published_or_final_version / Medicine / Master / Master of Research in Medicine

My PhD project investigated the regulation of human cardiac transient outward potassium current (Ito) by protein tyrosine kinases (PTKs) and the functional expression of transient receptor potential (TRP) channels in human atrial myocytes to make an advanced understanding of human cardiac electrophysiology and pathophysiology. The modulation of human cardiac Itoby PTKs was studied in human atrial myocytes and HEK 293 cells expressing hKv4.3 (coding human cardiac Ito). We found that the broad-spectrum PTK inhibitor genistein, the selective EGFR kinase inhibitor AG556, and the Src-family kinases inhibitor PP2 inhibited human atrial Itoand the inhibitory effect was countered by the protein tyrosine phosphatase (PTP) inhibitor orthovanadate. Similar results were observed in hKv4.3-HEK cells. Interestingly, tyrosine phosphorylation of hKv4.3channels was reduced by genistein, AG556, and PP2,and the reduction was antagonized by orthovanadate. The mutant Y136F of hKv4.3 lost the inhibitory response to AG556, whileY108F lost the response to PP2.The double mutant Y108F-Y136F hKv4.3 failed to respond to both AG556 and PP2, and exhibited a dramatic reduction of tyrosine phosphorylation. These results indicate that native cardiac Itois regulated by both EGFR and Src family kinases. In the second part, we studied whether TRPC channels would mediate the nonselective cation current described previously in human atrial myocytes. It was found that TRPC1 channel activator thapsigargin activated the current, and the effect was suppressed by La3+or prevented by intracellular anti-TRPC1 antibody. Endothelin-1 and angiotensin II stimulated the current, andthe effect was inhibited by La3+and/or 2-APB. RT-PCR and Western blot analysis revealed that in addition to the TRPC1 channels mediating the nonselective cation current, the components of store-operated Ca2+channels (SOCs), STIM1 and Orai1 were abundantly expressed in human atria. The interaction of TRPC1, STIM1, and Orai1 was confirmed by co-immunoprecipitation. Interestingly, we found that protein expression of TRPC1 and STIM1, but not Orai1, was up-regulated in human atria with atrial fibrillation. The third part of the project determined whether TRPM7 channels were expressed in human atrial myocytes, since this channel was reported in human atrial fibroblasts, conferring atrial fibrosis in human atria with atrial fibrillation. We found a TRPM7 -like current which was potentiated by acidic pH, and inhibited by La3+and 2-APB, and a Ca2+-activated TRPM4 current. RT-PCR and Western blot analysis confirmed the expression of TRPM7 and TRPM4 channels in human atria. Moreover, we found TRPM7 protein, but not TRPM4 protein was significantly up-regulated in human atria with atrial fibrillation, suggesting the potential participation of TRPM7 channels in atrial remodeling of human atria with atrial fibrillation. Collectively, this PhD thesis project has demonstrated for the first time that human cardiac Itois modulated by EGFR kinase and Src kinases via phosphorylating Y136and Y108, respectively. TRPC1 channels mediate the nonselective cation current and SOCs.TRPM7 channels are expressed in human atrial myocytes. The up-regulation of TRPC1, STIM1, and TRPM7 channels in human atria with atrial fibrillation suggest that they are likely involved in atrial electrical and/or structure remodeling in patients with atrial fibrillation. / published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

This thesis is an investigation into the possible genetic background underlying the liability of Standardbred racehorses (Std) to atrial fibrillation (AF). Pedigrees of 204 affected Std admitted to the Ontario Veterinary College Teaching Hospital (OVCTH) for treatment of AF born from 1978 to 2007 and of 1,017 randomly selected racing contemporaries (5 for each Std admitted). Estimates of heritability of AF were obtained using a linear threshold animal model. Marginal genetic contributions of ancestors to affected and control cohorts were determined and differences were tested. The estimate of heritability of AF in the Std was 14% on the observed scale. Eleven ancestors had significantly higher contributions to affected cohorts than to controls. Many of these ancestors appeared multiple times in the five-generation pedigrees of affected horses. Results strongly indicate a genetic predisposition to AF in the Std, with the arrhythmia particularly prevalent in one popular sire line.

