Altered Immunogenicity of Donor Lungs via Removal of Passenger Leukocytes Using Ex Vivo Lung Perfusion.

Passenger leukocyte transfer from the donor lung to the recipient is intrinsically involved in acute rejection. Direct presentation of alloantigen expressed on donor leukocytes is recognized by recipient T cells, promoting acute cellular rejection. We utilized ex vivo lung perfusion (EVLP) to study passenger leukocyte migration from donor lungs into the recipient and to evaluate the effects of donor leukocyte depletion prior to transplantation. For this purpose, female pigs received male left lungs either following 3 h of EVLP or retrieved using standard protocols. Recipients were monitored for 24 h and sequential samples were collected. EVLP-reduced donor leukocyte transfer into the recipient and migration to recipient lymph nodes was markedly reduced. Recipient T cell infiltration of the donor lung was significantly diminished via EVLP. Donor leukocyte removal during EVLP reduces direct allorecognition and T cell priming, diminishing recipient T cell infiltration, the hallmark of acute rejection.

Pharmacological normalization of circulation after acute brain death.

BACKGROUND: Circulatory instability is a serious problem after brain death in organ donors. The hypotension is often counteracted with infusion of large amounts of crystalloid solutions, which may impair lung function leading to rejection of the lungs as donor organs. The aim was to show that the circulation can be normalized pharmacologically for 24 h in pigs after total removal of the brain and brainstem by decapitation (between C2 and C3). METHODS: Twenty-four 40-kg pigs (n = 8 × 3) were included: non-decapitated, decapitated, and decapitated with pharmacological treatment. All animals got the same basal fluid supply and ventilation. The pharmacological treatment consisted of the neuronal monoamine reuptake blocker cocaine and low doses of noradrenaline and adrenaline. Desmopressin, triiodothyroxine, thyroxine and cortisol were also given. RESULTS: After decapitation, a catecholamine storm occurred, with an increase of noradrenaline and adrenaline by a factor of 79 and 298, respectively. Thirty minutes later, the pigs were hypotensive. The median time to the aortic pressure that was less than 40 mmHg was 9:09 h (range 5:50 to 22:01). After 6 h, the concentration of thyroid hormones and cortisol was significantly reduced. With pharmacological treatment of decapitated animals, the aortic pressure, renal blood flow, creatinine, urine production, liver function and blood gases did not differ significantly from the non-decapitated control animals. CONCLUSION: Pharmacological substitution of pituitary gland function, blockade of peripheral catecholamine neuronal reuptake and low doses of catecholamines normalize circulation in decapitated pigs throughout a 24-h observation period, whereas untreated decapitated pigs all develop severe circulatory collapse within 12 h.

Relation between cardiac dimensions and peak oxygen uptake.

BACKGROUND: Long term endurance training is known to increase peak oxygen uptake (VO2peak) and induce morphological changes of the heart such as increased left ventricular mass (LVM). However, the relationship between and the total heart volume (THV), considering both the left and right ventricular dimensions in both males and females, is not completely described. Therefore, the aim of this study was to test the hypothesis that THV is an independent predictor of VO2peak and to determine if the left and right ventricles enlarge in the same order of magnitude in males and females with a presumed wide range of THV. METHODS AND RESULTS: The study population consisted of 131 subjects of whom 71 were athletes (30 female) and 60 healthy controls (20 female). All subjects underwent cardiovascular MR and maximal incremental exercise test. Total heart volume, LVM and left- and right ventricular end-diastolic volumes (LVEDV, RVEDV) were calculated from short-axis images. was significantly correlated to THV, LVM, LVEDV and RVEDV in both males and females. Multivariable analysis showed that THV was a strong, independent predictor of (R2 = 0.74, p < 0.001). As LVEDV increased, RVEDV increased in the same order of magnitude in both males and females (R2 = 0.87, p < 0.001). CONCLUSION: Total heart volume is a strong, independent predictor of maximal work capacity for both males and females. Long term endurance training is associated with a physiologically enlarged heart with a balance between the left and right ventricular dimensions in both genders.

Respiratory indices by gas analysis and fat metabolism by indirect calorimetry in normal subjects and triathletes.

