Replacing risk-based early-onset-disease prevention with intrapartum group B streptococcus PCR testing.

OBJECTIVE: To evaluate the effect of a rapid PCR-based group B streptococcus (GBS) test on length of stay in hospital among newborns, antibiotic use, and GBS-early-onset-disease (EOD) incidence. METHODS: We conducted a before and after service evaluation including term deliveries between 1st January and 12th November 2014 (6688 deliveries). Length of stay in the hospital, GBS-EOD incidence and antibiotic use were evaluated. RESULTS: We recorded three confirmed and 74 possible cases of GBS-EOD in Phase 1, and 85 possible cases in Phase 2. In newborns with suspected infection, the introduction of the rapid test was related to a decreased length of stay on the pediatric care unit by 1.16 days (p = 0.01), and an increase in the length of stay on the mother-and-baby ward by 1.11 days (p < 0.001). No increase in antibiotics was noted. CONCLUSION: The introduction of a point of care test was associated with a reduction in length of stay in the pediatric care unit, without an increase in antibiotic use. This test could improve the accuracy of GBS colonization detection, and help to prevent intrapartum transmission as no verified GBS-EOD cases were recorded with the intrapartum PCR algorithm.

Viral RNA replication in association with cellular membranes.

All plus-strand RNA viruses replicate in association with cytoplasmic membranes of infected cells. The RNA replication complex of many virus families is associated with the endoplasmic reticulum membranes, for example, picorna-, flavi-, arteri-, and bromoviruses. However, endosomes and lysosomes (togaviruses), peroxisomes and chloroplasts (tombusviruses), and mitochondria (nodaviruses) are also used as sites for RNA replication. Studies of individual nonstructural proteins, the virus-specific components of the RNA replicase, have revealed that the replication complexes are associated with the membranes and targeted to the respective organelle by the ns proteins rather than RNA. Many ns proteins have hydrophobic sequences and may transverse the membrane like polytopic integral membrane proteins, whereas others interact with membranes monotopically. Hepatitis C virus ns proteins offer examples of polytopic transmembrane proteins (NS2, NS4B), a "tip-anchored" protein attached to the membrane by an amphipathic alpha-helix (NS5A) and a "tail-anchored" posttranslationally inserted protein (NS5B). Semliki Forest virus nsP1 is attached to the plasma membrane by a specific binding peptide in the middle of the protein, which forms an amphipathic alpha-helix. Interaction of nsP1 with membrane lipids is essential for its capping enzyme activities. The other soluble replicase proteins are directed to the endo-lysosomal membranes only as part of the initial polyprotein. Poliovirus ns proteins utilize endoplasmic reticulum membranes from which vesicles are released in COPII coats. However, these vesicles are not directed to the normal secretory pathway, but accumulate in the cytoplasm. In many cases the replicase proteins induce membrane invaginations or vesicles, which function as protective environments for RNA replication.

Thiol metabolism in preterm infants during the first week of life.

BACKGROUND: Oxidative stress is implicated in the pathogenesis of several complications of prematurity. The glutathione cycle is one of the most important intracellular antioxidant systems. The synthesis of glutathione may not be adequate in preterm neonates because of the low levels of cysteine available. The aim of this study was to evaluate cysteine and glutathione metabolism during the first week of life in preterm infants. METHODS: Plasma and erythrocyte thiol concentrations were measured in 78 preterm infants with a birthweight of 500-1500 g, and erythrocyte glutamate-cysteine ligase (GCL), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferases (GST) and glucose 6-phosphatedehydrogenase (G6PDH) in 26 infants with a birthweight of 1000-1500 g. RESULTS: The mean (SD) plasma glutathione concentration increased from day 0 to day 1 (14.9 (7.1) vs. 27.7 (11.9) micromol/L, p < 0.001), and then decreased. The plasma cysteine concentration changed in the opposite direction (172 (59) vs. 129 (42) micromol/L, p < 0.01). In infants with respiratory distress syndrome (RDS) the mean plasma glutathione concentration, but not cysteine, was lower on day 0 compared with infants without RDS (11.7 (5.2) vs. 21.4 (5.6) micromol/L, p < 0.01). Erythrocyte glutathione concentration decreased during the first week of life, whereas erythrocyte cysteine concentration increased significantly from day 3 to day 7 (p < 0.01). Erythrocyte cysteine and glutathione concentrations had a positive correlation. The GCL and GR activities did not change, but GST and G6PDH activities decreased during the first week (p < 0.01). GPx activity decreased until day 3 (p < 0.01) and was higher on day 0 and day 1 in infants with RDS. CONCLUSIONS: Very low birthweight infants have an initial increase in plasma glutathione and initial decrease in plasma cysteine level during the first week of life, and also a positive correlation between erythrocyte cysteine and glutathione levels.

