Ion channels are membrane proteins that facilitate electrical signaling in important physiological processes, such as the rhythmic contraction of the heart. KCNQ1 is the pore-forming subunit of a voltage-gated potassium channel that assembles with the β-subunit KCNE1 in the heart to generate the IKs current, which is critical to cardiac action potential repolarization and electrical conduction in the heart. Mutations in IKs subunits can cause potentially lethal arrhythmia, including long QT syndrome, short QT syndrome, and atrial fibrillation. Each channel consists of four voltage-sensing domains and a central pore through which ions permeate. Voltage-dependent gating occurs when movement of voltage sensors cause pore opening/closing through coupling mechanisms. Although KCNQ1 by itself is able to form a voltage-dependent potassium channel, its assembly with KCNE1 is essential to generating the physiologically critical cardiac IKs current, characterized by a delay in the onset of activation, an increase in current amplitude, and a depolarizing shift in the current-voltage relationship. KCNE1 is thought to have multiple points of contact with KCNQ1 that reside within both the voltage-sensing domain and the pore domain, allowing for extensive modulation of channel function. Atrial fibrillation is the most common cardiac arrhythmia and affects more than 3 million adults in the United States. Much rarer, genetic forms of atrial fibrillation have been associated with gain-of-function mutations in KCNQ1, such as two adjacent mutations, S140G and V141M. Both mutations drastically slow channel deactivation, which underlies their pathophysiology. Deactivation slowing causes accumulation of open channels in the context of repeated stimulation, which abnormally increases the repolarizing K+ current, excessively shortens the action potential duration, and predisposes to re-entry arrhythmia such as atrial fibrillation. Although both mutations are located in the voltage-sensing domain, their mechanisms of action remain unknown. Understanding the gating mechanisms underlying deactivation slowing may provide key insights for the development of mechanism-based pharmacologic therapies for arrhythmias associated with KCNQ1 mutations. In addition to gain-of-function mutations, molecular activators of KCNQ1 can slow deactivation and increase channel activity. An existing problem in the pharmacologic treatment of arrhythmia is that many antiarrhythmic drugs do not have specific targets and cause undesired side effects such as additional arrhythmia. Thus, developing mechanism-based therapies may optimize clinical treatment for patients with specific forms of channel dysfunction. Two KCNQ1 activators, ML277 and R-L3, have been previously shown to slow current deactivation, but the underlying gating mechanisms remain known. Although these modulators are unlikely to serve directly as antiarrhythmic therapy, investigating their mechanisms will likely provide fundamental insights on channel modulation and guide future efforts to develop personalized therapies for arrhythmia, such as congenital long QT syndrome. Given the central importance of deactivation slowing in both pathophysiology and pharmacology, we focused on investigating gating mechanisms that underlie deactivation slowing. To this end, we utilized voltage clamp fluorometry, a technique that simultaneously assays for voltage sensor movement and ionic current through the channel pore. In Chapter 1, we begin our study by examining the gating mechanisms of KCNQ1 atrial fibrillation mutations in the absence of KCNE1. We show that S140G slows voltage sensor deactivation, which indirectly slows current deactivation. On the other hand, V141M neither slows voltage sensor nor current deactivation. This is followed by Chapter 2, where we examine the gating mechanisms underlying deactivation slowing by atrial fibrillation mutations in the presence of KCNE1. We show that both S140G and V141M slow IKs deactivation by slowing pore closing and altering voltage sensor-pore coupling. Based on these findings, we proposed a molecular mechanism in which both mutations disrupt the orientation of KCNE1 relative to KCNQ1 and thus impede pore closing, implying that future efforts to modulate KCNQ1 function can benefit from targeting the β-subunit. Finally, in Chapter 3, we explore the gating mechanisms underlying deactivation slowing for two small-molecule activators of KCNQ1. We show that ML277 predominantly slows pore transitions, whereas R-L3 slows voltage sensor deactivation, which indirectly slows current deactivation. Taken together, these studies guide future efforts to develop mechanism-based therapies for arrhythmia.

