Pro-angiogenic Potential of Mesenchymal Stromal Cells Regulated by Matrix Stiffness and Anisotropy Mimicking Right Ventricles.

Capillary rarefaction is a hallmark of right ventricle (RV) failure. Mesenchymal stromal cell (MSC)-based therapy offers a potential treatment due to its pro-angiogenic function. However, the impact of RV tissue mechanics on MSC behavior is unclear, especially when referring to RV end-diastolic stiffness and mechanical anisotropy. In this study, we assessed MSC behavior on electrospun scaffolds with varied stiffness (normal vs failing RV) and anisotropy (isotropic vs anisotropic). In individual MSCs, we observed the highest vascular endothelial growth factor (VEGF) production and total tube length in the failing, isotropic group (2.00 ± 0.37, 1.53 ± 0.24), which was greater than the normal, isotropic group (0.70 ± 0.15, 0.55 ± 0.07; p < 0.05). The presence of anisotropy led to trends of increased VEGF production on normal groups (0.75 ± 0.09 vs 1.20 ± 0.17), but this effect was absent on failing groups. Our findings reveal synergistic effects of RV-like stiffness and anisotropy on MSC pro-angiogenic function and may guide MSC-based therapies for heart failure.

Current status of myocardial restoration via the paracrine function of mesenchymal stromal cells.

Mesenchymal stromal cells (MSCs) have been studied for nearly two decades as a therapy for myocardial restoration. An emerging direction to repair myocardium is through their paracrine function, which includes the utilization of MSC-derived conditioned medium or extracellular vesicles. In this review, we go over the unique characteristics of MSCs that make it suitable for "off the shelf," cell-free regenerative therapy, current MSC-derived cell-free approaches including their advantages and disadvantages, and the known mechanisms of action of the paracrine effect of MSCs. With a summary of the clinical trials and preclinical studies of MSC-derived cell-free therapy, we classify the aforementioned mechanisms into angiogenesis, immunomodulation, extracellular matrix remodeling, antiapoptosis, and antioxidation. Particularly, we discuss on ways researchers have worked toward enhancing these desired properties to improve the therapeutic outcomes and the investigation of mechanobiology involved in MSC paracrine function. Lastly, we bring up the remaining challenges in this arising field and suggestions for future directions to improve our understanding and control over the potential of MSC paracrine function for myocardial restoration.

Different Passive Viscoelastic Properties Between the Left and Right Ventricles in Healthy Adult Ovine.

Ventricle dysfunction is the most common cause of heart failure, which leads to high mortality and morbidity. The mechanical behavior of the ventricle is critical to its physiological function. It is known that the ventricle is anisotropic and viscoelastic. However, the understanding of ventricular viscoelasticity is much less than that of its elasticity. Moreover, the left and right ventricles (LV&RV) are different in embryologic origin, anatomy, and function, but whether they distinguish in viscoelastic properties is unclear. We hypothesized that passive viscoelasticity is different between healthy LVs and RVs. Ex vivo cyclic biaxial tensile mechanical tests (1, 0.1, 0.01 Hz) and stress relaxation (strain of 3, 6, 9, 12, 15%) were performed for ventricles from healthy adult sheep. Outflow track direction was defined as the longitudinal direction. Hysteresis stress-strain loops and stress relaxation curves were obtained to quantify the viscoelastic properties. We found that the RV had more pronounced frequency-dependent viscoelastic changes than the LV. Under the physiological frequency (1 Hz), the LV was more anisotropic in the elasticity and stiffer than the RV in both directions, whereas the RV was more anisotropic in the viscosity and more viscous than the LV in the longitudinal direction. The LV was quasi-linear viscoelastic in the longitudinal but not circumferential direction, and the RV was nonlinear viscoelastic in both directions. This study is the first to investigate passive viscoelastic differences in healthy LVs and RVs, and the findings will deepen the understanding of biomechanical mechanisms of ventricular function.

Biomechanical Properties and Mechanobiology of Cardiac ECM.

