Significance of Perfusate Lactic Acidosis in Cadaveric Renal Transplantation

Thirty-two human cadaver kidneys were assessed by continuous perfusion before transplantation. Altogether, 26 were transplanted. Ten kidneys functioned immediately, of which nine had excellent renal function at three months. Sixteen had delayed onset of function, of these only five had excellent function at three months. There was no difference in warm ischaemic interval between the two groups or in ice storage time. The immediate function group, however, were perfused for twice as long as the delayed function group. The two groups could not be separated on donor history, but they could be precisely separated during perfusion by measuring lactic acidosis and lactic acid dehydrogenase (LDH). The biggest single difference between the groups was in terms of donor preparation.     

Lidocaine-containing Euro-Collins solution prevents renal injury in the isolated perfused canine kidney exposed to prolonged cold ischemia

Previous studies have reported a decreased incidence of delayed graft function after cadaveric transplantation with the use of lidocaine pretreatment of the donor. We evaluated the effects of lidocaine on prolonged cold ischemia and reperfusion injury in a canine model of isolated kidney perfusion (IPK). The purpose of this study was to evaluate the renal function of isolated perfused canine kidneys after 48 h of cold storage with Euro-Collins (EC) solution or EC solution plus lidocaine. Isolated perfused canine kidneys were randomized into four groups which contained six kidneys: I) cold flush with EC solution and immediately reperfused, II) cold flush with EC solution plus lidocaine and immediately reperfused, III) 48 h of cold storage with EC and reperfusion, IV) 48 h of cold storage with EC solution plus lidocaine and reperfusion. The measured renal functions were glomerular filtration rate, urine production, perfusate flow, urinary lactic dehydrogenase (ULDH), Na reabsorptive capacity, and tissue MDA levels. Histological examination was performed after reperfusion. The tubular functions of kidneys preserved with EC solution containing lidocaine were better when compared with the kidneys preserved with EC alone. Tubular injury marker levels (ULDH) in group IV were significantly lower than in group III and lidocaine also reduced lipid peroxidation during reperfusion. This is in agreement with the histological results. The results of the present study can be taken as evidence of the cytoprotective effect of lidocaine, which may therefore be accepted as a useful agent for kidney preservation.     

Preservation of Human Kidneys for Transplantation: Preliminary Results with a Gambro Perfusion Machine

Twenty-six human cadaveric kidneys have been preserved for up to 36 hours by continuous perfusion with 4·5% albumin solution on a Gambro machine. Of these, 20 kidneys were transplanted, and six were discarded owing to poor perfusion characteristics during storage. Immediate function was obtained with four kidneys, of which two had been preserved for over 30 hours.Satisfactory renal function has been obtained with 17 kidneys, and three kidneys were lost owing to rejection or technical complications. It is our experience that human kidneys can be successfully preserved for up to 36 hours by this technique.     

Damaging effect of subzero temperature (-4 degrees C) on rabbit renal function

The effect of exposing rabbit kidneys to -4 degrees C for 1 hr in the unfrozen state was evaluated by means of measurement of tissue slice K/Na ratio and whole organ creatinine clearance. Freezing was prevented in one series (groups SC1-SC3) by supercooling with temperature monitoring and in a second series by a 2 M mixture of propylene glycol and glycerol. The latter agent was introduced prior to storage and later removed before the viability testing using a perfusion method (groups CPA1-CPA4). The results indicated a significant loss of slice and whole organ function during this short period of supercooling. The injury did not appear to result from either the rapidity of cooling or the formation of ice. There was some loss of function resulting from perfusion itself. Since this injury was evident in the whole organ but not in the tissue slice it may be ascribed to a vascular affect. When this damage was taken into account the data indicated that cryoprotective agents appeared to protect against any additional damage resulting from 1 hr storage at -4 degrees C.     

Measurement by HPLC of Desferrioxamine-Available Iron in Rabbit Kidneys to Assess the Effect of Ischaemia on the Distribution of Iron Within the Total Pool

A method for the determination of desferrioxamine-available iron in tissue fractions is described which involves incubation with desferrioxamine, extraction of desferrioxamine and its iron-bound form, ferrioxamine, and quantitation of these two forms of the drug by reversed-phase hplc analysis. Chelatable iron levels in the 1‐10µMolar region could be accurately and reproducibly measured using this technique.

The desferrioxamine-available iron levels in both the cortex and medulla of rabbit kidneys were significantly elevated (up to 2-fold) after the organs had been subjected to 2 hours warm ischaemia or 24 hours cold storage at 0°C In hypertonic citrate solution. There was no change in the total iron content of the tissues under these circumstances and thus a redistribution of intracellular iron to more available pools had presumably taken place as a result of ischaemia. This redistribution of iron may be an important factor in the initiation of peroxidative damage to cell membranes upon reperfusion of the organ with oxygen.     

