bluuebunny/crossref_metadata_2025 huggingface dataset:
Does caffeine confound relationships among adrenergic tone, blood pressure and sleep apnoea?
Authors: Bardwell  , Ziegler  , Ancoli‐Israel  , Berry  , Nelesen  , Durning  , Dimsdale
Published: March 2003

The purpose of this study was to determine whether caffeine consumption confounds the relationship among adrenergic tone, as measured by urinary norepinephrine (NE), blood pressure (BP) and obstructive sleep apnoea (OSA). Data were analysed using correlation and regression analysis, analysis of covariance and t‐tests. Subjects included normotensives and hypertensives with and without OSA: 38 men, 23 women, aged 30–60 y; 100–150% of ideal body weight; without other major illness. Patients were studied using polysomnography, caffeine consumption was assessed, 24‐h urinary NE levels were examined and ambulatory BP was recorded. Patients with OSA (N=27) reported significantly greater caffeine consumption than those without OSA (N=34) (295 vs. 103 mg, P=0.010), but caffeine was not significantly correlated with their ambulatory BP. In contrast, NE excretion correlated with caffeine consumption (r=0.24, P=0.041), apnoea severity (r=0.65, P < 0.001) and BP (r=0.34, P < 0.005). Significant OSA–NE and BP–NE relationships remained even after controlling for caffeine consumption. Patients with OSA consumed nearly three times the amount of caffeine as patients without OSA. While caffeine partially explains the increased adrenergic tone in patients with OSA and the relationship between BP and NE, it does not appear to contribute significantly to the relationship between OSA and elevated BP.
Temporal and stagewise distribution of high frequency EEG activity in patients with primary and secondary insomnia and in good sleeper controls
Authors: Perlis Michael L. , Kehr Elizabeth L. , Smith Michael T. , Andrews Patrick J. , Orff Henry , Giles Donna E.
Published: March 2003

In the present study, we evaluate the temporal and stagewise distribution of high frequency EEG activity (HFA) in primary and secondary insomnia. Three groups (n=9 per group) were compared: primary insomnia (PI), Insomnia secondary to major depression (MDD), and good sleeper controls (GS). Groups were matched for age, sex and body mass. Average spectral profiles were created for each sleep epoch. Grand averages were created for each NREM cycle and each stage of sleep after removing waking and movement epochs and epochs containing micro or miniarousals. It was found that HFA (in terms of relative power) tends to increase across NREM cycles, occurs maximally during stage 1 and during REM sleep, and that both these effects are exaggerated in patients with PI. In addition, HFA was found to be inversely associated with Delta activity and the three groups in our study appear to exhibit characteristic Delta/Beta patterns. Our data are consistent with the perspective that HFA is related to CNS arousal to the extent that Beta/Gamma activity occurs maximally during shallow stages of sleep and maximally in subjects with PI.
Time‐courses in renin and blood pressure during sleep in humans
Authors: CHARLOUX A. , PIQUARD F. , EHRHART J. , METTAUER B. , GENY B. , SIMON C. , BRANDENBERGER G.
Published: March 2003

We previously described a strong concordance between nocturnal oscillations in plasma renin activity (PRA) and the rapid eye movement (REM) and non‐REM (NREM) sleep cycles, but the mechanisms inducing PRA oscillations remain to be identified. This study was designed to examine whether they are linked to sleep stage‐related changes in arterial blood pressure (ABP). Analysis of sleep electroencephalographic (EEG) activity in the delta frequency band, intra‐arterial pressure, and PRA measured every 10 min was performed in eight healthy subjects. Simultaneously, the ratio of low frequency power to low frequency power + high frequency power [LF/(LF + HF)] was calculated using spectral analysis of R–R intervals. The cascade of physiological events that led to increased renin release during NREM sleep could be characterized. First, the LF/(LF + HF) ratio significantly (P < 10−4) decreased, indicating a reduction in sympathetic tone, concomitantly to a significant (P < 10−3) decrease in mean arterial pressure (MAP). Delta wave activity increased (P < 10−4) 10–20 min later and was associated with a lag of 0–10 min with a significant rise in PRA (P < 10−4). Rapid eye movement sleep was characterized by a significant increase (P < 10−4) in the LF/(LF + HF) ratio and a decrease (P < 10−4) in delta wave activity and PRA, whereas MAP levels were highly variable. Overnight cross‐correlation analysis revealed that MAP was inversely correlated with delta wave activity and with PRA (P < 0.01 in all subjects but one). These results suggest that pressure‐dependent mechanisms elicit the nocturnal PRA oscillations rather than common central processes controlling both the generation of slow waves and the release of renin from the kidney.
The impact of a nap opportunity during the night shift on the performance and alertness of 12‐h shift workers
Authors: Purnell M. T. , Feyer A.‐M. , Herbison G. P.
Published: March 2003

