Background: Once aortic stenosis (AS) is severe, patients develop symptoms at different stages. Indeed, symptom status may correlate poorly with the grade of valve narrowing. Multiple pathophysiological mechanisms, other than valvular load, may explain the link between AS and symptom severity. We aimed to describe the relationship between the severity of symptoms and the characteristics of a cohort of patients with severe AS already referred for aortic valve replacement (AVR). Methods: We analyzed 118 consecutive patients (70 ± 9 years, 55% men) with severe AS referred for AVR. We identified 84 patients with New York Heart Association (NYHA) I–II, and 34 with NYHA III–IV symptoms. Clinical and echocardiographic parameters were compared between these two groups. Left ventricular ejection fraction (LVEF), global longitudinal peak systolic strain (GLPS), NT-pro-B-type natriuretic peptide (BNP), and high-sensitive troponin T (hs-TNT) were determined at the time of admission. Results: AS severity was similar between groups. Compared with the NYHA I–II group, patients in NYHA III–IV group were older and more likely to have comorbidities, worse intracardiac hemodynamics and more LV damage. Variables independently associated with NYHA III–IV symptomatology were the absence of sinus rhythm, higher E/e0 ratio, and increased hs-TNT. GLPS showed a good correlation not only with hs-TNT as a marker of myocardial damage, but also with markers of increased afterload imposed on LV, being not directly related with advanced symptoms. Conclusions: Advanced symptoms in patients with severe AS referred for AVR are associated with worse intracardiac hemodynamics, absence of sinus rhythm, and more myocardial damage. It supports the concept of transition from adaptive LV remodeling to myocyte death as an important determinant of symptoms of heart failure. :Einführung................................................................................................3 Publikationsmanuskript...........................................................................12 Zusammenfassung..................................................................................19 Literaturverzeichnis..................................................................................24 Erklärung über die eigenständige Abfassung der Arbeit.........................28 Darstellung des eigenen Beitrags............................................................29 Curriculum vitae.......................................................................................30 Danksagung.............................................................................................36

Mechanistic details on the regulation of striated muscle contraction still need to be determined, particularly the specific structural locations of the elements comprising the thick and thin filaments. Of special interest is the location of the regulatory component, troponin, on the actin filament and how its presence influences the behavior of myosin binding to the thin filament. In the present study: (1) Luminescence resonance energy transfer was used to monitor potential conformational changes in the reconstituted thin filament between the C-terminal region of troponin T and myosin subfragment 1; (2) Location of troponin in previously derived atomic models of the acto-myosin complex was mapped to visualize specific contacts; and (3) Shortened tropomyosin was engineered and protein binding and ATPase assays were performed to study the effect of myosin binding close to the troponin complex. Analysis of the results suggest the following: (1) Irrespective of calcium levels, the C-terminal region of troponin T is located close to myosin loop 3 and a few actin helices that may perturb strong acto-myosin interactions responsible for force production. (2) Atomic models indicate myosin subfragment 1 cannot attain the post- powerstroke state due to the full motion of the lever arm being sterically hindered by troponin. (3) A shortened tropomyosin with five actin binding modules (instead of the native seven in muscle cells) binds actin contiguously in a head-to-tail manner and serves to increase the periodicity of troponin complexes on the actin filament. Such behavior eliminates the structure of the actin filament being responsible for the binding location of tropomyosin. (4) Differential behavior of myosin subfragment 1 i.e. (a) binding adjacent to troponin and (b) binding further away from troponin, is apparent as tropomyosin and troponin appear to govern the regions or "target zones" where myosin can bind productively along the actin filament. Physiologically, myosins able to bind close to troponin, but not participate in force production may function as mechanical sensors to attenuate or dampen the force generated from the so-called "target zones". Therefore, this could be a pseudo-regulatory mechanism that functions to protect the contractile apparatus from damage.

