Biology subjects

Jimenez-Vazquez, E. N.

Publications and source records attributed to Jimenez-Vazquez, E. N..

6 recordsLinked to original sources

bioRxiv2026

Acute Exposure to Perfluorooctanoic Acid (PFOA) During Cardiomyogenesis disrupts Transcriptional and Electrophysiological Profiles in Differentiated Myocytes

The early developmental environment plays a critical role in the etiology of cardiovascular diseases (CVDs), but underlying molecular mechanisms are poorly understood. Exposure to per and polyfluoroalkyl substances (PFAS) are linked to various CVDs, but effects of developmental PFAS exposures on the human heart remain unclear. Using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM), the objective of this study was to investigate the effects of PFAS exposure during cardiac differentiation on gene expression and function of cardiomyocytes. We exposed two hiPSC lines (one male and one female donor) to perfluorooctanoic acid (PFOA), a common and ubiquitous PFAS (0.05, 0.5, 5, 50, 100, 150, 200 M), followed by assessment of cellular number and pluripotency marker expression. PFOA exposure for 72 hours had no significant effects on hiPSC pluripotency, and modest inhibition of proliferation was observed only at the highest concentration. hiPSCs were then differentiated into ventricular cardiomyocytes in the continued presence or absence of PFOA (0, 0.5, 5, 50 M) using an established small molecules protocol. Optical mapping studies using voltage and calcium-sensitive dyes revealed dose and cell line-specific effects of PFOA on cardiomyocyte voltage and calcium dynamics that were still present 10 days after cessation of exposure. Patch clamping studies demonstrated small but significant reductions in repolarizing IKr currents with 5{micro}M PFOA exposure in cardiomyocytes from both donors. Using RNA-seq, we found that exposure to PFOA led to significant changes in transcriptional pathways related to lipids and lipoproteins in the female hiPSC-CM. In the male hiPSC-CM, we observed significant effects on developmental pathways and calcium homeostasis. Thus, we found that environmentally relevant PFOA exposure during cardiomyocyte differentiation affects the electrophysiological properties and transcriptome of hiPSC-CM even after cessation of exposure, with effects that differ by donor cell line. These findings provide direct experimental evidence that transient developmental exposure to PFOA can durably reprogram human cardiomyocyte function, supporting a developmental origin of PFAS-associated cardiovascular risk. Impact StatementThese studies demonstrate that exposure to environmentally relevant levels of PFOA during the differentiation of hiPSCs into cardiomyocytes alters cardiac gene expression and function, with effects that persist beyond cessation of exposure.

pharmacology and toxicology↗

bioRxiv2025

Sudden Death-Associated KCNH2 Variants Have Opposing Effects on hERG1NP Function

hERG1NP is a non-conducting subdomain of the full-length hERG1 channel that is expressed in the nuclei of developing cardiomyocytes. From the nucleus, hERG1NP modulates gating and expression of the full-length hERG1 channel. hERG1NP variants are linked with sudden death in the young, but the impact of these variants on hERG1NP activity has not been studied. To determine the effect of hERG1NP variants on hERG1NP activity, we measured hERG1NP intracellular localization and modulation of membrane current from the full-length hERG1a channel in HEK293 cells. The wildtype hERG1NP suppressed both hERG1a current (IhERG) and protein. We then screened six hERG1NP variants for changes in either intracellular targeting or modulation of hERG1a current. Two variants, R885C and R1047L, disrupted hERG1NP activity by altering nuclear transport or abolishing both IhERG current suppression and nuclear targeting, respectively. Two additional variants, G1036D and Q1068R, enhanced hERG1NP activity by depolarizing hERG1as voltage dependence of activation via accelerated channel deactivation and recovery from inactivation. Lastly, two variants had no effect: R1035W and R1069S. This work demonstrates that hERG1NP variants associated with sudden death in the young can trigger both loss-of-function and gain-of-function effects on hERG1NP. Additional work is needed to identify specific pathogenic mechanisms of hERG1NP variants.

cell biology↗

bioRxiv2022

hERG1 channel subunit composition mediates proton inhibition of IKr in hiPSC CMs

hERG1 conducts cardiac IKr and is critical for repolarization of the human heart. Reduced IKr causes long QT syndrome and increases the risk for cardiac arrhythmia and sudden cardiac death. At least two subunits combine to form functional hERG1 channels, hERG1a and hERG1b. Changes in hERG 1a/1b subunit abundance modulates IKr kinetics, magnitude, and drug sensitivity. Studies from native cardiac tissue have suggested that hERG1 subunit abundance is dynamically regulated, but the impact of altered subunit abundance on IKr and its response to external stressors is not well understood. Here, we used a substrate-driven hiPSC-CM maturation model to investigate how changes in relative hERG 1a/1b subunit abundance impact the response of native IKr to extracellular acidosis, a known component of ischemic heart disease and sudden infant death syndrome. IKr recorded from immature hiPSC-CMs display a two-fold greater inhibition by extracellular acidosis (pH 6.3) compared to matured hiPSC-CMs. qRT-PCR and immunocytochemistry demonstrated that hERG1a subunit mRNA and protein were upregulated, and hERG1b subunit mRNA and protein were downregulated in matured hiPSC-CMs compared to immature hiPSC-CMs. The shift in subunit abundance in matured hiPSC-CMs was accompanied by an increased in IKr density. Silencing the impact of hERG1b on native IKr kinetics by overexpressing a polypeptide identical to the hERG1a PAS domain reduced the magnitude of IKr proton inhibition in immature hiPSC-CMs to levels comparable to those observed in matured hiPSC-CMs. These data demonstrate that hERG1 subunit abundance is dynamically regulated and that hERG1 subunit abundance determines IKr sensitivity to protons in hiPSC-CMs.

