The ultimate treatment for patients with end-stage heart failure is heart transplantation. The number of donor hearts which are primarily procured from donation after brain death (DBD) donors is limited, but donation after circulatory death (DCD) donor hearts can increase the heart donor pool. However, ischemia and reperfusion injuries associated with the DCD process causes myocardial damage, limiting the use of DCD hearts in transplantation. Addressing this problem is critical in the exploration of DCD hearts as suitable donor hearts for transplantation. In this study, rat hearts were procured following the control beating-heart donor (CBD) or DCD donation process. Changes in mitochondria and cardiac function from DCD hearts subjected to 25 or 35 minutes of ischemia followed by 60 minutes of reperfusion were compared to CBD hearts. Following ischemia, rates of oxidative phosphorylation and calcium retention capacity were progressively impaired in DCD hearts compared to CBD hearts. Reperfusion caused additional mitochondrial dysfunction in DCD hearts. Developed pressure, inotropy and lusitropy, were significantly reduced in DCD hearts compared to CBD hearts. We, therefore, suggest that interventional strategies targeted before the onset of ischemia and at reperfusion could protect mitochondria, thus potentially making DCD hearts suitable for heart transplantation.
Background: Mitochondria contribute to ischemia and reperfusion injury (IRI) in donation after circulatory death (DCD) donor hearts. Cyclosporine A (CyA), an inhibitor of mitochondrial permeability transition pore (MPTP) opening, limits IRI. CyA administration at the onset of reperfusion is essential for therapeutic efficacy. DCD heart transplantation (HTx) is a unique setting for controlled administration of CyA. Hypothesis: CyA administered to DCD hearts during reperfusion is cardioprotective. Methods: Under general anesthesia Sprague Dawley rats underwent simulated clinical DCD setup with a global ischemia of 25 minutes. Control beating-heart donor (CBD) hearts served as controls (n=10). Hearts were reanimated ex-vivo with Krebs-Henseleit buffer (KH) perfusion at 37 o C for 90 minutes. DCD hearts were randomly assigned to receive CyA (0.5mM) or DMSO added to KH buffer for the first 15 minutes of perfusion (n=10 each). The hearts were collected to measure oxidative phosphorylation (OXPHOS) and calcium retention capacity (CRC). Additional CBD (n=5) and DCD (with or without CyA, n=15) hearts underwent heterotopic HTx in recipient rats to measure developed pressure (DP) and rate pressure product (RPP) at 48 hours. Results: Compared to CBD hearts, OXPHOS in DCD hearts was significantly reduced. CyA improved OXPHOS and was similar to CBD hearts. CRC was reduced in DCD hearts compared to CBD hearts (317 vs.578nM/mg) and was increased with CyA treatment (413 vs. 317nM/mg; p<0.05, Figure). Infarct size decreased significantly with CyA treatment (15% vs. 25%). In transplanted DCD hearts, DP and RPP were significantly reduced compared to CBD hearts (80 vs.153mmHg and 17639 vs. 29545mmHg*bpm). CyA treatment improved DP (124vs.80mmHg; p<0.05) and RPP (35491vs.17639mmHg*bpm; p<0.05) which was comparable to CBD hearts. Conclusions: CyA administered at reperfusion reduces mitochondrial damage, limits infarct size, and preserves function in transplanted DCD hearts.
Heart failure patients requiring total artificial heart (TAH) support often have concomitant renal insufficiency (RI). We sought to quantify renal function recovery in patients supported with TAH at our institution. Renal function data at 30, 90, and 180 days after TAH implantation were analyzed for patients with RI, defined as hemodialysis supported or an estimated glomerular filtration rate (eGFR) less than 60 ml/min/1.73 m. Between January 2008 and December 2013, 20 of the 46 (43.5%) TAH recipients (age 51 ± 9 years, 85% men) had RI, mean preoperative eGFR of 48 ± 7 ml/min/1.73 m. Renal function recovery was noted at each follow-up interval: increment in eGFR (ml/min/1.73 m) at 30, 90, and 180 days was 21 ± 35 (p = 0.1), 16.5 ± 18 (p = 0.05), and 10 ± 9 (p = 0.1), respectively. Six patients (30%) required preoperative dialysis. Of these, four recovered renal function, one remained on dialysis, and one died. Six patients (30%) required new-onset dialysis. Of these, three recovered renal function and three died. Overall, 75% (15 of 20) of patients' renal function improved with TAH support. Total artificial heart support improved renal function in 75% of patients with pre-existing significant RI, including those who required preoperative dialysis.