Pre-winter lipid stores in young-of-year Atlantic salmon along a north–south gradient

The pre-winter lipid stores of young-of-the-year (YOY, age 0 year) Atlantic salmon Salmo salar were analysed along a north–south gradient from c. 71 to 58° N, with winter conditions ranging from >200 days of ice cover to no ice. The rivers sampled in Northern Norway represent some of the most northerly S. salar rivers. There was an increase in lipid content with increasing latitude, and mean lipid content (size adjusted to common mass) for YOY in northern rivers were almost three times higher: 0·035 g compared to 0·013 g in southern rivers. The relationship was not sensitive to variation in sampling time or variation in YOY body size. The lipid stores, however, varied markedly between rivers and also between neighbouring rivers, indicating different strategies or opportunities for pre-winter lipid storage both at latitudinal and local scales.     

Measurement by HPLC of Desferrioxamine-Available Iron in Rabbit Kidneys to Assess the Effect of Ischaemia on the Distribution of Iron Within the Total Pool

A method for the determination of desferrioxamine-available iron in tissue fractions is described which involves incubation with desferrioxamine, extraction of desferrioxamine and its iron-bound form, ferrioxamine, and quantitation of these two forms of the drug by reversed-phase hplc analysis. Chelatable iron levels in the 1-10µMolar region could be accurately and reproducibly measured using this technique.

The desferrioxamine-available iron levels in both the cortex and medulla of rabbit kidneys were significantly elevated (up to 2-fold) after the organs had been subjected to 2 hours warm ischaemia or 24 hours cold storage at 0°C In hypertonic citrate solution. There was no change in the total iron content of the tissues under these circumstances and thus a redistribution of intracellular iron to more available pools had presumably taken place as a result of ischaemia. This redistribution of iron may be an important factor in the initiation of peroxidative damage to cell membranes upon reperfusion of the organ with oxygen.     

Brain-dead kidney donor: selection,care, and administration.

Over four and a half years a district general hospital provided 34 cadaveric kidneys for transplantation. All brain-dead patients were regarded as potential donors, flow charts being used to maintain circulation and urine formation and facilitate administration. With this system the time lapse between diagnosis of brain death and removal of kidneys ranged from three to six hours and ischaemia was minimised. It is concluded that adoption of the system by other hospitals of comparable size would result in enough good-quality kidneys to satisfy present needs, thus reducing the initial high failure rate and enabling more patients to be accepted for dialysis.     

Feeding Frequency and Appetite in Lean and Obese Prepubertal Children

To determine the effect of feeding frequency on appetite in normal weight (NW) and obese (OB) prepubertal children, we carried out a prospective, randomized interventional study of 18 NW and 17 OB children ages 6–10. Children received three or five feedings in random order on separate days. Total calories, carbohydrate, protein, and fat composition on each day were equal. Two hours following the last feeding, children were offered ice cream ad lib. The major outcome variable was kilocalories ice cream consumed. A visual analog scale to assess fullness was also administered before consumption of ice cream. We observed that OB children consumed 73.0 ± 37.4 kcal more after five feedings than after three feedings whereas the NW children consumed 47.1 ± 27.8 kcal less. There was significant interaction between meal pattern and weight group indicating that this change in ice cream consumption differed significantly between groups (P = 0.014 by two‐factor analysis). Ice cream intake/kg was less in OB compared to NW subjects (P = 0.012). Fullness ratings before ice cream did not differ by meal pattern or weight group. However, pre‐ice cream fullness predicted ice cream intake in NW but not OB children. In summary, OB and NW children differed in appetite response to meal frequency. Our data suggest that: (i) satiety in OB children is related more to proximity of calories (larger supper) than to antecedent distribution of calories and; (ii) NW children may be more prone to restrict intake based on subjective fullness.     