Virus-specific mRNA capping enzyme encoded by hepatitis E virus.

Hepatitis E virus (HEV), a positive-strand RNA virus, is an important causative agent of waterborne hepatitis. Expression of cDNA (encoding amino acids 1 to 979 of HEV nonstructural open reading frame 1) in insect cells resulted in synthesis of a 110-kDa protein (P110), a fraction of which was proteolytically processed to an 80-kDa protein. P110 was tightly bound to cytoplasmic membranes, from which it could be released by detergents. Immunopurified P110 catalyzed transfer of a methyl group from S-adenosylmethionine (AdoMet) to GTP and GDP to yield m(7)GTP or m(7)GDP. GMP, GpppG, and GpppA were poor substrates for the P110 methyltransferase. There was no evidence for further methylation of m(7)GTP when it was used as a substrate for the methyltransferase. P110 was also a guanylyltransferase, which formed a covalent complex, P110-m(7)GMP, in the presence of AdoMet and GTP, because radioactivity from both [alpha-(32)P]GTP and [(3)H-methyl]AdoMet was found in the covalent guanylate complex. Since both methyltransferase and guanylyltransferase reactions are strictly virus specific, they should offer optimal targets for development of antiviral drugs. Cap analogs such as m(7)GTP, m(7)GDP, et(2)m(7)GMP, and m(2)et(7)GMP inhibited the methyltransferase reaction. HEV P110 capping enzyme has similar properties to the methyltransferase and guanylyltransferase of alphavirus nsP1, tobacco mosaic virus P126, brome mosaic virus replicase protein 1a, and bamboo mosaic virus (a potexvirus) nonstructural protein, indicating there is a common evolutionary origin of these distantly related plant and animal virus families.

Elimination of phosphorylation sites of Semliki Forest virus replicase protein nsP3.

nsP3 is one of the four RNA replicase subunits encoded by alphaviruses. The specific essential functions of nsP3 remain unknown, but it is known to be phosphorylated on serine and threonine residues. Here we have completed mapping of the individual phosphorylation sites on Semliki Forest virus nsP3 (482 amino acids) by point mutational analysis of threonine residues. This showed that threonines 344 and 345 represented the major threonine phosphorylation sites in nsP3. Experiments with deletion variants suggested that nsP3 itself had no kinase activity; instead, it was likely to be phosphorylated by multiple cellular kinases. Phosphorylation was not necessary for the peripheral membrane association of nsP3, which was mediated by the N-terminal region preceding the phosphorylation sites. Two deletion variants of nsP3 with either reduced or undetectable phosphorylation were studied in the context of virus infection. Cells infected with mutant viruses produced close to wild type levels of infectious virions; however, the rate of viral RNA synthesis was significantly reduced in the mutants. A virus totally defective in nsP3 phosphorylation and exhibiting a decreased rate of RNA synthesis also exhibited greatly reduced pathogenicity in mice.