Critically ill patients requiring catecholamine infusion may require extracorporeal membrane oxygenation (ECMO) support. Initiating ECMO support may affect the circulating catecholamine levels, which directly influences the circulatory support. We measured the timed plasma levels of epinephrine, norepinephrine, and dopamine infused at a constant rate in pigs supported on the ECMO. Plasma levels of catecholamines decreased at 10 minutes after the initiation of ECMO, followed by a return to steady-state concentrations for the next 2 hours. Catecholamines levels in patients initiated on ECMO may follow similar changes. While ECMO provides essential life-sustaining circulatory support, it also introduces critical variables that directly alter the pharmacokinetics through drug distribution,1 drug adsorption,2 and drug elimination.3 Literature supporting the altered drug metabolism in patients supported with ECMO is available for different antibiotics,1 analgesics,4 and sedatives,5 commonly used in critical care units. However, the literature on the effects of ECMO on commonly used inotropes (epinephrine [EPI], dopamine [DOP]) and vasopressors (norepinephrine [NE]) is lacking. We examined the plasma levels of commonly used inotropes and vasopressor medications in pigs supported on the ECMO circuit. After institutional Institution Animal Care and Utilization Committee approval, six 30 kg (body surface area 0.77 m2) juvenile Yorkshire pigs were anesthetized and maintained on a ventilator with inhaled isoflurane. Under sterile technique, intravenous catheters were placed in the external jugular (EJ) and femoral veins, and an arterial line in the carotid artery. All drugs and fluids were given through the EJ vein, while all plasma samples were taken from the femoral vein, to avoid contamination from the infusion site. Pigs underwent median sternotomy, heparinized to achieve an activated clotting time between 175 and 200 seconds, ascending aorta and right atrial appendage were cannulated, with a 16 French wire reinforced arterial cannula and 32 French venous cannula (both Edwards Lifescience, Irvine, CA) for central ECMO. The ECMO circuit was made up of 3/8 inch tubing (Maquet Softline Coating, Hirrlingen, Germany), a Biomedicus BPX-80 pumphead (Medtronic, Minneapolis, MN), and a Maquet Quadrox ID adult oxygenator. In addition, a Maquet reservoir was in line for priming and volume supplementation. The circuit was primed with 400 ml of Plasma-lyte (Baxter International Inc., Deerfield, IL). After collecting pre-ECMO timed blood samples (below), ECMO support was initiated to maintain a flow of 2.2 L/m2 for a 30 kg pig for 2 hours.6 After vascular access was achieved, two baseline plasma samples were collected 15 minutes apart before administering any inotropes. The inotrope/vasopressor medications were prepared and mounted on a syringe pump programmed to set delivery rates as follows: epinephrine 0.05 μg/k/min, norepinephrine 0.05 μg/k/min, and dopamine at 5 μg/k/min. Next, all vasopressors were infused at the specified rates for 15 minutes; then, two more samples were collected at 15 minutes apart representing pre-ECMO drug levels. Timed plasma samples were collected again after initiation of ECMO at 5, 10, 15, 30 minutes, and then every 30 minutes for 2 hours. Catecholamine levels were measured using liquid chromatography tandem mass spectrometry (LC-MS/MS) with a lower limit of detection of 0.2 ng/mL for each analyte. The resulting concentrations at each time point were pooled for analysis. Prism 8.4 (Graph Pad) was used for statistical analyses. The median baseline catecholamine levels were 0.69 [0.44–1.23] ng/ml, 0.68 [0.52–0.71] ng/ml for NE, DOP, respectively, and below the limit of detection for EPI. Immediately before initiation of ECMO, the levels were 0.52 [0.34–0.79] ng/ml, 0.54 [0.46–0.71] ng/ml, and 0.09 [0.06–0.22] ng/ml for NE, DOP, and EPI, respectively, (Figure 1). After the initiation of ECMO support, plasma catecholamines numerically increased at 5 minutes but were not statistically different from baseline levels. A reduction in the catecholamine concentrations were observed at the 10 minute time point compared to the levels at 5 minute time point (p = 0.138, p = 0.045, p = 0.029 for NE, DOP, and EPI, respectively). These levels then returned to the baseline concentrations by 15 minutes and were sustained