京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第13354号 / 論医博第2200号 / 新制||医||1044(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 長船 健二, 教授 木村 剛, 教授 羽賀 博典 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第13380号 / 論医博第2214号 / 新制||医||1047(附属図書館) / 京都大学大学院医学研究科外科系専攻 / (主査)教授 山下 潤, 教授 木村 剛, 教授 浅野 雅秀 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Hypertrophic cardiomyopathy (HCM), a heart-related abnormality, is the most prevalent cause of sudden death in young athletes at sporting events. A cluster of cardiomyopathy mutations are localized in β-cardiac myosin at the N-terminal region of subfragment-2. Using resonance energy transfer probes, a synthetic peptide model system was developed to study stability of the coiled coil (S2 fragment) structure by determining monomer-dimer equilibrium of the peptide. Fluorescence resonance energy transfer and MacroModel software suite were used to obtain distance measurements along with measurement of coiled coil formation. The model peptide was used to characterize the effects of disease-causing-mutations and examine potential candidate drugs (polyamines) to counteract effects of mutations causing HCM. Distance measurements between donor and acceptor probes obtained by computational simulation and fluorescence resonance energy transfer (FRET) were consistent. Measurements also agreed with simulations of unlabeled wildtype, indicating coiled coil structural stability of the peptide. Interaction of the site-specific antibody with the peptide strongly inhibited dimerization and destabilized coiled coil structure of the peptide. Presence of negatively charged glutamate residues in the region of subfragment-2 strongly suggested a potential interaction site for positively charged polyamines. Binding of certain polyamines, such as poly-L-Lysine 11 residues and poly-D-Lysine 17 residues, demonstrated the ability to enhance dimerization and improve stability of the coiled coil structure, while some other polyamines were shown to have insignificant impact on the structure. In an attempt to characterize the effect of HCM-causing-mutations, peptides containing E924K mutation and lethal mutation E930 deletion were synthesized. Fluorescence resonance probes were conjugated to the mutant peptides to determine coiled coil formation. Results obtained from both dynamic simulations and resonance energy transfer experiments indicated that these mutations strongly inhibit dimerization, and thus, destabilize coiled coil structure of the peptide. Further experiments were conducted using heterodimers containing a chain of wildtype and a chain of mutant peptide. Both E924K & Edel930 mutations destabilized coiled coil formation and prevented dimerization. This peptide model system would provide a promising tool for drug development targeting HCM-causing-mutations along the S2 region of myosin.

Familial hypertrophic cardiomyopathy (HCM) is a genetic disease largely caused by a mutation in myosin binding protein C (MYBPC3) and it affects about 1:500 population leading to arrhythmic sudden death, heart failure, and atrial fibrillation. MYBPC3 activates calcium-induced actin-myosin filament sliding within the cardiac sarcomere, creating the force necessary for heart contraction. The underlying molecular mechanisms causing HCM phenotype remain elusive, therefore, there is an urgent need for a reliable in vitro human HCM model to investigate the pathogenesis of HCM. This study utilized isogenic human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with MYBPC3 gene mutation (wildtype, heterozygous, homozygous) and further micropatterned them into fiber-like structures on polyacrylamide hydrogels of physiological and fibrotic-like stiffnesses. Cells were cultured for an extended culture time up to 60 days and their morphology/attachment, contractility, and calcium transient were extensively and carefully evaluated. It was found that MYBPC3 knockout cells maintained the highest contraction amplitude, but had increased contraction, and relaxation durations, decreased calcium transient amplitude, as well as time to peak and decay times over the culture period in comparison to the isogenic wildtype. Overall, this study demonstrates that hiPSC-CMs can be successfully patterned and cultured for an extended time on hydrogels forming end-to-end connections, which can be served as a simple yet effective in vitro human model for studying mechanical dysfunction of HCM.

