Mind the model for end-stage liver disease: model for end-stage liver disease score as an indicator of hemoderivate transfusion and survival in liver transplantation.

The scarcity of liver donors has increased the necessity to closely select recipients to improve liver transplantation outcomes. If we were able to recognize those recipients with the best outcomes, then this could result in a better and more accurate selection of our donors. Hemoderivate transfusion is one of the main important factors to analyse. We reviewed the data of all of our liver transplant recipients from May 1998 to December 2013 and selected 888 patients with complete records. We divided these patients into 5 groups to get a better selection. We found differences between these groups with respect to the following: recipient age at the time of transplantation, percentage of patients with hepatocarcinoma, and those with hepatitis C virus (HCV)-related etiology. Also, intensive care unit (ICU) time and the need for retransplantation were distinctive factors with observable differences between our groups. With respect to model for end-stage liver disease (MELD) score, the groups were clearly defined by their mean MELD score, finding significant statistical differences between these groups with respect to this score. We also found a significant relationship between MELD scores and survival in the different groups. This is also the first review in which the MELD score and intraoperative transfusion requirements are well associated with patient and graft survival.

Ceratitis capitata brain adenylate cyclase and its membrane environment.

Adenylate cyclase activation by GTP and octopamine as well as basal activity (in the presence of Mg2+) have been studied as a function of membrane structure in plasma membranes from brain of the dipterous Ceratitis capitata. Benzyl alcohol and lidocaine, but not phenobarbital, inhibited the three activities to the same extent. Triton X-100-solubilized adenylate cyclase was also inhibited by benzyl alcohol and lidocaine, but not by phenobarbital. Results could be explained by an effect on the catalytic unit lipid environment, which would be maintained after solubilization, counteracting the effect of these drugs to facilitate lateral diffusion and coupling of adenylate cyclase components in the lipid bilayer. The observation that the insect adenylate cyclase is relatively insensitive to changes in bulk bilayer fluidity is strengthened by the absence of effect of phenobarbital on enzyme activities. Indeed, this compound was as active as lidocaine or benzyl alcohol in increasing bulk membrane fluidity. The response of C. capitata adenylate cyclase to changes in membrane fluidity is different from that recorded in mammalian systems. This may be functionally important and result from the fact that insects are not warm-blooded.

Effect of ethanol on arachidonic acid metabolism in macrophages.

The authors report preliminary data on the production of arachidonic acid metabolites by ethanol stimulated macrophages. They focus on the substantial implications for understanding the pathobiology of alcoholism.

Intracellular calcium and pH alterations induced by Escherichia coli endotoxin in rat hepatocytes.

In this study, the fluorescent Ca2+ probe fura-2 and the fluorescent pH indicator BCECF have been used to monitor cytosolic free Ca2+ and intracellular pH (pHi), respectively, in isolated and cultured hepatocytes treated with Escherichia coli O111:B4 endotoxin. Uptake of 45Ca2+ was also measured to study the effect of endotoxin on the extracellular calcium influx. Endotoxin treatment produced a progressive increase of cytosolic Ca2+ in a dose-dependent manner caused by both induction of a significant release of Ca2+ from intracellular stores and stimulation of the extracellular calcium influx. The perturbation of Ca2+ homeostasis by endotoxin may cause an abnormal stimulation of physiological processes, developing lethal cell injury. Endotoxin also produced a significant decrease in the pHi of hepatocytes which can justify important metabolic alterations during endotoxicosis.

Acetylated low-density lipoproteins promote the release and metabolism of arachidonic acid by murine macrophages.

We studied the effect of acetyl-LDL on the synthesis of eicosanoids by primary cultures of mouse-resident peritoneal macrophages. A dosis of 150 micrograms of acetyl-LDL proteins promotes a maximal release of arachidonic acid metabolites into extracellular medium. This process is very rapid and reaches the maximal value about ten minutes after stimulation. The pattern of arachidonic acid metabolites released is different from that obtained when the cells are stimulated with a phagocytosable particle like zymosan or calcium ionophore A23187. The data show that the metabolism of arachidonic acid by cycloxigenase is diminished. In an unusual way the amounts of prostaglandin PGE2, 6-keto-prostaglandin PGF1 alpha and thromboxane TXB2 synthesized are similar.