Helicase and capping enzyme active site mutations in brome mosaic virus protein 1a cause defects in template recruitment, negative-strand RNA synthesis, and viral RNA capping.

Brome mosaic virus (BMV) encodes two RNA replication proteins: 1a, which contains RNA capping and helicase-like domains, and 2a, which is related to polymerases. BMV 1a and 2a can direct virus-specific RNA replication in the yeast Saccharomyces cerevisiae, which reproduces the known features of BMV replication in plant cells. We constructed single amino acid point mutations at the predicted capping and helicase active sites of 1a and analyzed their effects on BMV RNA3 replication in yeast. The helicase mutants showed no function in any assays used: they were strongly defective in template recruitment for RNA replication, as measured by 1a-induced stabilization of RNA3, and they synthesized no detectable negative-strand or subgenomic RNA. Capping domain mutants divided into two groups. The first exhibited increased template recruitment but nevertheless allowed only low levels of negative-strand and subgenomic mRNA synthesis. The second was strongly defective in template recruitment, made very low levels of negative strands, and made no detectable subgenomes. To distinguish between RNA synthesis and capping defects, we deleted chromosomal gene XRN1, encoding the major exonuclease that degrades uncapped mRNAs. XRN1 deletion suppressed the second but not the first group of capping mutants, allowing synthesis and accumulation of large amounts of uncapped subgenomic mRNAs, thus providing direct evidence for the importance of the viral RNA capping function. The helicase and capping enzyme mutants showed no complementation. Instead, at high levels of expression, a helicase mutant dominantly interfered with the function of the wild-type protein. These results are discussed in relation to the interconnected functions required for different steps of positive-strand RNA virus replication.

Effects of palmitoylation of replicase protein nsP1 on alphavirus infection.

The membrane-associated alphavirus RNA replication complex contains four virus-encoded subunits, the nonstructural proteins nsP1 to nsP4. Semliki Forest virus (SFV) nsP1 is hydrophobically modified by palmitoylation of cysteines 418 to 420. Here we show that Sindbis virus nsP1 is also palmitoylated on the same site (cysteine 420). When mutations preventing nsP1 palmitoylation were introduced into the genomes of these two alphaviruses, the mutant viruses remained viable and replicated to high titers, although their growth was slightly delayed. The subcellular distribution of palmitoylation-defective nsP1 was altered in the mutant: it no longer localized to filopodial extensions, and a fraction of it was soluble. The ultrastructure of the alphavirus replication sites appeared normal, and the localization of the other nonstructural proteins was unaltered in the mutants. In both wild-type- and mutant-virus-infected cells, SFV nsP3 and nsP4 could be extracted from membranes only by alkaline solutions whereas the nsP2-membrane association was looser. Thus, the membrane binding properties of the alphavirus RNA replication complex were not determined by the palmitoylation of nsP1. The nsP1 palmitoylation-defective alphaviruses produced normal plaques in several cell types, but failed to give rise to plaques in HeLa cells, although they induced normal apoptosis of these cells. The SFV mutant was apathogenic in mice: it caused blood viremia, but no infectious virus was detected in the brain.

Putative RNA capping activities encoded by brome mosaic virus: methylation and covalent binding of guanylate by replicase protein 1a.