Introduction: Inflammatory cardiomyopathy (DCMi) represents a non-familial form of dilated cardiomyopathy (DCM) and endomyocardial biopsy (EMB) is crucial for its diagnosis. Aims: To assess the prevalence of DCMi in patients with DCM of unclear origin, to evaluate the significance of serological tests for antibodies against infectious cardiotrophic agents and to analyze the effect of specific therapy guided by EMB results. Methods: EMB was performed in 56 subjects (mean age 52 ± 10 years) with DCM of unclear etiology and left ventricular (LV) ejection fraction (EF) < 40% with a history of heart failure less than 1 year. EMB samples were analyzed by immunohistochemistry, polymerase chain reaction (PCR) and electron microscopy. Results: Immunohistochemical examination revealed myocardial inflammation in 26 patients (46%), the PCR method detected genome of microbial agents in 32 patients (57%). Electron microscopy showed the presence of particles of microbial agents in 41 patients (73%). Serological blood tests found no IgM antibody positivity against any of the investigated microbial agents. Targeted antibiotic therapy in patients with evidence of Borrelia burgdorferi (Bb) genome in the EMB led to a reduction in LV size, improvement of LV EF and alleviate symptoms of heart failure. Conclusion: DCMi...

Kardiomyopathien (KMP) sind bei Katzen die häufigste kardiovaskuläre Erkrankung und ein wichtiger Grund für Morbidität und Mortalität (FOX et al. 1995). Die morphologische und strukturelle Veränderung des Myokards entsteht definitionsgemäß in Abwesenheit von Klappenerkrankungen, kongenitalen Defekten oder koronaren Gefäßerkrankungen. Unterschieden werden können primäre und sekundäre Kardiomyopathien. Primäre Kardiomyopathien (1°KMP) sind genetisch oder idiopathisch bedingt. Anhand des Phänotyps können hypertrophe (HKMP), restriktive (RKMP), unklassifizierte (UKMP), dilatative (DKMP) und arrhythmogene rechtsventrikuläre (ARVC) Kardiomyopathie unterschieden werden. Sekundäre Kardiomyopathien (2°KMP) werden durch eine systemische Erkrankung ausgelöst. Als Ursachen werden insbesondere Hyperthyreose, systemische Hypertension, Myokarditis, Taurinmangel, Kortikosteroidapplikation, chronische Niereninsuffizienz (CNI), anhaltende Tachyarrhythmie, Diabetes mellitus, Anämie, Neoplasie, Anthrazykline, Hypersomatotropismus, Sepsis und Myopathie beschrieben. In der Literatur machen die sekundären Kardiomyopathien bisher noch einen relativ geringen Anteil der Kardiomyopathien aus. Die für diese Untersuchung ausgewerteten Daten von Katzen mit Myokarderkrankungen wurden innerhalb von 29 Monaten erhoben. Einschlusskriterien waren das Vorliegen des Signalements, der Anamnese, des Blutdrucks und einer standardisierten echokardiografischen Untersuchung. Außerdem musste zur Erfassung der häufigsten auslösenden Erkrankungen bei Patienten über neun Jahren das Gesamtthyroxin erfasst worden sein, bei Polyurie und Polydipsie die Parameter Harnstoff, Kreatinin und Glukose und bei blassen Schleimhäuten der Hämatokrit. Von den 106 Katzen, die den Einschlusskriterien entsprachen, machte die 2°KMP mit 50 Katzen (47%) den größten Anteil aus. Zweithäufigste Kardiomyopathie war die HKMP mit 39 Fällen (37%), gefolgt von UKMP (9 Katzen, 8%), RKMP (6 Katzen, 6%) und DKMP (2 Katzen, 2%). Das Signalement der Katzen mit 82 EKH (77%) eine signifikante Häufung dieser Rasse, die aber der Verteilung der Rassen im Patientengut entspricht (p<0,001). Im Mittel waren die Katzen 9 Jahre alt (1-17 Jahre). Die Katzen mit HKMP waren signifikant jünger als die Katzen der anderen Gruppen (p<0,001). Die Katzen über 12 Jahre zeigten signifikant häufiger eine 2°KMP als eine 1°KMP (p=0,001). Mit 74% (n=78) waren Kater signifikant häufiger betroffen als Kätzinnen (p=0,001). Die 28 Kätzinnen waren mit 64% (n=18) häufiger an einer 2°KMP erkrankt als an einer primären Kardiomyopathie (p=0,03). Beim Gewicht (mittleres Gewicht 4,7 kg, Median 4,2 kg; 2-9 kg) zeigten sich keine relevanten Unterschiede zwischen den Gruppen. Das häufigste Symptom war bei 37% der Katzen (n=38) Dyspnoe. Weitere respiratorische Probleme waren Husten (n=5, 5%) und Maulatmung (n=3, 3%). Bei sechs Tieren (6%) lag eine arterielle Thrombembolie mit Paraplegie vor. Aszites und subkutane Ödeme waren bei je zwei Tieren (2%) auffällig. 