The heart is comprised of cardiac cells and extracellular matrix (ECM) which function together to pump blood throughout the body, provide organs with nutrients and oxygen, and remove metabolic wastes. Cardiac ECM provides a scaffold to cardiac cells and contributes to the mechanical properties and function of the cardiac tissue. Recently, more evidence suggests that cardiac ECM plays an active role in cardiac remodeling in response to mechanical loads. To that end, we provide an overview of the structure and function of the heart and the currently available in vivo and ex vivo mechanical measurements of cardiac tissues. We also review the biomechanical properties of cardiac tissues including the myocardium and heart valves, with a discussion on the differences between the right ventricle and left ventricle. Lastly, we go into the mechanical factors involved in cardiac remodeling and review the mechanobiology of cardiac tissues, i.e., the biomechanical responses at the cellular and tissue level, with an emphasis on the impact on the cardiac ECM. The regulation of cardiac ECM on cell function, which is a new and open area of research, is also briefly discussed. Future investigation into the ECM deposition and the interaction of cardiac cells and ECM components for mechanotransduction can assist to understand cardiac remodeling and inspire new therapies for cardiac diseases.

Multiscale Contrasts Between the Right and Left Ventricle Biomechanics in Healthy Adult Sheep and Translational Implications.

Cardiac biomechanics play a significant role in the progression of structural heart diseases (SHDs). SHDs alter baseline myocardial biomechanics leading to single or bi-ventricular dysfunction. But therapies for left ventricle (LV) failure patients do not always work well for right ventricle (RV) failure patients. This is partly because the basic knowledge of baseline contrasts between the RV and LV biomechanics remains elusive with limited discrepant findings. The aim of the study was to investigate the multiscale contrasts between LV and RV biomechanics in large animal species. We hypothesize that the adult healthy LV and RV have distinct passive anisotropic biomechanical properties. Ex vivo biaxial tests were performed in fresh sheep hearts. Histology and immunohistochemistry were performed to measure tissue collagen. The experimental data were then fitted to a Fung type model and a structurally informed model, separately. We found that the LV was stiffer in the longitudinal (outflow tract) than circumferential direction, whereas the RV showed the opposite anisotropic behavior. The anisotropic parameter K from the Fung type model accurately captured contrasting anisotropic behaviors in the LV and RV. When comparing the elasticity in the same direction, the LV was stiffer than the RV longitudinally and the RV was stiffer than the LV circumferentially, suggesting different filling patterns of these ventricles during diastole. Results from the structurally informed model suggest potentially stiffer collagen fibers in the LV than RV, demanding further investigation. Finally, type III collagen content was correlated with the low-strain elastic moduli in both ventricles. In summary, our findings provide fundamental biomechanical differences between the chambers. These results provide valuable insights for guiding cardiac tissue engineering and regenerative studies to implement chamber-specific matrix mechanics, which is particularly critical for identifying biomechanical mechanisms of diseases or mechanical regulation of therapeutic responses. In addition, our results serve as a benchmark for image-based inverse modeling technologies to non-invasively estimate myocardial properties in the RV and LV.

The Interventricular Septum Is Biomechanically Distinct from the Ventricular Free Walls.

The interventricular septum contributes to the pumping function of both ventricles. However, unlike the ventricular wall, its mechanical behavior remains largely unknown. To fill the knowledge gap, this study aims to characterize the biaxial and transmural variation of the mechanical properties of the septum and compare it to the free walls of the left and right ventricles (LV/RV). Fresh hearts were obtained from healthy, adult sheep. The septal wall was sliced along the mid-line into two septal sides and compared to the epicardial layers of the LV- and RV-free walls. Biaxial tensile mechanical tests and constitutive modeling were performed to obtain the passive mechanical properties of the LV- and RV-side of the septum and ventricular walls. We found that both sides of the septum were significantly softer than the respective ventricular walls, and that the septum presented significantly less collagen than the ventricular walls. At low strains, we observed the symmetric distribution of the fiber orientations and a similar anisotropic behavior between the LV-side and RV-side of the septum, with a stiffer material property in the longitudinal direction, rather than the circumferential direction. At high strains, both sides showed isotropic behavior. Both septal sides had similar intrinsic elasticity, as evidenced by experimental data and constitutive modeling. These new findings offer important knowledge of the biomechanics of the septum wall, which may deepen the understanding of heart physiology.

Mechanical Considerations of Electrospun Scaffolds for Myocardial Tissue and Regenerative Engineering.

