DeMiguel, Garlappi e Uppal (2009) fizeram a comparação da regra 1/N ou de Talmud com 14 modelos de otimização que vieram depois do trabalho de Markowitz (1952). As conclusões mostraram que todos os modelos de alocação ótima analisados tiveram um desempenho inferior ao da regra de Talmud. Tu e Zhou (2011) propuseram uma combinação entre Markowitz e Talmud para que tal modelo superasse Talmud. Os resultados obtidos foram satisfatórios. A desconsideração dos eventos extremos (dependência assimétrica ou caudal) durante o processo de construção de carteiras poderá diminuir as habilidades dos gestores de ativos em reduzir o risco através da diversificação. A modelagem de cópulas sobre os retornos dos ativos nos permite calcular uma alternativa para medir a dependência dos ativos em eventos extremos através do índice de dependência caudal inferior. Hatherley e Alcock (2007) relataram que o modelo de Markowitz tende a subestimar as perdas potenciais que venham a ocorrer na presença de eventos extremos de mercado (crashes) para um determinado nível de retorno esperado. Verificamos se as estratégias com dependência caudal superaram Talmud, o modelo de Markowitz e o modelo de Tu e Zhou (2011) através da simulação de 1.000 carteiras com 3, 5, 10 e 20 ativos escolhidos ao acaso do índice DJIA no período de 03/1990 até 12/2012. Concluímos que os modelos de dependência caudal e o de Markowitz tiveram uma desempenho fora da amostra superior ao Talmud e ao modelo de Tu e Zhou (2011) para as carteiras com 3, 5, 10 e 20 ativos. A estratégia com dependência caudal superou Markowitz, em termos de retorno acumulado, em mais de 60% dos meses considerados em todas as análises. Os resultados apontam que a regra de Talmud deve ser descartada num contexto de construção de carteiras com ações frente à estratégia com dependência caudal. / DeMiguel, Garlappi and Uppal (2009) made a comparison of rule 1 / N or Talmud with most optimization techniques that followed the work of Markowitz (1952). The conclusions were devastating for all asset allocation models in the context of portfolios combined with other portfolios. Tu and Zhou (2011) proposed a combination between Markowitz and Talmud to overcome such a rule Talmud. The results were satisfactory. In the presence of extreme events, the Pearson correlation coefficient tends to increase in magnitude, making spurious results diversification based solely on this factor. The elimination of extreme events (asymmetric or tail dependence) during the portfolio construction process can reduce the skills of asset managers to reduce risk through diversification. The copula theory allows us to calculate an alternative to measure the dependence of extreme events in assets through the index lower tail dependence. Hatherley and Alcock (2007) reported that the Markowitz model tends to underestimate the potential losses that may occur in the presence of extreme market events (crashes) for a given level of expected return. We check that the strategies with tail dependence overcame Talmud rule, the Markowitz model and the model of Tu and Zhou (2011) by simulating 1,000 portfolios with 3, 5, 10 and 20 randomly selected assets from DJIA for the period 03/1990 until 12/2012. We conclude that models of tail dependence and Markowitz had more performance ex-ante than Talmud and the Tu and Zhou (2011) model for portfolios with 3, 5, 10 and 20 assets. Tail dependence models overcome Markowitz, in terms of cumulative return, in over 60% of months considered in the analysis. The results indicate that the Talmud rule should be discarded in a context of constructing portfolios with individual stocks ahead strategies with tail dependence.

