Search

Background Heart transplantation (HTx) surgically transects all connections to the heart, including the autonomic nerves. We prospectively examined signs, timing and consequences of early sympathetic and parasympathetic sinoatrial reinnervation, as well as explored indirect evidence of afferent cardiopulmonary reinnervation. Methods Fifty HTx recipients were assessed at 2.5, 6 and 12 months after HTx. For comparison, 50 healthy controls were examined once. Continuous, noninvasive recordings of hemodynamic variables and heart rate variability indices were carried out at supine rest, 0.2 Hz controlled breathing, 60 degrees head-up-tilt, during the Valsalva maneuver and during handgrip isometric exercise. Results In HTx recipients, supine low-frequency heart rate variability gradually increased; supine high-frequency variability did not change; heart rate variability indices during controlled breathing remained unaltered; heart rate responses during tilt and isometric exercise gradually increased; the tachycardia response during Valsalva maneuver increased while the bradycardia response remained unchanged; and indices of baroreflex sensitivity improved. Responses remained low compared to healthy controls. A negative correlation between indices of preload and heart rate response during head-up tilt emerged at 12 months. Conclusions Results suggest that sympathetic reinnervation of the sinoatrial node starts within 6 months after HTx and strengthens during the first year. No evidence of early parasympathetic reinnervation was found. Indirect signs of afferent reinnervation of cardiopulmonary low-pressure baroreceptors emerged at 12 months. Better sympathetic sinoatrial control improved heart rate responsiveness to orthostatic challenge and isometric exercise, as well as heart rate buffering of blood pressure fluctuations.

We have recently demonstrated that accelerometer-based pump thrombosis and thromboembolic events detection is feasible in vitro. This article focuses on detection of these conditions in vivo. In an open-chest porcine model (n = 7), an accelerometer was attached to the pump casing of an implanted HeartWare HVAD. Pump vibration was analyzed by Fast Fourier Transform of the accelerometer signals, and the spectrogram third harmonic amplitude quantified and compared with pump power. Interventions included injection of thrombi into the left atrium (sized 0.3-0.4 ml, total n = 35) and control interventions; pump speed change, graft obstruction, and saline bolus injections (total n = 47). Graft flow to cardiac output ratio was used to estimate the expected number of thrombi passing through the pump. Sensitivity/specificity was assessed by receiver operating characteristic curve. Graft flow to cardiac output ratio averaged 66%. Twenty-six of 35 (74%) thrombi caused notable accelerometer signal change. Accelerometer third harmonic amplitude was significantly increased in thromboembolic interventions compared with control interventions, 64.5 (interquartile range [IQR]: 18.8-107.1) and 5.45 (IQR: 4.2-6.6), respectively (p0.01). The corresponding difference in pump power was 3 W (IQR: 2.9-3.3) and 2.8 W (IQR: 2.4-2.9), respectively (p0.01). Sensitivity/specificity of the accelerometer and pump power to detect thromboembolic events was 0.74/1.00 (area under the curve [AUC]: 0.956) and 0.40/1.00 (AUC: 0.759), respectively. Persistent high third harmonic amplitude was evident at end of all experiments, and pump thrombosis was confirmed by visual inspection. The findings demonstrate that accelerometer-based detection of thromboembolic events and pump thrombosis is feasible in vivo and that the method is superior to detection based on pump power.