Brome mosaic virus (BMV) RNA replication is directed by two virus-encoded proteins, 1a and 2a. The amino-terminal half of 1a is a distant homolog of alphavirus nonstructural protein nsP1, which has been implicated in capping viral RNAs. In this study, we examined the enzymatic activities of BMV 1a expressed in yeast, where the protein is fully functional in RNA replication. 1a methylated GTP, dGTP, and the cap analogs GpppG and GpppA, using S-adenosylmethionine (AdoMet) as the methyl donor. Product analysis by nuclear magnetic resonance spectroscopy showed that 1a methylation was specific for guanine position 7. Additionally, 1a interacted with GTP to form a covalent 1a-m(7)GMP complex. This reaction was specific for GTP, required AdoMet, and was accompanied by transfer of (3)H-methyl from AdoMet to the covalent 1a-guanylate complex. The covalent complex could be immunoprecipitated by 1a antibodies. The 1a-m(7)GMP complex was inhibited in catalyzing further methyltransferase reactions. Mutation of conserved amino acids in the N-terminal half of 1a reduced both methyltransferase and covalent complex formation activities to very low or undetectable levels. Covalent 1a-guanylate complex formation took place in similar, AdoMet-dependent fashion in extracts of BMV-infected barley protoplasts. These results show that BMV 1a has activities similar to those of alphavirus nsP1, demonstrating conservation of these putative capping functions across a wide span of sequence divergence within the alphavirus-like superfamily. Conservation of this unusual combination of functions also supports the inference that the superfamily caps viral RNAs by an unusual pathway proceeding via a m(7)GMP intermediate.

Low trehalase activity is associated with abdominal symptoms caused by edible mushrooms.

BACKGROUND: The purpose of the study was to evaluate whether maldigestion of trehalose causes abdominal symptoms and which available diagnostic method best distinguishes intolerant from tolerant subjects. METHODS: A 25-g oral trehalose load test was performed in 64 subjects. The 19 experiencing clear symptoms constituted the trehalose-intolerant subjects. Changes from base-line levels of blood glucose, breath hydrogen, and methane and symptoms were recorded after the test. Trehalase activity was determined in serum and on a duodenal biopsy specimen obtained by endoscopy. RESULTS: Intolerant subjects were best differentiated from tolerant subjects by changes in breath gases (hydrogen and methane) and duodenal trehalase to sucrase ratio. The change in breath gases correlated inversely with duodenal trehalase activity, duodenal trehalase to sucrase ratio, and plasma trehalase activity. The correlation between serum and duodenal trehalase activities was on the order of 0.6. Two subjects were found to have trehalase deficiency. CONCLUSIONS: It is obvious that trehalose maldigestion can cause symptoms similar to those of lactose maldigestion and intolerance. Three factors control the genesis of symptoms: 1) the activity of small-bowel trehalase: if it is low, trehalose is maldigested and more trehalose is passed into the colon; 2) the maldigested trehalose, which causes osmotic water flow into the colon, resulting in loose stools and diarrhea; and 3) most importantly, the microflora of the colon, from which symptoms will arise if there are bacteria capable of producing gases from maldigested trehalose. If colonic bacteria cannot produce gases, then distention of the abdomen and intestinal gas expulsion as eructations and flatus will not occur.

Intracellular distribution of rubella virus nonstructural protein P150.

Antiserum prepared against an amino-terminal fragment of rubella virus (RUB) nonstructural polyprotein was used to study RUB-infected Vero cells. Replicase protein P150 was associated with vesicles and vacuoles of endolysosomal origin and later with large, convoluted, tubular membrane structures. Newly incorporated bromouridine was associated with the same structures and specifically with small membrane invaginations, spherules, indicating that these structures may be the sites of viral RNA synthesis.

Virus-specific capping of tobacco mosaic virus RNA: methylation of GTP prior to formation of covalent complex p126-m7GMP.

In capping cellular mRNAs, a covalent GMP-enzyme intermediate leads to formation of G(5')ppp(5')N at the 5' end of the RNA, which is modified by methylation catalyzed by guanine-7-methyltransferase. Here we show that isolated membranes from tobacco mosaic virus (TMV)-infected plant or insect cells expressing TMV replicase protein p126, synthesized m7GTP using S-adenosylmethionine (AdoMet) as the methyl donor, and catalyzed the formation of a covalent guanylate-p126 complex in the presence of AdoMet. The methyl group from AdoMet was incorporated into p126, suggesting that the complex consisted of m7GMP-p126. Thus, TMV and alphaviruses, despite their evolutionary distance, share the same virus-specific capping mechanism.