Submitted by FABRÍCIO ELY GOSSLER null (fabricio_ely8@hotmail.com) on 2017-02-09T16:24:59Z No. of bitstreams: 1 Fabrício E. Gossler-Dissertação - Unesp - Feis-PPGEE.pdf: 5023440 bytes, checksum: b5346eb35f509f2283b503acccf22ec3 (MD5) / Approved for entry into archive by LUIZA DE MENEZES ROMANETTO (luizamenezes@reitoria.unesp.br) on 2017-02-14T16:08:30Z (GMT) No. of bitstreams: 1 gossler_fe_me_ilha.pdf: 5023440 bytes, checksum: b5346eb35f509f2283b503acccf22ec3 (MD5) / Made available in DSpace on 2017-02-14T16:08:30Z (GMT). No. of bitstreams: 1 gossler_fe_me_ilha.pdf: 5023440 bytes, checksum: b5346eb35f509f2283b503acccf22ec3 (MD5) Previous issue date: 2016-12-19 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Existem diferentes tipos de funções wavelets que podem ser utilizadas na Transformada Wavelet. Na maioria das vezes, a função wavelet escolhida para a análise de um determinado sinal vai ser aquela que melhor se ajusta no domínio tempo-frequência do mesmo. Existem vários tipos de funções wavelets que podem ser escolhidas para certas aplicações, sendo que algumas destas pertencem a conjuntos específicos denominados de famílias wavelets, tais como a Haar, Daubechies, Symlets, Morlet, Meyer e Gaussianas. Nesse trabalho é apresentada uma nova família de funções wavelets geradas a partir de polinômios com coeficientes de Fibonacci (FCPs). Essa família recebe o nome de Golden, e cada membro desta é obtido por uma derivada de ordem n do quociente entre dois FCPs distintos. As Golden wavelets foram deduzidas através das observações de que, em alguns casos, a derivada de ordem n, do quociente entre dois FCPs distintos, resulta em uma função que possui as características de uma onda de duração curta. Como aplicação, algumas wavelets apresentadas no decorrer deste trabalho são utilizadas na classificação de arritmias cardíacas em sinais de eletrocardiograma, que foram extraídos da base de dados do MIT-BIH arrhythmia database. / There exist different types of wavelet functions that can be used in the Wavelet Transform. In most cases, the wavelet function chosen for the analysis of a given signal will be the one that best adjusts in the time-frequency domain of the same signal. There are many types of wavelet functions that can be chosen for certain applications, some of which belong to specific sets called wavelet families, such as Haar, Daubechies, Symlets, Morlet, Meyer, and Gaussians. In this work a new wavelet functions family generated from Fibonacci-coefficients polynomials (FCPs) is presented. This family is called Golden, and each member is obtained by the n-th derivative of the quotient between two distinct FCPs. The Golden wavelets were deduced from the observations that in some cases the n-th derivative of the quotient between two distinct FCPs results in a function that has the characteristics of a short-duration wave. As an application, some wavelets presented in the course of this work are used to cardiac arrhythmia classification in electrocardiogram signals, which were extracted from the MITBIH arrhythmia database. / CNPq: 130123/2015-3

Recent studies suggest that cardiomyocyte membrane microdomains, caveolae and transverse tubules, play a key role in cardiac arrhythmia. Mutation of caveolin-encoding genes CAV3, co-expressed with genes of caveolae ion channels, leads to a late persistent sodium currents and delayed repolarization stage, called LQT9 disease. A simplified three-current model is created to largely reduce the well-known Pandit rat ventricular myocyte model. The mathematical tractability of the three-current model allows us to conduct asymptotic analysis and efficiently estimate action potential duration. Improvement in the description of the mechanism for caveolae sodium current is incorporated into the three-current model utilizing a probability density approach for the four-state caveolae neck-channel coupling. The prolongation of action potentials and the formation of potential arrhythmia are shown to arise if caveolae neck open probability varies. A minimal model of the Ca2+ spatial distribution of CICR units illustrates the transverse tubule remodeling in failing myocyte causes dysfunction in the Ca2+ profile. With regards to discrimination of protein localization, which is widely used in biological experiments, the bagging pruned decision tree algorithm is tested to be one of the algorithms with best performance on the large data set, and it succeeds in extracting information to be highly predictive on test data. Parallel computation technique is applied to accelerate the speed of implementation in K-nearest neighbor learning algorithms on big data sets.

