Ryanodine receptor type 2: A molecular target for dichlorodiphenyltrichloroethane- And dichlorodiphenyldichloroethylene-mediated cardiotoxicity

Abstract

Dichlorodiphenyltrichloroethane (DDT) and its metabolite dichlorodiphenyl-dichloroethylene (DDE) are ubiquitously found in the environment and linked to cardiovascular diseases-with a majority of the work focused on hypertension. Studies investigating whether DDx can interact with molecular targets on cardiac tissue to directly affect cardiac function are lacking. Therefore, we investigated whether o,p0-DDT, p,p0-DDT, o,p0-DDE, or p,p0-DDE (DDx, collectively) can directly alter the function of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) by assessing their effect(s) on hiPSC-CMs Ca2þ dynamics. DDx (0.1-10 mM) affected hiPSC-CMs synchronous Ca2þ oscillation frequency in a concentration-dependent manner, with p,p0-DDT and p,p0-DDE also decreasing Ca2þ stores. HEK-RyR2 cells cultured under antibiotic selection to induce expression of wild-type mouse ryanodine receptor type 2 (RyR2) are used to further investigate whether DDx alters hiPSC-CMs Ca2þ dynamics through engagement with RyR2, a protein critical for cardiac muscle excitation-contraction coupling (ECC). Acute treatment with 10 mM DDx failed to induce Ca2þ release in HEK293-RyR2, whereas pretreatment with DDx (0.1-10 mM) for 12- or 24-h significantly decreased sarcoplasmic reticulum Ca2þ stores in HEK-RyR2 cells challenged with caffeine (1 mM), an RyR agonist. [3H]ryanodine-binding analysis using murine cardiac RyR2 homogenates further confirmed that all DDx isomers (10 mM) can directly engage with RyR2 to favor an open (leaky) confirmation, whereas only the DDT isomers (10 mM) modestly (10%) inhibited SERCA2a activity. The data demonstrate that DDx increases heart rate and depletes Ca2þ stores in human cardiomyocytes through a mechanism that impairs RyR2 function and Ca2þ dynamics. Impact Statement: DDT/DDE interactions with RyR2 alter cardiomyocyte Ca2þ dynamics that may contribute to adverse cardiovascular outcomes associated with exposures.