The purpose of this workplace evaluation was to assess the effects on performance, alertness and subsequent sleep of strategic napping on 12‐h overnight shifts. In a counterbalanced crossover design, 24 male aircraft maintenance engineers working in a forward rotating 12‐h shift pattern volunteered to take part in the study for two work weeks. During the experimental week, each subject was given the opportunity to take a 20‐min nap at work between 01:00 and 03:00 h on each of their two overnight shifts. On the control week no naps were taken on the night shifts. A computerized neurobehavioural test battery was employed to assess performance and subjective levels of fatigue at the beginning and end of each night shift, and pre‐ and postnap. Subjects were also asked to rate how near they had come to falling asleep while driving to and from work. The results revealed that taking a single 20‐min nap during the first night shift significantly improved speed of response on a vigilance task measured at the end of the shift compared with the control condition. On the second night shift there was no effect of the nap on performance. Taking a short nap during either night shift had no significant effect on subjective ratings of fatigue, the level of sleepiness reported while driving to and from work, or subsequent sleep duration and sleep quality. Overall the results suggest some promise for a short duration nap taken in the workplace to counteract performance deficits associated with the first night shift.
Evolution of sleep and sleep EEG after hemispheric stroke
Authors: Vock Jacqueline , Achermann Peter , Bischof Matthias , Milanova Milena , Müller Caroline , Nirkko Arto , Roth Corinne , Bassetti Claudio L.
Published: March 2003

The evolution of subjective sleep and sleep electroencephalogram (EEG) after hemispheric stroke have been rarely studied and the relationship of sleep variables to stroke outcome is essentially unknown. We studied 27 patients with first hemispheric ischaemic stroke and no sleep apnoea in the acute (1–8 days), subacute (9–35 days), and chronic phase (5–24 months) after stroke. Clinical assessment included estimated sleep time per 24 h (EST) and Epworth sleepiness score (ESS) before stroke, as well as EST, ESS and clinical outcome after stroke. Sleep EEG data from stroke patients were compared with data from 11 hospitalized controls and published norms. Changes in EST (>2 h, 38% of patients) and ESS (>3 points, 26%) were frequent but correlated poorly with sleep EEG changes. In the chronic phase no significant differences in sleep EEG between controls and patients were found. High sleep efficiency and low wakefulness after sleep onset in the acute phase were associated with a good long‐term outcome. These two sleep EEG variables improved significantly from the acute to the subacute and chronic phase. In conclusion, hemispheric strokes can cause insomnia, hypersomnia or changes in sleep needs but only rarely persisting sleep EEG abnormalities. High sleep EEG continuity in the acute phase of stroke heralds a good clinical outcome.
Periodic limb movement disorder of sleep in children
Authors: Crabtree Valerie Mclaughlin , Ivanenko Anna , O'Brien Louise Margaret , Gozal David
Published: March 2003

