Biology subjects

Boudina, S.

Publications and source records attributed to Boudina, S..

9 recordsLinked to original sources

bioRxiv2026

Perm1 Gene Therapy Mitigates PRDM16-Associated Cardiomyopathy

BackgroundPathogenic variants in PR domain containing 16 (PRDM16) cause pediatric and adult cardiomyopathies characterized by ventricular dilation, systolic dysfunction, and impaired metabolic maturation. Cardiac deficiency of PRDM16 alters metabolic gene expression and long-chain fatty acid (FA) metabolites. However, the downstream mediators involved are not well characterized. Furthermore, whether improving mitochondrial FA metabolism can prevent PRDM16-associated cardiomyopathy is currently unknown. MethodsIn vivo and in vitro approaches using patient-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) and mouse models with Prdm16 deletion/mutation were employed. Transcriptomics and proteomics analyses were conducted, and adeno-associated virus (AAV)-mediated therapy was tested. ResultsHere, we show that a defect in FA metabolism is an early hallmark of PRDM16 cardiac deficiency. We show, for the first time, that PERM1 is a direct downstream target of PRDM16 and is involved in the regulation of FA metabolism through coordinated action with PGC1. Most importantly, neonatal delivery of AAV9-Perm1 in cardiac-specific Prdm16 knockout (Prdm16 cKO) mice markedly improved contractile parameters, reduced left ventricular (LV) dilation, and extended survival. These cardioprotective effects of PERM1 gene therapy occurred independent of restoring FA oxidation. Transcriptional and proteomic analyses of AAV-Perm1-treated Prdm16 cKO mice demonstrated significant improvements in mitochondrial cristae architecture, preservation of sarcomere organization, reduced cardiomyocyte apoptosis, attenuated myocardial fibrosis, and diminished cardiac remodeling. ConclusionsWe identify PERM1 as a direct downstream effector of PRDM16 and uncover a previously unrecognized PRDM16-PGC1-PERM1 axis essential for FA metabolic regulation in the heart. Perm1 gene therapy ameliorated PRDM16-associated cardiomyopathy through post-transcriptional mechanisms involving preservation of mitochondrial and sarcomere integrity. The current study provides preclinical evidence suggesting that Perm1 gene therapy may be a promising therapeutic target to improve the cardiac outcomes of patients affected by pathogenic PRDM16 variants.

physiology↗

bioRxiv2026

Excessive Ca2+-dependent ER-mitochondrial contact stabilization by EFHD1 drives liver injury

Metabolic-associated steatohepatitis (MASH) involves hepatocyte damage that cannot be explained solely by lipid accumulation. Here, to discover injury-specific pathways, we focused on a gene of uncertain function, EF-Hand Domain Family Member D1 (EFHD1), identified in human genome-wide association studies of liver injury but not liver fat. We show that EFHD1, a Ca2+-dependent actin crosslinker, stabilizes endoplasmic reticulum-mitochondria contact sites (ERMCS), detecting spatiotemporal coincidence of inter-organellar proximity and ER Ca2+ release. During MASH, EFHD1 upregulation drives pathological mitochondrial fragmentation via excessive contact persistence. This structural failure promotes mitochondrial double-stranded RNA escape and activation of a maladaptive antiviral PKR-dependent stress response, a causal relationship also supported by Mendelian randomization in humans. Consequently, inhibiting EFHD1 in human and mouse models blunts hepatocyte damage. These findings identify EFHD1 as a Ca2+-dependent ERMCS stabilizer, reveal a hepatocyte-intrinsic injury pathway, and suggest EFHD1 inhibition as a therapeutic strategy.

cell biology↗

bioRxiv2025

EPHB2 promotes diet-induced MASH liver fibrosis

The EphB2 receptor tyrosine kinase is thought to participate in numerous fibroinflammatory disorders. In metabolic dysfunction-associated steatohepatitis (MASH), we find EphB2 becomes strongly overexpressed and overactive in hepatic stellate cells (HSCs) from humans with the disease and from mice fed liver-injuring high fat diets. Genetic deletion of EphB2 or inactivation of its tyrosine kinase catalytic domain suppressed diet-induced MASH fibrosis, while a kinase overactive point mutant displayed exacerbated steatosis and hepatic damage. Silencing EphB2 in primary HSCs dampened the ability of TGF-{beta}/SMAD signals to stimulate the transdifferentiation of stellate cells into profibrotic myofibroblasts, and HSC-specific deletion of the receptor, but not hepatocyte deletion, reduced liver scarring in multiple mouse models, even after fibrosis was established. Finally, a newly developed small molecule tetramerization inhibitor that targets EphB2-Ephrin receptor-ligand interactions effectively blunts inflammation and fibrosis in chemical and diet-induced liver injury models, demonstrating that therapeutically targeting EphB2 can counter MASH fibrosis.

