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Bovine pericardium (BP) is a vascular biomaterial used in cardiovascular surgery that is typically cross-linked for masking antigenicity and enhance stability. There is a need for biochemical evaluation of the tissue properties prior to implantation to ensure that quality and reliability standards are met. Here, engineered antigen removed BP (ARBP) that was cross-linked with 0.2% and 0.6% glutaraldehyde (GA), and further calcified in vitro to simulate graft calcifications upon implantation was characterized nondestructively using fluorescence lifetime imaging (FLIm) to identify regions of interest which were then assessed by Raman spectroscopy. We observed that the tissue fluorescence lifetime shortened, and that Raman bands at 856, 935, 1282, and 1682 cm-1 decreased, and at 1032 and 1627 cm-1 increased with increasing GA cross-linking. Independent classification analysis based on fluorescence lifetime and on Raman spectra discriminated between GA-ARBP and untreated ARBP with an accuracy of 91% and 66%, respectively. Pearson's correlation analysis showed a strong correlation between pyridinium cross-links measured with high-performance liquid chromatography and fluorescence lifetime measured at 380-400 nm (R = -0.76, p = 0.00094), as well as Raman bands at 856 cm-1 for hydroxy-proline (R = -0.68, p = 0.0056) and at 1032 cm-1 for hydroxy-pyridinium (R = 0.74, p = 0.0016). Calcified areas of GA cross-linked tissue showed characteristic hydroxyapatite (959 and 1038 cm-1) bands in the Raman spectrum and fluorescence lifetime shortened by 0.4 ns compared to uncalcified regions. FLIm-guided Raman imaging could rapidly identify degrees of cross-linking and detected calcified regions with high chemical specificity, an ability that can be used to monitor tissue engineering processes for applications in regenerative medicine.

The extracellular matrix architecture of bovine pericardium (BP) has distinct biochemical and biomechanical properties that make it a useful biomaterial in the field of regenerative medicine. Collagen represents the dominant structural protein of BP and is therefore intimately associated with the properties of this biomaterial. Enzymatic degradation of collagen molecules is critical for extracellular matrix turnover, remodeling and ultimately tissue regeneration. We present a quantitative, label-free and non-destructive method for monitoring changes in biochemical and biomechanical properties of BP during tissue degradation, based on multi-spectral fluorescence lifetime imaging (ms-FLIm). Strong correlations of fluorescence intensity ratio and average fluorescence lifetime were identified with collagen content, Young's Modulus and Ultimate tensile strength during collagenase degradation, indicating the potential of optically monitoring collagen degradation using ms-FLIm. The obtained results demonstrate the value of ms-FLIm to assess the quality of biomaterials in situ for applications in regenerative medicine.

New techniques able to monitor the maturation of tissue engineered constructs over time are needed for a more efficient control of developmental parameters. Here, a label-free fluorescence lifetime imaging (FLIm) approach implemented through a single fiber-optic interface is reported for nondestructive in situ assessment of vascular biomaterials. Recellularization processes of antigen removed bovine pericardium scaffolds with endothelial cells and mesenchymal stem cells were evaluated on the serous and the fibrous sides of the scaffolds, 2 distinct extracellular matrix niches, over the course of a 7 day culture period. Results indicated that fluorescence lifetime successfully report cell presence resolved from extracellular matrix fluorescence. The recellularization process was more rapid on the serous side than on the fibrous side for both cell types, and endothelial cells expanded faster than mesenchymal stem cells on antigen-removed bovine pericardium. Fiber-based FLIm has the potential to become a nondestructive tool for the assessment of tissue maturation by allowing in situ imaging of intraluminal vascular biomaterials.

