Hepatic cold hypoxia and oxidative stress: implications for ICAM-1 expression and modulation by glutathione during experimental isolated liver preservation

Cold preservation and reperfusion of liver during transplantation are necessary steps in the procedure but which are also associated with damage to the organ. One aspect of this damage is thought to concern up-regulation of inflammatory markers, such as the adhesion molecule intercellular adhesion molecule 1 (ICAM-1) on target cells in the liver. This aids sequestration of activated leucocytes, which promote inflammation, by a complex sequence of events, including free radical mediated damage. We have studied changes in ICAM-1 in rat liver as a consequence of cold preservation for various times, and also after warm reperfusion during isolated liver perfusion. We have also investigated the effects of the free radical scavenging agent (reduced glutathione-GSH) on the modulation of ICAM-1 expression after cold hypoxia and reperfusion. Livers were subjected to various regimes of cold preservation and reperfusion. Liver biopsies were taken at three time points (initial baseline on liver exposure; after organ flushing and post-storage at 0, 8, 16, and 24h cold hypoxia in University of Wisconsin solution; in the same livers after 1h warm reperfusion). The tissues were processed for frozen biopsy work, and frozen sections were stained using immunohistochemical methods, for blinded scoring by an independent observer. Positive controls were obtained by exposure to endotoxin lipopolysaccharide before liver flushing. ICAM-1 expression was low in control livers (0.33+/-0.21), and increased to near maximal (2.83+/-0.17) after endotoxin exposure. ICAM-1 expression increased progressively with cold preservation, reaching values of 1.17+/-0.31 and 1.83+/-0.31 after 16 and 24h, respectively (P<0.05 and 0.02 versus controls). Warm reperfusuion increased ICAM-1 expression in all flushed groups and with longer cold preservation was close to maximal (2.67+/-0.21 after 16h and 2.98+/-0.02 after 24h; P<0.001 in both cases). Addition of the free radical scavenger GSH prevented up-regulation of ICAM-1 in livers reperfused after flushing and cold storage for up to 8h; beyond this time, ICAM-1 expression still increased, such that by 24h cold preservation and reperfusion absence (2.98+/-0.02) or presence (2.67+/-0.21) made no difference. We conclude that liver ICAM-1 expression is demonstrably increased by progressive cold preservation and reperfusion, and is only marginally affected by addition of GSH during reperfusion. The model can be used to investigate other agents which might be more successful in preventing post-storage inflammatory damage.     

Allosteric activation of plasma membrane receptors--physiological implications and structural origins

Allosteric modulation of receptors has physiological not just pharmacological significance. Thus, the chemical context in which an agonist signal is received can have a major impact on the nature of the physiological response by modifying receptor sensitivity and/or maximal activity-even the nature of the signalling response. In addition, recognising that an endogenous activator is the allosteric modulator of a known receptor, rather than the agonist of a novel receptor, has the potential to solve, in dramatic fashion, key physiological questions. What is an allosteric modulator and why are allosteric effects on receptors so diverse and frequently complex? What is the scope of allosteric effects? Can the existence of endogenous modulators be predicted from a receptor's amino acid sequence? How should screening for endogenous allosteric modulators be undertaken? These questions form the framework of this mini-review on physiological and structural aspects of receptor allostery.     

Vitamin E and selenium supplementation to alleviate cold-stress-associated deterioration in egg quality and egg Yolk mineral concentrations of Japanese quails

The effects of vitamin E (dl-α-tocopheryl acetate) and selenium (Se; Na₂-SeO₃) on egg production, egg quality, and mineral content of egg yolk in Japanese quails reared under a low ambient temperature (6°C) were evaluated. Birds (n=300; 7 wk old) were randomly assigned to 12 treatment groups, 25 birds per group. The birds in a 3×2 factorial design received either three levels of vitamin E (125, 250, and 500 mg/kg diet) or two levels of selenium (0.1 or 0.2 mg/kg diet). After 2 wk on feed, six groups of the birds were maintained at 18°C (thermoneutral temperature [TN]), and the other half were acclimated over 3 d to a decreased environmental temperature of 6°C (cold stress [CS]). The performance, egg quality, and mineral content of egg yolk were not influenced by supplemental vitamin E and selenium in quails not exposed to cold stress (p≥0.09). Two hundred fifty and 500 mg vitamin E/kg diet compared with 125 mg/kg diet and higher dietary selenium inclusions (0.2 vs 0.1 mg/kg) resulted in a better body weight, egg production, and feed efficiency (p=0.01) in quails reared under CS. Similarly, egg weight, egg specific gravity, eggshell thickness, and Haugh unit were positively influenced with vitamin E (p=0.01) and selenium (p≤0.05) supplementation. Egg yolk concentrations of Zn, Fe, and Mn were higher with higher dietary vitamin E (p=0.01) and selenium (p=0.05). There was no interaction detected for parameters measured in the present study (p≥0.3). The results of the present study showed that a combination of 250 or 500 mg vitamin E and 0.2 mg selenium per kilogram of diet provides the greatest effects on performance and egg quality of Japanese quails reared under cold stress and suggest that such a supplementation can be considered as a protective management practice in Japanese quail diets to reduce the detrimental effects of cold stress.     