Background: Atrial fibrillation (AF) is the most common arrhythmia in the adult population, affecting about 5% of the population over 65 years. Occurrence of AF is an independent risk factor for stroke, and together with other cardiovascular risk factors (CHADS2/CHA2DS2- VASc), the stroke risk increases. Since AF is often paroxysmal and asymptomatic (silent) it may remain undiagnosed for a long time and many AF patients are not discovered before suffering a stroke. Aims: To estimate the prevalence of previously undiagnosed AF in an out-of-hospital population with CHADS2 ≥1, in patients with an enlarged left atrium (LA) and of total AF prevalence in sleep apnea (SA) patients, conditions that have been associated with AF. To compare the efficacy of short intermittent ECG with continuous 24h Holter ECG in detecting arrhythmias. Methods: Patients without known AF recorded 10−30 second handheld ECG (Zenicor-EKG®) registrations during 14−28 days at home, both regular, asymptomatic registrations twice daily and when having cardiac symptoms. Recordings were transmitted through the in-built SIM card to an internet-based database. Patients with palpitations or dizziness/presyncope referred for 24h Holter ECG were asked to additionally record 30-second handheld ECG registrations during 28 days at home. Results: In the out-of-hospital population with increased stroke risk, previously unknown AF was diagnosed in 3.8% of 928 patients. Comparing AF detection in patients with an enlarged LA versus normal LA showed that eleven of 299 patients had AF. Five of these had an enlarged LA (volume/BSA). No statistical difference in AF prevalence was found between patients with enlarged and normal LA, 3.3% and 3.2% respectively, (p = 0.974). AF occurred in 7.6% of 170 patients with sleep apnea, in 15% of patients with sleep apnea ≥60 years, and in 35% of patients with central sleep apnea. AF prevalence was also associated with severity of sleep apnea, male gender and diabetes. Comparing the efficacy of arrhythmia detection in 95 patients with palpitations or dizziness/presyncope with continuous 24h Holter and short intermittent ECG, 24h Holter found AF in two and AV-block II in one patient, resulting in 3.2% relevant arrhythmias detected. Short intermittent ECG diagnosed nine patients with AF, three with PSVT and one with AV-block II, in total 13.7% relevant arrhythmias. (p = 0.0094). Conclusions: Screening in the out-of-hospital patient population (mean age 69.8 years) yielded almost 4% AF, making it seem worthwhile to screen older patients with increased stroke risk for AF with this method. Screening patients with LA enlargement (mean age 73.1 years) did not result in higher detection rates compared with the general out-of-hospital population. AF occurred in 7.6% of patients with sleep apnea, (mean age 57.6 years) and was associated with severity of sleep apnea, presence of central sleep apnea, male gender, age ≥60 years, and diabetes. Short intermittent ECG is more effective in detecting relevant arrhythmias than 24h Holter ECG in patients with palpitations or dizziness/presyncope.

The costs for stroke management and reduced health related quality of life (QoL) can extend throughout life as mental and physical disabilities are common. The aim of this thesis was to quantify this stroke-related burden with data from Riks-Stroke (RS), the Swedish stroke register. Costs for hospital and primary care, secondary drug prevention, home and residential care services, and production losses were estimated for first-ever stroke patients registered in the RS. The present value lifetime costs were estimated from the expected survival and discounted by 3%. Quality of life was estimated with the EQ-5D instrument on a subset of patients at 3 months after the index event and mapped to patient-reported outcome measures in the RS. Standard descriptive and analytic (multivariate regressions) statistical methods were used. The life-time societal present value cost per patient in 2009 was approximately €69,000 whereof home and residential care due to stroke was 59% and indirect costs for productivity losses accounted for 21% (year 2009 prices). Women had higher costs than men in all age groups. Treatment at stroke units had a low incremental cost per life-year gained compared to patients who were not treated at such facilities. The estimated disutility from stroke was greatest for women and the oldest, and compared to 1997 the cost per patient increased after a revised assumption. Hospitalisation costs were stable while long-term costs for ADL support increased in part due to a changed age structure. Patients with atrial fibrillation (AF; 24%) had €367 higher inpatient costs compared to non-AF stroke patients €8,914 (P<0.01; year 2001 prices). As the index case fatality was higher among AF patients, the cost difference was higher for patients surviving the first 28 days. A multivariate regression showed that AF, diabetes, stroke severity, and death during the 3-year follow-up period were independent cost drivers. Three regression techniques (OLS, Tobit, CLAD) were chosen for mapping EQ-5D utilities to patient-reported outcome measures in the RS. The mean utility was overestimated with all models and had lower variance than the original data. In conclusion, total societal lifetime cost for 22,000 first-ever stroke patients in 2009 amounted to €1.5 billion (whereof production losses were €314 million). About 56,600 QALYs were lost due to premature death and disability. Including a preference-based QoL instrument in the RS would allow cost-utility analyses, but it is important to control for confounders in comparator arms to avoid bias.

