ATP-sensitive Potassium Channels and Cardiac Arrhythmia

PhDATP-sensitive potassium channels (KATP) open in response to metabolic challenge. They form of pore subunits (Kir6.1 or Kir6.2) and modulatory subunits (SUR1, SUR2A or SUR2B) and are ubiquitously expressed. Differential subunit composition between cardiac chambers was investigated, as were atrial anti-arrhythmic effects of KATP modulation. Selective pharmacology of KATP openers and inhibitors was confirmed in a heterologous expression system through whole-cell patch clamp. Isolated HL-1 cells (a murine atrial cardiomyocyte model) and murine atrial cardiomyocytes showed identical KATP pharmacological responses representing Kir6.2/SUR1 channels. Relative quantification of murine whole atrial RNA concurred, and was distinct from the ventricles (Kir6.2/SUR2). Human whole heart RNA from normal hearts exhibited a different pattern with no obvious chamber specificity. Kir6.1-/- and Kir6.2-/- mice demonstrated that both pore types contribute to electrophysiological parameters in isolated atrial cardiomyocytes, but Kir6.2 appears more important. In atrial tissue (Langendorff hearts), Kir6.2-/- more than Kir6.1-/- mice demonstrated increased effective refractory periods and reduced conduction velocity at baseline, and during hypoxia, compared to wildtype. A trend to reduced arrhythmogenicity was observed during programmed electrical stimulation in the Kir6.2-/- mouse. In syncytia of spontaneously beating HL-1 cells, KATP activation with diazoxide was met with rotational to uniform wavefront organisation and silencing of electrical activity in a dose-dependent manner, reversed with channel blockade. In Langendorff mouse hearts KATP inhibition reversed hypoxia induced slowing of spontaneous sinus node activation, but pharmacological activation alone did not, suggesting different mechanisms with hypoxic channel activation. Thus, both pore subunits contribute to the cardiac electrophysiology of murine atria, but Kir6.2 appears more important. HL-1 cells exhibit identical KATP pharmacology to murine atrial myocytes, which have a differential subunit composition compared to the ventricle. Any human cardiac KATP differential subunit expression needs further exploration. KATP activation and inhibition have anti-arrhythmic effects and this might be explored further clinically.Medical Research Council MR/L016230/1

The molecular genetics of familial cardiomyopathy

Introduction The cardiomyopathies are responsible for approximately 5.9 of 100,000 deaths in the general global population and in sub-Saharan Africa (SSA), these myocardial diseases are observed in 21.4% of patients with heart failure. The precise etiology of the cardiomyopathies is currently not well known and through our research we aim to contribute to the genetic landscape and bridge the gaps in knowledge for the different cardiomyopathies as SSA could provide some very important insights into the cardiomyopathies and identify other possible disease mechanisms. Methods Through next generation sequencing techniques such as whole exome sequencing and targeted resequencing we studied three South African families with severe cardiomyopathy. Clinical diagnosis and recruitment of cardiomyopathy patients into the study was done at Groote Schuur Hospital, Cape Town by a panel of experts. Next generation sequencing data was analysed and filtered through various stringent criteria and the final list of variants were validated through Sanger sequencing. Results In the first multi-generational family with severe dilated cardiomyopathy (DCM) (DCM 334), we identified a pathogenic DMPK c.1067C>T(p.P356L) variant in the proband and her affected father. We also screened a cohort of 542 cardiomyopathy probands though Sanger sequencing of the DMPK gene and identified the DMPK c.1477C>T(p.R493C) variant as a variant of unknown significance. We then investigated a three-generation family with four affected family members who were also affected with severe DCM (DCM343). We used whole exome sequencing and identified the pathogenic BAG3 c.925C>T (p.R309Ter) variant as the cause of disease within this family. Viral infection, anti-hypertensive medication and genetic modifiers in RYR1 and NEB contributed to the variable phenotype among the individuals with the BAG3 variant. Through targeted resequencing we also identified the same pathogenic BAG3 variant in 2 of the 634 cardiomyopathy probands screened. In the third family, we investigated a South African family affected with severe arrhythmogenic cardiomyopathy (ACM). We used whole exome sequencing and targeted resequencing in combination and identified the pathogenic PKP2 c.2197_2202InsGdelCACACC (p.H733Afs*8) as the cause of disease in the proband and his father. We also present evidence of the ALPK3 c.2701C>T(p.Q901Ter) variant modifying the phenotypic manifestation which correlates with the variable penetrance that is seen among ACM families. Conclusion Through this project, we have identified many firsts. To the best of our knowledge, we are the first to show that DMPK is associated with primary DCM in severely affected young patients. As a first for South Africa, we not only identified the pathogenic BAG3 variant in a family with severe DCM, but we also identified the same variant in two additional probands, raising the possibility of a founder effect. In the third and final family with ACM, we identified the pathogenic PKP2 variant as the cause of disease within this family with the novel ALPK3 variant acting as a possible modifier. Our research has added to what is currently known about the cardiomyopathies in Africa but there is still much work to be done as we believe we have just scratched the tip of the iceberg