Renal preservation by hypothermic perfusion. III. The lack of influence of pulsatile flow

Rabbit kidneys were perfused at 5 °C with a plasma-like solution containing dextran 70 and bovine serum albumin, and were autografted 24 hr later. One experimental group was perfused at a constant pressure of 40 mm Hg, while the second group was perfused with a pulsatile pressure having a root-mean-square (rms) equivalent of 40 mm Hg; the pulse pressure was 15 mm Hg and the pulse rate 60 min^?1 The behaviour of the two groups during perfusion and after transplantation was similar. It is concluded that pulsatile flow is without benefit during renal preservation by hypothermic perfusion.     

Isolated sheep heart hypothermic (5–13 °C) perfusion with fresh blood: Successful preservation for 24–72 hours with continuous strong ventricular activity

Isolated lamb hearts perfused with fresh whole blood at 10 and 13 °C in an ex vivo perfusion circuit continuously contracted at a rate of 15 to 20 times/min with a peak left ventricular systolic pressure (LVPSP) up to 70 mm Hg. These contractions persisted for the duration of the hypothermic study, up to three days with no change in vascular resistance. On rewarming to 38 °C, the hearts resumed regular and efficient contractions. Hearts perfused at 5 °C, however, exhibited no electrical or mechanical activity during hypothermic preservation and were uniformly poorly preserved.Quality of heart preservation was improved if, prior to final cooling, hearts were first rewarmed to 38 °C, followed by cooling. Change of the support animal, or interruption of flow of fresh blood into the perfusion circuit, resulted in cessation of ventricular contractions, ventricular fibrillation, and poor organ preservation.     

Pulsatile flow during cardiopulmonary bypass preserves postoperative microcirculatory perfusion irrespective of systemic hemodynamics

The onset of nonpulsatile cardiopulmonary bypass is known to deteriorate microcirculatory perfusion, but it has never been investigated whether this may be prevented by restoration of pulsatility during extracorporeal circulation. We therefore investigated the distinct effects of nonpulsatile and pulsatile flow on microcirculatory perfusion during on-pump cardiac surgery. Patients undergoing coronary artery bypass graft surgery were randomized into a nonpulsatile (n = 17) or pulsatile (n = 16) cardiopulmonary bypass group. Sublingual mucosal microvascular perfusion was measured at distinct perioperative time intervals using sidestream dark field imaging, and quantified as the level of perfused small vessel density and microvascular flow index (vessel diameter < 20 μm). Microcirculation measurements were paralleled by hemodynamic and free hemoglobin analyses. The pulse wave during pulsatile bypass estimated 58 ± 17% of the baseline blood pressure waveform. The observed reduction in perfused vessel density during aorta cross-clamping was only restored in the pulsatile flow group and increased from 15.5 ± 2.4 to 20.3 ± 3.7 mm/mm(2) upon intensive care admission (P < 0.01). The median postoperative microvascular flow index was higher in the pulsatile group [2.6 (2.5-2.9)] than in the nonpulsatile group [2.1 (1.7-2.5); P = 0.001]. Pulsatile flow was not associated with augmentation of free hemoglobin production and was paralleled by improved oxygen consumption from 70 ± 14 to 82 ± 16 ml·min(-1)·m(-2) (P = 0.01) at the end of aortic cross-clamping. In conclusion, pulsatile cardiopulmonary bypass preserves microcirculatory perfusion throughout the early postoperative period, irrespective of systemic hemodynamics. This observation is paralleled by an increase in oxygen consumption during pulsatile flow, which may hint toward decreased microcirculatory heterogeneity during extracorporeal circulation and preservation of microcirculatory perfusion throughout the perioperative period.     

