Biology subjects

Castillero, E.

Publications and source records attributed to Castillero, E..

4 recordsLinked to original sources

bioRxiv2025

In vivo imaging of reactive oxygen species after myocardial ischemia-reperfusion injury: a large animal multimodal imaging and transcriptomic study

BackgroundReactive oxygen species (ROS) contribute to myocardial ischemia-reperfusion injury (IRI), but in-vivo data on the spatial myocardial distribution and systemic effects of ROS after IRI remain limited. This multimodal CMR and PET/CT study aimed to non-invasively image ROS activity in a clinically-relevant swine model of IRI using [18F]ROStrace, a fluorine-18-labeled analogue of dihydroethidium (DHE), and to investigate regional changes in ROS activity in the infarcted myocardium during the subacute post-IRI phase. MethodsIRI was induced by percutaneous occlusion of the left anterior descending artery for 90 minutes in swine (N=9). CMR and whole-body PET/CT imaging with [18F]ROStrace were performed before myocardial infarction (MI) and 3-5 days post-MI to assess ROS in non-infarct myocardium, lungs, bone marrow, spleen and skeletal muscle. Late gadolinium enhanced CMR was performed to structurally characterize infarct regions. Post-MI, in vivo [18F]ROStrace signal in infarcted myocardium was compared with remote, non-infarcted myocardium and validated via ex vivo DHE fluorescent imaging. Bulk RNA-sequencing (RNA-seq) and Gene Ontology pathway analysis were conducted on biopsies from infarct and remote myocardial tissue to identify differentially expressed genes and pathways connected to oxidative stress. ResultsDuring the subacute phase following MI, [18F]ROStrace fractional uptake rate (FUR; min-1) was significantly increased in skeletal muscle, compared to baseline (0.011{+/-}0.003 vs 0.016{+/-}0.005, p=0.04), with a trend toward increased FUR in bone marrow (0.046{+/-}0.009 vs 0.056{+/-}0.011, p=0.12) and the left ventricular free wall (0.067{+/-}0.007 vs 0.073{+/-}0.010, p=0.15). Within the myocardium, [18F]ROStrace FUR ((min-1)/(mL/min/g)) was significantly higher in infarcted compared to non-infarcted myocardium regions (0.110{+/-}0.034, vs 0.148{+/-}0.035, p=0.0005). DHE staining confirmed elevated ROS levels in the infarcted myocardium. RNA-seq identified 8,707 differentially expressed genes between infarct and remote myocardium, with downregulated pathways in the infarct associated with mitochondrial function, cellular respiration, and metabolic adaptation. ConclusionThis study demonstrated MI ROS imaging using [18F]ROStrace using a whole-body PET/CT scanner and structural assessment with CMR. Systemic and myocardial increases in ROS activity were observed post-MI, accompanied by substantial molecular alterations in infarcted tissue. These findings show potential imaging strategies to evaluate therapeutic targets that can mitigate oxidative stress after MI.

bioengineering↗

bioRxiv2021

RAGE antagonist peptide mitigates AGE-mediated endothelial hyperpermeability and accumulation of glycoxidation products in human ascending aortas and in a murine model of aortic aneurysm

BackgroundAortic dissection and aneurysm are the result of altered biomechanical forces associated with structural weakening of the aortic wall caused by genetic or acquired factors. Current guidelines recommend replacement of the ascending aorta when the diameter is >5.5 cm in tricuspid aortic valve patients. Aortopathies are associated with altered wall stress and stiffness as well as endothelial cell dysfunction and synthetic vascular smooth muscle cell (VSMC) phenotype. We reported that these mechanisms are mediated by glycoxidation products [Reactive oxygen species (ROS) and Advance Glycation End products (AGE)]. This study addresses the role of glycoxidation on endothelial function and AGE-mediated aortic stiffness. Hypothesis and aimsHere we investigate how circulating glycation products infiltrate the aortic wall via AGE-mediated endothelial hyperpermeability and contribute to both VSMC synthetic phenotype and extracellular matrix (ECM) remodeling in vivo and ex vivo. We also study how RAGE antagonist peptide (RAP) can rescue the effect of AGEs in vitro and in vivo in eNOS-/- vs WT mice. Methods and resultsHuman ascending aortas (n=30) were analyzed for AGE, ROS, and ECM markers. In vitro glycation was obtained by treating VSMC or human and murine aortas with glyoxal. Endothelial permeability was measured under glycation treatment. Vascular stiffness was measured by a pressure myograph comparing wild-type mice {+/-} glyoxal. eNOS-/- mice, a model of increased endothelial permeability, were treated for 28 days with hyperlipidemic diet {+/-} Angiotensin II (1000ng/kg/min) with or without anti-glycation treatment (RAP 20mg/kg). Echo data of aortic diameter were collected. Murine vascular stiffness was measured by a pressure myograph (n=5/group). Glycoxidation products were detected in all human aortas independently of aortic diameter, with stronger accumulation on the lumen and the adventitia layer. AGEs increased endothelial permeability, induce synthetic phenotypic switch in human VSMCs, and inhibit cell migration. RAP pre-treatment rescue the effect of glyoxal on endothelial cells. Ex vivo glycation treatment of murine arteries impacted on ECM and increased stiffness. Aortic stiffness was higher in eNOS-/- vs WT mice. Ang II-mediated aortopathies results in aortic dilation, and AGE/ROS accumulation, which is rescued by RAGE antagonist peptide treatment of eNOS-/- mice. ConclusionsGlycoxidation reaction mediate EC permeability, VSMCs phenotype, and ECM remodeling leading to dysfunctional microstructure of the ascending aorta, altered vascular stiffness and increasing aortic susceptibility to dilation and rupture. Moreover, we show that RAP can mitigate AGE-mediated endothelial hyper-permeability in vitro and impact on ascending aneurysm in vivo

physiology↗

bioRxiv2020

Inhibition and down regulation of the serotonin transporter contribute to the progression of degenerative mitral regurgitation