To characterize periodic limb movement disorder (PLMD) in a cohort of prepubertal children we examined sleep‐related identifiable differences between children with PLMD and attention‐deficit/hyperactivity disorder (ADHD), PLMD alone, and age‐matched controls. Children were selected from a chart review of all children referred to a pediatric sleep medicine center and from a community survey of 5–7‐year‐old‐children. Polysomnography (PSG) and parental report data from all children identified as having periodic limb movement index (PLMI) >5 were reviewed and compared with a cohort of age‐matched controls. A total of 8.4% of children in the clinic‐referred sample, and 11.9% of the children recruited from the community had PLMI >5. Of those, 44.4% were identified as having ADHD. Children with PLMD had significantly lower percentage of rapid eye movement (REM) than control children (P < 0.001). Children in the PLMD/ADHD group had a significantly greater number of arousals associated with PLM (PLMa) than children with PLMD only (P < 0.05). While a relationship between ADHD and PLMD was observed, it was weaker than previous reports (Chervin, R. D. et al. Sleep, 2002; 25: 213; Chervin, R. D. and Archbold, K. H. Sleep, 2001; 24: 313; Picchietti et al. J. Child Neurol., 1999; 13: 588; Picchietti et al. Mov. Disord., 1999; 14: 1000; Picchietti and Walters Sleep, 1999; 22: 297). Children in the PLMD/ADHD group were more likely to have PLMas than were children with PLMD only. We postulate that rather than a direct relationship between ADHD and PLMD, this link may be mediated by the presence of reduced REM sleep and more importantly by the sleep fragmentation associated with PLM‐induced arousals.
Endocrine and metabolic responses in sheep during halothane and pentobarbitone anaesthesia with dobutamine infusion
Authors: Taylor
Published: March 2003

The study investigated the stimulus to pituitary‐adrenocortical activity (PACA) during halothane anaesthesia. Groups of six sheep were anaesthetized with thiopentone/halothane (TH group), acepromazine/thiopentone/halothane (ATH group) or pentobarbitone (P group). Dobutamine was infused in the TH and ATH groups to prevent hypotension (0.3–1.4 μg/kg/min) and in the P group at 0.05 μg/kg/min. Pulse rate, arterial blood gases and pressure (ABP) were measured and sequential blood samples taken for assay of cortisol, adrenocorticotrophic hormone (ACTH), arginine vasopressin (AVP), glucose and lactate. Pulse rate increased in all groups. Arterial blood pressure decreased by 13% in TH, by 24% in ATH and remained stable in P. All three groups developed hypercapnia and acidosis but were well oxygenated. Cortisol increased in all groups; with ATH the sevenfold rise occurred earlier than with either TH (sixfold rise) or P (fivefold rise). Adrenocorticotrophic hormone changes were as for cortisol but AVP increases were not consistent. Glucose and lactate were stable, but lactate was lowest with ATH. Dobutamine infusion failed to prevent hypotension during halothane anaesthesia and PACA appeared proportional to the hypotension. Dobutamine may have stimulated ACTH and cortisol release after 120 min. Halothane‐induced hypotension may cause adrenocortical activity but a direct effect of halothane cannot be ruled out.
Pharmacokinetic disposition and arthropathic potential of oral ofloxacin in dogs
Authors: YOSHIDA K. , YABE K. , NISHIDA S. , YAMAMOTO N. , OHSHIMA C. , SEKIGUCHI M. , YAMADA K. , FURUHAMA K.
Published: March 2003

We examined the relation between the pharmacokinetic disposition and arthropathic potential of ofloxacin, a new quinolone antibacterial agent, using both male immature (3‐month‐old) and mature (18‐month‐old) beagles. Ofloxacin was orally administered to these dogs at 20 mg/kg once daily for 8 consecutive days, and the animals were killed 2 h after the last treatment. Serum ofloxacin concentrations were repeatedly measured on days 1 and 7 by use of high‐performance liquid chromatography (HPLC), and pharmacokinetic parameters were calculated. In addition, on day 8, the drug concentrations in the joint synovial fluid and humeral and femoral condyles were measured. Clinico‐pathological tests of blood and serum or histopathological examination of bone specimens were also performed. Arthropathy was macroscopically observed in the cartilage surface of all immature dogs, but not in mature dogs. There were, however, no noticeable differences in pharmacokinetic parameters between the two age groups of dogs or between single and 7‐day treatments. In contrast to the occurrence of arthropathic lesions, the synovial fluid and condylar drug concentrations in immature dogs was equal to or lower than those in mature dogs, suggesting that the pharmacokinetic disposition of ofloxacin may not be essential for cartilage lesions.
Pharmacokinetics and milk discard times of pirlimycin after intramammary infusion: a population approach
Authors: Whittem
Published: March 2003