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<p>Worldwide, more than 2 million patients die within 30 days after noncardiac surgery anually. Postoperative ischemic myocardial injury is frequent, however, no consensus exists about its definition.</p> <p><strong>Objective: </strong>to develop a term Myocardial Injury after Noncardiac Surgery (MINS) caused by myocardial ischemia, requiring at least, troponin T (TnT) elevation, and with prognostic relevance at 30 days after surgery.</p> <p><strong>Methods: </strong>we performed a prospective study including 15,167 patients ³45 years-old undergoing noncardiac surgery, who had fourth-generation TnT measurements during the first 3 postoperative days. We undertook Cox regression analyses with 30-day mortality after surgery as the dependent variable, using different TnT thresholds, clinical features and several perioperative variables. Non-ischemic etiologies were excluded. Furthermore, we developed a scoring system to predict risk in MINS patients.</p> <p><strong>Results:</strong> MINS was defined as TnT ≥0.03 ng/mL with or without clinical features, and it was an independent predictor of 30-day mortality (adjusted HR 3.82, CI 95% 2.84-5.10). We determined that MINS incidence was 8%, its population attributable risk 33.7%, and 30-days mortality rate 9.6%. Patients did not experience ischemic symptoms in 84% of MINS cases. Additionally, we developed a scoring system in patients suffering MINS with 3 independent predictors of death (age ≥75 years, new ST elevation or left bundle branch block, and anterior location of ECG changes),</p> <p><strong>Conclusion: </strong>Among patients undergoing noncardiac surgery, we defined MINS based on a TnT threshold ≥0.03 ng/mL. Mostly, MINS patients were asymptomatic. Therefore, this strongly suggests the importance of a troponin monitoring during the first few days after surgery.</p> / Master of Health Sciences (MSc)

<p>Reduced kidney function around the time of surgery is an important risk factor for postoperative mortality. Despite this there is limited information on how reduced kidney function prior to surgery alters prognosis, what causes sudden decrements in kidney function after surgery (known as acute kidney injury), or how they might be avoided. The studies in this thesis inform these knowledge gaps. Chapter 2 describes the results of a post hoc analysis of the interaction between preoperative estimated glomerular filtration rate, a marker of kidney function, and postoperative cardiac troponin T, a marker of heart damage, for predicting 30-day mortality in a prospective cohort study of patients undergoing noncardiac surgery. Chapter 3 uses administrative and clinical data from a single centre to inform the risk of acute kidney injury after noncardiac surgery by concentrations of preoperative hemoglobin and change in postoperative hemoglobin. Chapter 4 uses the same data to determine a definition of intraoperative hypotension that is prognostic of acute kidney injury, myocardial injury and death. Chapter 5 describes a randomized controlled trial that compares a novel therapeutic procedure called remote ischemic preconditioning to a sham procedure in patients undergoing cardiac surgery.</p> / Doctor of Philosophy (PhD)

(1) Background: Highly sensitive cardiac troponin T (hs-cTnT) plays an essential role in the diagnosis of myocardial injury. The upper reference limit of the respective assay is generally applied, irrespective of age, renal function, or sex. We aimed to identify age-adjusted and sex-adjusted upper reference limits in relation to renal function in a large population-based cohort without cardiac diseases. (2) Methods: We included 5428 subjects of the population-based LIFE-Adult cohort, free of diagnosed cardiac diseases. Sex-adjusted and age-adjusted 99th percentiles for hs-cTnT in subjects with preserved renal function were obtained. (3) Results: The hs-cTnT values were higher in men of all age groups. In both sexes, an increasing age positively correlated with higher hs-cTnT values. Hs-cTnT weakly correlated with serum creatinine. The three-dimensional analysis of age, creatinine, and hs-cTnT showed no relevant additional effect of creatinine on hs-cTnT. In men aged above 60 and women above 70, the calculated 99th percentiles clearly exceeded the commonly applied thresholds. (4) Conclusion: Age and sex have a major impact on the serum concentration of hs-cTnT, while renal function does not. We propose to consider age-adjusted and sex-adjusted reference values.