Kinetic evidence of a rapid activation of phosphatidylcholine hydrolysis by Ki-ras oncogene. Possible involvement in late steps of the mitogenic cascade.

A novel phospholipase C specific for phosphatidylcholine has been shown to be activated by several agonists. Also, recent evidence suggests that transformation mediated by the ras oncogene possibly involves the activation of this novel phospholipid degradative pathway which would account for the increased diacylglycerol levels associated with transformation. Here we use a mutant of Ki-ras which is temperature-sensitive for transformation to investigate the kinetics of activation of the phosphodiesterase-mediated turnover of phosphatidylcholine. Upon shift to the permissive temperature, products of the activated phosphatidylcholine-specific phospholipase C were detected by 30 min and reached maximal levels by 1-2 h. These results suggest that the product of the ras oncogene rapidly activates the phosphodiesteratic hydrolysis of phosphatidylcholine. Furthermore, the fact that at least 4 h are required for serum to activate this phospholipase C strongly suggests that the ras oncogene product might be involved in late steps of the mitogenic signaling cascade.

Association of changes in lysophosphatidylcholine metabolism and in microsomal membrane lipid composition to the pulmonary injury induced by oleic acid.

Alterations in the lipid composition of lung microsomal membranes occur in oleic acid-induced respiratory distress. The marked decrease in the phosphatidylcholine/lysophosphatidylcholine molar ratio could be related with an altered metabolism of lysophosphatidylcholine in these membranes. Results revealed that the activity of phospholipase A increased whereas that of acyl-CoA:lysophosphatidylcholine acyltransferase decreased. Microsomal lysophospholipase activity remained unchanged. On the other hand, the microsomal enzyme system involved in the de novo synthesis of diacylglycerol was impaired, and cholinephosphotransferase activity was lowered. These changes in the activity of some membrane-bound enzymes were not caused by changes in the membrane lipid fluidity since lipid structural order parameter (SDPH) did not change and neither did the major factors on which the fluidity depends. The possible significance of microsomal lipid alterations in the pathogenesis of respiratory distress induced by oleic acid is discussed.

Effect of Escherichia coli endotoxin on ascorbic acid transport in isolated adrenocortical cells.

The active uptake of ascorbic acid by isolated rat adrenocortical cells increases with ascorbic acid concentration, depends on time and calcium, and is inhibited by ACTH concentrations required for maximal steroidogenesis. Lipopolysaccharide of Escherichia coli 0111:B4 modifies the ascorbic acid uptake in a calcium-dependent manner. At low calcium concentrations, lipopolysaccharide exerts a stimulatory effect on ascorbic acid transport and at high concentrations lipopolysaccharide produces a dose-dependent inhibitory effect. This inhibition of the ascorbic acid transport by the endotoxin can alter the ascorbic acid accumulation in the adrenal gland during endotoxin shock.

Adrenal gland function in reversible endotoxic shock.

Because adrenal participation in the defense mechanisms against endotoxic shock is essential for survival, adrenal gland function during reversible endotoxicosis was studied. The injection of E. coli LPS into rats produces an increase in plasma corticosteroids (maximum at 2-4 hr post-endotoxin injection) and ACTH levels (maximum at 2 hr post-endotoxin injection), which return to control values at the recovery phase. Nevertheless, ACTH-induced steroidogenesis in cells isolated from adrenal glands of endotoxemic rats is clearly impaired, even at 2 hr post-endotoxin injection when corticosteroid levels are maximal. During reversible endotoxic shock there is also a depletion of adrenal ascorbic acid (maximum at 2-4 hr post-endotoxin injection) and a decrease in adrenal cytochrome P-450 levels. These data suggest that impairment of the adrenal gland function could involve mechanisms at the receptor level (desensitization by the high plasma ACTH levels or a direct effect of LPS) and/or at post-receptor steps (decrease in adrenal cytochrome P-450 levels related to the diminution in adrenal ascorbic acid content).

