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Abstract Despite important advancements in addressing cardiovascular diseases (CVDs), there has been an overall lack of progress in the field, leading to a slower decline in the rate of CVDs related deaths, and even an increase for some risk groups (e.g. increase in stroke mortality) exacerbated by an aging and obese population. While a multi-faceted problem, this deceleration may be influenced by the preferred model systems utilized in translation research. Cardiac cell lines, although easier to handle, lack biological accuracy due to the unnatural modifications required for successful culture and may not recapitulate complex 3-dimensional structural and environmental factors. At the same time, whole animal experimentation provides unwanted complexity during initial scientific development. Alternatively, ex vivo perfusion of isolated rodent hearts provides the needed biological accuracy with decreased organismal complexity. This platform facilitates the evaluation of the isolated heart, without neuro-reflexes and/or humoral contributions, unveiling the direct effects of stimuli in heart function/homeostasis. This manuscript leverages the wide array of perfusion parameters (i.e. perfusate, flow rate, coronary pressures), to demonstrate the capability of ex vivo heart perfusion protocols to accommodate a large range of experimental needs. Through this work, it was determined that the use of physiological perfusion pressures leads to increased left ventricular (LV) pressures but results in a loss of function over time, making it ideal conditions for organ assessment. Conversely, lower-than-physiological perfusion pressures lead to decreased LV pressures but prevent loss of function over time, which is preferable when longer perfusion times are relevant to experimental needs. Similarly, the use of adenosine as a pharmacological intervention was found to decrease both edema formation and inflammatory responses. In contrast, the use of packed red blood cells as oxygen carriers appears to induce a pro-inflammatory response and cause greater cardiac damage, particularly when combined with low perfusion pressures. 

Abstract Thermal ablation provides minimally invasive treatment for cardiovascular and cerebrovascular conditions but risks damaging healthy tissues due to their low imaging contrast against diseased areas. This study introduces an adaptive thermal ablation probe leveraging anisotropic magnetic heating of magnetite nanorods pre‐aligned within a polymer substrate. During magnetic pre‐alignment, the nanorods form chain‐like aggregates, enhancing their magnetic anisotropy and minimizing demagnetization effects. Under an alternating magnetic field, these features create a distinct difference in heat generation along the aggregates’ easy and hard axes. This probe utilizes a bimorph structure incorporating a heating layer with aligned nanorods and an actuation layer containing NdFeB microparticles. Exposure to static and alternating magnetic fields induces probe bending, adjusting nanorod orientation to modulate heat generation and prevent overheating. In vitro experiments demonstrate successful thrombus phantom ablation in both fluid flow and porcine artery models while preserving tissue viability. This innovative approach advances thermal ablation technology by offering a safer, more precise, and adaptive solution with a high potential for clinical translation. 

BackgroundDonation after circulatory death (DCD) grafts are vital for increasing available donor organs. Gradual rewarming during machine perfusion has proven effective in mitigating reperfusion injury and enhancing graft quality. Limited data exist on artificial oxygen carriers as an effective solution to meet the increasing metabolic demand with temperature changes. The aim of the present study was to assess the efficacy and safety of utilizing a hemoglobin-based oxygen carrier (HBOC) during the gradual rewarming of DCD rat livers. MethodsLiver grafts were procured after 30 min of warm ischemia. The effect of 90 min of oxygenated rewarming perfusion from ice cold temperatures (4 °C) to 37 °C with HBOC after cold storage was evaluated and the results were compared with cold storage alone. Reperfusion at 37 °C was performed to assess the post-preservation recovery. ResultsGradual rewarming with HBOC significantly enhanced recovery, demonstrated by markedly lower lactate levels and reduced vascular resistance compared to cold-stored liver grafts. Increased bile production in the HBOC group was noted, indicating improved liver function and bile synthesis capacity. Histological examination showed reduced cellular damage and better tissue preservation in the HBOC-treated livers compared to those subjected to cold storage alone. ConclusionThis study suggests the safety of using HBOC during rewarming perfusion of rat livers as no harmful effect was detected. Furthermore, the viability assessment indicated improvement in graft function. 