BACKGROUND: The effects of exercise on different indices reflecting the metabolism have been of interest for a long time, and a relationship between anaerobic indices and maximal oxygen uptake has been established. The inter-relationship between different respiratory indices during an exercise test remains to be studied in order to understand differences between individuals. Therefore, the aim of this study was to determine three respiratory indices and investigate their inter-relationship in individuals with highly variable working capacity. A second aim was to investigate the fat metabolism at the VO(2) corresponding to the respiratory compensation point (Pq) in the different subjects using indirect calorimetry. METHODS: Sixty control subjects (20 female) and 18 triathletes (six female) performed an exercise test with gas analysis. Three respiratory indices, derivative crossing (Dx), point of crossing (Px) and respiratory compensation point (Pq), were calculated using a computerized method. Fat metabolism at Pq was calculated using indirect calorimetry. RESULTS: Two different sequences of the respiratory indices were found: Dx

The negative inotropic effect of esmolol on isolated cardiac muscle.

OBJECTIVE: Esmolol is an ultra-short-term acting beta adrenergic blocker for intravenous use. The most common side effect is hypotension, which is often manageable by careful titration of the dose. We speculated whether esmolol had a direct negative inotropic effect on the cardiac muscle. DESIGN: Papillary muscles and trabeculae were excised from guinea pig and pig hearts. Force production was recorded together with action potentials. Membrane currents were recorded in isolated myocytes. The effects of two concentrations of esmolol were tested (55 and 110 micromol/L). RESULTS: Esmolol reduced action potential duration and plateau voltage, and decreased force production of isolated cardiac muscle. Voltage-clamp experiments from a holding potential of -40 mV and a step change to 0 mV showed a reduction in the inward current due to esmolol. CONCLUSION: Apart from being a beta adrenergic blocker esmolol also exerts a direct negative inotropic effect on cardiac muscle due to its inhibition of the calcium current during the action potential.

Disastrous physiological effects of saline on the cell membrane demonstrated in cardiac cells.

OBJECTIVE: Saline is not an ideal storage solution. It has a low pH, no buffering capacity, and lacks other ions and nutrients. The objective was to explore the effects of storing cardiac muscle in saline. DESIGN: Guinea pig papillary muscles and ventricular myocytes were exposed to saline. The effects on action potential, membrane current, contraction and cell shortening were recorded in vitro at 35-37 degrees C. RESULTS: Saline caused transient hyperpolarization of the resting potential (-140 mV), prolonged duration of the action potential, and increased contraction amplitude, which was later reversed. The membrane resting potential depolarized after a few minutes to about -15 mV and the preparations became unexcitable. The depolarized preparations remained slightly contracted. Upon reperfusion both papillary muscles and cells became unstable and spontaneously active. Storing myocytes in saline for only 2 h resulted in excessive cell death. CONCLUSION: Saline is disastrous for the function of the heart muscle and leads to depolarization, sustained contraction and unexcitable tissue. Saline should not be used as a storage medium, even for short periods of time.

Working capacity and resting energy expenditure after ileal pouch-anal anastomosis.

BACKGROUND: The aim was to evaluate the working capacity and resting energy expenditure in patients who had undergone restorative proctocolectomy. METHODS: Of 72 patients operated on between April 1990 to September 1998, 51 were eligible and 38 participated in the study. Resting energy was assessed by indirect calorimetry, and working capacity by ergospirometry on an exercise bicycle. RESULTS: The median functional score was 2 (range 0-7). Oxygen uptake during rest was reduced for men compared with predicted values. The corresponding values for women were in keeping with predicted values. The median working capacity was 96 (range 59-102) per cent for women and 91 (range 51-113) per cent for men, compared with reference values of maximum workload based on age, height and sex. There was no correlation between functional score and any other variable measured. CONCLUSION: Patients who have undergone restorative proctocolectomy for ulcerative colitis have normal resting energy expenditure and working capacity.

Reference values for the physical work capacity on a bicycle ergometer for women between 20 and 80 years of age.