<p>Metadon je sintetski agonist opijatnih receptora koji se primenjuje u sklopu supstitucione terapije opijatnih zavisnika metadonom (STM) i u terapiji hroničnog bola. Dugoročna primena STM je praćena blagim, uglavnom prolaznim, neželjenim delovanjima. Međutim, metadon pripada grupi lekova koji mogu da prouzrokuju prolongaciju korigovanog QT intervala (QTc) u elektrokardiogramu (EKG-u) i povećaju rizik za nastanak potencijalno fatalnih aritmija tipa torsades de pointes. Opijatni zavisnici metadon najče&scaron;će koriste u kombinaciji sa benzodiazepinima, i ova kombinacija lekova predstavlja faktor rizika za nastanak smrtnog ishoda. Iako je najveći broj lekara upoznat sa rizikom za razvoj respiratorne depresije prilikom primene opijata u kombinacji sa benzodiazepinima, velika studija otkriva da su ventrikularne aritmije i srčani zastoj najče&scaron;će prijavljivana neželjena delovanja metadona, primenjenog u kombinaciji sa benzodiazepinima. Ciljevi ovoga radu su da se analizom smrtnih slučajeva povezanih sa upotrebom metadona (MRDs) tokom desetogodi&scaron;njeg perioda na teritoriji Vojvodine i sprovođenjem kliničkog ispitivanja kod opijatnih zavisnika na STM proceni kardiolo&scaron;ka bezbednost primene metadona, posebno u kombinaciji sa benzodiazepinima. Sprovedena je retrospektivna studija za određivanje karakteristika MRDs na teritoriji Vojvodine, kao i kliničko ispitivanje u kome su učestvovali opijatni zavisnici koji počinju sa STM. Snimanje EKG-a (za izračunavanje QTc intervala) i uzorkovanje krvi (za određivanje koncentracije metadona i diazepama i vrednosti troponina) je sprovedeno kod svih učesnika istraživanja u 5 vremenskih tačaka (pre početka primene STM, 8. i 15. dana i nakon 1. i 6. meseca primene STM). Koncentracije metadona i diazepama u serumu su određivane metodom tečne hromatografije sa masenom spektrometrijom (LC-MS). U Vojvodini je zapažena rastuća tendencija MRDs, ali ni jedan od umrlih nije bio na STM, i najverovatnije su samoinicijativno koristili metadon i benzodiazepine. Patohistolo&scaron;ki nalaz na srcu može govoriti u prilog kardiotoksičnosti metadona i njegove kombinacije sa benzodiazepinima, pogotovo kod slučajeva sa pronađenim akutnim miokardijalnim o&scaron;tećenjem. &Scaron;to se tiče hroničnih promena na srcu, ne postoji mogućnosti da se potvrdi niti opovrgne uloga psihostimulanasa. Detektovane koncentracije metadona i diazepama kod MRDs su bile u opsegu terapijskih (&lt;1 &mu;g/ml). Poredeći socio-demografske karakteristike opijatnih zavisnika koji su počeli sa STM u ovom istraživanju sa podacima iz sličnih studija sprovedenih &scaron;irom sveta, zapažena je sličnost u pogledu velikog broja karakteristika. Srednje doze metadona 8., 15. dana i nakon 1. i 6. meseca primene STM su bile 40,23&plusmn;17,11 mg, 47,11&plusmn;16,79 mg, 50,00&plusmn;17,55 mg i 78,63&plusmn;18,14 mg, dok su srednje doze diazepama u istim vremenskim tačkama bile 35,92&plusmn;10,47 mg, 33,89&plusmn;9,23 mg, 28,33&plusmn;11,55 mg i 28,12&plusmn;11,67 mg. Srednje koncentracije metadona su u posmatranim tačkama ispitivanja iznosile 153,44&plusmn;111,51 ng/ml, 157,43&plusmn;112,39 ng/ml, 176,77&plusmn;118,56 ng/ml i 342,86&plusmn;181,54 ng/ml, dok su srednje koncentracije diazepama bile 923,00&plusmn;537,89 ng/ml, 923,76&plusmn;739,96 ng/ml, 560,74&plusmn;436,72 ng/ml i 1045,32&plusmn;932,72 ng/ml. Dužina QTc intervala pre primene STM je bila 411,87&plusmn;27,22 ms, tj. 414,64&plusmn;29,38 ms 8. dana STM, 416,97&plusmn;26,39 15. dana, i 425,20&plusmn;17,71 ms nakon 1. meseca tj. 423,50&plusmn;14,72 ms nakon 6. meseca primene STM. Pokazan je statistički značajan porast dužine QTc intervala nakon 1. i nakon 6. meseca primene STM u odnosu na vrednost pre primene STM, kako u grupi svih ispitanika, tako i u podgrupi mu&scaron;kog pola. Pokazano je postojanje statistički značajne korelacije između koncentracije metadona i dužine QTc intervala nakon 15. dana, 1. i 6. meseca primene STM, kako kod svih ispitanika, tako i u podgrupi mu&scaron;kog pola. Ova korelacija ostaje statistički značajna i ukoliko se uključe i drugi faktori &ndash; koncentracija diazepama i dužina perioda upotrebe heroina, kod svih ispitanika i u podgrupi mu&scaron;kog pola nakon 15 dana i mesec dana primene STM, kao i u podgrupi mu&scaron;kog pola nakon 6. meseca STM. Iako nijedan pacijent nije prijavio neko neželjeno delovanje metadona na nivou kardiovaskularnog sistema, najveći broj pacijenata oba pola se nakon prvog meseca primene STM žalio na pojačano znojenje i opstipaciju. Koncentracije metadona i diazepama u uzorcima krvi kod MRDs se nalaze u rasponu koncentracija ovih lekova u krvi ispitanika koji su učestvovali u prospektivnoj studiji. Trećina umrlih je imala samo znake akutnog o&scaron;tećenja srca, dok do porasta troponina i vrednosti QTc intervala preko 500 ms nije do&scaron;lo ni kod jednog ispitanika iz prospektivne studije. Potrebno je sprovesti dalja istraživanja sa ciljem razja&scaron;njenja moguće uloge benzodiazepina u povećanju kardiotoksičnosti metadona kod opijatnih zavisnika na STM.