颈动脉窦内压的变化速率对反射性降压效应的影响

在成年狗身上制备右侧孤离颈动脉窦,借助灌流装置用饱和氧的任氏液对其进行灌流。灌流压为搏动性的。通过改变每搏泵出量、灌流管道的阻力和弹性来调节灌流压及其变化速率。用多导生理记录仪同步记录股动脉血压、窦内灌流压及其变化速率。本文主要观察窦内压的变化速率对降压反射的影响。在8只狗身上共进行了93次实验。结果表明,在窦内压相同的情况下,灌流压的上升速率愈快,降压效应愈明显,而其下降速率则无显著作用。已有资料证明搏动性窦内压所引起的降压效应较非搏动性压力更为明显。由此可见,狗的颈动脉窦压力感受器不仅对搏动性压力而且对其上升的变化速率也很敏感。     

Thin-film coupled fluid-solid analysis of flow through the Ahmed glaucoma drainage device

The Ahmed glaucoma valve (AGV) is a popular glaucoma drainage device, allowing maintenance of normal intraocular pressure in patients with reduced trabecular outflow facility. The uniquely attractive feature of the AGV, in contrast to other available drainage devices, is its variable resistance in response to changes in flow rate. As a result of this variable resistance, the AGV maintains a pressure drop between 7 and 12 mm Hg for a wide range of aqueous humor flow rates. In this paper, we demonstrate that the nonlinear behavior of the AGV is a direct result of the flexibility of the valve material. Due to the thin geometry of the system, the leaflets of the AGV were modeled using the von Kármán plate theory coupled to a Reynolds lubrication theory model of the aqueous humor flow through the valve. The resulting two-dimensional coupled steady-state partial differential equation system was solved by the finite element method. The Poisson's ratio of the valve was set to 0.45, and the modulus was regressed to experimental data, giving a best-fit value 4.2 MPa. Simulation results compared favorably with previous experimental studies and our own pressure-drop/flow-rate data. For an in vitro flow of 1.6 microL/min, we calculated a pressure drop of 5.8 mm Hg and measured a pressure drop of 5.2 +/- 0.4 mm Hg. As flow rate was increased, pressure drop rose in a strongly sublinear fashion, with a flow rate of 20 microL/min giving a predicted pressure drop of only 10.9 mm Hg and a measured pressure drop of 10.5 +/- 1.1 mm Hg. The AGV model was then applied to simulate in vivo conditions. For an aqueous humor flow rate of 1.5-3.0 microL/min, the calculated pressure drops were 5.3 and 6.3 mm Hg.     

Mechanical function and fatty acid oxidation in the neonatal pig heart with ischemia and reperfusion

We investigated mechanical function and exogenous fatty acid oxidation in neonatal pig hearts subjected to ischemia, followed by reperfusion. Isolated, isovolumically-beating hearts, from pigs 12 h to 2 days of age, were perfused with an erythrocyte-enriched (hematocrit approximately 15%) solution (37 degrees C). All hearts were studied for 30 min. with a perfusion pressure of 60 mmHg (pre-ischemia). One group of hearts (low-flow ischemia, N = 12) was then perfused for 30 min. with a perfusion pressure of approximately 12 mmHg. In the other group (no-flow ischemic arrest, N = 9), the perfusion pressure was zero for 30 min. Following ischemia in both groups, the perfusion pressure was restored to 60 mmHg for 40 min. (reperfusion). Pre-ischemia parameters for all hearts averaged: left ventricular peak systolic pressure, 99.0 +/- 2.0 mmHg; end diastolic pressure, 1.9 +/- 0.2 mmHg; coronary flow, 3.4 +/- 0.1 ml/min per g; myocardial oxygen consumption, 56.6 +/- 1.6 microliter/min per g and fatty acid oxidation, 33.4 +/- 1.4 nmol/min per g. During low-flow ischemia, hearts released lactate, and the corresponding parameters decreased to: 30.7 +/- 0.9 mmHg; 1.2 +/- 0.3 mmHg; 0.8 +/- 0.1 ml/min per g; 26.6 +/- 2.3 microliters/min per g and 12.9 +/- 1.1 nmol/min per g, respectively. Early in reperfusion in both groups, all parameters, except for fatty acid oxidation, exceeded pre-ischemia values, before recovering to near pre-ischemia values. Late in reperfusion, however, rates of fatty acid oxidation exceeded pre-ischemia rates by approximately 60%. Thus, the neonatal pig heart demonstrated similar recovery following 30 min of low-flow ischemia or no-flow ischemic arrest.(ABSTRACT TRUNCATED AT 250 WORDS)     

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage,Assessment, and Repair of Marginal Liver Grafts