Extrapancreatic organ impairment in caerulein induced pancreatitis.

BACKGROUND AND AIMS: Multiorgan function failures are the major fatal complications in acute pancreatitis. In this experiment, we studied 1) the manifestation and time course of extrapancreatic organ damage in an acute pancreatitis model and 2) whether the obstructive liver damage in this model is caused by the obstruction of common biliopancreatic duct compressed by oedematous pancreas. MATERIAL AND METHODS: 80 male Wistar rats were divided into two groups: control and caerulein groups (five subgroups in each group). In the caerulein group, the acute pancreatitis was induced by caerulein intraperitoneal injections. In the controls equal volume of saline was injected. Two subgroups, one in caerulein and one in control groups, had an intrapancreatic bile duct stent inserted transduodenally before the injections. The pancreas, liver, lung and kidney tissues and blood samples were obtained for the measurement or analysis of interstitial oedema, plasma amylase, alanine aminotransferase, bilirubin, urea, creatinine, alkaline phosphatase, lactate dehydrogenase, blood gas and electron microscopy at 1, 6, 12 and 24 hours after the last injection in unstented animals, and at 6 hours in stented animals. RESULTS: Lungs and kidney remained unchanged. Liver damage was found during the first 6-12 hours, manifest as increased plasma alanine aminotransferase and bilirubin and dilatation of bile canaliculi and hepatocyte damage in electron microscopy. The intrapancreatic bile duct stent did not resolve these changes. CONCLUSIONS: The liver may be the first evolved extrapancreatic organ in the early stage in this mild oedematous pancreatitis model and the hepatocyte damage is not caused by the obstruction of common biliopancreatic duct compressed by the oedematous pancreas.

Semliki Forest virus mRNA capping enzyme requires association with anionic membrane phospholipids for activity.

The replication complexes of all positive strand RNA viruses of eukaryotes are associated with membranes. In the case of Semliki Forest virus (SFV), the main determinant of membrane attachment seems to be the virus-encoded non-structural protein NSP1, the capping enzyme of the viral mRNAs, which has guanine-7-methyltransferase and guanylyltransferase activities. We show here that both enzymatic activities of SFV NSP1 are inactivated by detergents and reactivated by anionic phospholipids, especially phosphatidylserine. The region of NSP1 responsible for binding to membranes as well as to liposomes was mapped to a short segment, which is conserved in the large alphavirus-like superfamily of viruses. A synthetic peptide of 20 amino acids from the putative binding site competed with in vitro synthesized NSP1 for binding to liposomes containing phosphatidylserine. These findings suggest a molecular mechanism by which RNA virus replicases attach to intracellular membranes and why they depend on the membranous environment.

Guanosine nucleotide analogs as inhibitors of alphavirus mRNA capping enzyme.

The two virus-specific reactions in the capping of alphavirus RNAs, catalyzed by the replicase protein nsP1, are promising targets for developing virus-specific inhibitors. In this report, we have studied the effect of over 50 cap analogs on the guanine-7-methyltransferase and guanylyltransferase activities of Semliki Forest virus nsP1. Recombinant nsP1 was expressed in Escherichia coli and partially purified by flotation in a discontinuous sucrose gradient. The methyltransferase activity had a pH optimum between pH 6.5 and 7.1, and the apparent Km values were 1.9 mM for GTP, 6.0 microM for S-adenosyl-L-methionine and 170 microM for Mg2+. NsP1 methyltransferase was able to methylate efficiently GTP (relative activity 100%), GDP (16%), GpppG (35%), GppppG (50%) and less efficiently GpppA (12%), m2GTP (9%), and m2,2GTP (25%), but not m7GppG. The most potent inhibitors for nsP1 methyltransferase were et2m7GMP (Ki value 42 microM), m2,7GMP, (64 microM), m2,7GpppG (82 microM), m2et7GMP (105 microM), m2(2-phet)7GMP (194 microM) and m2GMP (386 microM). Of these compounds, m2GMP, m2et7GMP and m2(2-phet)7GMP showed competitive inhibition, whereas the others showed mixed type inhibition. All compounds that inhibited the methyltransferase activity inhibited also the guanylyltransferase activity of nsP1.