Renal preservation by hypothermic perfusion: 2. The influence of oxygenator design and oxygen tension

Rabbit kidneys have been perfused for 24 hr at 5 °C and tested by autotransplantation with immediate contralateral nephrectomy. The perfusate contained an extracellular balance of ions with dextran and bovine serum albumin. The circuit included a nonpulsatile pump, a 0.22-μm membrane filter, and an oxygenator.The efficiency of membrane, film, and bubble oxygenators was compared. Flat membrane oxygenators were found to be more efficient than tubing oxygenators, and although the bubble oxygenator was the most efficient it created problems with foaming. A simple film oxygenator provided the best compromise.Ten kidneys were perfused and transplanted in each of three experimental groups; a film oxygenator was used to provide partial pressures of oxygen of 676 mmHg, 137 mmHg, and 9 mmHg, respectively.There were no significant differences in behavior during perfusion, function after transplantation, or histology of transplanted kidneys in the three groups.The similarity of the results using a film oxygenator with those previously obtained with a membrane oxygenator indicates that complex membrane oxygenators are not necessary for preservation in our system. Moreover, the similarity of the results with oxygen tensions from 9 mmHg to 676 mmHg shows that deliberate oxygenation is also unnecessary.     

Hypothermic preservation of rabbit kidneys for 48 hours using low ionic strength solutions

Microwaves offer the prospect of rapid and uniform heating of frozen organs. This is significant in the context of cryopreservation, and particularly of vitrification, because microwave heating may help to avoid crystallization or recrystallization of ice during warming, minimize any effects of high cell density, and reduce thermal-mechanical stresses. Previous work has established a rationale for reducing the ionic strength of solutions used to prepare tissues for microwave heating, since this permits the use of lower frequencies, which makes heating more uniform, without increasing the risk of thermal runaway (T. P. Marsland, S. Evans, and D. E. Pegg, Cryobiology 24, 311-323, 1987). In this paper we report a study of two possible low ionic strength perfusates, in rabbit kidneys, using 48 hr of hypothermic storage and autotransplantation as the test system. This model was chosen because there is a great deal of basic information about it. Both a single-pass "flush" preservation solution and a solution designed for continuous perfusion gave excellent results. The continuous perfusion system, which would be the more suitable for introducing cryoprotectants, gave five of five surviving animals with peak serum creatinine levels of 353-555 mumol/liter normal histology in three cases, and only very minor damage in the other two. There would therefore seem to be no obstacle to the use of perfusates having a low ionic strength in renal cryopreservations studies.     

Ice encapsulation protects rather than disturbs the freezing lichen

Arctic and alpine terricolous lichens are adapted to harsh environments and are tolerant to extremely low temperatures when metabolically inactive. However, there are reports indicating that freezing can be lethal to metabolically active lichens. With a projected warmer and more unstable climate, winter precipitation at high latitudes will fall more frequently as rain, causing snowmelt and encapsulating terricolous lichens in ice or exposing them to large temperature fluctuations. Lichens are a major winter food source for reindeer in most parts of the circumpolar region. A laboratory experiment tested how three hydrated reindeer forage lichen species covered by snow, encapsulated in ice, or uncovered responded to storage at freezing temperatures and subsequent warming. Photosynthetic performance (maximal fluorescence of dark-adapted samples and net photosynthetic rates) was significantly lower in lichens not insulated by snow or ice, whereas there were few differences between the snow and ice treatments. It is suggested that snow and ice provide sufficiently moist environments to improve extracellular and reduce intracellular ice nucleation activity. Ice encapsulation, which is often lethal to vascular plants, did not have any negative effects on the studied lichens. The results indicate that complete snow and ice melt followed by refreezing can be detrimental to terricolous lichen ecosystems. Reduced lichen biomass will have a negative effect both on reindeer winter survival and the indigenous peoples who herd reindeer.     

Influence of curing times on the effectiveness of treatments with acetic acid on the control of P. digitatum on lemons

The restricted number of postharvest fungicides used in packing houses is leading to the selection of resistant strains of Penicillium digitatum (citrus green mould), one of the most common and serious pathogens during storage and marketing of lemons. Furthermore a growing concern for human health and a greater awareness for environmental conservation have multiplied the studies on new ecological technologies. Among the alternatives to synthetic postharvest fungicides, the use of acetic acid (classified as GRAS) together with a physical method such as curing, have led to encouraging results. In the present study is reported the combined use of curing, performed at reduced times compared to those reported to be effective, followed by acetic acid (AAC) treatments. Lemons of the variety "Limone di Massa" artificially inoculated with P. digitatum at a concentration of 10(4) spores/mL were cured for 0, 3, 6, 12 and 24 hours and then treated with three different concentrations of AAC (25, 50 and 75 microL/L) for 15 min. Fruit was then stored at 20 degrees C and 80% relative humidity (RH) for 9 days, when the number of decayed fruits was monitored. The same combined treatments were also carried out on naturally infected lemons, stored for 6 or 8 weeks at 5 degrees C and 90% RH. After 9 days of storage the lowest percentage of infected wounds, in artificially inoculated fruit, was 0% after 6 hours of curing followed by AAC fumigation performed at 50 microL/L, while lemons untreated or cured for three hours showed the worst results with 71.4 and 61.9% of rotted fruit respectively. In naturally infected lemons the best results were achieved with curing performed for 24 hours followed by AAC fumigation at 50 microL/L. In these cases the combined treatment reduced decay by the 91.0 and 66.5% after 6 or 8 weeks of storage respectively, if compared to untreated fruit. The weight loss was not affected by any of the treatments. These results show that a good control of green mould during storage could be achieved, on lemon fruit, by combining a reduced curing time of 24 hours to the effect of AAC. The best results were obtained after 6 week of storage even if a satisfactory control was observed after 8 weeks of storage.     