La fertilidad masculina puede ser medida mediante un espermatograma convencional, sin embargo este examen no incluye la valoración de la integridad del ADN espermático. Esta variable ha sido correlacionada con las tasas de fertilización, viabilidad y desarrollo del embrión, convirtiéndose en una herramienta de importancia clínica tanto para los programas de reproducción animal como los tratamientos de fertilidad asistida. El bioensayo Cometa es capaz de determinar de una manera exacta el valor de la integridad del ADN espermático, lamentablemente este examen no es de uso rutinario por su elevado costo de implementación ya que utiliza microscopia especializada y tinciones fluorescentes para evidenciar la migración del ADN. El objetivo de esta investigación fue evaluar la compatibilidad de las tinciones no fluorescentes Diffquik, Giemsa, de Feulgen y FastBlast en el bioensayo Cometa usando un método visual y automatizado. Se utilizaron 15 eyaculados previamente seleccionados de acuerdo al manual OMS 2010, para luego ser capacitados en búsqueda de homogeneidad adecuada para la experimentación. Cada muestra fue expuesta a una gradiente de Peróxido de hidrogeno (0, 10, 30,60 y 100 mM) por 1 hora a 4°C para luego evaluar el coeficiente de daño mediante el método visual y porcentaje de ADN en la cola mediante el método automatizado. Las pendientes de la regresión lineal en el método visual indican que los valores obtenidos por la tinción control SybrGreen (m=3,69) difieren con Giemsa (m=3,45) y Diffquik (m=2,57). En el método automatizado de igual manera SybrGreen (m=0.83), Giemsa (m=0,79) y Diffquik (m=0,77). Sin embargo SybrGreen es 1,06 veces más efectivo que Giemsa en el visual y 1,05 veces en el automatizado, sugiriendo una compatibilidad con el bioensayo cometa. De igual manera SybrGreen es 1,07 veces más efectivo que Diffquik en el visual y 1,44 veces en el automatizado, concluyendo una compatibilidad solo en el método visual.Male fertility can be measured by a conventional semen analysis, however, this examination does not include the assessment of sperm DNA integrity. This variable has been correlated with fertilization rates, embryo viability and development, becoming a tool of clinical importance for both animal breeding programs and assisted fertility treatments. Comet bioassay is able to determine an exact way the value of sperm DNA integrity, unfortunately this test is not routinely used because of its high cost of implementation because it uses specialized microscopy and fluorescent dyes to demonstrate DNA migration. The objective of this research was to evaluate the compatibility of non-fluorescent dyes Diffquik, Giemsa, Feulgen and Comet FastBlast in the bioassay using a visual and automated method. 15 ejaculates were used previously manually selected according to WHO 2010 and then be trained in finding adequate homogeneity for experimentation. Each sample was exposed to a hydrogen peroxide gradient (0, 10, 30,60 and 100 mM) for 1 hour at 4 ° C and then assess the damage coefficient by visual method and percentage of DNA in the tail by automated method. The slopes of the linear regressions on the visual method indicate that the values obtained by the SybrGreen Control staining (m = 3.69) differ with Giemsa (m = 3.45) and Diffquik (m = 2.57). In the same way automated method SybrGreen (m = 0.83), Giemsa (m = 0.79) and Diffquik (m = 0.77). However SybrGreen is 1.06 times more effective than Giemsa visual and 1.05 times in the automated, suggesting a comet support bioassay. Similarly SybrGreen is 1.07 times more effective than Diffquik visual and 1.44 times in the automated, concluding compatibility only in the visual method. Keywords:

This thesis investigates different aspects of the impact of extreme downside risk on stock returns. We first investigate the impact at market level, where the return of the stock market index is expected to be positively correlated to its tail risk. More specifically, we incorporate Markov switching mechanism into the framework of Bali et al. (2009) to analyse the relationship between risk and returns under different market regimes. Interestingly, although highly significant in calm periods, the tail risk-return relationship cannot be captured during turbulent times. This is puzzling since this is the time when the distress risk is most prominent. We show that this pattern persists under different modifications of the framework, including expanding the set of state variables and accounting for the non-iid feature of return process. We suggest that this result is due to the leverage and volatility feedback effects. To better filter out these effects, we propose a simple but effective modification to the risk measures which reinstates the positive extreme risk-return relationship under any state of market volatility. The success of our method provides insights into how extreme downside risk is factored into expected returns. In the second investigation, this thesis explores the impact of extreme downside risk on returns in a security level analysis. We demonstrate that a stock with higher tail risk exposure tends to experience higher average returns. Motivated by the limitations of systematic extreme downside risk measures in the literature, we propose two groups of new ‘co-tail-risk’ measures constructed from two different approaches. The first group is the natural development of canonical downside beta and comoment measures, while the second group is based on the sensitivity of stock returns on innovations in market systematic crash risk. We utilise our new measures to investigate the asset pricing implication of extreme downside risk and show that they can capture a significant positive relationship between this risk and expected stock return. Moreover, our second group of ‘co-tail-risk’ measures show a highly consistent performance even in extreme settings such as low tail threshold and monthly sample estimation. The ability of this measure to generate a number of observations given limited return data solves one of the most challenging problems in tail risk literature. In the last investigation, this thesis examines the influence of extreme downside risk on portfolio optimisation. It is motivated by the evidence in Chapter 4 regarding the size pattern of the extreme downside risk impact on stock returns where the impact is larger for small stocks. Accordingly, portfolio optimisation practice that focuses on tail risk should be more effective when applied to small stocks. In comparing the performance of mean-Expected Tail Loss against that of mean-variance across size groups of Fama and French’s (1993) sorted portfolios, we confirm this conjecture. Moreover, we further investigate the performance of different switching approaches between mean-variance and mean-Expected Tail Loss to utilise the suitability of these optimisation methods for specific market conditions. However, our results reject the use of any switching method. We demonstrate the reason switching could not enhance performance is due to the invalidity of the argument regarding the suitability of any optimisation method for a specific market regime.

