Pre-Analytical Determination of the Effect of Extended Warm or Cold Ischaemia on RNA Stability in the Human Ileum Mucosa

The use of banked human tissue, obtained with informed consent after elective surgical procedures, represents a powerful model for understanding underlying mechanisms of diseases or therapeutic interventions and for establishing prognostic markers. However, donated tissues typically have varying times of warm ischaemia in situ due to blood arrest or cold ischaemia due to procurement and transportation. Hence, before using these tissues, it is important to carry out pre-analytical studies to ensure that they are representative of the in vivo state. In particular, tissues of the gastrointestinal tract have been thought to have low RNA stability. Therefore, this study aimed to determine if extended warm or cold ischaemia times and snap-freezing or banking in RNA stabilization solution affects RNA integrity or gene expression in human ileum mucosa. In short, ileum mucosa was collected for up to 1.5 h and 6 h of simulated warm or cold ischaemia respectively. Subsequently, RNA integrity and gene expressions were determined. It was found that RNA integrity remained high over the course of warm and cold ischaemia examined and there were in general no significant differences between snap-freezing and banking in RNA stabilization solution. Following the same trend, there were in general no significant changes in gene expressions measured (MYC, HIF1α, CDX, HMOX1 and IL1β). In conclusion, RNA in the ileum mucosa is maintained at a high integrity and has stable gene expression over the examined time course of warm or cold ischaemia when banked in RNA stabilization solution or snap-frozen in liquid nitrogen. As the average warm and cold ischaemia times imposed by surgery and the process of tissue banking are shorter than the time period examined in this study, human ileum mucosa samples collected after surgeries could be used for gene expression studies.     

Experimental Appraisal of Factors Involved in Provision of Donor Livers

The results of a series of porcine orthotopic liver homotransplants have shown that hepatic damage is related almost entirely to the period of warm ischaemia following death. Low flow perfusion with modified plasma would seem to be a satisfactory method of preserving cadaveric livers for at least six hours.In man it would appear possible to use cadaver livers derived from patients who had or had not been treated with artificial ventilation during life, provided the period of warm ischaemia is not longer than 25 to 30 minutes.     

Subarachnoid haemorrhage: older patients have low cerebral blood flow.

Daily estimations of hemispheral cerebral blood flow using the xenon-133 inhalation technique was made in 116 patients during the first three weeks after subarachnoid haemorrhage. The patients'' cerebral perfusion on average remained less than the normal perfusion expected for their age (based on a single estimation of cerebral blood flow in 67 volunteers). On each separate day after subarachnoid haemorrhage cerebral blood flow was inversely related to the patient''s age. Older patients seem especially at risk of developing cerebral ischaemia after subarachnoid haemorrhage. The clinical outcome was more often unfavourable in older patients--that is, in those who tended to have the lowest cerebral blood flow. Present results support the view that episodes of low cerebral blood flow lead to a poor outcome after subarachnoid haemorrhage. Because of the risk of inducing cerebral ischaemia great care should be exercised by physicians administering hypotensive drugs to older patients after subarachnoid haemorrhage.     

Melatonin and its protective role in attenuating warm or cold hepatic ischaemia/reperfusion injury

Although the liver is the only organ with regenerative capacity, various injury factors induce irreversible liver dysfunction and end‐stage liver disease. Liver resection and liver transplantation (LT) are effective treatments for individuals with liver failure, liver cirrhosis and liver cancers. The remnant or transplanted liver tissues will undergo hepatic ischaemia/reperfusion (IR), which leads to oxidative stress, inflammation, immune injury and liver damage. Moreover, systemic ischaemia induced by trauma, stroke, myocardial ischaemia, haemorrhagic shock and other injury factors also induces liver ischaemia/reperfusion injury (IRI) in individuals. Hepatic IRI can be divided into warm IRI, which is induced by liver surgery and systemic ischaemia, and cold IRI, which is induced by LT. Multiple studies have shown that melatonin (MT) acts as an endogenous free radical scavenger with antioxidant capacity and is also able to attenuate hepatic IRI via its anti‐inflammatory and antiapoptotic capacities. In this review, we discuss the potential mechanisms and current strategies of MT administration in liver surgery for protecting against warm or cold hepatic IRI. We highlight strategies to improve the efficacy and safety of MT for attenuating hepatic IRI in different conditions. After the potential mechanisms underlying the interactions between MT and other important cellular processes during hepatic IR are clarified, more opportunities will be available to use MT to treat liver diseases in the future.     