AimsHeart valve disease attributed to serotonin (5HT) has been observed with 5HT-secreting carcinoid tumors and in association with medications, such as the diet drug, Dexfenfluoramine, a serotonin transporter (SLC6A4) inhibitor and 5HT receptor (HTR) 2B agonist. HTR2B signaling upregulates TGF{beta}-1 resulting in increased production of extracellular matrix proteins. SLC6A4 internalizes 5HT, limiting HTR signaling. Selective 5HT reuptake inhibitors (SSRI), widely used antidepressants, target SLC6A4, thus enhancing HTR signaling. However, 5HT and SLC6A4 mechanisms have not been previously associated with degenerative mitral regurgitation (MR). The present studies investigated the hypothesis that both dysregulation of SLC6A4 and inhibition of SLC6A4 contribute to the pathophysiology of MR. Methods and ResultsHere we report SLC6A4 related studies of 225 patients with MR requiring surgery. A multivariate analysis showed that SSRI use in MR patients was associated with surgery at a younger age, indicating more rapidly progressive MR (p=0.0183); this was confirmed in a national dataset (p<0.001). Aspirin use by MR patients was associated with surgery at an older age (p=0.0055). Quantitative reverse transcriptase PCR of MR leaflet RNA from 44 patients, and 20 normal mitral leaflets from heart transplant recipients, demonstrated down regulation in MR of both SLC6A4 and vesicular monoamine transporter-2 (SLC18A2), that packages 5HT (p<0.001). Human mitral valve interstitial cells cultivated with Fluoxetine, a SSRI, demonstrated down regulation of SLC6A4 and upregulation of HTR2B, compared to untreated, in cells from both normal and MR leaflets. Platelet 5HT studies in healthy subjects without heart disease used ADP-induced activation to model MR-associated activation. Fluoxetine significantly increased platelet activation and plasma 5HT levels, while Aspirin inhibited ADP platelet activation. ConclusionsDown regulation and inhibition of SLC6A4 influences MR through enhanced HTR signaling. SSRI may further influence MR through inhibition and down regulation of SLC6A4, upregulation of HTR2B, and increased platelet release of 5HT. Translational PerspectiveDegenerative mitral valve regurgitation (MR) affects millions, and there is no medical therapy for this disease. MR becomes progressively worse, and for severe MR, the only option is cardiac surgery. Serotonin (5HT) is best known as a neurotransmitter. However, 5HT secreting carcinoid tumors cause a cardiac valve disorder in many cases, and 5HT related medications, such as the diet drug Fenfluoramine, have been associated with the development of cardiac valve disease. The present paper presents evidence that diminished serotonin transporter (SLC6A4) expression and inhibition, lead to increased 5HT receptor signaling, contributing to the progression of MR.

physiology↗

bioRxiv2019

Mitral valve leaflet response to ischemic mitral regurgitation: From gene expression to tissue remodeling

AimsIschemic mitral regurgitation is frequently observed following myocardial infarction and is associated with higher mortality and poor clinical prognosis if left untreated. Accumulating evidence suggests that mitral valve leaflets actively remodel post-myocardial infarction, yet the cellular mechanisms underlying these responses and how this affects tissue function remain largely unknown. We sought to elucidate mitral valve remodeling post myocardial infarction at the tissue, cellular, and transcriptomic levels. Methods and ResultsThe mechanical behavior of ovine mitral valve leaflets pre- and 8 weeks post- myocardial infarction reveal a significant decrease in radial direction extensibility, which essentially eliminated the mechanical anisotropy typically observed in healthy mitral valves. Quantitative histology and ultrastructural assessment by transmission electron microscopy revealed altered leaflet composition and architecture at 8 weeks post-myocardial infarction. Assessment of the mitral valve interstitial cell nuclear aspect ratio, a metric of cellular deformation, revealed that they were on average rounder following myocardial infarction. RNA sequencing indicated that YAP-induced genes were elevated at 4 weeks post-myocardial infarction and genes related to extracellular matrix organization were some of the most downregulated in sheep with IMR compared to sheep without ischemic mitral regurgitation at 4 weeks post-myocardial infarction. Additionally, RNA sequencing revealed the possible recruitment of immune cells in this remodeling process due to the drastic elevation of CXCL9 and CLEC10A. ConclusionsOur multiscale assessment revealed significant mechanical and microstructural changes due to myocardial infarction. RNA sequencing provided a baseline for global gene expression changes in response to myocardial infarction with and without ischemic mitral regurgitation and suggests YAP-induced mechanotransduction, altered expression of extracellular matrix-related genes, and recruitment of immune cells as mechanisms contributing to altered mitral valve biomechanics post-myocardial infarction.

bioengineering↗