A sample of women (n=87) uniformly distributed in age (20-80 years) was randomly selected from the municipal population register and conventional clinical exercise tests were performed on a bicycle. The load started at 30 W and increased (5 W/30 s) until exhaustion. Care was taken to perform a maximum exercise test. The women were required to be cardiovascularly healthy and not on regular medication to be included in the study. The maximum work load was found to be dependent on age and height. The following equation described the maximum load. Max load (W)=(137.7 * Height (m) - 23.1)/(1 + exp (0.064 * (Age (years) - 75.9))). The upper and lower limits of the reference interval were 120 and 80% of the predicted load, respectively. The upper and lower limits for maximum heart rate were 110 and 90% of the maximum heart rate given by 190.2/(1 + exp (0.0453 * (Age - 107.5))). The reference value for maximum systolic blood pressure was taken as the interval between the two lines 153.3 + 0.281 * Age and 172.0 + 1.13 * Age. The reference values presented for work capacity are higher than those normally used in Sweden.

High-frequency components in ECG analysed in guinea-pig Langendorf preparations.

High-frequency components in ECG during global ischaemia were studied in isolated guinea-pig hearts perfused ad modum Langendorf. Electrocardiograph recordings were carried out from the epicardial surface both in normo- and low-flow perfusion. After bandpass filtering (5-500 Hz), signal-averaging, was undertaken. The high-frequency components either increased or decreased after low-flow perfusion was instituted. Root-mean-square voltage (RMS) of the depolarization signal correlated poorly with the signal amplitude, but highly with the first and second derivative, i.e. the velocity and the acceleration of the signal. It is concluded that high-frequency components are not pathological phenomena per se, but reflect the shape of the original electrocardiographic signal.

Arrhythmia as a result of poor intercellular coupling in the sinus node: a simulation study.

The effects of reduced intercellular coupling in the sinus node were investigated by means of simulations. Coupling was reduced both uniformly, and by introducing localized interaction blocks. In either case, model sinus node element networks typically splitted into frequency domains. These were defined as groups of neighbour elements which all attained the same mean firing frequency. In systems, simulating the vicinity of an impulse outlet to the atrium, the sinus node elements often splitted into two domains, one slowly firing just inside the outlet, and one normally firing large domain in the sinus node interior. This two-domain situation was analysed using a two-element system. Wenckebach conduction and advanced (m:1) exit blocks were seen, together with more odd block patterns and slow chaotic rhythms. The two-domain situation appeared also when two discrete outlets were considered. The slow domains around each outlet synchronized via the atrium. However, if there were some degree of exit block through one of the outlets only, brady-tachy like rhythms could be simulated due to a re-entrant circuit including both sinus node and atrial tissue. In conclusion, poor coupling in the sinus node seems to be sufficient to produce most arrhythmias in the sick sinus syndrome

Simulated sinoatrial exit blocks explained by circle map analysis.

In an accompanying study, it was seen that most cardiac arrhythmias that were simulated during poor intercellular coupling in the sinus node, were the same as those obtained in a two-element system in which one element suffered from a strong leakage current. This element corresponds to the sinus node periphery and is thus the one which feeds the atrium. In this paper, the interior element was replaced by a periodic stimulator. The dynamics of the peripheral element is then determined by its phase response curve. Phase response curves for sinus node elements subject to leakage were simulated for many different amplitudes of depolarizing stimuli. Simulations with circle maps based on these curves produced the same sequence of progressing levels of exit block as stimulus strength decreased, as did the two-element system when coupling strength was reduced. The bifurcations of the circle maps leading to the observed rhythms were identified. We found that the essential qualities of the phase response curves were determined by generally accepted properties of membrane currents. This suggests that the observed rhythms and bifurcations are generic.

Tolerability of concurrent use of nicotine gum and smoking in healthy volunteers.

