Porcine Ventricular Tissue Research Articles

Highly Efficient Cardiac Differentiation and Maintenance by Thrombin-Coagulated Fibrin Hydrogels Enriched with Decellularized Porcine Heart Extracellular Matrix.

Biochemical and biophysical properties instruct cardiac tissue morphogenesis. Here, we are reporting on a blend of cardiac decellularized extracellular matrix (dECM) from porcine ventricular tissue and fibrinogen that is suitable for investigations employing an in vitro 3D cardiac cell culture model. Rapid and specific coagulation with thrombin facilitates the gentle inclusion of cells while avoiding sedimentation during formation of the dECM-fibrin composite. Our investigations revealed enhanced cardiogenic differentiation in the H9c2 myoblast cells when using the system in a co-culture with Nor-10 fibroblasts. Further enhancement of differentiation efficiency was achieved by 3D embedding of rat neonatal cardiomyocytes in the 3D system. Calcium imaging and analysis of beating motion both indicate that the dECM-fibrin composite significantly enhances recovery, frequency, synchrony, and the maintenance of spontaneous beating, as compared to various controls including Matrigel, pure fibrin and collagen I as well as a fibrin-collagen I blend.

The effects of inhomogeneities on scroll-wave dynamics in an anatomically realistic mathematical model for canine ventricular tissue

Ventricular tachycardia (VT) and ventricular fibrillation (VF) are lethal rhythm disorders, which are associated with the occurrence of abnormal electrical scroll waves in the heart. Given the technical limitations of imaging and probing, the in situ visualization of these waves inside cardiac tissue remains a challenge. Therefore, we must, perforce, rely on in silico simulations of scroll waves in mathematical models for cardiac tissue to develop an understanding of the dynamics of these waves in mammalian hearts. We use direct numerical simulations of the Hund–Rudy-Dynamic (HRD) model, for canine ventricular tissue, to examine the interplay between electrical scroll-waves and conduction and ionic inhomogeneities, in anatomically realistic canine ventricular geometries with muscle-fiber architecture. We find that millimeter-sized, distributed, conduction inhomogeneities cause a substantial decrease in the scroll wavelength, thereby increasing the probability for wave breaks; by contrast, single, localized, medium-sized (≃ cm) conduction inhomogeneities, exhibit the potential to suppress wave breaks or enable the self-organization of wave fragments into stable, intact scrolls. We show that ionic inhomogeneities, both distributed or localized, suppress scroll-wave break up. The dynamics of a stable rotating wave is not affected significantly by such inhomogeneities, except at high concentrations of distributed inhomogeneities, which can cause a partial break up of scroll waves. Our results indicate that inhomogeneities in the canine ventricular tissue are less arrhythmogenic than inhomogeneities in porcine ventricular tissue, for which an earlier in silico study (Majumder et al., 2016) has shown that the inhomogeneity-induced suppression of scroll waves is a rare occurrence.

652The effect of varying irrigation flow rate during irrigated radiofrequency ablation on optimising lesion shape

Abstract Funding Acknowledgements Investigator Sponsored Research grant from Boston Scientific Introduction Irrigated catheters are the standard tool for radiofrequency (RF) ablation in the left atrium and ventricles. However, pathological studies of irrigated RF lesions show a "tear-drop" shape, with the widest diameter some depth below the endocardial surface and relative endocardial sparing. This requires overlap of lesions to achieve contiguity at the endocardial surface. There has been little investigation into the effect of altering irrigation rate on lesion shape and volume. Purpose To test the hypothesis that varying the irrigation flow rate would optimise lesion shape by minimising endocardial sparing while maintaining lesion depth. Methods In an ex vivo animal heart model, irrigated ablation lesions were performed in porcine ventricular tissue at 30W using an Intellatip MiFi OI catheter with 5 different irrigation protocols: A. fixed rate (30ml/min); B. continuous reduction (30 to 2ml/min over 30s); C. continuous increase (2 to 30ml/min over 30s); D. stepwise reduction (30ml/min (10s) to 16ml/min (10s) to 2ml/min (10s)); E. stepwise increase (2ml/min (10s) to 16ml/min (10s) to 30ml/min (10s). Contact force (10g) and ablation duration (30s) were constant. Steam pops during ablation were recorded. Tissue sections were stained with triphenyltetrazolium chloride (TTC) after ablation to allow accurate measurement of lesion boundaries. Surface diameter, lesion depth, maximum diameter, and depth at maximum diameter were measured using Vernier Calipers to calculate lesion volume. Pictures of lesions were analysed further by ImageJ software to measure the degree of endocardial sparing. The optimal protocol was further tested against fixed-rate irrigation at 20W – 40W. Results 10-20 lesions were performed for each irrigation protocol. Of the 4 experimental protocols, continuous reduction (protocol B) resulted in the most optimal lesion shape (Figure). With this protocol, endocardial sparing area was significantly reduced compared to fixed-rate irrigation (1.61 vs. 2.64mm2, P < 0.0001), with a trend towards an increase in surface diameter (9.25 vs. 8.46mm, P = 0.08). There was no significant difference in lesion depth (5.35 vs. 4.77mm), lesion volume (374 vs. 332mm3) or maximum diameter (10.3 vs. 10.8mm). Steam pop occurred in 1 of 20 (5%) lesions in each of protocols A and B. Significantly reduced endocardial sparing with preserved volume/depth was consistent for continuous reduction when compared to fixed-rate irrigation across power settings of 20W (17ml/min), 30 W (17ml/min or 30ml/min) and 40W (30ml/min) (P < 0.0001). Conclusions Continuous reduction in irrigation flow rate from 30 to 2ml/min during irrigated RF ablation results in reduced endocardial sparing with preserved lesion depth and volume when compared to fixed-rate irrigation across power settings of 20 – 40W. This may allow for greater lesion spacing while maintaining endocardial contiguity and merits further investigation to improve irrigated RF ablation efficiency. Abstract Figure