Coenzyme Q10 supplementation and recovery from ischemia in senescent rat myocardium.

Many studies have suggested that parenteral administration of coenzyme Q10 (Q10) protects the myocardium of young experimental animals from post-ischemic reperfusion injury. Although parenteral administration, in contrast to per os supplementation, seems to elevate coenzyme Q concentrations in heart tissue, it is not suitable for prophylactic use. In addition, the incidence of ischemic events is greatest in older age. We studied the effect of Q10 supplementation on myocardial postischemic recovery in 18-month-old Wistar rats. The treated group (n=9) received 10 mg/kg/day of Q10 for 8 weeks in their chow while the normal chow of the control group (n=9) contained less than 0.5 mg/kg/day of Q10. The treatment clearly elevated plasma Q10 concentration (286 +/- 25 micromol/l and 48 +/- 30 micromol/l, treated and controls, respectively, p<0.0001) but neither Q9 nor Q10 concentrations in heart tissue were affected by the supplementation. The isolated perfused hearts were subjected to 20 minutes of ischemia and 30 minutes of reperfusion. The preischemic values of developed pressure (DP) but not contractility (+DP/delta t) and relaxation (-DP/delta t) were improved by Q10 supplementation (p=0.034, p=0.057 and p=0.13, respectively) while in postischemic recovery no differences were observed between the groups (p>0.05 at all time points). Also, in myocardial flow, myocardial oxygen consumption (MVO2) and myocardial aerobic efficiency (DP/MVO2) the groups did not differ at any time points. Although dietary Q10 supplementation clearly elevated plasma Q10 concentrations in senescent rats, the coenzyme Q contents in heart tissue and myocardial recovery from ischemia were not affected. However, it is possible that the site of action for the reported beneficial effects of Q10 is in the coronary endothelium rather than myocardium itself.

Pharmacokinetics of intravenous N-acetylcysteine in pre-term new-born infants.

BACKGROUND: Reactive oxygen species have been considered to play a role in several clinical complications in pre-term infants. The aim of this study was to determine the pharmacokinetics of intravenous N-acetylcysteine in pre-term neonates. This information is needed to evaluate the use of N-acetylcysteine as an antioxidant in this patient group. METHODS: N-acetylcysteine was infused intravenously in ten patients (gestational age 24.9-31.0 weeks, weight 500-1384 g) for 24 h (3.4-4.6 mg/kg/h), starting 2.0-11.2 h from birth (study I) and in six patients (gestational age 25.9-29.7 weeks, weight 520-1335 g) for 6 days (0.3-1.3 mg/kg/h), starting at the age of 24 h (study II). Arterial plasma N-acetylcysteine and cyst(e)ine concentrations were determined from timed samples taken during (study I and II) and after (study I) the N-acetylcysteine infusion. RESULTS: In study I, the mean elimination half-life of N-acetylcysteine was 11 h (range 7.8-15.2 h). The mean plasma clearance of N-acetylcysteine was 37 ml/kg/h (range 13-62 ml/kg/h) and the mean volume of distribution was 573 ml/kg (range 167-1010 ml/kg). The plasma clearance and volume of distribution correlated with weight (r = 0.81, P < 0.01, and r = 0.78, P < 0.01, respectively) and with gestational age (r = 0.71, P < 0.05, and r = 0.64, P < 0.05, respectively). In study II, the steady-state concentration of N-acetylcysteine was reached in 2-3 days in five of six patients during a constant infusion. CONCLUSIONS: The pharmacokinetics of N-acetylcysteine in pre-term infants depend markedly on weight and gestational age. The elimination of N-acetylcysteine is much slower in pre-term new-borns than in adults.