Although nicotine replacement therapy (NRT) is mainly licensed as an aid for smoking cessation, many smokers alternate NRT with cigarettes. This study evaluated the cardiovascular tolerability of nicotine gum plus smoking. This open, three-way, randomized, multiple-dose, crossover study involved 19 healthy adult volunteer smokers. Three treatments (placebo gum+cigarette vs. nicotine 4-mg gum+unlit cigarette vs. nicotine 4-mg gum+cigarette) were each administered hourly seven times during one study day. Plasma nicotine, ECG variables (ST60 and ST(slope)), heart rate, and blood pressure were measured at rest and during/after 5 min exercise tests. Mean plasma nicotine levels were higher with gum plus smoking than with either gum or smoking alone. After the final administration, mean levels with gum plus smoking were 39.5 (range 27.6-54.4) ng/ml vs. 19.6 (11.8-30.1) ng/ml with gum and 22.6 (6.6-36.6) ng/ml with smoking alone. No signs of myocardial ischaemia were observed during concomitant use of gum plus smoking, despite high plasma nicotine levels. Decreases of ST60 and ST(slope) were observed in all treatment groups during exercise but were asymptomatic. Concomitant gum plus smoking did not increase the incidence of extrasystoles or arrhythmia. No serious adverse events occurred during the study. In conclusion, there were no signs of myocardial ischaemia during multiple submaximal exercise tests in healthy volunteers while smoking and using nicotine 4-mg gum.

Day-to-day variation in oxygen consumption at submaximal loads during ergometer cycling by adolescents.

The day-to-day variation in oxygen consumption (VO2) during ergometer cycling by 20 healthy adolescents, 10 females and 10 males, was measured using indirect calorimetry. The two sets of measurements were performed on two consecutive days. Great care was taken to minimize possible disturbing factors. Cycling started at 50 and 100 W for female and male adolescents, respectively. The load was increased at a rate of 5 W 30 s(-1). In order to reach steady state, the load was kept constant for 3.5 min twice during the cycling session, at 100 and 130 W for the females and at 130 and 160 W for the males. Cycling continued until exhaustion. The maximal loads were 196 W (mean) and 271 W (mean) for females and males, respectively. At the maximal loads the day-to-day variation (+/-2 SD) in oxygen consumption (VO2) was +/-330 ml min(-1) for females and 390 ml min(-1) for males. At the submaximal loads the day-to-day variation in heart rate (HR) was 9.3 beats min(-1) (+/-2 SD) (coefficient of variation, CV=3.4% at 130 W) for both sexes. The day-to-day variation in oxygen consumption (VO2) was +/-199 ml min(-1) (+/-2 SD) at the different submaximal loads and did not differ between female and male adolescents (CV=5.7% at 130 W). This natural day-to-day variation must be taken into consideration when using a submaximal ergometer cycling test for the evaluation of physical capacity in the two sexes.

[Ca2+]i following extrasystoles in guinea-pig trabeculae microinjected with fluo-3 - a comparison with frog skeletal muscle fibres.

Force production of cardiac muscle is highly dependent on the interval between the excitations. The aim was to investigate relations between intracellular calcium ([Ca2+]i) and force when a stimulus protocol, with three extrasystoles (ESs) at various intervals, was used. The relation between [Ca2+]i and force was compared with that in frog skeletal muscle fibre. Fluo-3 was microinjected into thin cardiac trabeculae to monitor [Ca2+]i. During steady-state [Ca2+]i consisted of a rapid rise (phase 1) that lasted until peak dF/dt (rate of force development) and was followed by a slower rise (phase 2) that coincided with the action potential and had a peak after peak force. The decline in [Ca2+]i outlasted the duration of the contraction. As the ES intervals were prolonged, there was a gradual restitution of force and of the amplitude and rate of rise of phase 1 [Ca2+]i. Peak dF/dt was linearly related to the amplitude of phase 1 [Ca2+]i during restitution and potentiation of force. Skeletal muscle fibres were loaded with fluo-3-AM. From [Ca2+]i the amount of calcium bound to troponin ([Ca-T]) as a function of time was estimated. Force production of the skeletal muscle fibre could be predicted from [Ca-T] when the signal was delayed (time constant 36 ms). This finding indicates that the recorded [Ca2+]i in skeletal muscle represents activator calcium. In cardiac muscle probably only phase 1 [Ca2+]i represents activator calcium. Phase 2 [Ca2+]i probably represents calcium entry during the action potential and does not activate the contractile system to any significant extent.

Vectorcardiographic loop alignment and the measurement of morphologic beat-to-beat variability in noisy signals.