Heart failure (HF) following myocardial infarction (MI) is associated with high incidence of cardiac arrhythmias. Development of therapeutic strategy requires detailed understanding of electrophysiological remodeling. However, changes of ionic currents in ischemic HF remain incompletely understood, especially in translational large-animal models. Here, we systematically measure the major ionic currents in ventricular myocytes from the infarct border and remote zones in a porcine model of post-MI HF. We recorded eight ionic currents during the cell's action potential (AP) under physiologically relevant conditions using selfAP-clamp sequential dissection. Compared with healthy controls, HF-remote zone myocytes exhibited increased late Na+ current, Ca2+-activated K+ current, Ca2+-activated Cl- current, decreased rapid delayed rectifier K+ current, and altered Na+/Ca2+ exchange current profile. In HF-border zone myocytes, the above changes also occurred but with additional decrease of L-type Ca2+ current, decrease of inward rectifier K+ current, and Ca2+ release-dependent delayed after-depolarizations. Our data reveal that the changes in any individual current are relatively small, but the integrated impacts shift the balance between the inward and outward currents to shorten AP in the border zone but prolong AP in the remote zone. This differential remodeling in post-MI HF increases the inhomogeneity of AP repolarization, which may enhance the arrhythmogenic substrate. Our comprehensive findings provide a mechanistic framework for understanding why single-channel blockers may fail to suppress arrhythmias, and highlight the need to consider the rich tableau and integration of many ionic currents in designing therapeutic strategies for treating arrhythmias in HF.

Current clinically used electronic implants, including cardiac pacing leads for epicardial monitoring and stimulation of the heart, rely on surgical suturing or direct insertion of electrodes to the heart tissue. These approaches can cause tissue trauma during the implantation and retrieval of the pacing leads, with the potential for bleeding, tissue damage, and device failure. Here, we report a bioadhesive pacing lead that can directly interface with cardiac tissue through physical and covalent interactions to support minimally invasive adhesive implantation and gentle on-demand removal of the device with a detachment solution. We developed 3D-printable bioadhesive materials for customized fabrication of the device by graft-polymerizing polyacrylic acid on hydrophilic polyurethane and mixing with poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) to obtain electrical conductivity. The bioadhesive construct exhibited mechanical properties similar to cardiac tissue and strong tissue adhesion, supporting stable electrical interfacing. Infusion of a detachment solution to cleave physical and covalent cross-links between the adhesive interface and the tissue allowed retrieval of the bioadhesive pacing leads in rat and porcine models without apparent tissue damage. Continuous and reliable cardiac monitoring and pacing of rodent and porcine hearts were demonstrated for 2 weeks with consistent capture threshold and sensing amplitude, in contrast to a commercially available alternative. Pacing and continuous telemetric monitoring were achieved in a porcine model. These findings may offer a promising platform for adhesive bioelectronic devices for cardiac monitoring and treatment. Editor's summary: Temporary epicardial pacing leads provide short-term monitoring and stimulation for cardiac electrophysiological abnormalities. Clinically adopted pacing leads involve puncture of the cardiac tissue by direct electrode insertion or suture. Deng et al. developed a 3D-printed adhesive polymeric pacing lead that supported minimally invasive implantation on the epicardial surface and on-demand detachment to reduce the potential for tissue damage. In rat and porcine models, the adhesive pacing lead demonstrated strong adhesion to cardiac tissue and did not cause macroscopic tissue damage or bleeding with adhesion or on-demand removal. The device integrated with existing clinical pacing equipment and provided a stable electrical interface for cardiac monitoring and pacing for 10 to 14 days in rat and porcine models. These studies provide proof of principle for the use of printable adhesive electronics for cardiac pacing applications. —Molly Ogle [ABSTRACT FROM AUTHOR]

Extracellular matrix (ECM) has emerged as a dynamic tissue component, providing not only structural support, but also functionally participating in a wide range of signaling events during development, injury, and disease remodeling. Investigation of dynamic changes in cardiac ECM proteome is challenging due to the relative insolubility of ECM proteins, which results from their macromolecular nature, extensive post-translational modification (PTM), and tendency to form protein complexes. Finally, the relative abundance of cellular and mitochondrial proteins in cardiac tissue further complicates cardiac ECM proteomic approaches. Recent developments of various techniques to enrich and analyze ECM proteins are playing a major role in overcoming these challenges. Application of cardiac ECM proteomics in disease tissues can further provide spatial and temporal information relevant to disease diagnosis, prognosis, treatment, and engineering of therapeutic candidates for cardiac repair and regeneration.