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for liver transplantation die without receiving an organ transplant or are delisted for disease progression. One strategy to increase the donor pool is the utilization of marginal grafts, such as fatty livers, grafts from older donors, or donation after cardiac death (DCD). The current preservation technique of cold static storage is only poorly tolerated by marginal livers resulting in significant organ damage. In addition, cold static organ storage does not allow graft assessment or repair prior to transplantation.These shortcomings of cold static preservation have triggered an interest in warm perfused organ preservation to reduce cold ischemic injury, assess liver grafts during preservation, and explore the opportunity to repair marginal livers prior to transplantation. The optimal pressure and flow conditions, perfusion temperature, composition of the perfusion solution and the need for an oxygen carrier has been controversial in the past.In spite of promising results in several animal studies, the complexity and the costs have prevented a broader clinical application so far. Recently, with enhanced technology and a better understanding of liver physiology during ex vivo perfusion the outcome of warm liver perfusion has improved and consistently good results can be achieved.This paper will provide information about liver retrieval, storage techniques, and isolated liver perfusion in pigs. We will illustrate a) the requirements to ensure sufficient oxygen supply to the organ, b) technical considerations about the perfusion machine and the perfusion solution, and c) biochemical aspects of isolated organs.     

Hypoxic pulmonary vasoconstriction during steady and pulsatile flow in ferrets

We examined the effects of hypoxia and pulsatile flow on the pressure-flow relationships in the isolated perfused lungs of Fitch ferrets. When perfused by autologous blood from a pump providing a steady flow of 60 ml/min, the mean pulmonary arterial pressure rose from 14.6 to 31.3 Torr when alveolar PO2 was reduced from 122 to 46 Torr. This hypoxic pressor response was characterized by a 10.1-Torr increase in the pressure-axis intercept of the extrapolated pressure-flow curves and an increase in the slope of these curves from 130 to 240 Torr X l-1 X min. With pulsatile perfusion from a piston-type pump, mean pulmonary arterial pressure increased from 17.5 to 36.3 Torr at the same mean flow. This hypoxic pressor response was also characterized by increases in the intercept pressure and slope of the pressure-flow curves. When airway pressure was raised during hypoxia, the intercept pressure increased further to 25 +/- 1 Torr with a further increase in vascular resistance to 360 Torr X l-1 X min. Thus, in contrast to the dog lung, in the ferret lung pulsatile perfusion does not result in lower perfusion pressures during hypoxia when compared with similar mean levels of steady flow. Since the effects of high airway pressure and hypoxia are additive, they appear to act at or near the same site in elevating perfusion pressure.     

An improved isolated working rabbit heart preparation using red cell enhanced perfusate

The performance of isolated working rabbit hearts perfused with Krebs-Henseleit (KH) buffer was compared with those in which the buffer was supplemented with washed human red blood cells (KH + RBC) at a hematocrit of 15 percent. When perfused with KH alone at 70 cm H2O afterload and paced at 240 beats/minute, coronary flow was more than double, whereas aortic flow was 40-60 percent of that in hearts perfused with KH + RBC, regardless of left atrial filling pressures (LAFP). Peak systolic pressure reached a plateau at 120 mm Hg in KH + RBC, but at 95 mm Hg in the KH group. Stroke work, however, was similar in the two groups. Despite the high coronary flow, oxygen uptake by hearts perfused with KH was substantially less and did not respond to increases in LAFP as in those perfused with KH + RBC. There was a 20 percent drop in ATP and glycogen content after 90 minutes' perfusion. In contrast, isolated hearts perfused with RBC-enriched buffer remained stable for at least 150 minutes. Irrespective of the perfusate, triacylglycerol content of the muscle remained at similar levels throughout the course of study. Increasing RBC in the perfusate from 15 percent to 25 percent had no additional effect on cardiac performance or oxygen consumption. Our findings demonstrate that in the isolated working rabbit heart inclusion of RBC in the perfusate improves mechanical and metabolic stability by providing an adequate oxygen supply.     