The measurement of subtle morphologic beat-to-beat variability in the electrocardiogram (ECG)/vectorcardiogram (VCG) is complicated by the presence of noise which is caused by, e.g., respiration and muscular activity. A method was recently presented which reduces the influence of such noise by performing spatial and temporal alignment of VCG loops. The alignment is performed in terms of scaling, rotation and time synchronization of the loops. Using an ECG simulation model based on propagation of action potentials in cardiac tissue, the ability of the method to separate morphologic variability of physiological origin from respiratory activity was studied. Morphologic variability was created by introducing a random variation in action potential propagation between different compartments. The results indicate that the separation of these two activities can be done accurately at low to moderate noise levels (less than 10 microV). At high noise levels, the estimation of the rotation angles was found to break down in an abrupt manner. It was also shown that the breakdown noise level is strongly dependent on loop morphology; a planar loop corresponds to a lower breakdown noise level than does a nonplanar loop.

Effects of [Na+]o, [Ca2+]o and cyclopiazonic acid on decline of post-extrasystolic potentiation and twitch kinetics in guinea-pig and human myocardial preparations.

The decline of post-extrasystolic potentiation was investigated in atrial and ventricular muscle of guinea-pigs and in human atrial muscle. This decline is described in terms of the recirculation fraction (RF) for calcium in myocardial cells. Under control conditions, the mean values for RF were as follows: 0.61 +/- 0.03 in the guinea-pig atrium, 0.32 +/- 0.05 in the guinea-pig ventricle and 0.51 +/- 0.10 in the human atrium. Upon increasing calcium concentrations within the range of 0.5-10 mM or decreasing the sodium to 70%, the steady-state force and recirculation fraction increased concomitantly in all three types of muscles. Cyclopiazonic acid (CPA), in a 5-20 microM concentration, decreased force in the guinea-pig preparations and decreased RF. The effects of CPA were great in atrial muscle, 20 microM CPA decreasing RF from 0.61 to 0. 39. In ventricular muscle, this effect was small and not statistically significant. When changing calcium or sodium concentrations, increased force was accompanied by slower relaxation in atrial muscle, but an unchanged or slightly faster relaxation in ventricular muscle. In guinea-pig myocardium, CPA (5-20 microM) prolonged the time to peak force (TPF), but it slowed relaxation only in the ventricle. Reducing the calcium outflow by Na/Ca exchange increased RF as expected, and reducing the SR calcium re-uptake decreased RF. These interventions were also expected to reduce the rate of decline of intracellular calcium, but relaxation was not consistently prolonged. This indicates that factors other than the SR calcium pump and sarcolemmal Na/Ca exchange determine, or at least influence, the rate of relaxation.

Day-to-day variation in oxygen consumption and energy expenditure during submaximal treadmill walking in female adolescents.

The day-to-day variation in oxygen consumption (VO2) and energy expenditure (EE) during horizontal treadmill walking was measured using indirect calorimetry in 20 female adolescents (mean age 17.3 years). Two different walking speeds were used: 5 km h-1 and an individually convenient speed of 3.0 km h-1 (mean). The two sets of measurements were performed on 2 consecutive days, and great care was taken to minimize possible disturbing factors. The mean VO2 was 919 ml min-1 at 5 km h-1 and 622 ml min-1 at the individual speed, and the mean values of EE were 4.5 kcal min-1 and 3.1 kcal min-1 respectively. The individual day-to-day variation in VO2 (at 5 km h-1) was between -11.7% and +12.6% of the mean VO2. The coefficient of variation (CV) was 6.4% when values were calculated in ml min-1 kg-1. The energy expenditure varied somewhat less between the 2 days (CV = 5.7%). The corresponding value for EE when walking at the individual speed was 7.2%, and the mean day-to-day variation in VO2 was 7.5% (CV). The rate of perceived exertion according to Borg's scale was lower on day 2 (11.9) compared with day 1 (13.0) when walking at 5 km h-1. There was no difference in heart rate between the 2 days. It is concluded that EE varies somewhat less than VO2 on successive days, probably because of an interchangeable relationship between breathing gases, depending on which substrate is used for combustion. When using VO2 and EE for evaluation of physical capacity, the day-to-day variation in the measurements must be taken into consideration.