Preserved vascular reactivity of rat renal arteries after cold storage

In cultured renal tubular cells hypothermia results in cell damage caused by iron-dependent formation of reactive oxygen species. It is unknown whether cold preservation affects function of renal vessels. Rat renal arcuate arteries were stored in a physiological salt solution at 4 degrees C for 24h and compared to control arteries (not stored). To some of the stored arteries the iron chelator 2,2'-dipyridyl was added. Endothelium-independent vasoconstriction was assessed by cumulative concentration-response curves for potassium and phenylephrine in a small vessel myograph. Endothelium-independent vasodilation was assessed with sodium nitroprusside and endothelium-dependent vasodilation with histamine. Cold storage for 24h did not affect vascular reactivity of renal small arteries and no influence of the iron chelator was seen. Since 24h of cold storage considerable damages renal tubular cells both in vitro and after kidney transplantation, these results suggest that renal arteries are less sensitive to cold-induced damage than tubular cells.     

The acquisition of neural fate in the chick

Neural development in the chick embryo is now understood in great detail on a cellular and a molecular level. It begins already before gastrulation, when a separation of neural and epidermal cell fates occurs under the control of FGF and BMP/Wnt signalling, respectively. This early specification becomes further refined around the tip of the primitive streak, until finally the anterior-posterior level of the neuroectoderm becomes established through progressive caudalization. In this review we focus on processes in the chick embryo and put classical and more recent molecular data into a coherent scenario.     

Exogenous Carbohydrate Utilisation by Explants of Brassica napus Cultured in vitro

In 5; non-coding regions of genes for phytochrome A from horseradish (ArPHYAs) were fused with the 35S promoter of cauliflower mosaic virus in the antisense direction (CaMV35SantiArPHYAs) and introduced into horseradish hairy roots. Phytochrome levels of proximal areas in many hairy roots that had been transformed with CaMV35SantiArPHYAs decreased to levels of about one-half to one-quarter those of control hairy roots. The extent of the light-induced formation of adventitious shoots from hairy roots with less than half of the control level of phytochrome was lower than in the controls and not different between the three ArPHYAs. In contrast, the efficiency of phytochrome on the extent of shoot formation differed in hairy roots transformed with CaMV35SantiArPHYAs when phytochrome levels were more than 0.02 (;(;A) g^–1). The efficiency of ArPHYA3 to initiate shoot formation was greatest and that of ArPHYA2 was smallest. Furthermore, previous reports on hairy roots overexpressing ArPHYAs showed a similar efficiency of phytochrome on shoot formation. These results indicate that the ArphyA1 and/or ArphyA3 play major roles in the light-induced formation of adventitious shoots and that ArphyA2 has a minor role.     

Temperature acclimation in overwintering nymphs of a cockroach,Periplaneta japonica: walking on ice

Nymphs of the Yamato cockroach, Periplaneta japonica, showed a seasonal change in the ability to move at low temperature. Laboratory-reared nymphs buried in ice for 15 min-3 h took about 600 s to get up at 25 °C after being placed with their dorsal surface on the floor of a Petri dish. The mean time to get up at 25 °C (recovering time) after 15 or 60-min burial in ice was relatively long in early autumn, decreased rapidly in autumn, reached a minimal level in winter (<100 s) and increased in spring. Temperature was responsible for this seasonal change in physiology, and neither photoperiod nor diapause status was important. Recovering time depended on the acclimation temperature and its duration. For a 10-day exposure in a range of 0-25 °C, the lower the temperature the shorter the recovering. Longer periods of acclimation at these temperatures tended to shorten recovering time. The acquisition of the ability to move on ice is a seasonal phenomenon unique to P. japonica. No comparable response was found in three other cockroaches including two sub-tropical species (Periplaneta americana and Opisthoplatia orientalis) and a temperate species (Blattela nipponica).