People living with heart failure (PLHF) should be screened for symptoms at every healthcare visit since they are 3 times more likely to experience ventricular arrhythmias. This quality improvement project (QIP) compared 3 self-administered HF symptoms questionnaires to determine the best screening tool for a tertiary hospital arrhythmia devices clinic. The instruments included the Minnesota Living with Heart Failure Questionnaire (MLHFQ), the Kansas City Cardiomyopathy Questionnaire (KCCQ), and the Self-Reported Heart Failure Symptoms (SHEFS) questionnaire. For a 30-day period, 76 people were eligible to participate in the QIP, with 55 participants included in the final analysis (72.5% participation). The questionnaires were compared and assessed with the gold standard laboratory test for HF (NT-proBNP) for sensitivity and specificity. For HF, the SHEFS was the most sensitive (83%) compared to the NT-proBNP, but the MLHFQ was most specific (89%). When compared to the MLHFQ as the standard, SHEFS was 71% sensitive, and 73% specific for HF. Similarly, when compared to the KCCQ, the SHEFS was both, 75% specific and sensitive in identifying HF. However, the rate of correlation to a positive or negative NT-proBNP test results was the highest for the SHEFS (87%). All 3 questionnaires were statistically significant in predicting admission to hospital for HF in the past 6 months (p = 0.02 to 0.03). Finally, given the shortest length and simplicity of use, the SHEFS was selected by the stakeholders to be the standard screening tool for the clinic. This project contributes to positive social change by providing the first reported comparison in the literature to implement questionnaires in a clinic to assess symptoms for PLHF attending an arrhythmia devices clinic.

<p>During the last years, drugs that prolong the repolarisation phase of the myocardial action potential, due to inhibition of the rapid component of the delayed-rectifying potassium channel (I_Kr) have been in focus. In addition to arrhythmogenic potential, selective Ikr-blockers have also been shown to be embryotoxic and teratogenic in animal studies. The aim of this thesis was to investigate a theory that these developmental toxic results from pharmacologically induced episodes of embryonic cardiac arrhythmias leading to hypoxia related damage in the embryo. Almokalant (ALM) was used as a model compound for selective Ikr-blockers. ALM induced embryonic cardiac arrhythmia, and in similarity with results obtained by maternal hypoxia, ALM induced embryonic death and growth retardation in both rats, and mice. </p><p>The theory of a hypoxia-related mechanism was strengthened by the results that ALM induce phase specific external and visceral defects (e.g. cleft lip/palate, distal digital, cardiovascular, and urogenital defects), and that the skeletal defects (not shown before) showed a clear trend; the later the treatment the more caudal was the site of the defect, which is in accordance with results from maternal hypoxia induced by e.g. lowering of the O₂ content in the air. The spin trapping agent PBN decreased almokalant induced malformations, suggesting that the defects mainly are caused by reoxygenation damage after episodes of severe embryonic dysrhythmia, rather than "pure hypoxia".</p><p>Sotalol was tested in a third species, the rabbit who expresses functional I_Kr channels both in the embryo and in the adult, where it induced developmental toxicity, and indicating that the embryo is more sensitive than the adult towards arrhythmia caused by I_Kr-blockers. </p>