11% der Katzen (n=12) hatten einen anderen Vorstellungsgrund: bei sieben Katzen lag eine Harnabsatzstörung (feline lower urinary tract disease) und bei fünf ein Trauma vor. Bei der kardialen Auskultation waren 53 (50%) Katzen mit einem systolischen Herzgeräusch vom Lautstärkegrad 1-4/6 auffällig. Signifikant häufiger als in den anderen Gruppen war bei den Katzen mit HKMP ein Herzgeräusch zu verzeichnen (p <0,001). Bei 48 (45%) Katzen lag ein Galopprhythmus vor, bei 11 der Tiere (10%) in Kombination mit einem Herzgeräusch. Die Katzen mit Galopprhythmus waren signifikant häufiger dekompensiert als die mit Herzgeräusch oder ohne abnormen Auskultationsbefund (p=0,001). Die kardiogene Dyspnoe als häufigster Vorstellungsgrund (n=38) wurde in 45% (n=17) der Fälle durch ein Lungenödem, in 26% (n=10) durch einen Pleuraerguss und in 29% (n=11) durch die Kombination beider bedingt. Katzen mit RKMP hatten signifikant häufiger einen Pleuraerguss als Katzen der anderen Gruppen (p=0,001). Von den Katzen mit UKMP lag hingegen häufiger ein Lungenödem vor (p=0,016). Aszites trat bei keiner Katze mit HKMP und RKMP auf. Alle Katzen mit RKMP und DKMP gehörten der ISACHC-Klasse 3 (International Small Animal Cardiac Health Council) an (p<0,001). Sieben der neun Katzen mit UKMP (78%) entsprachen ebenfalls dieser Klasse. Die Katzen mit RKMP zeigten die am stärksten ausgeprägte linksatriale Dilatation (LADs 27,4 mm; 23,8-30,5 mm), die mit 2°KMP die am wenigsten ausgeprägte (19,2 mm; 13,4-28,2 mm). Innerhalb der 2°KMP (n=50) lag bei 19 Katzen eine Hyperthyreose, bei 16 CNI und bei 15 Tieren systemische Hypertension vor. Weitere Ursachen waren Glukokortikoidgabe (n=4), Anämie (n=2), Diabetes mellitus (n=2) und schwere Lungenerkrankung oder Pyothorax (n=4). Eine Kombination dieser Erkrankungen bestand bei 24% (n=12) der Katzen. Unter Vernachlässigung potentiell auslösender Erkrankungen ergab eine Klassifizierung der 50 Katzen mit 2°KMP anhand des Phänotyps bei 68% (n=34) eine HKMP. Eine UKMP machte 18% (n=9) und eine RKMP 14% (n=7) aus. Hyperthyreose auch in Kombination mit anderen Erkrankungen war in 84% (16/19) ebenso wie systemische Hypertension in 80% (12/15) mit linksventrikulärer Hypertrophie assoziiert. Mit 47% übersteigt die Häufigkeit der 2°KMP deutlich die Angaben in der Literatur. Zusätzliche Fälle können als 1°KMP fehlinterpretiert worden sein, während eine erworbene Begleiterkrankung neben einer 1°KMP fälschlicherweise als 2°KMP gewertet sein kann. In jedem Fall betonen die Ergebnisse dieser Untersuchung aber die Wichtigkeit weiterführender diagnostischer Schritte, um Erkrankungen mit Auswirkungen auf das Myokard zu identifizieren. Inwieweit eine kausale Therapie die Rückbildung myokardialer Veränderungen erlaubt, müssen weitere Untersuchungen klären.

Context:Broken heart syndrome or Takotsubo cardiomyopathy (TC) is a disorder characterized by transient left ventricular apical ballooning that almost invariably precedes emotional or physical stress. Although the patients with chronic kidney disease on hemodialysis have shown to exhibit sustained activity of sympathetic nervous system, the presentation of TC in these patients is a rare entity with few case reports in the literature. Case Report: A 75-year-old female with past medical history of end-stage renal disease presented with chest pressure and heaviness that started during her maintenance hemodialysis session. Electrocardiogram showed ST elevation and T wave inversion in V3-V6 leads. Emergent left heart catheterization was done that showed normal coronaries and akinesis of apical left ventricle wall consistent with TC. She was started on maximal medical management and underwent hemodialysis the next day without recurrence of the symptoms. Conclusion: TC may an underdiagnosed entity in patients on hemodialysis. However, it should be considered in the differential diagnosis in hemodialysis patients, particularly who presents with chest pain and/or symptoms.