throughout the remaining 2 hours.Figure 1.: Catecholamine concentrations at sampling timepoints.In pigs supported with catecholamine infusions held at a constant rate when initiated on ECMO support showed a transient decrease in circulating catecholamines at 10 minutes followed by a return to steady-state concentrations by 15 minutes. In critically ill patients, maintaining therapeutic levels of inotropes/vasopressors is crucial in providing adequate circulatory support and monitoring the signs of improvement. Extracorporeal membrane oxygenation support is well known to alter drug availability of antibiotics and analgesics1,4; however, literature examining the effect of ECMO on catecholamine levels is sparse. In an ex vivo single-dose close-loop crystalloid-based ECMO circuit, Mehta et al.7 demonstrated a drop of dopamine and epinephrine levels from the baseline by 10% and 16% at 30 minutes after initiation of ECMO and a further decrease of 53% and 23% by 3 hours. When they tested the epinephrine levels in a similar setup, but with blood-based circuit, they only noticed a drop of 7% at 3 hours. These observations are in line with our study findings, where the blood-based circuit did decrease the catecholamine levels transiently but returned to steady-state level in less than an hour. The key factors associated with decreased drug levels on initiation of ECMO are higher lipid solubility, higher protein binding, and use of roller pump with silicone fiber oxygenator.1 Lipid solubility of a drug is standardized with octanol/water partition constant (logP).8 Drugs with higher logP such as fentanyl (4.05) compared to morphine (0.89) loose up 96% of steady-state level in 24 hours when placed on ECMO.9 The logP of epinephrine, norepinephrine, dopamine (−1.37, −1.24, and −0.98) are low. As we might expect based on this chemical property, we did not find a significant drop in drug concentration over the 2 hours of ECMO flow. The majority of current ECMO pumps are centrifugal pumps with hollow-fiber membrane oxygenators, like the one used for this study. These pumps have features known to decrease drug adsorption.1,9 Monitoring catecholamines levels in our swine ECMO model provide a real-life picture of changes in catecholamine levels to be observed in patients placed on ECMO. Because the catecholamine levels remain steady after a brief decrease in levels after initiating the ECMO, sustained hemodynamic support from catecholamine is expected in patients initiated on ECMO. A lack of sustained support as expected from catecholamine infusion is unlikely the result of drug adsorption to the ECMO circuit.
Background Donation after circulatory death (DCD) donors can expand the heart donor pool for transplantation, which primarily is dependent on donation after brain death (DBD) donors. Inherent to the DCD process is ischemia and reperfusion injury, predominantly mediated by interleukin‐1 (IL‐1) and IL‐18, the downstream mediators of inflammasome. Hypothesis Pharmacologic blockage of IL‐1 or IL‐18 with recombinant IL‐1 receptor antagonist (IL‐Ra) or IL‐18 binding protein (IL‐18 BP) prior to initiation of DCD process is superior to the myocardial protection with IL‐1 or IL‐18 blockade given at reperfusion alone. Methods Following clinical protocol, DCD mice model was developed with in‐situ warm ischemia time maintained at 40 minutes. Mice (strain C57bl6/j) were randomly assigned to DCD control, DCD IL‐1Ra early and late or DCD IL‐18 BP early and late groups (n = 8–10 each). Pharmacologic inhibition of IL‐1 and IL‐18 was obtained with recombinant IL‐1Ra or IL‐18BP, respectively. Anesthetized mice were ventilated while monitoring heart with EKG and echocardiography. After muscle paralysis with vecuronium, ventilatory support was terminated and cardiac asystole observed. Hearts were procured and reanimated for 90 minutes on a Langendorff system with Krebs Henseleit (KH) buffer at 37°C. Physiologic parameters including heart rate, perfusion rate, developed pressure (DP), +/− dP/dt were obtained at 15‐minute intervals. After reanimation, hearts were collected for molecular and histological analysis. In early inhibition groups IL‐1 receptor antagonist (10 mg/kg) or IL‐18 binding protein (1 mg/kg) were given intraperitoneally, 30 minutes prior to DCD process (termination of ventilation). In the late inhibition groups IL‐1Ra and IL‐BP, 1μg/ml