DECELLULARIZED VENTRICULAR MATRIX AS A MODEL FOR STUDYING HUMAN MESENCHYMAL CELL PHENOTYPES

As a result of pioneering research unveiling the influence of extracellular environment in determining stem/progenitor cell differentiation, a myriad of cellular scaffolds have been developed in order to optimize the delivery of these cells in a diseased environment as a technique to improve their efficacy in stem cell therapy. Although a variety of scaffolds are being studied in different contexts, the most physiologically relevant ones are those which most closely resemble the environment to be treated. Our lab has a keen interest in cardiovascular disease and we have recently optimized a technique that decellularizes and reseeds isolated sections of healthy porcine ventricular tissue with primary human atrial cells.

Bipolar irrigated radiofrequency ablation: A therapeutic option for refractory intramural atrial and ventricular tachycardia circuits

Irrigated radiofrequency (RF) ablation can be insufficient to eliminate intramurally located septal atrial flutter (AFL) and ventricular tachycardia (VT) circuits. Bipolar ablation between 2 ablation catheters may be considered for such circuits. To evaluate the utility of bipolar irrigated ablation to terminate arrhythmias resistant to unipolar ablation. In vitro: Bipolar and sequential unipolar RF ablation lesions were placed on porcine ventricular tissue in a saline bath to assess for lesion transmurality. Clinical: 3 patients with atypical septal flutter (AFL), 4 patients with septal VT, and 2 with left ventricle free-wall VT, all of whom failed sequential unipolar RF ablation, underwent bipolar RF ablation using irrigated catheters placed on either surface of the interatria/interventricular septum and left ventricle free-wall, respectively. In vitro: Bipolar RF was found to be more likely to achieve transmural lesions (82% vs 33%; P = .001) and could do so in tissues with thicknesses of up to 25 mm. Clinical: All 5 AFLs (3 patients) were successfully terminated with bipolar RF. In follow-up, AFL recurred in 2 of the 3 patients and atrial fibrillation and AFL recurred in 1 of the 3. All 3 thereafter underwent repeat procedures with successful maintenance of sinus rhythm in 2 of the 3 patients (6-month follow-up). In the VT subgroup, 5 of 6 septal VTs and 2 of 3 free-wall VTs were terminated successfully during ablation. In follow-up (12 months), 2 of the 4 patients in the septal bipolar group and 1 of the 2 patients in the free-wall group remained free of VT. Bipolar RF can be used to terminate arrhythmias in select patients with tachyarrhythmias.

Modulation of material properties of a decellularized myocardial matrix scaffold.

Injectable materials offer the potential for minimally invasive therapy for myocardial infarction (MI), either as an acellular scaffold or as a cell delivery vehicle. A recently developed myocardial matrix hydrogel, derived from decellularized porcine ventricular tissue, has the potential to aid in cardiac repair following an MI. Herein, we set out to study the effects of cross-linking on the cardiac hydrogel stiffness, degradation properties, cellular migration, and catheter injectability in vitro. Cross-linking increased stiffness, while slowing degradation and cellular migration through the gels. Additionally, the cross-linked material was pushed through a clinically relevant catheter. These results demonstrate that the material properties of myocardial matrix can be tuned via cross-linking, while maintaining appropriate viscosity for catheter injectability.

Long electrodes for radio frequency ablation: comparative study of surface versus intramural application

There is increasing use of radio frequency (RF) ablation with long electrodes in the intraoperative treatment of atrial fibrillation. Nevertheless, the disparity in the lesion geometry in both depth and width is the major pitfall in the use of RF currents. The objective of this study was to differentiate the shape and size of long lesions created by three surface application electrodes (SAE) and two intramural electrodes (IE). The SAE included a standard multi-polar catheter, and two standard electrosurgical pencils. The IE consisted of a needle and a wire both intramurally buried. The lesions were created on fresh fragments of porcine ventricular tissue. The IE created lesions with a curved prism-like shape around the electrode body, with homogeneous characteristics along the lesion trajectory. On the contrary, the lesions created with the SAE were in the shape of an hourglass. They showed a different geometry between the central zone and the edge zone ( p<0.001 for depth and surface width). Electrical impedance evolution was recorded during the RF heating. We observed a slow decrease of the impedance in all the electrodes, except in the wire electrode. In conclusion, the results suggest that the IE might be a more suitable option than SAE when it is necessary to create long and homogeneous thermal lesions.

