Atze van der Pol

Background: Clinical presentation of phospholamban (PLN) R14del cardiomyopathy is highly variable, from asymptomatic carriers to rapidly progressing heart failure. which cannot be fully explained by its known disturbance of sarcoplasmic reticulum calcium handling. Nuclear calcium signalling, shaped by sarcoplasmic reticulum calcium release, plays a key role in regulating gene expression and the cellular response to mechanical stress. However, how the R14del variant affects nuclear signalling under mechanical loading remains unclear. Objective: To uncover the impact of the PLN R14del variation on mechanosensing, nuclear calcium handling and its link to the variable clinical representation of this cardiomyopathy. Methods: Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying the PLN R14del variant, an isogenic control, and wildtype control were subjected to mechanical stimulation by cyclic stretch or cultured on substrates with physiological (10-20 kPa) and pathological (65-85 kPa) stiffness. Nuclear morphology, cytoskeletal organisation and nuclear body formation were assessed via immunofluorescence. Traction force microscopy was used to assess mechanical stress and force generation. Cardiac tissues from PLN R14del cardiomyopathy patients, dilated cardiomyopathy patients, and healthy controls were analysed under mechanical loading (at left ventricular assist device implantation) and unloading (at heart transplantation). Living myocardial slices from a PLN R14del patient were maintained under overloading or physiological loading conditions. Results: PLN R14del hiPSC-CMs showed impaired nuclear calcium handling and, under mechanical stress, exhibited impaired mechanotransduction and elevated nuclear bodies. A strain-avoidance response was triggered but failed to prevent apoptosis. Nuclear abnormalities were confirmed in patient myocardial slices and cardiac biopsies which normalized upon relative mechanical unloading. Conclusion: PLN R14del increases nuclear susceptibility to mechanical stress, linking nuclear dysfunction to new disease insights and highlighting potential for mechanobiology-based therapeutic interventions. Condensed PLN R14del cardiomyopathy presents with highly variable clinical severity, which cannot be fully explained by its known disturbance of sarcoplasmic reticulum calcium handling. We investigated whether the R14del variant impairs nuclear calcium signalling and mechanosensing, thereby contributing to disease heterogeneity. Using PLN R14del, isogenic control, and wildtype hiPSC-CMs subjected to cyclic stretch and substrates of physiological and pathological stiffness, we demonstrate that PLN R14del cardiomyocytes exhibit impaired nuclear calcium handling, elevated nuclear damage, disrupted cytoskeletal organisation, and failed mechanotransduction under mechanical stress. Although a strain-avoidance response was triggered, it was insufficient to prevent apoptosis. These findings were confirmed in living myocardial slices and cardiac biopsies from PLN R14del patients, where nuclear abnormalities were most pronounced under mechanical loading and reversed upon unloading. Together, these data establish increased nuclear susceptibility to mechanical stress as a novel pathological feature of PLN R14del cardiomyopathy, offering new insight into disease heterogeneity and highlighting mechanobiology-based therapeutic opportunities.

Background: Administrating extracellular matrix (ECM) to restore cardiac function post-myocardial infarction (MI) shows promise, however study variability obscures its true impact. We therefore conducted a systematic review and meta-analysis of preclinical studies to assess the effects of ECM treatments on cardiac function and tissue homeostasis post-MI. Methods: We searched PubMed and SCOPUS from inception to June 28, 2024, for animal studies describing ECM treatment post-MI (pre-registered on PROSPERO, CRD42022368400). Random effects meta-analyses compared ECM treatment to controls regarding left ventricular ejection fraction (LVEF), fractional shortening, infarct size, stroke volume, and left ventricular wall thickness. Subgroup analyses examined the influence of sex, species, ECM source, and administration method. Funnel plots and Egger’s regression assessed publication bias. Results: We identify 88 articles which meet our inclusion criteria. These studies describe the use of rats (51%), mice (38%), and pigs (11%). 44% of studies use males, 34% females, 5% both sexes, and 17% did not report sex. Most studies employ permanent MI models (85%) over ischemia reperfusion models (15%), and deliver ECM via intramyocardial injection (59%), cardiac patch (39%), cardiac sleeve (1%), or osmotic pump (1%). Our meta-analysis demonstrates that ECM treatment significantly improves LVEF (MD: 10.9%, 95% CI: [8.7%;13.0%]; p = 8.057e-24), fractional shortening (MD: 8.2%, 95% CI: [5.6%; 10.9%]; p = 1.751e-09), stroke volume (SMD 0.6, 95% CI: [0.2;1.0], p = 0.004), left ventricular wall thickening (SMD 1.2, 95% CI: [0.9; 1.5], p = 1.321e-17), while reducing infarct size (-11.7%, 95% CI: [-14.7%;-8.6%], p = 3.699e-14). We find no significant differences between the various subgroups and no indication of publication bias. Conclusions: ECM-based treatments significantly enhance cardiac function and tissue homeostasis in preclinical post-MI models, supporting further research toward clinical translation.