Tgf-β, Wnt, and Fgf signaling pathways are required for many developmental processes. Here, I investigated the requirement of these signaling pathways during tail regeneration and limb development in the Mexican axolotl (Ambystoma mexicanum). Using small chemical inhibitors during tail regeneration, I found that the Tgf-β signaling pathway was required from 0-24 and 48-72 hours post tail amputation (hpa), the Wnt signaling pathway was required from 0-120 hpa, and the Fgf signaling pathway was required from 0-12hpa. Tgf-β1 was upregulated after amputation and thus may mediate Tgf-β signaling pathway during tail regeneration. Both Smad-mediated and non-Smad mediated Tgf-β signaling were activated as early as 1hpa. Smad-mediated Tgf-β signaling via activated pSmad2 and pSmad3, and via phosphorylated Erk and Akt. Two different Tgf-β signaling pathway inhibitors, SB505124 and Naringenin, differentially regulated pSmad2, pSmad3, p-Erk, and p-Akt, while SB505124 and Naringenin both inhibited tail regeneration; only SB505124 reduced cell proliferation. Wnt/β-Catenin signaling was increased and was enhanced by Wnt-C59. Disruption of the Wnt signaling pathway directly or indirectly activated Erk and Akt signaling. Disruption of the Fgf signaling pathway decreased p-Erk and increased p-Akt. All three signaling pathways affected cell proliferation and mitosis during tail regeneration. The Wnt pathway inhibitor Wnt-C59 prevented forelimb bud outgrowth. The critical window for Wnt signaling regulating forelimb bud outgrowth was approximately developmental stage 40-42. Wnt signaling ligand Wnt3a and tight junction protein Zo-1 were expressed in the epidermis of the forelimb bud and both were down-regulated by Wnt-C59. Moreover, both Wnt and Fgf signaling pathways affected cell proliferation and mitosis of mesodermal cells during forelimb bud outgrowth. Overall, my results show that Tgf-β, Wnt, and Fgf signaling pathways are required for axolotl tail regeneration. All three pathways affect Erk and Akt signaling and guide cell proliferation and mitosis. The Wnt signaling pathway is required for forelimb bud outgrowth, and it appears to regulate expression of Wnt3a and Zo1, and control cell proliferation and mitosis of mesodermal cells underlying the forelimb epidermis. These data enrich understanding of signaling network dynamics that underlie tissue regeneration and vertebrate limb development.

Let (X1, X2, X3) be a tri-variate normal vector with a non-singular co-variance matrix ∑ , where for i ≠ j, ∑ij < 0 . It is shown here that it is then possible to determine the three means, the three variances and the three correlation coefficients based only on the knowledge of the probability density function for the minimum variate Y = min{X1 , X2 , X3 }. We will present a method for identifying the nine parameters which consists of careful determination of the asymptotic orders of various bivariate tail probabilities.

<p>We model Normal Inverse Gaussian distributed log-returns with the assumption of stochastic volatility. We consider different methods of parametrization of returns and following the paper of Lindberg, [21] we</p><p>assume that the volatility is a linear function of the number of trades. In addition to the Lindberg’s paper, we suggest daily stock volumes and amounts as alternative measures of the volatility.</p><p>As an application of the models, we perform Value-at-Risk and Expected Tail Loss predictions by the Lindberg’s volatility model and by our own suggested model. These applications are new and not described in the</p><p>literature. For better understanding of our caluclations, programmes and simulations, basic informations and properties about the Normal Inverse Gaussian and Inverse Gaussian distributions are provided. Practical applications of the models are implemented on the Nasdaq-OMX, where we have calculated Value-at-Risk and Expected Tail Loss</p><p>for the Ericsson B stock data during the period 1999 to 2004.</p>