Contrasting behavior between two populations of an ice‐obligate predator in East Antarctica

The Austral autumn–winter is a critical period for capital breeders such as Weddell seals that must optimize resource acquisition and storage to provision breeding in the subsequent spring. However, how Weddell seals find food in the winter months remains poorly documented. We equipped adult Weddell seals after their annual molt with satellite‐relayed data loggers at two sites in East Antarctica: Dumont D'Urville (n = 12, DDU) and Davis (n = 20). We used binomial generalized mixed‐effect models to investigate Weddell seals’ behavioral response (i.e., “hunting” vs. “transit”) to physical aspects of their environment (e.g., ice concentration). Weddell seal foraging was concentrated to within 5 km of a breathing hole, and they appear to move between holes as local food is depleted. There were regional differences in behavior so that seals at Davis traveled greater distances (three times more) and spent less time in hunting mode (half the time) than seals at DDU. Despite these differences, hunting dives at both locations were pelagic, concentrated in areas of high ice concentration, and over areas of complex bathymetry. There was also a seasonal change in diving behavior from transiting early in the season to more hunting during winter. Our observations suggest that Weddell seal foraging behavior is plastic and that they respond behaviorally to changes in their environment to maximize food acquisition and storage. Such plasticity is a hallmark of animals that live in very dynamic environments such as the high Antarctic where resources are unpredictable.     

Freeze-drying and room temperature storage of coconut pollen

Experiments to determine the conditions necessary for room temperature storage of coconut pollen, particularly of the ‘Jamaica Tall’ variety, are described. Preliminary experiments using the “Speedivac 5 PS freeze-drier” over a wide range of drying times indicated that 15 minutes was better than the longer periods tried, and room temperature storage for periods in excess of three months was recorded. Pre-cooling with dry ice and acetone was found to be unnecessary. Centrifuging during drying lowered drying efficiency and good viability was retained only when initial pollen moisture content was low The effect of longer drying periods with centrifuging have not yet been investigated. The optimum residual moisture content for successful storage at room temperature was found to be in the range 3.5–10.0%. Germination was sometimes improved by exposing the pollen to moist air for two to four hours. Viable seed has been obtained following pollination with freeze-dried pollen; the results of further test pollinations are to be reported later.     

Histopathological changes in gerbil liver and kidney after aluminum subchronic intoxication

Young gerbil livers and kidneys were analyzed by means of light and electron microscope to assess the histopathological changes caused by prolonged systemic aluminum (Al) administration. The experimental group was injected with AlCl3 i.p. for 5 weeks, while litter mates received PBS as sham-injected controls or served as untouched controls. Mortality occurred in 33% of experimental and 12.5% of sham-injected groups. The animals were perfused intracardially with 1% glutaraldehyde plus 1% paraformaldehyde and samples of liver and kidneys were processed for aluminum and iron histochemistry and conventional light- and transmission electron microscopy. White deposits composed of cellular debris appeared on the surface of liver and kidneys and in the mesentery as a consequence of Al treatment. Adherences of Glisson capsule to the diaphragm, as well as scattered small foci of hepatocyte necrosis with non-caseificant microgranulomas and mild portal inflammation, developed in the experimental group. Sham-injected animals also exhibited these granulomas but to a lesser degree. Al deposits were found in experimental animal granulomas and inside macrophages cytoplasm scattered throughout the liver. Iron deposition appeared in pericentral hepatocytes of experimental animals, in granulomas and in portal spaces of the three groups of animals. Ultrastructurally, hepatocytes of experimental animals showed mitochondria hyalinization, disintegration of endoplasmic reticulum and clustering of ribosomes. Phagolysosomes appeared larger and occurred more frequently in both hepatocytes and Kupffer cells of experimental animals. In 2 out of the 6 experimental animals studied, tubular atrophy was present in the renal cortical region, the kidneys of the remaining animals appearing normal. Al and iron were found very occasionally in the kidney parenchyma of experimental animals, while isolated mesangial cells showed iron deposits in a few glomeruli of both experimental and the two groups of control animals.     