The switching of the fibroblast phenotype to myofibroblast is a hallmark of a wide variety of tissue pathologies. This phenotypical switch is known to be influenced not only by humoral factors such as TGF-β, but also by mechanical and physical cues in the cellular environment, and is accompanied by distinctive changes in cell morphology. However, the causative link between these cues, the concomitant morphological changes, and the resulting phenotypic switch remain elusive. Here, we use protein micropatterning to spatially control dermal fibroblast adhesion without invoking exogenous mechanical changes and demonstrate that varying the spatial configuration of focal adhesions (FAs) is sufficient to direct fibroblast phenotype. We further developed an automated morphometry analysis pipeline, which revealed FA eccentricity as the primary determinant of cell-state positioning along the spectrum of fibroblast phenotype. Moreover, linear fibronectin patterns that constrain the FAs were found to promote a further phenotype transition, characterized by dispersed expression of alpha-smooth muscle actin, pointing to an interesting possibility of controlling fibroblast phenotype beyond the canonical fibroblast–myofibroblast axis. Together, our study reveals that the spatial configuration of adhesion to the cellular microenvironment is a key factor governing fibroblast morphotype and phenotype, shedding new light on fibroblast phenotype regulation.

Purpose of Review: Cardiac tissue regenerative strategies have gained much traction over the years, in particular those utilizing hydrogels. With our review, and with special focus on supporting post-myocardial infarcted tissue, we aim to provide insights in determining crucial design considerations of a hydrogel and the implications these could have for future clinical use. Recent Findings: To date, two hydrogel delivery strategies are being explored, cardiac injection or patch, to treat myocardial infarction. Recent advances have demonstrated that the mechanism by which a hydrogel is gelated (i.e., physically or chemically cross-linked) not only impacts the biocompatibility, mechanical properties, and chemical structure, but also the route of delivery of the hydrogel and thus its effect on cardiac repair. Summary: With regard to cardiac regeneration, various hydrogels have been developed with the ability to function as a delivery system for therapeutic strategies (e.g., drug and stem cells treatments), as well as a scaffold to guide cardiac tissue regeneration following myocardial infarction. However, these developments remain within the experimental and pre-clinical realm and have yet to transition towards the clinical setting.

Tissue homeostasis is perturbed by stressful events, which can lead to organ dysfunction and failure. This is particularly true for the heart, where injury resulting from myocardial infarction or ischemic heart disease can result in a cascading event ultimately ending with the loss of functional myocardial tissue and heart failure. To help reverse this loss of healthy contractile tissue, researchers have spent decades in the hopes of characterizing a cell source capable of regenerating the injured heart. Unfortunately, these strategies have proven to be ineffective. With the goal of truly understanding cardiac regeneration, researchers have focused on the innate regenerative abilities of zebrafish and neonatal mammals. This has led to the realization that although cells play an important role in the repair of the diseased myocardium, inducing cardiac regeneration may instead lie in the composition of the extra cellular milieu, specifically the extra cellular matrix. In this review we will briefly summarize the current knowledge regarding cell sources used for cardiac regenerative approaches, since these have been extensively reviewed elsewhere. More importantly, by revisiting innate cardiac regeneration observed in zebrafish and neonatal mammals, we will stress the importance the extra cellular matrix has on reactivating this potential in the adult myocardium. Finally, we will address how we can harness the ability of the extra cellular matrix to guide cardiac repair thereby setting the stage of next generation regenerative strategies.

As the heart matures during embryogenesis from its foetal stages, several structural and functional modifications take place to form the adult heart. This process of maturation is in large part due to an increased volume and work load of the heart to maintain proper circulation throughout the growing body. In recent years, it has been observed that these changes are reversed to some extent as a result of cardiac disease. The process by which this occurs has been characterized as cardiac foetal reprogramming and is defined as the suppression of adult and re‐activation of a foetal genes profile in the diseased myocardium. The reasons as to why this process occurs in the diseased myocardium are unknown; however, it has been suggested to be an adaptive process to counteract deleterious events taking place during cardiac remodelling. Although still in its infancy, several studies have demonstrated that targeting foetal reprogramming in heart failure can lead to substantial improvement in cardiac functionality. This is highlighted by a recent study which found that by modulating the expression of 5‐oxoprolinase (OPLAH, a novel cardiac foetal gene), cardiac function can be significantly improved in mice exposed to cardiac injury. Additionally, the utilization of angiotensin receptor neprilysin inhibitors (ARNI) has demonstrated clear benefits, providing important clinical proof that drugs that increase natriuretic peptide levels (part of the foetal gene programme) indeed improve heart failure outcomes. In this review, we will highlight the most important aspects of cardiac foetal reprogramming and will discuss whether this process is a cause or consequence of heart failure. Based on this, we will also explain how a deeper understanding of this process may result in the development of novel therapeutic strategies in heart failure.