Allopurinol Inhibits Lipid Peroxidation in Warm Ischaemic and Reperfused Rabbit Kidneys

Rabbit kidneys were subjected to 120min of warm ischaemia or to 120min of warm ischaemia followed by 60min reperfusion with blood in vivo before being removed, homogenised and incubated at 37°C for 90min. Lipid extracts were obtained and monitored for Schiff base (fluorescence emission 400-450 nm, excited at 360 nm), thiobarbituric acid (TBA)-reactive material (emission 553 nm, excited at 515 nm) and diene conjugates (absorbance at 237 nm). Samples removed before incubation were assayed for reduced glutathione (GSH) and oxidised glutathione (GSSG) to provide an index of glutathione redox activity (GSH : GSSG). Allopurinol injected systemically i.v. (a) 15mins before kidneys were clamped. (b) 15mins before they were reperfused or (c) as two injections (before clamping and before reperfusion) significantly inhibited these biochemical markers of lipid peroxidation. Administration before reperfusion had a markedly more pronounced effect than when allopurinol was given before warm ischaemia only. It is concluded that allopurinol is probably effective because of its ability to inhibit xanthine oxidase and consequently lipid peroxidation during reperfusion rather than by preventing loss of purine nucleotides from hypoxic cells during ischaemia.     

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.     

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.     

Influence of Pre-, Intra- and Post-Operative Parameters of Donor Liver on the Outcome of Isolated Human Hepatocytes

The aim of the present study was to analyse, retrospectively on a large panel of patients (149), the influence of the donor liver characteristics on the outcome of human hepatocyte isolation obtained from resected liver biopsies from surgical waste after hepatectomy. Among the pre-operative parameters, the type of disease, age and sex of the patient, previous chemotherapy, alcohol or tobacco consumption did not affect the yield, viability, attachment rate and function of the isolated human hepatocytes. Pre-operative biological and anatomopathological data indicated that, while mild steatosis (≤10% steatotic hepatocytes) did also not affect the outcome of hepatocyte isolation, stronger steatosis (>10% steatotic hepatocytes) tended to decrease hepatocyte yield. Cholestasis, as assessed by γ-glutamyl transferase serum values, significantly negatively correlated with the percentage of digested liver and the yield of viable cells. Intra-operative clamping time, that is, warm ischaemia, longer than 30 min was found to decrease both the percentage of digested liver and cell yield. Among the post-operative parameters, the percentage of digested liver decreased when biopsy weights were higher than 100 g, the use of glue tended to increase both the percentage of digested tissue and the yield of viable cells. In conclusion, human diseased livers appear to be a valuable source of isolated functional human hepatocytes. We recommend, for an optimal isolation, to use liver biopsies weighing less than 100 g, to glue the section surfaces of the biopsies and to avoid the use of moderate steatotic livers (>10% steatotic hepatocytes) and cholestatic livers, as well as livers undergoing warm ischaemia or clamping during resection due to the decrease in cell yield. This revised version was published online in July 2006 with corrections to the Cover Date.     

Abundance,territory sizes,and pairing success of male Henslow's Sparrows in restored warm‐ and cool‐season grasslands

Henslow's Sparrows (Ammodramus henslowii) are known to breed in restored grasslands consisting of either warm‐ or cool‐season grasses, but additional information is needed concerning their breeding biology in these two types of grasslands. We compared the abundance, territory sizes, and pairing success of male Henslow's Sparrows in grasslands in west‐central Missouri in 2010 and 2011 using a paired‐treatment design, where warm‐ and cool‐season grasslands were located in close proximity. Selection indices indicated no apparent preference by male Henslow's Sparrows for either type of grassland, and the territory sizes and pairing success of males in cool‐ and warm‐season grasslands did not differ. In addition, we found no significant differences in habitat structure between these grassland types. Thus, our results suggest that the warm‐ and cool‐season grasslands in our study provided suitable habitat for Henslow's Sparrows. Other investigators have drawn similar conclusions for Henslow's Sparrows and for grassland birds in general, with the structural characteristics and, for some species, the size of grasslands being more important for grassland birds than plant species composition.     

Assessment of a test of renal viability.

A test of renal viability using the uptake of 125I-iodohippurate by kidney biopsy specimens has been developed. It is applicable to all kidneys irrespective of the method of storage. The uptake of 125I-iodohippurate in experimental kidneys correlated well with warm or cold ischaemia time and subsequent renal function. The test was used for human cadaver kidneys offered for transplantation and there was good correlation between iodohippurate uptake and warm ischaemic time. With this test, prediction of renal function was accurate in 85% of human cadaver kidneys transplanted. Pulsatile perfusion had no effect on cadaver kidneys as measured by this test.     

