes forcing the State of Texas to conduct Total Maximum Daily Load (TMDL) assessments of stream nutrients such as nitrogen (N) and phosphorus (P). One suggested Best Management Practice (BMP) is the export of phosphorus (P) through turfgrass sod produced with composted dairy manure from an impaired rural watershed to an urban watershed. The manure-grown sod releases P slowly and would not require additional P fertilizer for up to 20 years in the receiving watershed. This would eliminate P application to the sod and improve the water quality of urban streams. The Soil and Water Assessment Tool (SWAT) was used to model a typical suburban watershed that would receive the transplanted sod. The objective of the modeling was to determine the water quality changes due to the import of sod transplanted from turf fields and grown with composted dairy manure. The SWAT model was calibrated to simulate historical flow and sediment and nutrient loading to Mary's Creek. The total P stream loading to Mary's Creek was lower when manure-grown sod was imported instead of commercial sod grown with inorganic fertilizers. Yet, flow, sediment yield, and total N yield increased equally for both cases at the watershed outlet. The SWAT simulations indicate that a turfgrass BMP can be used effectively to import manure P into an urban watershed and reduce in-stream P levels when compared to sod grown with inorganic fertilizers.

A total maximum daily load (TMDL) assessment in the Upper North Bosque River (UNBR) has mandated reductions of soluble reactive phosphorus (SRP). The large concentrations of dairies in the UNBR watershed have been identified as a source of the SRP. Agricultural best management practices (BMPs) can be used to reduce in-stream loads of manure nutrients from confined dairy feeding operations (CAFOs). A new BMP utilizes turfgrass sod to export composted dairy manure nutrients out of the impaired watershed in a sustainable manner. Previous plot-scale experiments have showed that 46 to 77% of applied phosphorus (P) and 36 to 47% of applied nitrogen (N) were removed in a single sod harvest. Two, 1.4 ha turfgrass fields were instrumented to measure runoff flow and sediment and nutrient transport. One turfgrass field was topdressed with composted dairy manure and fertilizer N and the other with fertilizer N only. A total of 3.5% of the applied manure P and 3.1% of applied manure N were lost in the surface runoff over a 1.5 year period. The runoff data from the experimental fields were used to calibrate and validate Soil and Water Assessment Tool (SWAT) model simulations of flow, sediment, organic, and mineral nutrients. The Nash-Sutcliffe model fit statistic was greater than 0.6 for flow, sediment, and nutrients during the calibration period and greater than 0.3 during the validation period. Research results indicated that turfgrass sod can be used to export composted dairy manure out of impaired watersheds to improve water and soil quality.

No description available.