The LETR-Principle: a novel method to assess electrode-tissue contact in radiofrequency ablation.

Stable electrode-tissue contact is crucial for successful radiofrequency ablation of cardiac tachyarrhythmias. In this in vitro study, a custom-made radiofrequency generator was used to evaluate the correlation between tip temperature response to a minimal radiofrequency power delivery (Low Energy Temperature Response: LETR-Principle) and electrode-tissue contact as well as lesion size. A battery-powered radiofrequency generator (LETR-Box, 500 kHz, 0.1 to 0.3 W) could measure the temperature increase at the tip electrode with 0.01 degrees C accuracy. The device was tested in vitro using isolated porcine ventricular tissue. For various electrode-tissue settings (i.e., 0 to 0.89 N contact force), the temperature increase (deltaT) due to 0.1-W power delivery for 10 seconds was recorded. Subsequently, for the same electrode-tissue contact, a temperature-controlled radiofrequency ablation was performed (70 degrees C target temperature, 50-W maximum output, 30 sec). Thereafter, the lesion size was measured histologically. To prove the safety of the applied LETR-Principle, the tissue was inspected microscopically after continuous radiofrequency power delivery of 0.3 W for 1 hour with high contact pressure (1.33 N). The delivery of 0.1-W radiofrequency power resulted in an average deltaT of 0.18 degrees +/- 0.13 degrees C. During temperature-controlled radiofrequency ablation, the tip temperature was 59 degrees +/- 8.5 degrees C, resulting in a lesion depth of 4.8+/-0.6 mm. The correlation coefficient between deltaT and contact force was 0.97 and 0.81, respectively, for lesion depth. No lesion was microscopically visible after power delivery of 0.3 W for 1 hour with 1.33 N contact pressure. The LETR-Principle safely indicates electrode-tissue contact and lesion depth under in vitro conditions and can be useful for catheter positioning during radiofrequency ablation procedures.

Effects of saterinone and its enantiomers R(+)-saterinone and S(-)-saterinone on the phosphodiesterase isoenzymes from ventricular tissue of failing human hearts and porcine hearts.

Stereoisomers often exhibit differences in their pharmacological activities. Therefore, the phosphodiesterase inhibitory effects of the cardiotonic agent saterinone in form of the racemate were investigated in comparison with the inhibitory properties of its enantiomers R(+)- and S(-)-saterinone. For this purpose the phosphodiesterase isoenzymes from ventricular tissue of failing human hearts and porcine hearts were separated by DEAE-sepharose anion exchange chromatography. Four different phosphodiesterase isoenzymes were isolated from failing human myocardium and designated phosphodiesterase I-IV. Three phosphodiesterase isoenzymes could be separated from ventricular tissue of porcine hearts. A Ca2+/calmodulin stimulated phosphodiesterase I was not detectable in porcine myocardium. In failing human hearts racemic saterinone was a potent inhibitor of phosphodiesterase III (IC50 0.02 mumol/l) and IV (IC50 0.03 mumol/l) as compared to the inhibition of phosphodiesterase I (IC50 37.3 mumol/l) and II (IC50 51.4 mumol/l). In comparison with the racemate, R(+)- and S(-)-saterinone showed only slight differences in their phosphodiesterase inhibitory effects. R(+)-saterinone inhibited phosphodiesterase III slightly but significantly more potently and selectively than did S(-)-saterinone. Compared to the inhibition of phosphodiesterase I and II both enantiomers were similarly potent and selective inhibitors of phosphodiesterase III and IV. Similar results were obtained in porcine hearts. It is concluded that the racemate saterinone and its enantiomers (R(+)- and S(-)-saterinone are virtually equipotent concerning the inhibition of phosphodiesterase isoenzymes isolated from failing human hearts or porcine ventricular tissue. The enantiomers of saterinone did not exhibit distinct stereoselectivity in their phosphodiesterase inhibitory effects.

Immunohistochemical identification of Purkinje fibers and transitional cells in a terminal portion of the impulse-conducting system of porcine heart.

The ultrastructure of porcine ventricular tissue was studied by electron microscopy and immunocytochemical techniques. Electron-dense specific granules were found in both Purkinje fibers and transitional cells in the ventricular walls, and were positively stained by the immunogold staining method using an antiserum against atrial natriuretic polypeptide (ANP). This suggests that both the Purkinje fibers and transitional cells display the same specific granules as atrial cardiocytes containing ANP. These results demonstrate that Purkinje fibers and two types of transitional cells, in addition to the ordinary ventricular cardiocytes, can be identified in porcine ventricular wall tissue.