The biogeography of the beachflea, Orchestia gammarellus (Crustacea, Amphipoda, Talitridae), in the North Atlantic with special reference to Iceland: a morphometric and genetic study

The beachflea [Orchestia gammarellus (Pallas)] is a trans‐Atlantic amphipod inhabiting the littoral fringe. In Iceland, its distribution is temperature‐limited; it has recently colonized Iceland's relatively warm south‐western coast, and in the cooler north‐west, several small populations inhabit isolated warm springs. We address two questions: (i) Do the warm spring populations show evidence of long‐term residence in Iceland, or of recent colonization? (ii) For the new south‐western Icelandic populations, can the source population be determined? We sequenced COI for 22 populations in Iceland, Nova Scotia, the Faroe Islands, the British Isles, Norway and Sweden. Morphometric analysis of a subset of populations assessed 16 continuous and five discrete characters. Genetically, we found a star phylogeny: a common haplotype was found at all sites except two neighbouring warm spring populations, and all haplotypes were within two base pairs of this common haplotype. Morphometrically, almost all populations examined differed significantly in some characters; however, the warm spring populations differed slightly more from each other than did other populations. Although the origins of the Icelandic populations could not be well resolved, our data are consistent with a recent European origin.     

Invasive external otitis: review of 12 cases.

Invasive external otitis is an infection caused by Pseudomonas aeruginosa that often occurs in elderly people with diabetes. Twelve cases that illustrate the problems associated with the clinical recognition and successful outcome of the condition were reviewed. The patients'' average age was 62.5 years, and they had been ill for an average of 1.8 months before admission to hospital. Predisposing factors included diabetes, swimming in a warm climate and the use of a hearing aid. Radionuclide bone scanning and surgical exploration revealed pathognomonic findings. Initial therapy was often suboptimal: one or more relapses occurred in seven of the patients. All of the patients were cured without relapse after a minimum of 4 weeks of therapy with tobramycin plus an anti-Pseudomonas penicillin. The average duration of the illness was 3.9 months. The outcome in invasive external otitis should be excellent if the condition is diagnosed early and appropriate therapy is instituted.     

Prevalence of asymptomatic myocardial ischaemia in diabetic subjects.

OBJECTIVE--To compare the prevalence of silent myocardial ischaemia associated with coronary artery disease in diabetic subjects with that in controls of similar age and sex. DESIGN--A controlled study in which subjects with positive findings on exercise electrocardiography, 24 hour electrocardiographic recording, or dynamic thallium scintigraphy (diabetics only) underwent coronary angiography. SETTING--Academic medical centre; referral based cardiology clinic. SUBJECTS--136 Diabetic subjects, of whom 72 (33 women, 39 men (mean age 46.0] were insulin dependent and 64 (19 women, 45 men (mean age 49.3] non-insulin dependent. 80 Controls matched for age and sex; all were clients of the Occupational Health Service of Oulu University Central Hospital or the State Occupational Health Service Station in Oulu in whom diabetes had been excluded by a glucose tolerance test. INTERVENTIONS--Any subject showing signs of myocardial ischaemia was referred for cardiac catheterization. MAIN OUTCOME MEASURES--Exercise electrocardiography and 24 hour electrocardiographic recording were regarded as positive if there were ST depressions of greater than or equal to 1 mm that were planar or downsloping and persisted for 0.08 seconds after the J point. Thallium tomographic imaging. With cardiac catheterisation, coronary artery lesions were classified as significant in half or more of the vessel lumen was narrowed, or insignificant if such narrowing was less than half. RESULTS--40 (29%) diabetes and four (5%) controls had positive results in one or more of the non-invasive tests. Coronary angiography was performed on 34 of the diabetics (six refused); 12 had significant coronary artery narrowing; seven had unimportant atherosclerosis; 15 had patent coronary arteries. Among the controls only one had unimportant atherosclerosis; the other three had patent arteries. CONCLUSIONS--These results confirm the high prevalence of asymptomatic myocardial ischaemia in diabetics. Non-invasive screening of diabetic subjects, however, does not seem justified because of the low preset probability of the presence of the disease and the inaccuracy of the available test methods.     