<p>The Antiepilptic drugs (AEDs) phenytoin (PHT), carbamazepine (CBZ), phenobarbital (PB), tri- and dimethadione (TMD and DMD) are known teratogens having a common malformation pattern in human and animal studies. This thesis was designed chiefly to test a hypothesis correlating the teratogenicity of these AEDs to episodes of pharmacologically induced embryonic arrhythmia and hypoxia-reoxygenation damage.</p><p>Effects on the embryonic heart were studied both after maternal administration in mice and in</p><p>mouse embryos cultured in vitro. Only AEDs, correlated with the same type of malformation as could be induced by episodes of interrupted oxygen supply to the embryo (e.g. cleft palate) caused concentration dependent bradycardia and arrhythmia. PHT and DMD had the highest potential and affected embryonic heart at clinically relevant concentration, followed by CBZ, TMD and PB. Valproate and vigabatrin not associated with hypoxia-related malformations caused neither arrhythmia nor severe bradycardia.</p><p>The results showed that the embryonic heart is extremely susceptible to PHT and DMD only</p><p>during a restricted period of development, between gestational days 9-13 (weeks 5-9 of human pregnancy).An observed genetic susceptibility to react with arrhythmia at low concentrations when exposed to PHT or to external stress, could explain why A/J strain of mice is more susceptible to develop cleft palate compared to other strains. High activities of reactive oxygen species (ROS) capturing antioxidant enzymes observed in untreated A/J embryos supported this assumption. The potential to cause embryonic arrythmia by an AED was related to the potential to inhibit the rapid component of the delayed rectifier potassium channel (I _kr ).A marked I _kr blocking activity (70%)of DMD in voltage clamping studies was observed. The I _kr inhibition occurred at similar concentrations, which causes severe arrhythmia.</p><p>The idea of a relation between teratogenicity and arrhythmia, resulting in ischemia followed by reperfusion and generation of ROS was supported by mechanistic studies. Pre-treatment with the spin-trapping agent PBN, which has the capacity to capture ROS, markedly reduced the incidence of PHT and DMD-induced cleft palate. In utero exposure to teratogenic doses of DMD and PHT resulted in hemorrhages in the embryonic palatal region. The same type of haemorrhage in the palatal region precedes orofacial clefts induced by episodic hypoxia.</p>

During the last years, drugs that prolong the repolarisation phase of the myocardial action potential, due to inhibition of the rapid component of the delayed-rectifying potassium channel (IKr) have been in focus. In addition to arrhythmogenic potential, selective Ikr-blockers have also been shown to be embryotoxic and teratogenic in animal studies. The aim of this thesis was to investigate a theory that these developmental toxic results from pharmacologically induced episodes of embryonic cardiac arrhythmias leading to hypoxia related damage in the embryo. Almokalant (ALM) was used as a model compound for selective Ikr-blockers. ALM induced embryonic cardiac arrhythmia, and in similarity with results obtained by maternal hypoxia, ALM induced embryonic death and growth retardation in both rats, and mice. The theory of a hypoxia-related mechanism was strengthened by the results that ALM induce phase specific external and visceral defects (e.g. cleft lip/palate, distal digital, cardiovascular, and urogenital defects), and that the skeletal defects (not shown before) showed a clear trend; the later the treatment the more caudal was the site of the defect, which is in accordance with results from maternal hypoxia induced by e.g. lowering of the O2 content in the air. The spin trapping agent PBN decreased almokalant induced malformations, suggesting that the defects mainly are caused by reoxygenation damage after episodes of severe embryonic dysrhythmia, rather than "pure hypoxia". Sotalol was tested in a third species, the rabbit who expresses functional IKr channels both in the embryo and in the adult, where it induced developmental toxicity, and indicating that the embryo is more sensitive than the adult towards arrhythmia caused by IKr-blockers.