</p> / <p>Methadone is a synthetic agonist of opioid receptors which is used in methadone maintenance tratment (MMT) of opiate addicts as well as in the treatment of chronic pain. A long-term use of MMT is followed by mild, mostly transient, adverse effects. However, methadone belongs to a group of medicines which can provoke a prolongation of QTc (corrected QT) interval in electrocardiogram (ECG) and thus increase the risk from the development of potentially fatal arrhythmias &ndash; torsades de pointes. Moreover, methadone is widely associated with benzodiazepines use in heroin addicts, and this combination is considered as a risk factor for lethal outcome. Despite the fact that most of health care professionals are aware of possible respiratory depressant effect of methadone and benzodiazepines co-administration, recently published data reveal that ventricular arrhythmia and cardiac arrest are currently the most frequent adverse event attributed to methadone and benzodiazepine co-medication. The aim of this study is to assess cardiac safety of methadone use, especially in combination with benzodiazepines, by analyzing characteristics of methadone-related deaths (MRDs) during 10-year period as well as by conducting a clinical trial among opiate addicts in MMT. A retrospective study to determine the characteristics of MRDs in Vojvodina, as well as a clinical trial in which participated opiate addicts at the start of MMT were performed. ECG (to calculate QTc interval) and blood sampling (to determine methadone and diazepam concentrations and troponin values) were performed in all study participants at five time points (before the introduction of MMT, on 8th, on 15th day, after 1 and 6 months of MMT). Methadone and diazepam concentrations in serum were determined by using liquid chromatography-mass spectrometry (LC-MS). An increasing tendency of MRDs was observed in the region of Vojvodina, but none of the victims were under healthcare professionals&rsquo; control, and, most commonly, they used methadone and benzodiazepines, on their own initiative. Pathohistological findings in the heart in MRDs might support cardiac adverse effects of methadone and its combination with benzodiazepines, especially in cases with acute myocardial damage. As for the chronic heart changes, we can neither confirm nor exclude the role of psychostimulants. Detected concentrations of methadone and diazepam were in therapeutic range (&lt;1 &mu;g/ml). Comparing socio-demographic characteristics of opiate addicts who started with MMT in this study with data from similar studies conducted worldwide, the similarity in terms of large number of features was observed. The mean methadone dose on the 8th, 15th days, and after 1 and 6 months of MMT was 40.23&plusmn;17.11 mg, 47.11&plusmn;16.79 mg, 50.00&plusmn;17.55 mg and 78.63&plusmn;18.14 mg, respectively, while the mean diazepam dose at the same time points was 35.92&plusmn;10.47 mg, 33.89&plusmn;9.23 mg, 28.33&plusmn;11.55 mg and 28.12&plusmn;11.67 mg, respectively. The mean methadone concentration at observed time points was 153.44&plusmn;111.51 ng/ml, 157.43&plusmn;112.39 ng/ml, 176.77&plusmn;118.56 ng/ml and 342.86&plusmn;181.54 ng/ml, respectively, while the mean diazepam concentration was 923.00&plusmn;537.89 ng/ml, 923.76&plusmn;739.96 ng/ml, 560.74&plusmn;436.72 ng/ml and 1045.32&plusmn;932.72 ng/ml, respectively. The length of QTc interval before the introduction of MMT was 411.87&plusmn;27.22 ms, 414.64&plusmn;29.38 ms on the 8th day of MMT, 416.97&plusmn;26.39 on the 15th day of MMT, after 1 month of MMT 425.20&plusmn;17.71 ms and after 6 months of MMT 423.50&plusmn;14.72 ms. There was a statistically significant increase in the length of QTc interval after 1 and 6 months of MMT in comparison to the value before the application of MMT, within the whole group of patients and in the subgroup of men. A statistically significant correlation between the concentration of methadone and QTc interval length after 15 days, 1 and 6 months of MMT, both in the whole group and in the subroup of men was observed. The correlation remained statistically significant if the other factors, such as concentration of diazepam and the length of heroin use, were included, in all