Smith fir population structure and dynamics in the timberline ecotone of the Sejila Mountain, Tibet, China

Smith fir (Abies georgei var. smithii), which grows on cool aspects with elevations ranging between 3600 m and 4400 m, is a major native dominant alpine tree species in Southeast Tibet, China. The smith fir population structure, dynamics and characteristics were investigated in the timberline ecotone of the Sejila Mountain. Results indicate that the minimum crown closure (≤20%) was reached at 4320 m above sea level, where two types of alpine species exist, smith fir and blackseed savin (Sabina saltuaria). On the warm aspects, blackseed savin is a dominant species. Forest line was formed gradually with a wider timberline ecotone. While on the cool aspects, smith fir is a dominant species. Forest line was formed clearly with a narrow timberline ecotone. Furthermore, the upper limits of the distribution of the alpine species were 4570 m and 4390 m on warm and cool aspects, respectively. The timberline ecotone widths of the two species on the warm and cool aspects were 250 m and 70 m, respectively. The optimal distribution of smith fir on the cool aspects was from 3700 m to 3800 m. The smith fir's diameter class distribution was of reversed “J” shape and its density was about 380 stem·hm?2, while the age structure appeared to be of pyramid-shape, suggesting an expanding population. Finally, the static life table and the survivorship curve showed that the populations had experienced severe environmental selection during their first 20 years, self-thinning between 60 to 100 years, and environmental changes related mortality at later stages. The smith fir's physiological life span was around 200 years, and its maximum age was about 400 years old.     

Regulation of autophagy protects against liver injury in liver surgery-induced ischaemia/reperfusion

Transient ischaemia and reperfusion in liver tissue induce hepatic ischaemia/reperfusion (I/R) tissue injury and a profound inflammatory response in vivo. Hepatic I/R can be classified into warm I/R and cold I/R and is characterized by three main types of cell death, apoptosis, necrosis and autophagy, in rodents or patients following I/R. Warm I/R is observed in patients or animal models undergoing liver resection, haemorrhagic shock, trauma, cardiac arrest or hepatic sinusoidal obstruction syndrome when vascular occlusion inhibits normal blood perfusion in liver tissue. Cold I/R is a condition that affects only patients who have undergone liver transplantation (LT) and is caused by donated liver graft preservation in a hypothermic environment prior to entering a warm reperfusion phase. Under stress conditions, autophagy plays a critical role in promoting cell survival and maintaining liver homeostasis by generating new adenosine triphosphate (ATP) and organelle components after the degradation of macromolecules and organelles in liver tissue. This role of autophagy may contribute to the protection of hepatic I/R-induced liver injury; however, a considerable amount of evidence has shown that autophagy inhibition also protects against hepatic I/R injury by inhibiting autophagic cell death under specific circumstances. In this review, we comprehensively discuss current strategies and underlying mechanisms of autophagy regulation that alleviates I/R injury after liver resection and LT. Directed autophagy regulation can maintain liver homeostasis and improve liver function in individuals undergoing warm or cold I/R. In this way, autophagy regulation can contribute to improving the prognosis of patients undergoing liver resection or LT.     

Weather Effects on Mobile Social Interactions: A Case Study of Mobile Phone Users in Lisbon,Portugal

The effect of weather on social interactions has been explored through the analysis of a large mobile phone use dataset. Time spent on phone calls, numbers of connected social ties, and tie strength were used as proxies for social interactions; while weather conditions were characterized in terms of temperature, relative humidity, air pressure, and wind speed. Our results are based on the analysis of a full calendar year of data for 22,696 mobile phone users (53.2 million call logs) in Lisbon, Portugal. The results suggest that different weather parameters have correlations to the level and character of social interactions. We found that although weather did not show much influence upon people''s average call duration, the likelihood of longer calls was found to increase during periods of colder weather. During periods of weather that were generally considered to be uncomfortable (i.e., very cold/warm, very low/high air pressure, and windy), people were found to be more likely to communicate with fewer social ties. Despite this tendency, we found that people are more likely to maintain their connections with those they have strong ties with much more than those of weak ties. This study sheds new light on the influence of weather conditions on social relationships and how mobile phone data can be used to investigate the influence of environmental factors on social dynamics.     

Age and growth of sailfish Istiophorus platypterus (Shaw in Shaw and Nodder, 1792) from Mazatlan, Sinaloa, Mexico

