Engineered Extracellular Vesicle-Based Therapies for Valvular Heart Disease.

Introduction: Valvular heart disease represents a significant burden to the healthcare system, with approximately 5 million cases diagnosed annually in the US. Among these cases, calcific aortic stenosis (CAS) stands out as the most prevalent form of valvular heart disease in the aging population.  CAS is characterized by the progressive calcification of the aortic valve leaflets, leading to valve stiffening. While aortic valve replacement is the standard of care for CAS patients, the long-term durability of prosthetic devices is poor, calling for innovative strategies to halt  or reverse disease progression. Here, we explor the potential use of novel extracellular vesicle (EV)-based nanocarriers for delivering molecular payloads to the affected valve tissue. This approach aims to reduce inflammation and potentially promote resorption of the calcified tissue. Methods: Engineered EVs loaded with the reprogramming myeloid transcription factors, CEBPA and Spi1, known to mediate the transdifferentiation of committed endothelial cells into macrophages. We evaluated the ability of these engineered EVs to deliver DNA and transcripts encoding CEBPA and Spil into calcified aortic valve tissue obtained from patients undergoing valve replacement due to aortic stenosis. We also investigated whether these EVs could induce the transdifferentiation of endothelial cells into macrophage-like cells. Results: Engineered EVs loaded with CEBPA + Spi1 were successfully derived from human dermal fibroblasts. Peak EV loading was found to be at 4 h after nanotransfection of donor cells.  These CEBPA + Spi1 loaded EVs effectively transfected aortic valve cells, resulting in the successful induction of transdifferentiation, both in vitro with  endothelial cells and ex vivo with valvular endothelial cells, leading to the development of anti-inflammatory macrophage-like cells. Conclusions: Our findings highlight the potential of engineered EVs as a next generation nanocarrier to target aberrant calcifications on diseased heart valves. This development holds promise as a novel therapy for high-risk patients who may not be suitable candidates for valve replacement surgery. Supplementary Information: The online version contains supplementary material available at 10.1007/s12195-023-00783-x.

Flow Dynamics in Anomalous Aortic Origin of a Coronary Artery in Children: Importance of the Intramural Segment.

This study aims to assess the differences in pressure, fractional flow reserve (FFR) and coronary flow (with increasing pressure) of the proximal coronary artery in patients with anomalous aortic origin of a coronary artery with a confirmed ischemic event, without ischemic events, and before and after unroofing surgery, and compare to a patient with normal coronary arteries. Patient-specific flow models were 3D printed for 3 subjects with anomalous right coronary arteries with intramural course, 2 of them had documented ischemia, and compared with a patient with normal coronaries. The models were placed in the aortic position of a pulse duplicator and precise measurements to quantify FFR and coronary flow rate were performed from the aortic to the mediastinal segment of the anomalous right coronary artery. In an ischemic model, a gradual FFR drop (emulating that of pressure) was shown from the ostium location (∼1.0) to the distal intramural course (0.48). In nonischemic and normal patient models, FFR for all locations did not drop below 0.9. In a second ischemic model prior to repair, a drop to 0.44 was encountered at the intramural and mediastinal intersection, improving to 0.86 postrepair. There is a difference in instantaneous coronary flow rate with increasing aortic pressure in the ischemic models (slope 0.2846), compared to the postrepair and normal models (slope >0.53). These observations on patient models support a biomechanical basis for ischemia and potentially sudden cardiac death in aortic origin of a coronary artery, with a drop in pressure and FFR in the intramural segment, and a decrease in coronary flow rate with increasing aortic pressure, with both improving after corrective surgery.

Assessment of transfer of morphological characteristics of Anomalous Aortic Origin of a Coronary Artery from imaging to patient specific 3D Printed models: A feasibility study.