and 1μg/ml respectively were added to KH buffer. Coronary sinus samples were analyzed for cardiac troponin‐I (cTnI) levels. Results Heart rate and perfusion flow rates were comparable in all groups. In IL‐1 inhibition group, myocardial contractility was better (improved developed pressure, dP/dt compared to control group, however the differences attained statistical significance for DP and −ve dP/dt only). No additional incremental benefit was noted with early inhibition of IL‐1. In IL‐18 group, there was better physiologic function of heart compared to control (DP, +/−dP/dt and rate pressure product, all P = <0.05). Compared to late IL‐18 inhibitor group the early IL‐18 inhibitor group did not have significant improvements in myocardial function parameters. Markers of cell injury were significantly better with IL‐18 inhibition compared to control and IL‐1 inhibition groups. Conclusion Pharmacologic blockade of IL‐1 or IL‐18 is protective to the DCD mice hearts, with better protection with IL‐18 inhibition compared to IL‐1 inhibition. Early treatment with IL‐1Ra or IL‐18BP did not afford any additional benefits compared to what is observed with late inhibition given at reanimation. Our study findings are relevant to the clinical practice as only late interventions are allowed in clinic DCD process. Support or Funding Information Work supported by Merit Review Award and American Association grant to Dr. Quader DCD Control DCD IL‐1 Inhibition DCD IL‐18 Inhibition N=10 Early IL‐1Ra N = 13 Late IL‐1Ra N = 8 Early IL‐18BP N = 8 Late IL‐18BP N=9 Functional Data Heart Rate b/min mean±SEM 396±14 418±13 401±15 445±20 440±10 Perfusion rate ml/min 1.8±0.1 2.2±0.1 2.2±0.3 2.8±0.2 2.4±0.1 Developed pressure mmHg mean±SEM 78.7±5.8 99.3±6.5 # 94.1±9.3 99.5±8.1 # 108.3±6.5 # Rate Pressure Product 29,233±3,293 37,344±3,005 34,198±3,634 41781±4327 # 43082±3191 # Rate of positive developed pressure dt/dp mmHg/ms mean±SEM 3256±266 3595±481 3272±695 4923±44 # 4744±204 # Rate of negative developed pressure −dt/dp mmHg/ms mean±SEM −2278±18 −2991±19 # −2992±28 # −3256±29 # −3632±29 # Makers of myocyte damage Eluate troponin level ng/ml 7.79±0.98 6.16±1.12 7.68±0.85 4.21±0.67 # 3.82±0.65 # TUNEL assay 2.09±0.53 2.13±0.54 2.02±0.76 0.66±0.18 # 0.99±0.23 DCD = Donation after Circulatory Death, IL‐IRa = IL‐1 receptor antagonist, IL‐18 BP IL‐18 receptor binding protein, SEM = Standard Error of Mean, # P<0.05 vs DCD control
Introduction: Hearts by donation after circulatory death (DCD) have the potential to increase the number of transplantable hearts. However, the DCD process promotes ischemia and reperfusion injury (IRI). Mitochondrial damage and NLRP3 inflammasome activation contribute to the DCD heart damage. Blockade of electron transport chain (ETC) with the reversible inhibitor amobarbital (AMO) or the NLRP3 inhibitor 16773340 (NLRP3-I) can mitigate IRI. Hypothesis: AMO and/or NLRP3-I given to the DCD hearts at the moment of heterotopic heart transplantation afford lasting protection to the DCD hearts. Methods: We induced the DCD process in anesthetized rats that were intubated and ventilated by paralyzing the animal with vecuronium bromide and removing the ventilatory support. Treatment groups were AMO (1 mM), NLRP3-I, AMO+NLRP3-I, and vehicle (N=8-10/group). The DCD hearts were procured after 25 min of in-body ischemia then flushed with ice-cold cardioplegia solution supplemented or not with the AMO. NLRP3-I was administered orally (100 mg/kg) to the recipient before HTx and once a day. Control beating-heart donors (CBD) were procured without ischemia by giving the cardioplegia. The donor hearts underwent heterotopic transplantation in the abdomen of recipients. After 14 days, the function of the DCD heart was measured using an intraventricular balloon. Results: The developed pressure (DP), the peak -dP/dt, and the rate pressure product (RPP) of DCD hearts were significantly reduced compared to the CBD controls (63±7% vs 99±8%; -851±113 vs -1221±123; 20,729±2,268 vs 11,874±1,987, respectively; all p<0.05). AMO, NLRP3-I, and AMO+NLRP3-I preserved the DCD heart function (all parameters, p<0.03). Double treatment AMO+NLRP3-I trended toward a better improvement without reaching statistical significance over single treatments. Conclusions: Inhibition of ETC with AMO alone or combined with NLRP3 inhibition preserved the function of DCD hearts.
