Browsing by Subject "mechanical beating analysis"

Long QT syndrome, LQTS, is a congenital or acquired cardiac disorder characterized by prolonged cardiac repolarization phase. It is observed as a prolonged QT period in electrocardiodiagraph and can cause life-threatening specific ventricular tachycardia, torsades de pointes. Hundreds of mutations in 15 genes (LQT1-15) are linked to congenital LQTS. Worldwide prevalence of congenital LQTS gene mutations is from 1:2000 to 1:5000. However, the prevalence in Finland is much higher due to four founder mutations that alone occur in one out of 250 individuals. Acquired LQTS is often drug-induced and the most common cause for the withdrawal of drugs on the market. Carriers of LQTS mutations are more susceptible to acquired LQTS than normal population. LQTS-specific cardiomyocytes can thus provide a thorough model for drug cardiotoxicity screening, better insight into disease mechanisms and assist in drug development. This thesis was a part of a bigger project concentrating on validation of LQT2-specific cell lines that could be used for the purposes mentioned above. Induced pluripotent stem (iPS) cell technology enables creation of disease-specific pluripotent stem cell lines, which can be differentiated into any cell type. In this thesis, two LQT2-specific iPS cell lines derived from a clinically symptomatic 44-year-old female were used. She is heterozygous for Finnish founder mutation L552 in KCNH2 gene, which encodes the α-subunit of the cardiac rapidly activating potassium rectifier channel. iPS cells were first verified to express pluripotency markers and to form embryoid bodies containing all germ layers. iPS cells were then differentiated into cardiomyocytes by culturing them with END-2 cells and mechanical beating of the cardiomyocytes was assessed from video recordings. Single LQT2-specific cardiomyocytes showed LQT2-related phenotype in vitro with 43% of single LQT2 cardiomyocytes showing abnormal beating patterns and prolonged contraction time. This phenotype was rescued in LQT2-specific cardiomyocyte clusters. Finally, the expression ratios of wild type and mutated KCNH2 alleles were compared between cardiomyocytes derived from the female and her son, a carrier of the same mutation but with asymptomatic phenotype. Cardiomyocytes from both individuals expressed KCNH2 alleles with the ratio between 1:2 and 1:1 (wt:mut), thus allelic imbalance does not explain differences in the clinical phenotypes. All in all, the results of this thesis suggest that after further validation, mainly electrophysiology studies, these cell lines are most likely suitable to be applied for disease modeling, cardiotoxicity screening and finding new therapies for LQT2.