Rationale: Human-induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CM) are increasingly being used for modeling heart disease and are under development for regeneration of the injured heart. However, incomplete structural and functional maturation of hiPSC-CM, including lack of T-tubules, immature excitation–contraction coupling, and inefficient Ca-induced Ca release remain major limitations. Objective: Thyroid and glucocorticoid hormones are critical for heart maturation. We hypothesized that their addition to standard protocols would promote T-tubule development and mature excitation–contraction coupling of hiPSC-CM when cultured on extracellular matrix with physiological stiffness (Matrigel mattress). Methods and Results: hiPSC-CM were generated using a standard chemical differentiation method supplemented with T3 (triiodothyronine) and/or Dex (dexamethasone) during days 16 to 30 followed by single-cell culture for 5 days on Matrigel mattress. hiPSC-CM treated with T3+Dex, but not with either T3 or Dex alone, developed an extensive T-tubule network. Notably, Matrigel mattress was necessary for T-tubule formation. Compared with adult human ventricular cardiomyocytes, T-tubules in T3+Dex-treated hiPSC-CM were less organized and had more longitudinal elements. Confocal line scans demonstrated spatially and temporally uniform Ca release that is characteristic of excitation–contraction coupling in the heart ventricle. T3+Dex enhanced elementary Ca release measured by Ca sparks and promoted RyR2 (ryanodine receptor) structural organization. Simultaneous measurements of L-type Ca current and intracellular Ca release confirmed enhanced functional coupling between L-type Ca channels and RyR2 in T3+Dex-treated cells. Conclusions: Our results suggest a permissive role of combined thyroid and glucocorticoid hormones during the cardiac differentiation process, which when coupled with further maturation on Matrigel mattress, is sufficient for T-tubule development, enhanced Ca-induced Ca release, and more ventricular-like excitation–contraction coupling. This new hormone maturation method could advance the use of hiPSC-CM for disease modeling and cell-based therapy.

Transplant-associated coronary artery disease (TxCAD) appears to be initiated by endothelial cell activation and inflammation involving inflammatory cytokines and chemokines. Osteoprotegerin (OPG) and receptor activator of nuclear Factor-kappaB ligand (RANKL) have been implicated in cardiovascular disease progression and we measured the expression of these mediators in serum and myocardial biopsies taken serially during the first year after heart transplantation (HTx), relating them to the development of TxCAD. Serum OPG as well as myocardial gene expression of RANK and OPG, but not RANKL, were highest early after HTx and declined progressively. Importantly, patients who develop TxCAD or experience episodes of acute rejection showed a lower myocardial RANKL expression throughout the first year after transplantation than patients without these complications. Our findings may suggest an unrecognized role RANKL in maintaining myocardial and/or endothelial integrity and suggest that RANKL should be further investigated as a parameter that may predict development of TxCAD.

Background Whereas complement is a key mediator of hyperacute xenograft rejection, its role in acute vascular rejection (AVR) is a matter of controversy. AVR is associated with de novo synthesis of endothelial cell-derived inflammatory mediators, including the leukocyte-recruiting adhesion molecule E-selectin. Here we investigate the role and mechanism of complement in human serum-induced porcine endothelial cell activation. Methods An in vitro xenotransplantation method was designed using porcine aortic endothelial cells stimulated with human serum in microculture wells. E-selectin expression was measured by cell-enzyme immunoassay. Complement inhibitors acting at different levels in the cascade were investigated for their effect on E-selectin expression. Results E-selectin was strongly induced by normal human serum but not by heat-inactivated serum. Compstatin, a synthetic C3 inhibitor, markedly reduced human serum-induced E-selectin expression. Purified C1-inhibitor suppressed E-selectin induction completely, indicating activation through the classical or lectin pathway. Furthermore, a monoclonal antibody (mAb) that inhibits cleavage of C5 or another mAb that blocks the function of C7, completely inhibited the expression of serum-induced E-selectin, consistent with the terminal C5b-9 complement complex being the mediator of the endothelial cell activation. Inhibition of the alternative pathway using a novel antifactor D mAb did not reduce E-selectin expression. Conclusion Human serum-induced expression of porcine E-selectin is totally complement dependent, induced by a C1-inhibitor regulated pathway and mediated through the terminal complement complex. The data may have implications for therapeutic strategies, particularly of C1-inhibitor and anti-C5 mAb, to protect against endothelial cell activation and subsequent AVR of porcine xenografts.

Background. The possibility that human cytomegalovirus (HCMV) may infect porcine endothelial cells (ECs) was investigated. This may be relevant during xenotransplantation of porcine cells or organs into human recipients. Methods. HCMV was inoculated into low-passage porcine ECs. Replication of virus was detected by development of characteristic cytopathogenic effect. Appearance of immediate early, early, and late antigens was studied by immunocytochemical staining. Infectious virus was detected in human fibroblast cells. Presence of HCMV RNA was studied by Northern Blot and reverse transcriptase polymerase chain reaction. Results. All parameters indicated that a fresh clinical HCMV isolate productively infects porcine ECs. The same cells do not fully support replication of the laboratory strain Ad 169. Conclusion. Our results may indicate the possibility of cross-species infectivity of HCMV to porcine cells.