Transthyretin aggregates affect viability and electrical properties of cardiomyocytes

Senile systemic amyloidosis (SSA), is a sporadic disease affecting aged people whose main symptom is a severe cardiomyopathy associated with arrhythmias. SSA is characterized by the presence of extracellular amyloid fibrillar aggregates of transthyretin (TTR), a plasma protein carrying the thyroid hormone and the retinol binding protein. The aggregates are deposited in several tissues and, together with their oligomeric precursors, are responsible for tissue functional impairment. To date, liver and heart transplantation are the only medical treatments of SSA; accordingly, a thorough investigation of the molecular basis of cell/tissue functional and viability impairment induced by TTR aggregates is expected to provide knowledge needed to identify new pharmacological targets and to develop novel therapeutic strategies. We studied the effect on electrophysiology and viability of HL−1 cardiomyocytes of prefibrillar and fibrillar aggregates of TTR supplemented to the culture media. Only the prefibrillar aggregates were able to interact with the cell membrane and were internalized. This resulted in a moderate impairment of cell viability at the lowest aggregate concentration (10 M). In the same cells exposed to TTR prefibrillar aggregates, the cytosolic calcium content showed a slow, progressive rise over time; it did not reach a steady state level and came back to its basal levels upon TTR removal from bath solution. By the patch-clamp technique we investigated the effect of the enhanced intracellular calcium on the electrical properties of isolated mouse ventricular myocytes. Action potential recordings were performed at increasing rate of stimulation (0.5, 1 and 2 Hz) before and after application of TTR prefibrillar aggregates. The results showed a progressive prolongation of the action potential that was associated with a marked increase of the duration of the plateau phase; eventually, early and delayed afterdepolarizations occurred. These effects were seen at any frequency of stimulation. Altogether, our data indicate the presence of electrical abnormalities in the exposed cells with pro-arrhythmic potential induced by aggregated TTR. On-going investigation will assess the ionic basis of the altered intracellular calcium and clarify the mechanisms of TTR aggregate cytotoxicity particularly at the level of the cell membrane, the endoplasmic reticulum and the mitochondria. To our knowledge, this is the first mechanistic demonstration of a direct proarrhythmic effect of TTR aggregates in cardiomyocytes, a possible cause of SSA cardiomyopathy.