Fluctuations in rat testicular interstitial oxygen tensions are linked to testicular vasomotion: persistence after repair of torsion

Testicular microvascular blood flow is known to exhibit vasomotion, which has been shown to be significantly altered in the short term following the repair of testicular torsion. This loss of vasomotion may ultimately be responsible for the loss of spermatogenesis observed after testicular torsion in rats. In the present study, testicular vasomotion and interstitial oxygen tensions were simultaneously measured prior to, during, and at various time points after repair of testicular torsion in the rat. Testicular torsion was induced by a 720 degrees rotation of the testis for 1 h. Laser-Doppler flowmetry and an oxygen electrode were used to simultaneously measure vasomotion and interstitial oxygen tensions (PO(2)), respectively. Pretorsion control testes had a mean blood flow of 16.3 +/- 1.3 perfusion units (PU) and displayed vasomotion with a cycle frequency of 12 +/- 0.2 cycles per minute and a mean amplitude of 4.2 +/- 0.3 PU. Mean testicular interstitial PO(2) was 12.5 +/- 2.6 mm Hg, which displayed a cyclical variation of 11.9 +/- 0.4 cycles per minute with a mean amplitude of 2.8 +/- 0.8 mm Hg. During the torsion period, both mean blood flow and interstitial PO(2) decreased to approximately zero. Upon detorsion, mean microvascular blood flow and mean interstitial PO(2) values returned to values that were not significantly different from pretorsion values within 30 min; however, vasomotion and PO(2) cycling did not return, even after 24 h. It was 7 days after the repair of torsion before a regular pattern of vasomotion and PO(2) cycling returned. These results demonstrate for the first time a correlation between testicular vasomotion and interstitial PO(2) cycling, and this correlation persists after the repair of testicular torsion.     

Organ vessel functions by pulsatile flow

It was found that changes of resistance, capacity and exchange vessel functions of the skeletal muscle depended on values of amplitude and frequency of perfusion blood flow in acute experiments on cats. Amplitude-frequency characteristics of hemodynamic parameters of these functions were nonlinear. Ranges of amplitude and frequency variation with significant shifts of these parameters were determined. Conditions of perfusion (constant flow or constant pressure) changed volume and direction of haemodynamic parameter shifts under the amplitude-frequency modulation of the pulsatile blood flow. The pulsatile blood flow oscillations exerted maximum influence on hemodynamic parameters of exchange vessel function. The author has proposed the wave regulation conception of organ vessel functions.     

Cardiac dynamics of the Langendorff perfused heart

The Langendorff perfused heart is studied in a closed system with (i) automatic regulations to maintain constancy of the perfusion column (Krebs-Henseleit + 0.5% albumin or 25-30% washed erythrocyte suspension), (ii) continuous recording of rate, coronary flow, and supravalvular aortic pressure. A microcomputer with software interface is used for storage treatment and on-line analysis of the recorded variables. In 38 preparations perfused with Krebs-Henseleit, minimal diastolic (61.2 +/- 2.8 mm Hg) is significantly below and peak systolic (98.7 +/- 3.6 mm Hg) significantly above perfusion pressure (80 mm Hg). Pressure difference between minimal diastolic and peak systolic (delta P) is 37.5 +/- 1.8 mm Hg. Increases in perfusion pressure will be associated with increases of coronary flow and delta P, which is also increased by isoprenaline administration. Oxygen consumption decreased by 76% when perfusion pressure was lowered from 80 to 60 mm Hg in hearts perfused with a 30% erythrocyte suspension. All of these experimental results were interpreted as indicating that delta P measured in this system resulted from an ejected volume (x acceleration) from the heart. The ejected volume corresponds to a valvular leak caused by the rigid nature of the system which is devoid of aortic compliance. delta P may be considered an index of left ventricular performance, an indication that the Langendorff preparation studied under the present conditions is a working heart. A 100-microliter volume constant infusion syringe for time administration of cardioactive drugs may be inserted at the base of the perfusion column to obtain dose-response effects.     

Effect of variations in pressure and flow on the geometry of isolated canine carotid arteries

A study is described in which the effects of hemodynamics on arterial geometry are investigated in vitro. A novel perfusion apparatus is employed to deliver pulsatile flow through excised canine carotid arteries under carefully controlled conditions. Data of perfused vessel diameter and arterial wall thickness are derived from the radial displacement of the pulsating vessel as measured using a scanning laser micrometer whose accuracy is determined to be 0.0125 mm (0.0005 in). The results of 30 perfusion experiments suggest that the hemodynamic variables of transmural pressure, pulse pressure and flow rate influence vessel size and radial strain. The physiologic implications of these findings are discussed.     