physiology↗

bioRxiv2022

KCNH2 encodes a nuclear-targeted polypeptide that mediates hERG1 channel gating and expression

KCNH2 encodes hERG1, the voltage-gated potassium channel that conducts the rapid delayed rectifier potassium current (IKr) in human cardiac tissue. hERG1 is one of the first channels expressed during early cardiac development, and its dysfunction is associated with intrauterine fetal death, sudden infant death syndrome, cardiac arrhythmia, and sudden cardiac death. Here, we identified a novel hERG1 polypeptide (hERG1NP) that is targeted to the nuclei of immature cardiac cells, including hiPSC-CMs and neonatal rat cardiomyocytes. The nuclear hERG1NP immunofluorescent signal is diminished in matured hiPSC-CMs and absent from adult rat cardiomyocytes. Antibodies targeting distinct hERG1 channel epitopes demonstrated that the hERG1NP signal maps to the hERG1 distal C-terminal domain. KCNH2 deletion using CRISPR simultaneously abolished IKr and the hERG1NP signal in hiPSC-CMs. We then identified a putative nuclear localization sequence (NLS) within the distal hERG1 C-terminus, 883-RQRKRKLSFR-892. Interestingly, the distal C-terminal domain was targeted almost exclusively to the nuclei when overexpressed HEK293 cells. Conversely, deleting the NLS from the distal peptide abolished nuclear targeting. Similarly, blocking or {beta}1 karyopherin activity diminished nuclear targeting. Finally, overexpressing the putative hERG1NP peptide in the nuclei of HEK cells significantly reduced hERG1a current density, compared to cells expressing the NLS-deficient hERG1NP or GFP. These data identify a developmentally regulated polypeptide encoded by KCNH2, hERG1NP, whose presence in the nucleus indirectly modulates hERG1 current magnitude and kinetics.

cell biology↗

bioRxiv2022

SNTA1 GeneRescues Ion Channel Function in Cardiomyocytes Derived from Induced Pluripotent Stem Cells Reprogrammed from Muscular Dystrophy Patients with Arrhythmias

Patients with cardiomyopathy of Duchenne Muscular Dystrophy (DMD) are at risk of developing life-threatening arrhythmias, but the mechanisms are unknown. We aimed to determine the role of cardiac ion channels controlling cardiac excitability in the mechanisms of arrhythmias in DMD patients. To test whether cardiac dystrophin mutations lead to defective NaV1.5-Kir2.1 channelosomes and arrhythmias, we generated iPSC-CMs from two hemizygous DMD males, a heterozygous female, and two unrelated controls. Two Patients had abnormal ECGs with frequent runs of ventricular tachycardia. iPSC-CMs from all DMD patients showed abnormal action potential profiles, slowed conduction velocities, and reduced sodium (INa) and inward rectifier potassium (IK1) currents. Membrane NaV1.5 and Kir2.1 protein levels were reduced in hemizygous DMD iPSC-CMs but not in heterozygous iPSC-CMs. Remarkably, transfecting just one component of the dystrophin protein complex (1-syntrophin) in hemizygous iPSC-CMs restored channelosome function, INa and IK1 densities and action potential profile. We provide the first demonstration that iPSC-CMs reprogrammed from skin fibroblasts of DMD patients with cardiomyopathy have a dysfunction of the NaV1.5-Kir2.1 channelosome, with consequent reduction of cardiac excitability and conduction. Altogether, iPSC-CMs from patients with DMD cardiomyopathy have a NaV1.5-Kir2.1 channelosome dysfunction, which can be rescued by the scaffolding protein 1-syntrophin to restore excitability.

cell biology↗

bioRxiv2021

From atoms to cells: bridging the gap between potency, efficacy, and safety of small molecules directed at a membrane protein

Membrane proteins constitute a substantial fraction of the human proteome, thus representing a vast source of therapeutic drug targets. Indeed, newly devised technologies now allow targeting "undruggable" regions of membrane proteins to modulate protein function in the cell. Despite the advances in technology, the rapid translation of basic science discoveries into potential drug candidates targeting transmembrane protein domains remains challenging. We address this issue by harmonizing single molecule-based and ensemble-based atomistic simulations of ligand-membrane interactions with patient-derived induced pluripotent stem cell (iPSC)-based experiments to gain insights into drug delivery, cellular efficacy, and safety of molecules directed at membrane proteins. In this study, we interrogated the pharmacological activation of the cardiac Ca2+ pump (Sarcoplasmic reticulum Ca2+-ATPase, SERCA2a) in human iPSC-derived cardiac cells as a proof-of-concept model. The combined computational-experimental approach serves as a platform to explain the differences in the cell-based activity of candidates with similar functional profiles, thus streamlining the identification of drug-like candidates that directly target SERCA2a activation in human cardiac cells. Systematic cell-based studies further showed that a direct SERCA2a activator does not induce cardiotoxic pro-arrhythmogenic events in human cardiac cells, demonstrating that pharmacological stimulation of SERCA2a activity is a safe therapeutic approach targeting the heart. Overall, this novel platform encompasses organ-specific drug potency, efficacy, and safety, and opens new avenues to accelerate the bench-to-patient research aimed at designing effective therapies directed at membrane protein domains.

pharmacology and toxicology↗