Oxidized Low-Density Lipoprotein Associates with Ventricular Stress in Young Adults and Triggers Intracellular Ca2+ Alterations in Adult Ventricular Cardiomyocytes

Oxidized low-density lipoprotein (oxLDL) is associated with cardiac damage and causes injury to multiple cell types. We aimed to investigate the role of oxLDL in ventricular stress. We first examined the association between circulating oxLDL and N-terminal pro-brain natriuretic peptide (NT-proBNP), a marker of myocardial stress, in young subjects (30–50 years) with or without stable coronary artery disease (SCAD). oxLDL and NT-proBNP were significantly higher in subjects at high cardiovascular risk (CVR) than in subjects at low CVR and were associated independently of traditional CVR factors and C-reactive protein. Furthermore, the levels of oxLDL and NT-proBNP were significantly lower in subjects with SCAD than in peers at high CVR. To determine the intracellular mechanisms involved in the cardiac effects of oxLDL, we analyzed the in vitro effect of oxLDL on intracellular Ca2+ handling in adult rat ventricular cardiomyocytes using confocal microscopy. Acute challenge of adult ventricular cardiomyocytes to oxLDL reduced systolic Ca2+ transients and sarcoplasmic reticulum Ca2+ load. Moreover, diastolic spontaneous Ca2+ leak increased significantly after acute exposure to oxLDL. Thus, we demonstrate that oxLDL associates with NT-proBNP in young subjects, and can directly induce Ca2+ mishandling in adult ventricular cardiomyoyctes, predisposing cardiomyocytes to cardiac dysfunction and arrhythmogenicity.

In vitro and in vivo investigation of cardiotoxicity associated with anticancer proteasome inhibitors and their combination with anthracycline

Although proteasome inhibitors (PIs) are modern targeted anticancer drugs, they have been associated with a certain risk of cardiotoxicity and heart failure (HF). Recently, PIs have been combined with anthracyclines (ANTs) to further boost their anticancer efficacy. However, this raised concerns regarding cardiac safety, which were further supported by several in vitro studies on immature cardiomyocytes. In the present study, we investigated the toxicity of clinically used PIs alone (bortezomib (BTZ), carfilzomib (CFZ)) as well as their combinations with an ANT (daunorubicin (DAU)) in both neonatal and adult ventricular cardiomyocytes (NVCMs and AVCMs) and in a chronic rabbit model of DAU-induced HF. Using NVCMs, we found significant cytotoxicity of both PIs around their maximum plasma concentration (cmax) as well as significant augmentation of DAU cytotoxicity. In AVCMs, BTZ did not induce significant cytotoxicity in therapeutic concentrations, whereas the toxicity of CFZ was significant and more profound. Importantly, neither PI significantly augmented the cardiotoxicity of DAU despite even more profound proteasome-inhibitory activity in AVCMs compared with NVCMs. Furthermore, in young adult rabbits, no significant augmentation of chronic ANT cardiotoxicity was noted with respect to any functional, morphological, biochemical or molecular parameter under study, despite significant inhibition of myocardial proteasome activity. Our experimental data show that combination of PIs with ANTs is not accompanied by an exaggerated risk of cardiotoxicity and HF in young adult animal cardiomyocytes and hearts.

Abstract 333: Maturation Of ß-adrenergic Receptor (ß-AR) Signaling In Human Induced Pluripotent Stem Cell-derived Cardiomyocytes (hipsc-cms)

hiPSC-CMs are a novel model system for cardiovascular diseases, with proven utility in cardiac channelopathies, however, how accurately hiPSC-CMs recapitulate the signaling pathways regulating contractility and remodeling of adult cardiomyocytes remains to be determined. As ß-ARs are key regulators of both contractility and remodeling, we examined ß-AR signaling at different stages of hiPSC-CM maturation. hiPSCs, derived from healthy controls, were studied at 0, 14, 30, and 60d after cardiac induction under isoproterenol (ISO) stimulation ± ß1 or ß2 antagonists. Flag-tagged ß-ARs were transfected into hiPSC-CMs for trafficking studies. Compared to the ß1:ß2 ratio of 70:30 in adult ventricular cardiomyocytes, ß2-AR expression was higher in early stage (d14-30) hiPSC-CMs; ß1-ARs increased with maturation (d14-d60), reflective of the normal developmental pattern. Downstream signaling also matured from d14-60, regulating both function (PLB and TnI) and remodeling (ERK). The majority of cAMP generation and PKA activation at d30 was through ß2-ARs; by d60-90, ß1-AR signaling contributed equally. In contrast to studies in rodent cells, ß2-ARs did not show compartmentalization of PKA signaling. Between d30-60, components of the SR (RYR, PLB and CSQ) and caveolae (Cav3) matured, suggesting development of functional compartmentalization machinery. Finally, hiPSC-CMs showed ß2-AR downregulation and internalization in response to ISO, an important component of the normal cardiomyocyte response to pathologic stimulation. Thus, hiPSC-CM ß-AR signaling matures with time after cardiac induction, recapitulating many of the features of post-natal and several of adult cardiomyocytes. hiPSC-CM models of human cardiomyopathies are a reasonable platform for studying ß-AR signaling. Furthermore, the maturation of ß-AR signaling can be used as a read-out for hiPSC-CM maturity.