BACKGROUND AND OBJECTIVE: This study aims to determine the accuracy of patient specific 3D printed models in capturing pathological anatomical characteristics derived from CT angiography (CTA) in children with anomalous aortic origin of a coronary artery (AAOCA). METHODS & MATERIALS: Following institutional regulatory approval, a standardized protocol for CTA of AAOCA was utilized for imaging. Blood volume of the aorta and coronaries were segmented from the DICOM images. A total of 10 models from 8 AAOCA patients were created, including 2 post-operative models. Mechanical properties of Agilus30 a flexible photopolymer coated with a thin layer of parylene, polyurethane (PU) and silicone and native aortic tissue from a postmortem specimen were compared. AAOCA models with wall thicknesses of 2mm aorta and 1.5mm coronaries were 3D printed in Agilus30 and coated with PU. CT of the printed models was performed, and 3D virtual models were generated. Transfer of anatomical characteristics and geometric accuracy were compared between the patient model virtual models. RESULTS: Dynamic modulus of Agilus30 at 2mm thickness was found to be close to native aortic tissue. Structured reporting of anatomical characteristics by imaging experts showed good concordance between patient and model CTA Comparative patient and virtual model measurements showed Pearson's correlation (r) of 0.9959 for aorta (n=70) and 0.9538 for coronaries (n=60) linear, and 0.9949 for aorta (n=30) and 0.9538 for coronaries (n=30) cross-sectional, dimensions. Surface contour map mean difference was 0.08 ± 0.29mm. CONCLUSIONS: Geometrically accurate AAOCA models preserving morphological characteristics, essential for risk stratification and decision-making, can be 3D printed from a patient's CTA.

Effect of Left and Right Coronary Flow Waveforms on Aortic Sinus Hemodynamics and Leaflet Shear Stress: Correlation with Calcification Locations.

Coronary flow induces hemodynamic alterations in the aortic sinus region. The objectives of this study are to: (1) investigate the differences among sinus hemodynamics and leaflet wall shear stresses engendered by the left versus right versus non-coronary flow and (2) correlate respective wall shear stresses with leaflet calcification in patients. A left heart simulator flow loop with a tunable coronary circuit provided physiological coronary flow waveforms corresponding to the left coronary cusp case (LCC), right coronary cusp case (RCC), and non-coronary cusp case (NCC). High spatio-temporal resolution particle image velocimetry was conducted to quantify leaflet wall shear stress and sinus vorticity fields and to measure aortic leaflet tip kinematics. Thirty-one patients with severe calcific aortic valve disease were segmented from CT data for the calcific volumes in their respective left, right, and non-coronary cusps. Leaflet tip position during systole shows the RCC has a wider leaflet opening compared to LCC and NCC. Velocity and vorticity fields combined with leaflet position data show that sinus vorticity is diminished (peak ~ 43 s-1) in the LCC while RCC and NCC maintain high vorticity (~ 1200 and ~ 950 s-1 respectively). WSS magnitudes greater than 0.3 Pa show 20 and 81% greater occurrences in the LCC and RCC respectively compared to NCC. Significant differences [X2 (2, n = 31) = 7.31, p = 0.0258] between the calcification levels in each cusp of the patient population. Coronary flow differences between LCC, RCC, and NCC show significant impact on leaflet kinematics and sinus flow hemodynamics. Clinical data correlations of the coronary flow cases indicate the left coronary cusp has a higher likelihood of calcification compared to the right.

Selective light-triggered release of DNA from gold nanorods switches blood clotting on and off.

Blood clotting is a precise cascade engineered to form a clot with temporal and spatial control. Current control of blood clotting is achieved predominantly by anticoagulants and thus inherently one-sided. Here we use a pair of nanorods (NRs) to provide a two-way switch for the blood clotting cascade by utilizing their ability to selectively release species on their surface under two different laser excitations. We selectively trigger release of a thrombin binding aptamer from one nanorod, inhibiting blood clotting and resulting in increased clotting time. We then release the complementary DNA as an antidote from the other NR, reversing the effect of the aptamer and restoring blood clotting. Thus, the nanorod pair acts as an on/off switch. One challenge for nanobiotechnology is the bio-nano interface, where coronas of weakly adsorbed proteins can obscure biomolecular function. We exploit these adsorbed proteins to increase aptamer and antidote loading on the nanorods.