Methodological considerations for the human platelet 5-HT2A receptor binding kinetic assay

Analysis of an extensive database of human platelet 5-HT2A receptor binding assays has been conducted in order to identify factors that may affect the assay results. Despite anecdotal reports that storage of frozen platelet pellets may affect 5-HT2A binding affinity and capacity, no quantitative study has been reported in the literature. Analysis of binding data for 373 frozen samples with a storage time up to three years is presented in this paper. It is shown that prolonged storage significantly decreases binding. The loss of binding capacity begins in the first six month of storage. Bmax declines by half after 17 month. The impact of storage time on the binding affinity is much smaller. There is only about 20% increase in the value of affinity K(D) during the half-life of Bmax. Differences in sample storage time may partly explain discrepancies in results between different research groups. Nonspecific binding due to binding to filter material diminishes accuracy and reliability of the binding assays as a result of a decrease in the ratio of specific to nonspecific ratio. A data analysis based on our suggested mathematical model shows that this effect depends on tissue concentration in test tube and becomes pronounced when the concentration is below 0.1 mg protein/ml (at 0.2 nM of ligand). Above 0.1 mg protein/ml, percentage of specific to total binding exceeds 65%, which is an acceptable level for the ratio. The majority of the binding studies reported in the literature employed a tissue concentration more than 0.5 mg/ml, well above the minimal limit sufficient for a reliable assay. However, development of microassays to conserve precious tissue must take the limit into consideration.     

Antipyrine,paracetamol, and lignocaine elimination in chronic liver disease.

The plasma half lives of antipyrine, paracetamol, and lignocaine given by mouth were measured in 23 patients with stable chronic liver diseases of varying severity. Fifteen patients received all three drugs and 19 at least two. The half life of paracetamol was abnormally prolonged in nine out of 17 patients (mean 2-9 hours, normal 2-0 hours), of antipyrine in 10 out of 19 patients (mean 30-4 hours, normal 12-0 hours), and of lignocaine in 19 out of 21 patients (mean 6-6 hours, normal 1-4 hours). Prolongation of the half lives of all three drugs was significantly correlated with an increase of the vitamin-K1-corrected prothrombin time ratio and a reduction in serum albumin concentration. There was no correlation with serum bilirubin concentration or serum alanine aminotransferase activity. This suggests that impaired drug elimination was related to depressed hepatic protein synthesis. Considerable prolongation of the half life of one drug was invariably associated with delayed elimination of the others. The half life of lignocaine, however, was always the most prolonged and was a highly sensitive indicator of hepatic dysfunction. The pharmacokinetic characteristics of a drug as well as the severity of liver disease should be taken into account when considering drug dosage in patients with chronic liver disease.     

Effect of sulphur dioxide (SO2) and heat (35°C) stressors on plasma glucocorticoids and thyroxine levels in mice

The objective of this study was to determine acute effects of SO₂ and simultaneous exposure to SO₂ and heat on plasma glucocorticoids and thyroxine levels. Male mice (160) were divided into four treatment groups: control, heat, SO₂, SO₂ and heat. These groups were further divided into four time exposures, 30 min, 1, 12 and 24 hours. The effect of heat and SO₂ as compared to heat or SO₂ alone appears to be a more rapid and prolonged elevation of glucocorticoids, while thyroxine levels showed a more rapid depression compared to heat and a greater depression as compared to the SO₂ treatment group.     

A preliminary investigation into the roles played by the rectal gland and kidneys in the osmoregulation of the striped dogfish Poroderma africanum.

Presented here are the results of a preliminary investigation into ionic and osmotic regulation by the kidneys and rectal gland of the striped dogfish, Poroderma africanum. Fish with ligated rectal glands showed an increase in blood concentration of sodium and chloride within a short time period, reaching a maximum after four days. The blood concentration of the two ions then decreased over the following ten days. Control animals showed relatively unchanged blood-sodium and chloride levels, over the entire 14-day period. After salt loading, both control animals and those with ligated rectal glands showed initial rise in blood sodium and chloride levels, but these returned towards initial values within seven hours of injection. Comparison of the two groups indicates that the rectal gland may control blood-chloride levels more so than -sodium, although its action as a salt regulator does not seem very pronounced in either case. Urine and rectal gland fluid, were collected as a compound fluid, from normal fish, and the estimated cloacal salt loss is discussed. Urine from normal fish was also collected separately and was analysed for its contribution to salt loss. Results are discussed and compared with previous relevant findings.