pathology↗

bioRxiv2025

N-acetylcarnosine attenuates age-associated declines in multi-organ systems to improve survival

Histidine containing dipeptides (HCDs) such as N-acetylcarnosine are endogenous metabolites that are ergogenic and mitigate metabolic dysfunction. We previously demonstrated that short-term N-acetylcarnosine treatment is highly efficacious in protecting muscle atrophy induced by disuse. Here we demonstrate that a 6-months treatment of N-acetylcarnosine attenuates a broad spectrum of age-associated maladies and improved survival by [~]50% in female mice. A comprehensive survey of organ systems revealed that N-acetylcarnosine prevents decline in adiposity, diastolic function, vasodilation, muscle strength, and bone density. Together, N-acetylcarnosine substantially delays the onset of system-wide end-stage pathology to prolong lifespan. As an endogenously present metabolite, treatment with N-acetylcarnosine may be a safe and promising intervention to promote healthy aging in humans.

physiology↗

bioRxiv2023

A Fluorogenic-Based Assay to Measure Chaperone-Mediated Autophagic Activity in Cells and Tissues

ObjectivePathologies including cardiovascular diseases, cancer, and neurological disorders are caused by the accumulation of misfolded / damaged proteins. Intracellular protein degradation mechanisms play a critical role in the clearance of these disease-causing proteins. Chaperone mediated autophagy (CMA) is a protein degradation pathway that employs chaperones to bind proteins, bearing a unique KFERQ-like motif, for delivery to a CMA-specific Lysosome Associated Membrane Protein 2a (LAMP2a) receptor for lysosomal degradation. To date, steady-state CMA function has been assessed by measuring LAMP2A protein expression. However, this does not provide information regarding CMA degradation activity. To fill this dearth of tools / assays to measure CMA activity, we generated a CMA-specific fluorogenic substrate assay. MethodsA KFERQ-AMC [Lys-Phe-Asp-Arg-Gln-AMC(7-amino-4-methylcou-marin)] fluorogenic CMA substrate was synthesized from Solid-Phase Peptide Synthesis. KFERQ-AMC, when cleaved via lysosomal hydrolysis, causes AMC to release and fluoresce (Excitation:355 nm, Emission:460 nm). Using an inhibitor of lysosomal proteases, i.e., E64D [L-trans-Epoxy-succinyl-leucylamido(4-guanidino)butane)], responsible for cleaving CMA substrates, the actual CMA activity was determined. Essentially, CMA activity = (substrate)fluorescence - (substrate+E64D) fluorescence. To confirm specificity of the KFERQ sequence for CMA, negative control peptides were used. ResultsHeart, liver, and kidney lysates containing intact lysosomes were obtained from 4-month-old adult male mice. First, lysates incubated with KFERQ-AMC displayed a time dependent (0-5 hour) increase in AMC fluorescence vs. lysates incubated with negative control peptides. These data validate the specificity of KFERQ for CMA. Of note, liver exhibited the highest CMA (6-fold; p<0.05) > kidney (2.4-fold) > heart (0.4-fold) at 5-hours. Second, E64D prevented KFERQ-AMC degradation, substantiating that KFERQ-AMC is degraded via lysosomes. Third, cleavage of KFERQ-AMC and resulting AMC fluorescence was inhibited in Human embryonic kidney (HEK) cells and H9c2 cardiac cells transfected with Lamp2a vs. control siRNA. Further, enhancing CMA using Lamp2a adenovirus upregulated KFERQ degradation. These data suggest that LAMP2A is required for KFERQ degradation. Conclusion. We have generated a novel assay for measuring CMA activity in cells and tissues in a variety of experimental contexts. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/571785v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@7d43d8org.highwire.dtl.DTLVardef@1f11b2eorg.highwire.dtl.DTLVardef@2fa165org.highwire.dtl.DTLVardef@10d276d_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO Proposed mechanism of KFERQ-AMC degradation via CMA C_FIG

biochemistry↗

bioRxiv2021

Lipid hydroperoxides promote muscle atrophy through lysosomal amplification

Reactive oxygen species (ROS) accumulation is a cardinal feature of skeletal muscle atrophy. ROS refers to a collection of radical molecules whose cellular signals are vast, and it is unclear which downstream consequences of ROS are responsible for the loss of muscle mass and strength. Here we show that lipid hydroperoxides (LOOH) are increased with age and disuse, and the accumulation of LOOH by deletion of glutathione peroxidase 4 (GPx4) is sufficient to augment muscle atrophy. LOOH promoted atrophy in a lysosomal-dependent, proteasomal-independent manner. In young and old mice, genetic and pharmacologic neutralization of LOOH or their secondary reactive lipid aldehydes robustly prevented muscle atrophy and weakness, indicating that LOOH-derived carbonyl stress mediate age- and disuse-induced muscle dysfunction. Our findings provide novel insights for the role of LOOH in sarcopenia including a therapeutic implication by pharmacologic suppression.