BackgroundAlthough the incidence of acute death related to coronary artery disease has decreased with the advent of new interventional therapies, myocardial infarction remains one of the leading causes of death in the US. Current animal models developed to replicate this phenomenon have been associated with unacceptably high morbidity and mortality. A new model utilizing the first diagonal branch of the left anterior descending artery (D1-LAD) was developed to provide a clinically relevant lesion, while attempting to minimize the incidence of adverse complications associated with infarct creation.MethodsEight Yucatan miniature pigs underwent percutaneous embolization of the D1-LAD via injection of 90 µm polystyrene micro-spheres. Cardiac structure and function were monitored at baseline, immediately post-operatively, and at 8-weeks post-infarct using transthoracic echocardiography. Post-mortem histopathology and biochemical analyses were performed to evaluate for changes in myocardial structure and extracellular matrix (ECM) composition respectively. Echocardiographic data were evaluated using a repeated measures analysis of variance followed by Tukey's HSD post hoc test. Biochemical analyses of infarcted to non-infarcted myocardium were compared using analysis of variance.ResultsAll eight pigs successfully underwent echocardiography prior to catheterization. Overall procedural survival rate was 83% (5/6) with one pig excluded due to failure of infarction and another due to deviation from protocol. Ejection fraction significantly decreased from 69.7 ± 7.8% prior to infarction to 50.6 ± 14.7% immediately post-infarction, and progressed to 48.7 ± 8.9% after 8-weeks (p = 0.011). Left ventricular diameter in systole significantly increased from 22.6 ± 3.8 mm pre-operatively to 30.9 ± 5.0 mm at 8 weeks (p = 0.016). Histopathology showed the presence of disorganized fibrosis on hematoxylin and eosin and Picro Sirius red stains. Collagen I and sulfated glycosaminoglycan content were significantly greater in the infarcted region than in normal myocardium (p = 0.007 and p = 0.018, respectively); however, pyridinoline crosslink content per collagen I content in the infarcted region was significantly less than normal myocardium (p = 0.048).ConclusionSystolic dysfunction and changes in ECM composition induced via embolization of the D1-LAD closely mimic those found in individuals with chronic myocardial infarction (MI), and represents a location visible without the need for anesthesia. As a result, this method represents a useful model for studying chronic MI.

Knowledge and resources derived from veterinary medicine represent an underused resource that could serve as a bridge between data obtained from diseases models in laboratory animals and human clinical trials. Naturally occurring disease in companion animals that display the defining attributes of similar, if not identical, diseases in humans hold promise for providing predictive proof of concept in the evaluation of new therapeutics and devices. Here we outline comparative aspects of naturally occurring diseases in companion animals and discuss their current uses in translational medicine, benefits, and shortcomings. Last, we envision how these natural models of disease might ultimately decrease the failure rate in human clinical trials and accelerate the delivery of effective treatments to the human clinical market.

Aims In heart failure (HF), pulmonary venous hypertension (PVH) produces pulmonary hypertension (PH) with remodeling of pulmonary veins (PV) and arteries (PA). In a porcine PVH model, we performed proteomic-based bioinformatics to investigate unique pathophysiologic mechanisms mediating PA and PV remodeling. Methods and results Large PV were banded (PVH, n  = 10) or not (Sham, n  = 9) in piglets. At sacrifice, PV and PA were perfusion labelled for vessel-specific histology and proteomics. The PA and PV were separately sampled with laser-capture micro-dissection for mass spectrometry. Pulmonary vascular resistance [Wood Units; 8.6 (95% confidence interval: 6.3, 12.3) vs. 2.0 (1.7, 2.3)] and PA [19.9 (standard error of mean, 1.1) vs. 10.3 (1.1)] and PV [14.2 (1.2) vs. 7.6 (1.1)] wall thickness/external diameter (%) were increased in PVH (P  < 0.05 for all). Similar numbers of proteins were identified in PA (2093) and PV (2085) with 94% overlap, but biological processes differed. There were more differentially expressed proteins (287 vs. 161), altered canonical pathways (17 vs. 3), and predicted upstream regulators (PUSR; 22 vs. 6) in PV than PA. In PA and PV, bioinformatics indicated activation of the integrated stress response and mammalian target of rapamycin signalling with dysregulated growth. In PV, there was also activation of Rho/Rho-kinase signalling with decreased actin cytoskeletal signalling and altered tight and adherens junctions, ephrin B, and caveolae-mediated endocytosis signalling; all indicating disrupted endothelial barrier function. Indeed, protein biomarkers and the top PUSR in PV (transforming growth factor-beta) suggested endothelial to mesenchymal transition in PV. Findings were similar in human autopsy specimens. Conclusion These findings provide new therapeutic targets to oppose pulmonary vascular remodeling in HF-related PH. [ABSTRACT FROM AUTHOR]