Dissociation between force and [Ca2+]i during extra systoles in guinea-pig ventricular muscle microinjected with fura-2.

Thin trabeculae were dissected from the right ventricle of guinea-pig heart and stimulated to contract isometrically at 0.5 Hz (26 degrees C). Rapid and transient changes of force were obtained by inducing three extra systoles (ES1-3) at 450-ms intervals. The two regular contractions (P1-2) following (ES1-3) were potentiated. Fura-2 salt was microinjected into the preparation to monitor intracellular calcium ([Ca2+]i). Three distinct phases of [Ca2+]i were seen: (1) a rapid rising phase to about 200 nmol L(-1), (2) a slower rising phase to a peak at 400 nmol L(-1), and (3) a slow decline to about 50 nmol L(-1). During ES1, there was a discrepancy between force, which decreased, and peak [Ca2+]i, which increased to 600 nmol L(-1). It is likely that the increased [Ca2+]i during the extra systoles reflects increased sarcolemmal calcium inflow, causing post-extra-systolic potentiation. Ryanodine (1-2 microM) was added to inhibit the intracellular calcium release and thus reduce the intracellular [Ca2+]i gradients following excitation. Ryanodine inhibited phase 1 of [Ca2+]i and abolished post-extra-systolic potentiation. There was a close relationship between dF/dt and [Ca2+]i with ryanodine during control and ES1-3. It is likely that fura-2 reports a spatially averaged [Ca2+]i and that phase 1 of the signal therefore apparently underestimates activator calcium in the close vicinity of the contractile elements.

Properties of spiral waves in a piece of isotropic myocardium.

Tachyarrhythmias of the heart can be due to the presence of one or more spiral waves of electrical activity. Spiral waves were simulated using a previously described ionic model of cardiac action potentials in a 75 x 75 network of compartments. The compartments were connected by means of resistors and made isotropic in order to catch basic properties of spiral waves. The cross-field stimulation technique was used to generate single or double spiral waves. The analysis showed that a spiral wave was created when the second excitation front became critically curved, in the wake of the preceding wave, so that decremental propagation occurred. A spiral wave could also be generated from a wave circulating around an obstacle when the obstacle size was suddenly reduced. The spiral waves steadily circled around an area with excitable but unexcited cells. An undisturbed spiral wave in the isotropic medium circled around in a stable pathway, but drifted along the borders of cells made non-excitable. An excitation within an existing spiral wave could generate new spiral waves that interacted with each other and formed complex excitation patterns. A sudden prolongation of the refractory period reduced the central area with unexcited cells in the spiral pathway but only slightly prolonged the revolution time. A further prolongation of the refractory period extinguished the spiral wave when the tip of the spiral wave invaded refractory areas. The described ionic compartment model could accurately produce spiral waves with properties in line with experimental results reported by others.

Beat-to-beat QRS variability in the 12-lead ECG and the detection of coronary artery disease.

The aim of this article was to study beat-to-beat QRS variability in patients with ischemia and old myocardial infarction using the 12-lead resting electrocardiogram (ECG). The variability analysis was based on beats that have been synchronized in time with an iterative alignment technique. The QRS variability was measured in patients submitted for myocardial scintigraphy. Those with a normal myocardial scintigraphy (called NO, n = 34, mean age 57 years, 23 women) were compared with a group with both myocardial infarction and exercise-induced ischemia (called ISCINF, n = 27, mean age 57 years, 5 women). The mean QRS variability was somewhat smaller in lead I in ISCINF than in NO, and there was no statistically significant difference in QRS variability among the groups in leads II, III, and V1-V6. Using a multivariate approach, the joint variability in leads I, II, II, and V1-V6 was used for calculating receiver operating characteristics based on a leave-one-out procedure. The sensitivity for detecting coronary artery disease was 75% at a specificity of 50%. It is concluded that beat-to-beat QRS variability in the 12-lead ECG does not discriminate between the presence and absence of coronary artery disease sufficiently well for clinical purposes.