physiology↗

bioRxiv2021

Activating P2Y1 receptors improves function in arteries with repressed autophagy

ObjectiveThe importance of endothelial cell (EC) autophagy to vascular homeostasis is evolving. Earlier we reported that purinergic 2Y1 receptor (P2Y1-R) activation rejuvenates shearstress induced nitric oxide (NO) generation in bovine aortic endothelial cells that is otherwise compromised after pharmacological and genetic autophagy repression. Here we determined the translational and functional relevance of these findings. Approach and ResultsFirst we assessed translational relevance using older humans and mice that exhibit blunted EC autophagy at rest together with impaired arterial function vs. appropriate controls. Rhythmic handgrip exercise elevated radial artery shear rate similarly in adult and older males for 60-min. Compared to baseline, autophagy initiation, p-eNOSS1177 activation, and NO generation, occurred in radial artery ECs from adult but not older subjects. Regarding mice, indexes of autophagy and p-eNOSS1177 activation were robust in ECs from adult but not older mice in response to 60-min treadmill-running. Next we questioned whether an inability to initiate EC autophagy precipitates arterial dysfunction. Age-associated reductions in intraluminal flow-mediated vasodilation observed in older vs. adult mice were recapitulated in arteries from adult mice by : (i) NO synthase inhibition; (ii) autophagy impairment using 3-methyladenine (3-MA); (iii) EC Atg3 depletion (iecAtg3KO mice); (iv) P2Y1-R blockade; and (v) germline depletion of P2Y1-Rs. Importantly, P2Y1-R activation using 2-methylthio-ADP (2-Me-ADP) improved vasodilatory capacity in arteries from : (i) adult mice treated with 3-MA; (ii) adult iecAtg3KO mice; and (iii) older animals with repressed EC autophagy. ConclusionsArterial dysfunction concurrent with pharmacological, genetic, and age-associated EC autophagy compromise is improved by activating P2Y1-Rs.

physiology↗

bioRxiv2021

Late-in-life treadmill-training rejuvenates autophagy, protein aggregate clearance, and function in mouse hearts.

ABSTRACT AND KEYWORDSThere is evidence for a progressive decline of protein quality control mechanisms during the process of cardiac aging. This enables the accumulation of protein aggregates and damaged organelles that contribute to age-associated cardiac dysfunction. Macroautophagy (referred to as autophagy) is the process by which post-mitotic cells such as cardiomyocytes clear defective proteins and organelles. We hypothesized that late-in-life exercise training improves autophagy, protein aggregate clearance, and function that is otherwise dysregulated in hearts from old vs adult mice. As expected, 24-month old male C57BL/6J mice (old) exhibited : (i) repressed autophagosome formation and protein aggregate accumulation in the heart; (ii) systolic and diastolic dysfunction; and (iii) reduced exercise capacity, vs. 8-month old (adult) mice (all p< .05). Separate cohorts of 21 month old mice completed a 3-month progressive resistance treadmill-running program (old-ETR) that improved (all < .05) : (i) body composition; (ii) exercise capacity; and (iii) soleus muscle citrate synthase activity, vs. age-matched mice that did not train (old-SED). Importantly, (iv) protein expression of autophagy markers indicated trafficking of the autophagosome to the lysosome increased, (v) protein aggregate clearance improved, and (vi) overall function was enhanced (all p<0.05), in hearts from old-ETR vs. old- SED mice. Dietary maneuvers and pharmacological interventions shown to elevate basal autophagy are reported to mitigate / reverse age-associated cardiac dysfunction. Here we show the first evidence that a physiological intervention initiated late-in-life improves autophagic flux, protein aggregate clearance, and overall function in mouse hearts.

physiology↗

bioRxiv2019

Mitochondrial PE potentiates respiratory enzymes to amplify skeletal muscle aerobic capacity

Exercise capacity is a strong predictor of all-cause mortality. Skeletal muscle mitochondrial respiratory capacity, its biggest contributor, adapts robustly to changes in energy demands induced by contractile activity. While transcriptional regulation of mitochondrial enzymes has been extensively studied, there is limited information on how mitochondrial membrane lipids are regulated. Herein, we show that exercise training or muscle disuse alters mitochondrial membrane phospholipids including phosphatidylethanolamine (PE). Addition of PE promoted, whereas removal of PE diminished, mitochondrial respiratory capacity. Surprisingly, skeletal muscle-specific inhibition of mitochondrial-autonomous synthesis of PE caused a respiratory failure due to metabolic insults in the diaphragm muscle. While mitochondrial PE deficiency coincided with increased oxidative stress, neutralization of the latter did not rescue lethality. These findings highlight the previously underappreciated role of mitochondrial membrane phospholipids in dynamically controlling skeletal muscle energetics and function.

physiology↗