ABSTRACT We report on the design and fabrication of a novel circular pillar array as an interfacial barrier for microfluidic microphysiological systems (MPS). Traditional barrier interfaces, such as porous membranes and microchannel arrays, present limitations due to inconsistent pore size, complex fabrication and device assembly, and lack of tunability using a scalable design. Our pillar array overcomes these limitations by providing precise control over pore size, porosity, and hydraulic resistance through simple modifications of pillar dimensions. Serving as an interface between microfluidic compartments, it facilitates cell aggregation for tissue formation and acts as a tunable diffusion barrier that mimics diffusion in vivo. We demonstrate the utility of barrier design to engineer physiologically relevant cardiac microtissues and a heterotypic model with vasculature within the device. Its tunable properties offer significant potential for drug screening/testing and disease modeling, enabling comparisons of drug permeability and cell migration in MPS tissue with or without vasculature. 

Abstract Brief pulses of electric field (electroporation) and/or tensile stress (mechanoporation) have been used to reversibly permeabilize the plasma membrane of mammalian cells and deliver materials to the cytosol. However, electroporation can be harmful to cells, while efficient mechanoporation strategies have not been scalable due to the use of narrow constrictions or needles which are susceptible to clogging. Here we report a high throughput approach to mechanoporation in which the plasma membrane is stretched and reversibly permeabilized by viscoelastic fluid forces within a microfluidic chip without surface contact. Biomolecules are delivered directly to the cytosol within seconds at a throughput exceeding 250 million cells per minute. Viscoelastic mechanoporation is compatible with a variety of biomolecules including proteins, RNA, and CRISPR-Cas9 ribonucleoprotein complexes, as well as a range of cell types including HEK293T cells and primary T cells. Altogether, viscoelastic mechanoporation appears feasible for contact-free permeabilization and delivery of biomolecules to mammalian cells ex vivo. 

Abstract Heart failure, a leading global health challenge, affects over 23 million people worldwide, with heart transplantation being the gold standard for end-stage disease. However, the scarcity of viable donor hearts presents a significant barrier, with only one-third of available grafts used due to stringent selection criteria. Machine perfusion technologies, particularly normothermic machine perfusion (NMP), offer promise in improving graft preservation and assessment, yet their full potential for predicting transplantability remains underexplored. This study investigates three assessment methods to enhance human heart evaluation during NMP, focusing on mitochondrial function, left ventricular (LV) performance, and inflammatory markers. First, resonance Raman spectroscopy (RRS) is employed to assess mitochondrial redox state as a proxy for metabolic competency, offering a non-invasive and dynamic evaluation of mitochondrial function during ex vivo preservation. Second, LV function is quantified using intraventricular balloons, providing critical insights into graft viability and performance. Third, inflammatory markers and endothelial activation are assessed from perfusate to predict post-transplant outcomes. These methods were tested on human donor hearts declined for transplantation, preserved via static cold storage (SCS) and subsequently assessed with NMP in Langendorff mode. The results demonstrate that these parameters can be easily integrated into existing clinical perfusion workflows and hold potential for improving heart transplantation outcomes by enhancing graft selection and optimizing donor heart use. Future studies will further validate these biomarkers across different preservation techniques and evaluate their clinical applicability. 

Abstract Cryopreservation of biological matter nigh‐universally requires cooling organic aqueous solutions through the metastable supercooled temperature regime. However, for many solutions of interest, very little thermophysical data exists at these temperatures, hampering the meaningful design of cryopreservation protocols or interrogation of cryopreservation processes. Digital holography interferometry (DHI) has recently emerged as a promising technique for measuring and characterizing the physical properties of liquids. Herein, DHI is proposed as a thermal imaging and characterization tool capable of measuring optical properties, such as temperature‐dependent refractive index, and providing high‐resolution two‐dimensional (2D) visualization of temperature distribution in the supercooling regime. This approach overcomes the limitations of traditional techniques such as thermocouples and thermal cameras. This work describes a DHI platform instrumented with a custom thermoelectric supercooling apparatus, demonstrates 2D temperature mapping in the supercooled regime, and measures the refractive index of water, 10% dimethyl sulfoxide (DMSO), and 49% DMSO solutions at sub‐273 K temperatures. Overall, DHI is demonstrated as a noninvasive optical tool with high resolution for studying supercooled liquids, potentially enhancing cryopreservation protocols and providing new thermo‐optical information on cryoprotective agents at sub‐273 K temperatures. 