The Antiepilptic drugs (AEDs) phenytoin (PHT), carbamazepine (CBZ), phenobarbital (PB), tri- and dimethadione (TMD and DMD) are known teratogens having a common malformation pattern in human and animal studies. This thesis was designed chiefly to test a hypothesis correlating the teratogenicity of these AEDs to episodes of pharmacologically induced embryonic arrhythmia and hypoxia-reoxygenation damage. Effects on the embryonic heart were studied both after maternal administration in mice and in mouse embryos cultured in vitro. Only AEDs, correlated with the same type of malformation as could be induced by episodes of interrupted oxygen supply to the embryo (e.g. cleft palate) caused concentration dependent bradycardia and arrhythmia. PHT and DMD had the highest potential and affected embryonic heart at clinically relevant concentration, followed by CBZ, TMD and PB. Valproate and vigabatrin not associated with hypoxia-related malformations caused neither arrhythmia nor severe bradycardia. The results showed that the embryonic heart is extremely susceptible to PHT and DMD only during a restricted period of development, between gestational days 9-13 (weeks 5-9 of human pregnancy).An observed genetic susceptibility to react with arrhythmia at low concentrations when exposed to PHT or to external stress, could explain why A/J strain of mice is more susceptible to develop cleft palate compared to other strains. High activities of reactive oxygen species (ROS) capturing antioxidant enzymes observed in untreated A/J embryos supported this assumption. The potential to cause embryonic arrythmia by an AED was related to the potential to inhibit the rapid component of the delayed rectifier potassium channel (I kr ).A marked I kr blocking activity (70%)of DMD in voltage clamping studies was observed. The I kr inhibition occurred at similar concentrations, which causes severe arrhythmia. The idea of a relation between teratogenicity and arrhythmia, resulting in ischemia followed by reperfusion and generation of ROS was supported by mechanistic studies. Pre-treatment with the spin-trapping agent PBN, which has the capacity to capture ROS, markedly reduced the incidence of PHT and DMD-induced cleft palate. In utero exposure to teratogenic doses of DMD and PHT resulted in hemorrhages in the embryonic palatal region. The same type of haemorrhage in the palatal region precedes orofacial clefts induced by episodic hypoxia.

The evidence that treatment of life-threatening arrhythmia (LTA) with an Implantable Cardioverter Defibrillator (ICD) can prolong life is convincing. Living with a lifelong heart disease will gradually influence the everyday life and encompasses some or all aspects of life. In order to influence health outcomes, the impact of the ICD must be considered in a broader context including not only the physical, but also the psychological and social functioning of the individual. The general aim of this thesis was to describe everyday life in recipients living with an ICD in a longterm perspective. The aim in Paper I was to describe changes in the life situation of recipients’ with an ICD over a period of 1 year. The aim in Paper II was to describe quality of life (QOL) and uncertainty in recipients who have an ICD and to predict QOL at long-term follow-up. Fifty-six recipients participated (I) and 35 of these recipients, who had survived at least five years, were further included (II). The Quality of Life Index-Cardiac version (I, II), Mishel Uncertainty in Illness Scale-Community version (I, II), Patient ICD Questionnaire (I) and multiple regression analysis (II) were used. Higher scores indicate higher QOL and uncertainty. The questionnaires were completed before implantation, three and twelve months after implantation (I) and also five years after implantation i.e. long-term follow up (II). At the long-term follow up, the average ICD recipient had lived with an ICD for six years and nine months (6.9 years). The results showed the overall QOL and QOL in the health/functioning domain were unchanged over time. QOL in the socio-economic (p= .002) and psychological/spiritual domains (p= .012) decreased in the first year. From baseline to long-term follow up, the QOL in the family domain (p= .011) and overall uncertainty (p= .002) decreased. Uncertainty related to the information decreased at year 1 in relation to baseline (p= .001). The aim in Paper III was to illuminate the main concern of recipients living with an ICD and how they handle this in their daily life. Sixteen recipients who had lived with an ICD between six to twenty-four months were interviewed. Data was collected and analysed in a simultaneous process according to guidelines for classical grounded theory. In the analysis, a substantive theory was generated explaining the main concern of ICD recipients and how they handle this in their daily life. The core category, labelled “Striving to resume command”, illuminates the main concern of ICD recipients. To manage this main concern, the recipients used the following strategies: Economizing resources, Distracting oneself, Submitting to one’s fate and Re-evaluating life. The aim in Paper IV was to explore relationships between OQL, coping strategies, anxiety, depression and perceived control in recipients living with an ICD and to compare those having received an ICD less or more than one year ago and those with a primary or secondary preventive indication. A cross-sectional, correlational, multicenter design was used, and 147 recipients who had lived with an ICD between six to twenty-four months completed Quality of Life Index-Cardiac version, Jalowiec Coping Scale, Hospital Anxiety and Depression Scale and Control Attitude Scale. The results showed that anxiety, depression and perceived control were predictors of QOL. Anxiety was also a predictor of coping with optimistic coping being the most used coping strategy. There was no relationship between QOL and coping. No differences were found in QOL, coping, anxiety, depression and perceived control between recipients implanted either on a primary or secondary preventive indication or having the device less or more than one year. In this thesis, it was concluded that the ICD recipients strived to resume command over their life (III) and the more control the recipients perceived the more satisfied they were with their QOL (IV) and the more symptoms of anxiety, depression and uncertainty they experienced the less satisfied they were with their QOL (II, IV). Coping strategies were used more frequently by ICD recipient perceiving more anxiety (IV). QOL was fairly good 6,9 years after implantation and ICD recipients felt less uncertain once they had passed the first year of their illness.