Age and growth of the sailfish Istiophorus platypterus were determined for the area off Mazatlan coast, Sinaloa, in the Gulf of California, between September 2002 and August 2003. The lower jaw-fork length and total weight of 572 specimens were measured, and the fourth dorsal spine was collected to determine age. The monthly variation of the sample size displayed a well-defined seasonal pattern, which peaked during the warm period (May?CNovember) and declined during the rest of the year (temperate period). Significant differences were detected in the size structure by sex during the temperate period, with females displaying larger sizes and greater abundance (Female:Male = 3.35:1). In the warm period the size structure was similar, with the sex ratio reaching F:M = 0.73:1. This suggests a different sex-related recruitment in the fishing zone, with males moving more actively than females. While female size remained relatively unchanged over the year, male size increased during the warm period. Age was estimated using the number of growth rings on the cross-sectioned fourth dorsal spine, after the number of absorbed growth rings in the vascularized zone had been estimated. Nine age groups were identified; group-5 was the most abundant, representing 31% of the catch. The trend of the monthly change in the percentage of opaque-edge spines and the average of the marginal increase rate indicated that the formation of growth rings displayed an annual pattern. The von Bertalanffy growth model was adequately fitted to age and back-calculated length data. Significant differences were detected when growth was compared between sexes; females grew faster than males.     

Tree biodiversity of warm drylands is likely to decline in a drier world

Warm drylands represent 19% of land surfaces worldwide and host ca. 1100 tree species. The risk of decline due to climate aridification of this neglected biodiversity has been overlooked despite its ecological and societal importance. To fill this gap, we assessed the risk of decline due to climate aridification of tree species in warm drylands based on spatialized occurrence data and climate models. We considered both species vulnerability and exposure, compared the risk of tree species decline across five bioregions and searched for phylogenetic correlates. Depending on the future climate model, from 44% to 88% of warm drylands' tree species will undergo climate aridification with a high risk of decline even under the most optimistic conditions. On a regional scale, the rate of species that will undergo climate aridification in the future varies from 21% in the Old World North, to 90% in Australia, with a risk of decline confirming the high level of risk predicted at the global scale. Using generalized linear mixed models, we found that, species more exposed to climate aridification will be more at risk, but also that species vulnerability is a key driver of their risk of decline. Indeed, the warm drylands specialist species will be less at risk due to climate aridification than species being marginal in warm drylands. We also found that the risk of decline is widespread across the main clades of the phylogeny and involves several evolutionary distinct species. Estimating a high risk of decline for numerous tree species in all warm drylands, including emblematic dryland endemics, our work warns that future increase in aridity could result in an extensive erosion of tree biodiversity in these ecosystems.     

Physiological Distribution of Digoxin in Human Heart

Using the recipient''s human heart removed at cardiac transplantation, the distribution of digoxin at both the cellular and subcellular level has been studied. In the presence of diffuse histological myocardial abnormalities tissue digoxin is decreased, but the subcellular distribution, presumably reflecting binding to a possible receptor site, is uniform. When the histological abnormality is focal then digoxin distribution is uniform.These results suggest that in the presence of myocardial ischaemia plasma digoxin concentrations may not reflect total myocardial levels accurately.     

Pre- and post-harvest influences on seed dormancy status of an Australian Goodeniaceae species, Goodenia fascicularis

Background and Aims

The period during which seeds develop on the parent plant has been found to affect many seed characteristics, including dormancy, through interactions with the environment. Goodenia fascicularis (Goodeniaceae) seeds were used to investigate whether seeds of an Australian native forb, harvested from different environments and produced at different stages of the reproductive period, differ in dormancy status.

Methods

During the reproductive phase, plants were grown ex situ in warm (39/21 °C) or cool (26/13 °C) conditions, with adequate or limited water availability. The physiological dormancy of resulting seeds was measured in terms of the germination response to warm stratification (34/20 °C, 100 % RH, darkness).

Key Results

Plants in the cool environment were tall and had high above-ground biomass, yet yielded fewer seeds over a shorter, later harvest period when compared with plants in the warm environment. Seeds from the cool environment also had higher viability and greater mass, despite a significant proportion (7 % from the cool-wet environment) containing no obvious embryo. In the warm environment, the reproductive phase was accelerated and plants produced more seeds despite being shorter and having lower above-ground biomass than those in the cool environment. Ten weeks of warm stratification alleviated physiological dormancy in seeds from all treatments resulting in 80–100 % germination. Seeds that developed at warm temperatures were less dormant (i.e. germination percentages were higher) than seeds from the cool environment. Water availability had less effect on plant and seed traits than air temperature, although plants with reduced soil moisture were shorter, had lower biomass and produced fewer, less dormant seeds than plants watered regularly.

Conclusions

Goodenia fascicularis seeds are likely to exhibit physiological dormancy regardless of the maternal environment. However, seeds collected from warm, dry environments are likely to be more responsive to warm stratification than seeds from cooler, wetter environments.Key words: Goodenia fascicularis, Goodeniaceae, Australia, physiological dormancy, seeds, temperature, soil moisture, maternal influence, climate