Extrapancreatic multiorgan injury in a severe sublethal acute pancreatitis model.

In order to develop a new severe but sublethal acute pancreatitis model for the study of clinically relevant extrapancreatic multiorgan injury, we have induced acute pancreatitis in a rat model by intraductal injection with low dose and moderate concentration of bile acid under low pressure. We examined the structural and functional features in the pancreas, lung, liver and kidney. The animals were divided into two groups: the bile acid injection group and the control group. In the bile acid injection group, acute necrotizing pancreatitis was induced by intraductal administration of 0.2 ml of 2.0% bile acid under 30 cm H2O pressure, while the controls underwent the sham operation. The two groups were divided into six subgroups (8 rats for each) and sacrificed at 12, 24, 36, 48, 72 and 144 h, respectively. The pancreatitis induced hyperamylasemia, ascites, pancreatic oedema, haemorrhage, acinar cell necrosis and extensive fat necrosis without early mortality. Accompanied with the pancreatic injury, the function and histologic changes have developed continuously in the kidney and liver for 72 and 144 h in the bile acid injection animals respectively. No pancreatitis associated pulmonary changes were found. Taking into account the results with the two previously developed models of pancreatitis, we conclude that the extrapancreatic injury in acute pancreatitis is found in the liver, kidney and lung, in that order, depending on the severity of pancreatitis. The present sublethal pancreatitis model, in comparison with the two previously studied acute pancreatitis models, is perfect for pathogenetic and therapeutic study of liver and renal changes in acute necrotizing pancreatitis.

Extrapancreatic organ manifestations in a moderately severe acute pancreatitis model.

In order to study the extent of clinically relevant extrapancreatic organ injury in a moderately severe pancreatitis model, we examined the structural and functional features of the pancreas, lung, liver and kidney in a rat model simulating gallstone pancreatitis. The animals were divided into three groups: the low-ligation group, the high-ligation group and the control group, and sacrificed at 6, 24, 42, 60 and 96 h. In the low-ligation group, moderately severe acute pancreatitis was induced by the ligation of the common biliopancreatic duct plus intralipid intragastric injection, while controls underwent the ligation of the bile duct above the pancreas (the high-ligation group) or only sham operation (the control group) with fat injection. The pancreatitis induced hyperamylasemia, pancreatic oedema, haemorrhage, acinar cell necrosis and extensive fat necrosis. Accompanied with a peak value of serum amylase activity 24 h after the induction, the kidney changes developed, characterized by decrease in urine output, increase in serum urea and creatinine, and proximal convoluted tubular damage under electron microscope. There were no pancreatitis associated lung or liver changes. These results suggest that this model can be used to study the pathogenesis and therapy of renal injury during acute moderately severe pancreatitis.

An improved biotonometric method for determination of haemoglobin oxygen equilibrium curves.

A new biotonometric method for the determination of haemoglobin oxygen equilibrium curves is described. The procedure is based on the mixing of an oxygen-consuming organism, the yeast cell (Saccharomyces cerevisiae), with an oxidized blood/plasma mixture. The yeast cell consumes oxygen at a uniform rate, thus reducing the partial pressure of oxygen in the mixture. This in turn induces the dissociation of oxygen in a characteristic manner. The study of the whole blood samples from 26 healthy volunteer subjects gave the following results: p50 = 3.63 kPa +/- 0.23 kPa (mean +/- 1 S.D.); Hill slope n = 2.44 +/- 0.16; and CO2 Bohr factor = -0.47 +/- 0.11. For the within-run imprecision the coefficients of variation for the different parameters were: p50 C.V. = 4.4%; Hill slope n C.V. = 4.7%; and CO2 Bohr factor C.V. = 19%. The determination can be carried out with simple equipment: a blood gas analyzer with a coupled recorder.