Heat shock protein A12B (HSPA12B) is predominately expressed in endothelial cells (ECs) and has been reported to protect against cardiac dysfunction from endotoxemia or myocardial infarction. This study investigated the mechanisms by which endothelial HSPA12B protects polymicrobial sepsis–induced cardiomyopathy. Wild-type (WT) and endothelial HSPA12B knockout (HSPA12B–/–) mice were subjected to polymicrobial sepsis induced by cecal ligation and puncture (CLP). Cecal ligation and puncture sepsis accelerated mortality and caused severe cardiac dysfunction in HSPA12B–/– mice compared with WT septic mice. The levels of adhesion molecules and the infiltrated immune cells in the myocardium of HSPA12B–/– septic mice were markedly greater than in WT septic mice. The levels of microRNA-126 (miR-126), which targets adhesion molecules, in serum exosomes from HSPA12B–/– septic mice were significantly lower than in WT septic mice. Transfection of ECs with adenovirus expressing HSPA12B significantly increased miR-126 levels. Increased miR-126 levels in ECs prevented LPS-stimulated expression of adhesion molecules. In vivo delivery of miR-126 carried by exosomes into the myocardium of HSPA12B–/– mice significantly attenuated CLP sepsis increased levels of adhesion molecules, and improved CLP sepsis–induced cardiac dysfunction. The data suggest that HSPA12B protects against sepsis-induced severe cardiomyopathy via regulating miR-126 expression which targets adhesion molecules, thus decreasing the accumulation of immune cells in the myocardium.

Takotsubo cardiomyopathy, also known as “broken heart syndrome,” is a transient left ventricular dysfunction associated with stress (usually emotional) induced myocardial injury and stunning. It often presents as myocardial infarction on surface electrocardiogram (EKG). Diagnosis is made by coronary angiography, which rules out coronary artery disease and shows pathognomonic apical ballooning. In this article, we present a case of a 72-year-old woman who initially presented with an ST segment elevation myocardial infarction on EKG. Coronary angiography showed severe left anterior descending artery and diagonal lesions requiring percutaneous coronary intervention. Post–percutaneous coronary intervention, EKG changes resolved. The next day, the patient developed recurrent chest pain and her EKG showed diffuse T-wave inversion in precordial leads with reemerging ST segment elevations concerning for stent thrombosis. The patient underwent repeat emergent coronary angiography, which showed patent stents and findings consistent with takotsubo cardiomyopathy.

Isolated noncompaction of the left ventricular myocardium (INVM) is characterized by the presence of numerous prominent trabeculations and deep intertrabecular recesses within the left ventricle, sometimes also affecting the right ventricle and interventricular septum. Familial occurrence of this disorder was described previously. We present a family in which 6 affected individuals demonstrated X-linked recessive inheritance of this trait. Affected relatives presented postnatally with left ventricular failure and arrhythmias, associated with the pathognomonic echocardiographic findings of INVM. The usual findings of Barth syndrome (neutropenia, growth retardation, elevated urinary organic acids, low carnitine levels, and mitochondrial abnormalities) were either absent or found inconsistently. Fetal echocardiograms obtained between 24-30 weeks of gestation in 3 of the affected males showed a dilated left ventricle in one heart, but were not otherwise diagnostic of INVM in any of the cases. Four of the affected individuals died during infancy, one is in cardiac failure at age 8 months, and one is alive following cardiac transplant at age 9 months. The hearts from infants who died or underwent transplantation appeared, on gross examination, to be enlarged, with coarse, deep ventricular trabeculations and prominent endocardial fibroelastosis. Histologically, there were loosely organized fascicles of myocytes in subepicardial and midmyocardial zones of both ventricles, and the myocytes showed thin, often angulated fibers with prominent central clearing and reduced numbers of filaments. Markedly elongated mitochondria were present in some ventricular myocytes from one specimen, but this finding was not reproducible. Genetic linkage analysis has localized INVM to the Xq28 region, where other myopathies with cardiac involvement have been located.