The evidence that treatment of life-threatening arrhythmia (LTA) with an Implantable Cardioverter Defibrillator (ICD) can prolong life is convincing. Living with a lifelong heart disease will gradually influence the everyday life and encompasses some or all aspects of life. In order to influence health outcomes, the impact of the ICD must be considered in a broader context including not only the physical, but also the psychological and social functioning of the individual. The general aim of this thesis was to describe everyday life in recipients living with an ICD in a longterm perspective. The aim in Paper I was to describe changes in the life situation of recipients’ with an ICD over a period of 1 year. The aim in Paper II was to describe quality of life (QOL) and uncertainty in recipients who have an ICD and to predict QOL at long-term follow-up. Fifty-six recipients participated (I) and 35 of these recipients, who had survived at least five years, were further included (II). The Quality of Life Index-Cardiac version (I, II), Mishel Uncertainty in Illness Scale-Community version (I, II), Patient ICD Questionnaire (I) and multiple regression analysis (II) were used. Higher scores indicate higher QOL and uncertainty. The questionnaires were completed before implantation, three and twelve months after implantation (I) and also five years after implantation i.e. long-term follow up (II). At the long-term follow up, the average ICD recipient had lived with an ICD for six years and nine months (6.9 years). The results showed the overall QOL and QOL in the health/functioning domain were unchanged over time. QOL in the socio-economic (p= .002) and psychological/spiritual domains (p= .012) decreased in the first year. From baseline to long-term follow up, the QOL in the family domain (p= .011) and overall uncertainty (p= .002) decreased. Uncertainty related to the information decreased at year 1 in relation to baseline (p= .001). The aim in Paper III was to illuminate the main concern of recipients living with an ICD and how they handle this in their daily life. Sixteen recipients who had lived with an ICD between six to twenty-four months were interviewed. Data was collected and analysed in a simultaneous process according to guidelines for classical grounded theory. In the analysis, a substantive theory was generated explaining the main concern of ICD recipients and how they handle this in their daily life. The core category, labelled “Striving to resume command”, illuminates the main concern of ICD recipients. To manage this main concern, the recipients used the following strategies: Economizing resources, Distracting oneself, Submitting to one’s fate and Re-evaluating life. The aim in Paper IV was to explore relationships between OQL, coping strategies, anxiety, depression and perceived control in recipients living with an ICD and to compare those having received an ICD less or more than one year ago and those with a primary or secondary preventive indication. A cross-sectional, correlational, multicenter design was used, and 147 recipients who had lived with an ICD between six to twenty-four months completed Quality of Life Index-Cardiac version, Jalowiec Coping Scale, Hospital Anxiety and Depression Scale and Control Attitude Scale. The results showed that anxiety, depression and perceived control were predictors of QOL. Anxiety was also a predictor of coping with optimistic coping being the most used coping strategy. There was no relationship between QOL and coping. No differences were found in QOL, coping, anxiety, depression and perceived control between recipients implanted either on a primary or secondary preventive indication or having the device less or more than one year. In this thesis, it was concluded that the ICD recipients strived to resume command over their life (III) and the more control the recipients perceived the more satisfied they were with their QOL (IV) and the more symptoms of anxiety, depression and uncertainty they experienced the less satisfied they were with their QOL (II, IV). Coping strategies were used more frequently by ICD recipient perceiving more anxiety (IV). QOL was fairly good 6,9 years after implantation and ICD recipients felt less uncertain once they had passed the first year of their illness.