We model Normal Inverse Gaussian distributed log-returns with the assumption of stochastic volatility. We consider different methods of parametrization of returns and following the paper of Lindberg, [21] we assume that the volatility is a linear function of the number of trades. In addition to the Lindberg’s paper, we suggest daily stock volumes and amounts as alternative measures of the volatility. As an application of the models, we perform Value-at-Risk and Expected Tail Loss predictions by the Lindberg’s volatility model and by our own suggested model. These applications are new and not described in the literature. For better understanding of our caluclations, programmes and simulations, basic informations and properties about the Normal Inverse Gaussian and Inverse Gaussian distributions are provided. Practical applications of the models are implemented on the Nasdaq-OMX, where we have calculated Value-at-Risk and Expected Tail Loss for the Ericsson B stock data during the period 1999 to 2004.

This thesis demonstrates the feasibility of methods developed to increase the quality of the crescent bond together with the tail bond quality. Low pull force of the crescent bond limits the usage of insulated Au wire in microelectronics assembly. Premature break of the tail which results in the stoppage of the bonding machine is one of obstacles to overcome for Cu wire. The primary focus of this thesis is to understand the tail and crescent bonding process and then to propose methodologies to improve thermosonic wire bonding processes when Cu and insulated Au wires are used. Several series of experiments to investigate the crescent and tail bonding processes are performed on auto bonders. Cu and insulated Au wires with diameters of 25mm are bonded on the diepads of Ag leadframes. For simplicity, wire loops are oriented perpendicular to the ultrasonic direction. It was found that the crescent bond breaking force by pulling the wire loop (pull force) with insulated Au wire is about 80 % of that of bare Au wire. A modification of the crescent bonding process is made to increase the pull force with insulated Au wire. In the modified process, an insulation layer removing stage (cleaning stage) is inserted before the bonding stage. The cleaning stage consists of a scratching motion (shift) toward to the ball bond in combination with ultrasound. Bonds are then made on the fresh diepad with the insulation removed from the contact surface of the insulated Au wire. This process increases the pull force of the crescent bond up to 26% which makes it comparable to the results obtained with bare Au wire. An online tail breaking force measurement method is developed with a proximity sensor between wire clamp and horn. Detailed understanding of tail bond formation is achieved by studying tail bond imprints with scanning electron microscopy and energy dispersive x-ray analysis. Descriptions are given of the dependence of the tail breaking force on the bonding parameters, metallization variation, and cleanliness of the bond pad. Simultaneous optimization with pull force and tail breaking force can optimize the Cu wire bonding process both with high quality and robustness. It is recommended to first carry out conventional pull force optimization followed by a minimization of the bonding force parameter to the lowest value still fulfilling the pull force cpk requirement. The tail bond forms not only under the capillary chamfer, but also under the capillary hole. The tail breaking force includes both the interfacial bond breaking strength and the breaking strength of the thinned portion of the wire that will remain at the substrate as residue. Close investigations of the tail bond imprint with scanning electron microscopy indicate the presence of fractures of the substrate indicating substrate material being picked up by Cu wire tail. Pick up is found on Au and Cu wires, but the amount of pick up is much larger on Cu wire. The effect on the hardness of the subsequently formed Cu free air ball (FAB) as investigated with scanning electron microscopy and micro - hardness test shows that Cu FABs containing Au and Ag pick ups are softer than those without pick up. However, the hardness varies significantly more with Au pick up. The amount of Au pick up is estimated higher than 0.03 % of the subsequently formed FAB volume, exceeding typical impurity and dopant concentrations (0.01 %) added during manufacturing of the wire.