木犀草素改善低温下离体大鼠心脏保存效果的研究

目的:研究木犀草素是否能改善心脏停搏保存液(UW液)对离体大鼠心脏的低温保存效果。方法:将40只成年SD大鼠随机分成4组(n=10):对照组(UW组)、7.5μmol/L木犀草素小剂量组,15μmol/L木犀草素中剂量组及30μmol/L木犀草素大剂量组。利用Langendorff离体心脏灌流法,观察心脏在4℃含或不含木犀草素的UW液中保存12 h复灌60 min后心脏功能及超微结构变化,比较心脏冠脉流量(CF)、心肌含水量及冠脉流出液中磷酸肌酸激酶(CK)的释放量。结果:与对照组比较,添加木犀草素后,复灌期心脏的收缩功能(LVPSP,+dp/dtmax)与心脏舒张功能(-dp/dtmax)、冠脉流量在多个复灌时间点均优于对照组,心率在复灌60 min时也显著优于对照组;复灌过程中磷酸肌酸激酶的漏出量及低温保存后心脏超微结构的损伤也均明显低于对照组;随灌注时间延长木犀草素组心脏结构和功能的改善有剂量依赖性趋势;木犀草素对心肌含水量没有影响。结论:木犀草素能显著改善UW液对离体大鼠心脏的低温保存效果,对心脏有明显的保护作用,以30μmol/L的木犀草素大剂量组作用最显著。     

Chronic and intermittent hypoxia induce different degrees of myocardial tolerance to hypoxia-induced dysfunction

Chronic hypoxia (CH) is believed to induce myocardial protection, but this is in contrast with clinical evidence. Here, we test the hypothesis that repeated brief reoxygenation episodes during prolonged CH improve myocardial tolerance to hypoxia-induced dysfunction. Male 5-week-old Sprague-Dawley rats (n = 7-9/group) were exposed for 2 weeks to CH (F(I)O(2) = 0.10), intermittent hypoxia (IH, same as CH, but 1 hr/day exposure to room air), or normoxia (N, F(I)O(2) = 0.21). Hearts were isolated, Langendorff perfused for 30 min with hypoxic medium (Krebs-Henseleit, PO(2) = 67 mmHg), and exposed to hyperoxia (PO(2) = 670 mm Hg). CH hearts displayed higher end-diastolic pressure, lower rate x pressure product, and higher vascular resistance than IH. During hypoxic perfusion, anaerobic mechanisms recruitment was similar in CH and IH hearts, but less than in N. Thus, despite differing only for 1 hr daily exposure to room air, CH and IH induced different responses in animal homeostasis, markers of oxidative stress, and myocardial tolerance to reoxygenation. We conclude that the protection in animals exposed to CH appears conferred by the hypoxic preconditioning due to the reoxygenation rather than by hypoxia per se.     

Ex Situ Normothermic Machine Perfusion of Donor Livers

In contrast to conventional static cold preservation (0-4 °C), ex situ machine perfusion may provide better preservation of donor livers. Continuous perfusion of organs provides the opportunity to improve organ quality and allows ex situ viability assessment of donor livers prior to transplantation. This video article provides a step by step protocol for ex situ normothermic machine perfusion (37 °C) of human donor livers using a device that provides a pressure and temperature controlled pulsatile perfusion of the hepatic artery and continuous perfusion of the portal vein. The perfusion fluid is oxygenated by two hollow fiber membrane oxygenators and the temperature can be regulated between 10 °C and 37 °C. During perfusion, the metabolic activity of the liver as well as the degree of injury can be assessed by biochemical analysis of samples taken from the perfusion fluid. Machine perfusion is a very promising tool to increase the number of livers that are suitable for transplantation.     