Auxiliary liver by transplanted frozen-thawed hepatocytes.

Among the therapeutic alternatives to orthotopic liver transplantation, hepatocyte transplantation (HT) offers the best potential in a number of liver diseases, mainly inborn errors of metabolism. Nevertheless, HT presents several inconveniences such as the scarce knowledge of the functionality of the transplanted hepatocytes, which has given rise to controversy about the specificity or unspecificity of the transplant, and the lack of a suitable system for preserving the cells. This study was designed to test a system for cryopreserving hepatocytes and to assess their functionality over prolonged periods after their ectopic transplantation. A medium and a freezing schedule which are reproducible and yield elevated viability have been used, and a number of hepatospecific parameters have been assessed: the activity of ornithine carbamoyltransferase--an enzyme of primary importance in the urea cycle--lipogenesis, gluconeogenesis, glucose-6-phosphatase and cytochrome oxidase activities, the presence of albumin--as an index of plasma protein synthesis--and IDA uptake and metabolism, showing the UDP-glucuronyl transferase activity. As dedifferentiation markers, gamma-glutamyl transpeptidase and alpha-fetoprotein have been studied. From the results, it can be deduced that hepatocytes can be cryopreserved and transplanted and that under these conditions they maintain hepatic features for a long time. Following transplantation, several specific liver functions appear or are enhanced in the spleen. Freshly isolated and cryopreserved transplanted hepatocytes have similar behaviors, although a difference in the expression of the function can be observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Morphological studies of cytotoxic lesions in reversible endotoxic shock.

Reversible endotoxic shock was induced in adult rats by intravenous injection of E. coli 0111:B4 lipopolysaccharide (LPS) and the progression of metabolic and morphological alterations was evaluated. Serum samples and biopsies from adrenal gland, liver and lung were studied at different times after LPS injection. Histological changes in these tissues were observed after endotoxin administration, coinciding with both the acute-phase and the recovery-phase of shock (24-72h after LPS injection). Signs of tissue regeneration can be correlated with the regression of some serum parameters to their normal values. All these results indicate that in this experimental model of endotoxic shock, a reversible status was established, which will allow further studies of the endotoxic pathophysiological mechanisms in vivo, avoiding the complexity of the non-reversible process.

Microsomal membrane fluidity and phosphatidylcholine synthesis in rabbit lung under high oxygen tension.

Phosphatidylcholine metabolism and membrane fluidity were studied in microsomes isolated from rabbit lung, which had been exposed to high oxygen tension for 30 min. In these microsomes the incorporation of [3H]-palmitate into phosphatidylcholine increased whereas the incorporation of [14C]-glycerol and [14C]-choline from CDP-[methyl-14C]-choline remained unchanged in comparison to the control microsomes. The enhanced [3H]-palmitate incorporation may be explained by an increase of the specific activity of acyl-CoA:lysophosphatidylcholine acyltransferase which was measured in microsomes from hyperoxic lung. Although microsomal parameters influencing membrane fluidity, such as the cholesterol/phospholipid molar ratio, unsaturation degree of phospholipid acyl chains and lipid/protein ratio, are altered after oxygen treatment in vivo, no change of fluorescence polarization (PDPH) and lipid structural order parameter (SDPH) could be measured. Probably, the membrane maintains its fluidity by counteracting effects on different factors on which the fluidity depends.

Comparison between intra- and extracellular surfactant in respiratory distress induced by oleic acid.

The present study compares the phospholipid distribution and protein content in bronchoalveolar lavage, purified extracellular surfactant and lamellar bodies isolated from rabbits killed at intervals of 2.5, 12 and 24 h after oleic acid administration. The data suggest that the alteration of pulmonary surfactant could be partially due to the type II cell response to the injury.