Advances in cardiovascular research have identified oxidative stress as an important pathophysiological pathway in the development and progression of heart failure. Oxidative stress is defined as the imbalance between the production of reactive oxygen species (ROS) and the endogenous antioxidant defence system. Under physiological conditions, small quantities of ROS are produced intracellularly, which function in cell signalling, and can be readily reduced by the antioxidant defence system. However, under pathophysiological conditions, the production of ROS exceeds the buffering capacity of the antioxidant defence system, resulting in cell damage and death. Over the last decades several studies have tried to target oxidative stress with the aim to improve outcome in patients with heart failure, with very limited success. The reasons as to why these studies failed to demonstrate any beneficial effects remain unclear. However, one plausible explanation might be that currently employed strategies, which target oxidative stress by exogenous inhibition of ROS production or supplementation of exogenous antioxidants, are not effective enough, while bolstering the endogenous antioxidant capacity might be a far more potent avenue for therapeutic intervention. In this review, we provide an overview of oxidative stress in the pathophysiology of heart failure and the strategies utilized to date to target this pathway. We provide novel insights into modulation of endogenous antioxidants, which may lead to novel therapeutic strategies to improve outcome in patients with heart failure.

The prevalence of heart failure with a preserved ejection fraction (HFpEF) is increasing, but therapeutic options are limited. Oxidative stress is suggested to play an important role in the pathophysiology of HFpEF. However, whether oxidative stress is a bystander due to comorbidities or causative in itself remains unknown. Recent results have shown that depletion of 5-oxoprolinase (OPLAH) leads to 5-oxoproline accumulation, which is an important mediator of oxidative stress in the heart. We hypothesize that oxidative stress induced by elevated levels of 5-oxoproline leads to the onset of a murine HFpEF-like phenotype. Oplah full body knock-out (KO) mice had higher 5-oxoproline levels coupled to increased oxidative stress. Compared to wild-type littermates (WT), KO mice had increased cardiac and renal fibrosis with concurrent elevated left ventricular filling pressures, impaired LV relaxation, yet a normal left ventricular ejection fraction (LVEF). Following the induction of cardiac ischemia/reperfusion (IR) injury, 52.4% of the KO mice died compared to only 15.4% of the WT mice (p < 0.03). Furthermore, KO mice showed a significantly increased atrial, ventricular, kidney, and liver weights compared to WT mice (p < 0.05 for all). Cardiac and renal fibrosis were more pronounced following cardiac IR injury in the KO mice and these mice developed proteinuria post IR injury. To further address the link between 5-oxoproline and HFpEF, 5-oxoproline was measured in the plasma of HFpEF patients. Compared to healthy controls (3.8 ± 0.6 µM), 5-oxoproline levels were significantly elevated in HFpEF patients (6.8 ± 1.9 µM, p < 0.0001). Furthermore, levels of 5-oxoproline were independently associated with more concentric remodeling on echocardiography. Oxidative stress induced by 5-oxoproline results in a murine phenotype reminiscent of the clinical manifestation of HFpEF without the need for surgical or pharmacological interference. Better understanding of the role of oxidative stress in HFpEF may potentially lead to novel therapeutic options.

In response to heart failure (HF), the heart reacts by repressing adult genes and expressing fetal genes, thereby returning to a more fetal-like gene profile. To identify genes involved in this process, we carried out transcriptional analysis on murine hearts at different stages of development and on hearts from adult mice with HF. Our screen identified Oplah, encoding for 5-oxoprolinase, a member of the γ-glutamyl cycle that functions by scavenging 5-oxoproline. OPLAH depletion occurred as a result of cardiac injury, leading to elevated 5-oxoproline and oxidative stress, whereas OPLAH overexpression improved cardiac function after ischemic injury. In HF patients, we observed elevated plasma 5-oxoproline, which was associated with a worse clinical outcome. Understanding and modulating fetal-like genes in the failing heart may lead to potential diagnostic, prognostic, and therapeutic options in HF.