patients and in the subgroup of men after 15 days and one month of MMT as well as in the subgroup of men after 6 months of MMT. Although none of the patients reported any cardiac adverse effect of methadone, the majority of them complained of sweating and constipation after the first month of MMT. Concentrations of methadone and diazepam in blood samples in MRDs were within the range of concentrations of these drugs in blood of patients who participated in the prospective study. In one third of MRDs only signs of acute myocardial damage were detected, while an increase in troponin values and the length of QTc interval over 500 ms did not occur in any patient in the prospective study. Further studies could clarify the possible role of benzodiazepines in the increasing cardiotoxicity of methadone in opiate addicts in MMT.</p>

Die ersten identifizierten Krankheitsgene der dilatativen Kardiomyopathie (DCM) kodierten alle für Proteine des Zytoskeletts. Deshalb wurde DCM als Erkrankung des Zytoskeletts bezeichnet. Bei der hypertrophen Kardiomyopathie (HCM) wurden bisher mehr als 250 Mutationen in neun Sarkomerprotein-Genen beschrieben. Deshalb wurde HCM als Erkrankung des Sarkomers bezeichnet. In den letzten Jahren wurde dieses Konzept durch Entdeckung von Mutationen in Sarkomerprotein-Genen bei DCM jedoch in Frage gestellt. Vor diesem Hintergrund haben wir die Sarkomerprotein-Gene beta-MHC und Troponin T bei 46 nicht verwandten DCM-Patienten untersucht. Das systematische Mutationsscreening wurde mit Hilfe von SSCP-Analyse und DNA-Sequenzierung durchgeführt. Im beta-MHC-Gen konnten wir die zwei Missense-Mutationen Ala223Thr und Ser642Leu bei zwei jungen Patienten identifizieren. Beide Mutationen wurden weder bei 136 HCM-Patienten noch bei 88 Kontrollen gefunden. Mit dem Editor for Structural Alignment of Proteins (STRAP) wurden die Mutationen auf die Proteinstruktur des Myosins projiziert. Hier ist erkennbar, dass Ala223Thr in der oberen 50 kDa Domäne und Ser642Leu in der Aktin-Myosin-Bindungsregion liegt. Der Austausch von Alanin zu Threonin könnte die Raumstruktur des Proteins verändern, Thermostabilität verringern und die Proteinfaltung und somit die Proteinmotilität beeinträchtigen. In der Aktin-Myosin-Bindungsregion liegt neben Ser642Leu die bereits bekannte DCM-assoziierte Mutation Ser532Pro. Durch eine Verminderung der Krafterzeugung könnten die beiden Mutationen zu DCM führen. Ferner wurden die zwei stummen Mutationen IVS11+23A>T und Asp376Asp und sechs Polymorphismen identifiziert. Im Troponin T-Gen wurden keine Mutationen, jedoch sechs Polymorphismen beobachtet. Es ergab sich kein Anhaltspunkt auf eine funktionelle Relevanz der stummen Mutationen oder Polymorphismen. Wir konnten also bestätigen, dass Mutationen in Sarkomerprotein-Genen sowohl zu HCM als auch zu DCM führen können. / All of the initially identified disease-causing genes in dilated cardiomyopathy (DCM) encoded proteins of the cytoskeleton. Therefore DCM has been termed a disease of the cytoskeleton. In hypertrophic cardiomyopathy (HCM) more than 250 mutations in nine sarcomeric protein genes have been identified so far. Therefore HCM has been termed a disease of the sarcomere. However, in the last few years this concept has been queried by findings of mutations in sarcomeric protein genes in DCM. According to this consideration we screened the sarcomeric protein genes beta-MHC and troponin T in 46 patients with DCM. Systematic mutation screening was done using SSCP analysis and DNA sequencing. In the beta-MHC gene we identified the two missense mutations Ala223Thr and Ser642Leu in two young patients. Both mutations were neither found in 136 HCM patients nor in 88 controls. Using the Editor for Structural Alignment of Proteins (STRAP) the mutations were projected onto the protein structure of myosin. Ala223Thr turned out to be localized in the upper 50 kDa domain and Ser642Leu in the actin-myosin-interface region. The exchange from alanine to threonine might alter the spatial structure of the protein, decrease its thermostability and affect the protein folding and thus the protein motility. Closely to Ser642Leu the DCM-associated mutation Ser532Pro is located in the actin-myosin-interface region. By a decrease in force production both mutations might cause DCM. Furthermore we identified the two silent mutations IVS11+23A>T and Asp376Asp and six polymorphisms. In the troponin T gene no mutations but six polymorphisms were detected. No evidence was found for functional relevance of the silent variants or polymorphisms. Thus, we could confirm that mutations in sarcomeric protein genes can lead to both HCM and DCM.