Involvement of cytochrome b5 in the cytotoxic response to Escherichia coli lipopolysaccharide.

Cytotoxic lesions, induced by Gram-negative lipopolysaccharides (LPS), occur mainly in liver where the microsomal compartment of hepatocytes is involved in the detoxification mechanisms as well as in the biosynthesis of different active metabolites. The alterations induced by LPS from E. coli 0111:B4 on cytochrome b5 and its correlation with cytochrome P450, have been studied using an in vivo reversible endotoxic shock model and 24 h non-replicative hepatocyte monolayers. Results show that cytochrome b5 is directly affected by LPS that induces also a membrane damage with an active release of lactate dehydrogenase (LDH). The increase of cytochrome b5 levels may enhance the efficiency of the electron transport, thus facilitating the cytochrome P450-associate oxidations and reactions involved in the repair mechanisms of membranes.

Lipopolysaccharide-induced insulin resistance in monolayers of cultured hepatocytes.

In order to clarify the endotoxin effect on the hepatic removal of insulin, the influence of lipopolysaccharide (LPS) from E. coli 0111:B4 on the insulin binding and endocytosis in cultured hepatocytes from adult male rats has been investigated. LPS decreases both processes in a time and temperature-dependent manner, showing a major effect at short time and low temperature, according to the characteristics of LPS binding and uptake.

A possible new class of octopamine receptors coupled to adenylate cyclase in the brain of the dipterous Ceratitis capitata. Pharmacological characterization and regulation of 3H-octopamine binding.

Octopamine exerts its effects in insects through interaction with at least two classes of receptors, designated octopamine-1 and octopamine-2. Octopamine-2 receptors are positively coupled to adenylate cyclase, while octopamine-1 receptors are not coupled to this enzyme system. Ceratitis capitata brain appears to have octopamine receptors as unique aminergic receptors coupled to adenylate cyclase. These receptors show some pharmacological analogies with respect to octopamine-2 receptors, however they should constitute a new class of octopamine receptors. C. capitata brain octopamine receptors have also been characterized by [3H]octopamine-binding studies, exhibiting similar regulatory mechanisms to other receptors coupled to adenylate cyclase activation.

Induction of reversible shock by Escherichia coli lipopolysaccharide in rats. Changes in serum and cell membrane parameters.

Reversible endotoxic shock was induced in adult rats by i.v. injection of Escherichia coli O111:B4 lipopolysaccharide (1.6 mg/100 g). The shock progression was evaluated by measuring serum glucose levels as well as activities of aspartate aminotransferase (GOT) and alkaline phosphatase in serum. A rapid increase of serum glucose levels occurs, after LPS injection, followed by hypoglycaemia (minimum values at 6 h) with progressive reversion to control values. Serum GOT activity increased (twofold) 6 h after endotoxin administration and returned to control values at 72 h. No appreciable changes occurred in serum alkaline phosphatase activity. Endotoxaemia produced a decrease in the cytochrome P-450 levels in all target organs considered: lung, adrenal glands and liver. The progressive decrease in the serum albumin concentration as well as changes of the physical properties of the plasma membranes observed in vivo, can not be explained only by direct interaction of endotoxin with the target organs, underlining the importance of serum mediators in the induction of the shock response.

Ca2+-induced changes in the secondary structure of a 60 kDa phosphoinositide-specific phospholipase C from bovine brain cytosol.

The purification to homogeneity of a 60 kDa phosphoinositide-specific phospholipase C from bovine brain cytosol is reported here. This enzyme exhibits the same properties, in terms of response to Ca2+, as does the cytosolic activity in a variety of cell types. We show here that Ca2+ does not appear to modulate the binding of the enzyme to the substrate, but induces dramatic changes in its secondary structure. Therefore we suggest that a decrease in the alpha-helix content of this enzyme correlates with its ability to be activated by Ca2+.