Abstract Vascularized composite allotransplantations are complex procedures with substantial functional impact on patients. Extended preservation of VCAs is of major importance in advancing this field. It would result in improved donor-recipient matching as well as the potential for ex vivo manipulation with gene and cell therapies. Moreover, it would make logistically feasible immune tolerance induction protocols through mixed chimerism. Supercooling techniques have shown promising results in multi-day liver preservation. It consists of reaching sub-zero temperatures while preventing ice formation within the graft by using various cryoprotective agents. By drastically decreasing the cell metabolism and need for oxygen and nutrients, supercooling allows extended preservation and recovery with lower ischemia–reperfusion injuries. This study is the first to demonstrate the supercooling of a large animal model of VCA. Porcine hindlimbs underwent 48 h of preservation at − 5 °C followed by recovery and normothermic machine perfusion assessment, with no issues in ice formation and favorable levels of injury markers. Our findings provide valuable preliminary results, suggesting a promising future for extended VCA preservation. 

Abstract BackgroundPercutaneous coronary interventions (PCIs) within left main coronary arteries are high-risk procedures that require optimization of interactions between stent(s) and diseased vessels. Optical Coherence Tomography (OCT) is a widely accepted tool that enhances physicians’ ability to assess proper stent appositions during clinical procedures. The primary aim of this study was to develop complementary post-procedure imaging methodologies to better assess and interpret outcomes of left main PCI procedures, utilizing both reanimated and perfusion-fixed human hearts. MethodsPCIs were performed while obtaining OCT scans within the left main anatomies of six human hearts. Subsequently, each heart was scanned with a micro-CT scanner with optimized parameters to achieve resolutions up to 20 µm. Scans were reconstructed and imported into a DICOM segmentation software to generate computational models of implanted stents and associated coronary vessels. 2D images from OCT that were obtained during PCIs were compared to the 3D models generated from micro-CT reconstructions. In addition, the 3D models were utilized to create virtual reality scenes and enlarged 3D prints for development of “mixed reality” tools relative to bifurcation stenting within human left main coronary arteries. ResultsWe developed reproducible methodologies for post-implant analyses of coronary artery stenting procedures. In addition, we generated high-resolution 3D computational models, with ~ 20-micron resolutions, of PCIs performed within reanimated and perfusion-fixed heart specimens. ConclusionsGenerated computational models of left main PCIs performed in isolated human hearts can be used to obtain detailed measurements that provide further clinical insights on procedural outcomes. The 3D models from these procedures are useful for generating virtual reality scenes and 3D prints for physician training and education. 

Abstract Red blood cell (RBC) transfusions facilitate many life-saving acute and chronic interventions. Transfusions are enabled through the gold-standard hypothermic storage of RBCs. Today, the demand for RBC units is unfulfilled, partially due to the limited storage time, 6 weeks, in hypothermic storage. This time limit stems from high metabolism-driven storage lesions at +1-6 °C. A recent and promising alternative to hypothermic storage is the supercooled storage of RBCs at subzero temperatures, pioneered by our group. Here, we report on long-term supercooled storage of human RBCs at physiological hematocrit levels for up to 23 weeks. Specifically, we assess hypothermic RBC additive solutions for their ability to sustain supercooled storage. We find that a commercially formulated next-generation solution (Erythro-Sol 5) enables the best storage performance and can form the basis for further improvements to supercooled storage. Our analyses indicate that oxidative stress is a prominent time- and temperature-dependent injury during supercooled storage. Thus, we report on improved supercooled storage of RBCs at −5 °C by supplementing Erythro-Sol 5 with the exogenous antioxidants, resveratrol, serotonin, melatonin, and Trolox. Overall, this study shows the long-term preservation potential of supercooled storage of RBCs and establishes a foundation for further improvement toward clinical translation.