Ten-hour preservation of guinea pig isolated hearts perfused at low flow with air-saturated Lifor solution at 26{degrees}C: comparison to ViaSpan solution

There is no suitable solution to preserve hearts for longer than 5 h between donor explant and recipient implant. Lifor is a fully artificial preservation medium containing both a nonprotein oxygen and nutrient carrier (nanoparticles) and cellular nutrients, including amino acids and sugars. We proposed that recirculated Lifor solution would satisfactorily preserve guinea pig isolated hearts perfused at low flow with no added O(2) at room temperature for 10 h. Hearts were isolated from 21 guinea pigs and perfused with Krebs-Ringer (KR) solution (97% O(2) and 3% CO(2)) at 37 degrees C. Heart rate, inflow and outflow O(2) tension, coronary flow, left ventricular pressure (LVP), and maximal and minimal rate of change in LVP (dLVP/dt) were measured. After baseline measurements, hearts were perfused with recirculated Lifor or ViaSpan equilibrated with room air at 15% of control flow at 26 degrees C for 10 h. Hearts were then perfused at 100% flow with KR for 2 h at 37 degrees C. A time control (untreated) group was perfused only with KR solution for 15 h. Lifor arrested and protected hearts against diastolic contracture and maintained a low O(2) extraction. Compared with time controls, Lifor led to a higher developed LVP and coronary flow; %O(2) extraction and cardiac efficiency were similar between these two groups. Hearts similarly treated with ViaSpan exhibited diastolic contracture and lower %O(2) extraction during treatment and, upon reperfusion with KR, exhibited continued diastolic contracture, no return of heart rate or contractility, low coronary flow, low %O(2) extraction, and marked infarction. For long-term cardiac protection, a suitable preservation solution recirculated at low flow and room temperature without supplemental O(2) would reduce the support apparatus required for transport. Lifor was far superior to ViaSpan in meeting these requirements.     

Response of rat coronary circulation to carbon monoxide and nitrogen hypoxia

The effects of nitrogen (N2) or carbon monoxide (CO) hypoxia on coronary flow were assessed in the isolated nonworking rat heart perfused via the aorta with oxygenated (95% O2-5% CO2) Kreb's Henseleit solution. After 30 min, the hearts were challenged with solutions containing either CO (10% CO-85% O2-5% CO2) or N2 (10% N2-85% O2-5% CO2) for 2 min (Challenge I). After recovery in oxygenated solution, the hearts were challenged with the alternate test solution (Challenge II). There were no significant differences in heart rate or pulse pressure between the hearts challenged with CO or N2. Coronary flow was significantly higher in the hearts challenged with CO regardless of the challenge sequence. Coronary flows (ml X min-1 X g dry wt) in the CO- and N2-treated hearts, respectively, were 61.5 +/- 4.5 and 52.9 +/- 2.3 after Challenge I, and 64.3 +/- 2.6 and 56.4 +/- 3.0 after Challenge II. Because PO2 and oxygen content were the same in both test solutions, the results suggest that CO has a direct effect on coronary artery vascular smooth muscle.     

Lipid peroxidation and cell viability in isolated hepatocytes in a redesigned oxystat system: evaluation of the hypothesis that lipid peroxidation, preferentially induced at low oxygen partial pressures, is decisive for CCl4 liver cell injury

An oxystat system is described which is capable of maintaining steady-state oxygen partial pressures (PO2) at levels between 0.1 and 300 mm Hg for hours or even days in incubations of respiring cells. The system was used to study effects of the hepatotoxin carbon tetrachloride (CCl4) on lipid peroxidation and cell viability in isolated hepatocytes from phenobarbital-pretreated rats at various steady-state PO2. At PO2 below 35 mm Hg, with a maximum effect at 7 mm Hg, CCl4 induced an immediate lipid peroxidation, the rate of which slowed down during further incubation. AT PO2 between 35 and 70 mm Hg, CCl4 initially induced only slight lipid peroxidation, while there was a significant increase in lipid peroxidation after approximately 30 min. At PO2 above 100 mm Hg, no lipid peroxidation was induced by CCl4. At PO2 of 70 mm Hg and below, with the maximum effect at 3 mm Hg, CCl4 also induced marked losses of cell viability. Under anaerobic conditions and at PO2 greater than 70 mm Hg, CCl4 was without effect on the viability of the liver cells. Cells isolated from the pericentral area of the liver lobule showed more lipid peroxidation and loss of cell viability than cells from the periportal area of the lobule. These results provide further evidence for the decisive role of lipid peroxidation, preferentially induced at low PO2, in CCl4 liver injury.