Biomaterials to facilitate the restoration of cardiac tissue is of emerging importance. While there are many aspects to consider in the design of biomaterials, mechanical properties can be of particular importance in this dynamically remodeling tissue. This review focuses on one specific processing method, electrospinning, that is employed to generate materials with a fibrous microstructure that can be combined with material properties to achieve the desired mechanical behavior. Current methods used to fabricate mechanically relevant micro-/nanofibrous scaffolds, in vivo studies using these scaffolds as therapeutics, and common techniques to characterize the mechanical properties of the scaffolds are covered. We also discuss the discrepancies in the reported elastic modulus for physiological and pathological myocardium in the literature, as well as the emerging area of in vitro mechanobiology studies to investigate the mechanical regulation in cardiac tissue engineering. Lastly, future perspectives and recommendations are offered in order to enhance the understanding of cardiac mechanobiology and foster therapeutic development in myocardial regenerative medicine.

Establishment of adult right ventricle failure in ovine using a graded, animal-specific pulmonary artery constriction model.

BACKGROUND: Right ventricle failure (RVF) is associated with serious cardiac and pulmonary diseases that contribute significantly to the morbidity and mortality of patients. Currently, the mechanisms of RVF are not fully understood and it is partly due to the lack of large animal models in adult RVF. In this study, we aim to establish a model of RVF in adult ovine and examine the structure and function relations in the RV. METHODS: RV pressure overload was induced in adult male sheep by revised pulmonary artery constriction (PAC). Briefly, an adjustable hydraulic occluder was placed around the main pulmonary artery trunk. Then, repeated saline injection was performed at weeks 0, 1, and 4, where the amount of saline was determined in an animal-specific manner. Healthy, age-matched male sheep were used as additional controls. Echocardiography was performed bi-weekly and on week 11 post-PAC, hemodynamic and biological measurements were obtained. RESULTS: This PAC methodology resulted in a marked increase in RV systolic pressure and decreases in stroke volume and tricuspid annular plane systolic excursion, indicating signs of RVF. Significant increases in RV chamber size, wall thickness, and Fulton's index were observed. Cardiomyocyte hypertrophy and collagen accumulation (particularly type III collagen) were evident, and these structural changes were correlated with RV dysfunction. CONCLUSION: In summary, the animal-specific, repeated PAC provided a robust approach to induce adult RVF, and this ovine model will offer a useful tool to study the progression and treatment of adult RVF that is translatable to human diseases.

Detection of Pentosidine Cross-Links in Cell-Secreted Decellularized Matrices Using Time Resolved Fluorescence Spectroscopy.

Hyperglycemia-mediated, nonenzymatic collagen cross-links such as pentosidine (PENT) can have deleterious effects on cellular interactions with the extracellular matrix (ECM). Present techniques to quantify PENT are limited, motivating the need for improved methods to study the accumulation and contribution of PENT toward diabetic clinical challenges such as impaired bone healing. Current methods for studying PENT are destructive, laborious, and frequently employ oversimplified collagen films that lack the complexity of the native ECM. The primary goal of this study was to evaluate the capacity of time-resolved fluorescence spectroscopy (TRFS) to detect PENT in cell-secreted ECMs possessing enhanced compositional complexity. To demonstrate an application of this method, we assessed the response of human mesenchymal stem cells (MSCs) to cross-linked substrates to explore the role of detected PENT on osteogenic differentiation. We exposed MSC-secreted decellularized matrices (DMs) to 0.66 M ribose for 2 weeks and used TRFS to detect the accumulation of PENT. Ribose treatment resulted in a 30 nm blue shift in peak fluorescence emission and a significant decrease in average lifetime compared to that of control DMs (4.4 ± 0.3 ns vs 3.5 ± 0.09 ns). Evaluation of samples with high performance liquid chromatography (HPLC) confirmed that changes in observed fluorescence were due to PENT accumulation. A strong correlation was found between TRFS parameters and the HPLC measurement of PENT, validating the use of TRFS as an alternative method of PENT detection. Osteoblastic gene expression was significantly reduced in MSCs seeded on ribose DMs at days 7 and 14. However, no significant differences in calcium deposition were detected between control and ribose DMs. These data demonstrate the efficacy of nondestructive fluorescence spectroscopy to examine the formation of nonenzymatic collagen cross-links within biomimetic culture platforms and showcase one example where an improved biomimetic substrate can be used to probe cell-ECM interactions in the presence of collagen cross-links.