This project set out to derive a prediction rule based on preoperative clinical variables to identify patients with high risk of developing atrial fibrillation following cardiac surgery. Methods: Prospectively collected data from a perioperative database was corroborated with chart review to identify eligible patients who had non-emergent surgery in 2010. Details on 28 preoperative variables were collected and significant predictors (p<0.2) were inserted into multivariable logistic regression and recursive partitioning. Results: 305 (30.5%) of 999 patients developed new onset postoperative atrial fibrillation. Eleven variables were significantly associated with atrial fibrillation, however, both final models included only three: left atrial dilatation, mitral valve disease and age. Bootstrapping with 5000 samples confirmed that both final models provide consistent predictions. Coefficients from the logistic regression model were converted into a simple seven point predictive score. Conclusions: This simple risk score can identify patients at higher risk of developing atrial fibrillation after cardiac surgery.

No description available.

Atrial fibrillation (AF) is a complex, multifaceted arrhythmia. Pathogenesis of AF is associated with multiple aetiologies and the mechanisms by which it is sustained and perpetuated are similarly diverse. In particular, regional heterogeneity in the electrophysiological properties of normal and pathological tissue plays a critical role in the occurrence of AF. Understanding AF in the context of electrophysiological heterogeneity requires cell-specific ionic models of electrical activity which can then be incorporated into models on larger temporal and spatial scales. Biophysically-based models have typically dominated the study of cellular excitability providing detailed and precise descriptions in the form of complex mathematical formulations. However, such models have limited applicability in multidimensional simulations as the computational expense is too prohibitive. Simplified mathematical models of cardiac cell electrical activity are an alternative approach to these traditional biophysically-detailed models. Utilizing this approach enables the embodiment of cellular excitation characteristics at minimal computational cost such that simulations of arrhythmogensis in atrial tissue are conceivable. In this thesis, a simplified, generic mathematical model is proposed that characterizes and reproduces the action potential waveforms of individual cardiac myocytes. It incorporates three time-dependent ionic currents and an additional time-independent leakage current. The formulation of the three time-dependent ionic currents is based on 4-state Markov schemes with state transition rates expressed as nonlinear sigmoidal functions of the membrane potential. Parameters of the generic model were optimized to fit the action potential waveforms of the Beeler-Reuter model, and, experimental recordings from atrial and sinoatrial cells of rabbits. A nonlinear least-squares optimization routine was employed for the parameter fits. The model was successfully fitted to the Beeler-Reuter waveform (RMS error: 1.4999 mV) and action potentials recorded from atrial tissue (RMS error: 1.3398 mV) and cells of the peripheral (RMS error: 2.4821 mV) and central (RMS error: 2.3126 mV) sinoatrial node. Thus, the model presented here is a mathematical framework by which a wide variety of cell-specific AP morphologies can be reproduced. Such a model offers the potential for insights into possible mechanisms that contribute to heterogeneity and/or arrhythmia.

Diss. (sammanfattning) Stockholm : Karol. inst., 2002. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 5 uppsatser.