Protein arginine methyltransferase 1 (PRMT1) is a key posttranslational modification enzyme that catalyzes the methylation of specific arginine residues in histone and nonhistone protein substrates, regulating diverse cellular processes such as transcriptional initiation, RNA splicing, DNA repair, and signal transduction. Recently the essential roles of PRMT1 in cancer and cardiovascular complications have intrigued much attention. Developing effective PRMT inhibitors therefore is of significant therapeutic value. The research on PRMT inhibitor development however is greatly hindered by poor understanding of the biochemical basis of protein arginine methylation and lack of effective assays for PRMT1 inhibitor screening. Herein, we report our effort in the kinetic mechanism study as well as the fluorescent probe and inhibitor development for PRMT1. New fluorescent reporters were designed and applied to perform single-step analysis of substrate binding and methylation of PRMT1. Using these reporters, we performed transient-state fluorescence measurements to dissect the rate constants along the PRMT1 catalytic coordinate. The data give evidence that the chemistry of methyl transfer is the major rate-limiting step, and that binding of the cofactor SAM or SAH affects the association and dissociation of H4 with PRMT1. Importantly, we identified a critical kinetic step suggesting a precatalytic conformational transition induced by substrate binding. On the other hand, we discovered a type of naphthyl-sulfo (NS) compounds that block PRMT1- mediated arginine methylation at micromolar potency through a unique mechanism: they directly target the substrates but not PRMT enzymes for the observed inhibition. We also found that suramin, an anti-parasite and anti-cancer drug bearing similar functional groups, effectively inhibited PRMT1 mediated methylation. These findings about novel PRMT inhibitors and their unique inhibition mechanism provide a new way for chemical regulation of protein arginine methylation. Addionally, to dissect the interplaying relationship between different histone modification marks, we investigated how individual lysine acetylations and their different combinations at the H4 tail affect Arg-3 methylation in cis. Our data reveal that the effect of lysine acetylation on arginine methylation depends on the site of acetylation and the type of methylation. While certain acetylations present a repressive impact on PRMT-1 mediated methylation (type I methylation), lysine acetylation generally is correlated with enhanced methylation by PRMT5 (type II dimethylation). In particular, Lys-5 acetylation decreases activity of PRMT1 but increases that of PRMT5. Furthermore, hyperacetylation increases the content of ordered secondary structures of H4 tail. These findings provide new insights into the regulatory mechanism of Arg-3 methylation by H4 acetylation, and unravel that complex intercommunications exist between different posttranslational marks in cis.

This thesis demonstrates the feasibility of methods developed to increase the quality of the crescent bond together with the tail bond quality. Low pull force of the crescent bond limits the usage of insulated Au wire in microelectronics assembly. Premature break of the tail which results in the stoppage of the bonding machine is one of obstacles to overcome for Cu wire. The primary focus of this thesis is to understand the tail and crescent bonding process and then to propose methodologies to improve thermosonic wire bonding processes when Cu and insulated Au wires are used. Several series of experiments to investigate the crescent and tail bonding processes are performed on auto bonders. Cu and insulated Au wires with diameters of 25mm are bonded on the diepads of Ag leadframes. For simplicity, wire loops are oriented perpendicular to the ultrasonic direction. It was found that the crescent bond breaking force by pulling the wire loop (pull force) with insulated Au wire is about 80 % of that of bare Au wire. A modification of the crescent bonding process is made to increase the pull force with insulated Au wire. In the modified process, an insulation layer removing stage (cleaning stage) is inserted before the bonding stage. The cleaning stage consists of a scratching motion (shift) toward to the ball bond in combination with ultrasound. Bonds are then made on the fresh diepad with the insulation removed from the contact surface of the insulated Au wire. This process increases the pull force of the crescent bond up to 26% which makes it comparable to the results obtained with bare Au wire. An online tail breaking force measurement method is developed with a proximity sensor between wire clamp and horn. Detailed understanding of tail bond formation is achieved by studying tail bond imprints with scanning electron microscopy and energy dispersive x-ray analysis. Descriptions are given of the dependence of the tail breaking force on the bonding parameters, metallization variation, and cleanliness of the bond pad. Simultaneous optimization with pull force and tail breaking force can optimize the Cu wire bonding process both with high quality and robustness. It is recommended to first carry out conventional pull force optimization followed by a minimization of the bonding force parameter to the lowest value still fulfilling the pull force cpk requirement. The tail bond forms not only under the capillary chamfer, but also under the capillary hole. The tail breaking force includes both the interfacial bond breaking strength and the breaking strength of the thinned portion of the wire that will remain at the substrate as residue. Close investigations of the tail bond imprint with scanning electron microscopy indicate the presence of fractures of the substrate indicating substrate material being picked up by Cu wire tail. Pick up is found on Au and Cu wires, but the amount of pick up is much larger on Cu wire. The effect on the hardness of the subsequently formed Cu free air ball (FAB) as investigated with scanning electron microscopy and micro - hardness test shows that Cu FABs containing Au and Ag pick ups are softer than those without pick up. However, the hardness varies significantly more with Au pick up. The amount of Au pick up is estimated higher than 0.03 % of the subsequently formed FAB volume, exceeding typical impurity and dopant concentrations (0.01 %) added during manufacturing of the wire.