A population pharmacokinetic approach was used to analyse milk concentration data to determine whether milk discard times and the clearance of intramammary infusions of pirlimycin could be adequately predicted by readily available demographic variables. Milk samples were collected at 12 hourly milking intervals after dosing with pirlimycin during product development from both normal cows (primary data) and cows with naturally occurring mastitis (validation data) and pirlimycin concentration was determined by microbial inhibition assay. The data were analysed by the conditional estimation/maximum likelihood population approach within the computer program PPharm and fitted a two compartment open model. Bayesian estimates of individual parameters allowed solutions for each cow, predicting the time after last dosing by which milk concentration reached the target safe concentration. From this population of times, the 95% confidence interval of the 99th percentile was defined as the milk discard time. After elimination of one very low producing outlier, the calculated discard time agreed with the label recommendation of 36 h (3 milkings, USA) after the last dose. Milk pirlimycin clearance was strongly and positively correlated to the logarithm of the kilograms of milk produced in 24 h at time of dosing (r2 = 0.939). Agreement was strong at most time points between predicted and measured pirlimycin concentrations in milk from cows with mastitis. This alternative method for determining milk discard times was compared to existing recommendations.
Withdrawal time estimation of veterinary drugs: extending the range of statistical methods
Authors: Fisch R. D.
Published: March 2003

In order to use a drug in a food producing animal, evidence has to be provided that after a certain withdrawal time, drug residues in tissues, such as muscle meat, fat, liver, kidney etc., are below a given maximum residue limit (MRL), for a majority of animals. Several statistical methods, both regression based and nonparametric based methods, have been proposed, each relying on different sets of assumptions, which may or may not hold for the specific data situation. The purpose of this paper is to enrich the range of methods, i.e. to provide approaches for situations where current methods are inappropriate. Bayesian methods, using Markov chain Monte Carlo, are proposed to derive inference on the parameters of interest.
Pharmacokinetics of ketoprofen enantiomers after intravenous administration of racemate in camels: effect of gender
Authors: Al Katheeri N. A. , Wasfi I. A. , Lambert M. , Saeed A. , Khan I. A.
Published: March 2003

The pharmacokinetics of ketoprofen (KP) enantiomers were studied in ten female and eight male camels after a single intravenous dose (2.0 mg/kg) of racemic KP. A high performance liquid chromatographic (HPLC) method was developed for the quantitation of the R‐ and S‐enantiomers without derivatization of the samples using a S,S‐Whelk‐01 chiral stationary phase column. The data collected (median and range) were as follows: the areas under the curve to infinity (AUC) (μg/mL per h) were 22.4 (13.5–29.7) and 19.8 (13.8–22.1) for R‐ and S‐KP, respectively, in female camels while the corresponding values in male camels were 16.0 (12.9–22.4) and 14.4 (11.0–19.3). In both sexes, the AUC for the R‐enantiomer was significantly larger than that of the S‐enantiomer. Total body clearances (Clt) were 44.6 (33.7–74.1) and 50.6 (45.2–72.4) mL/kg per h for R‐ and S‐KP, respectively, in female camels and were 62.8 (44.6–77.8) and 69.6 (51.8–91.1) mL/kg per h for R‐ and S‐KP, respectively, in male camels. In both sexes of camels, the Clt values for R‐KP were significantly lower than its corresponding antipode. The steady‐state volumes of distribution (Vss) were 97.9 (82.8–147.2) and 102.0 (90.1–169.0) mL/kg for R‐ and S‐KP, respectively, in female camels and were significantly different from each other, while the respective values in male camels were 151.5 (105.3–222.3) and 154.0 (114.7–229.0) mL/kg but were not significantly different from each other. The volumes of distribution (area) followed a similar pattern, where the values for R‐ and S‐KP in female camels were 118.5 (95.6–195.2) and 137.6 (115.8–236.2) mL/kg, respectively, and the respective values in male camels were 215.6 (119.1–270.1) and 229.1 (143.3–277.4) mL/kg.

The elimination half‐lives (t1/2β) were 1.88 (1.42–2.34) h and 1.83 (1.67–2.26) h for R‐ and S‐KP, respectively, in female camels and were significantly different from each other, while the corresponding values in male camels were 2.11 (1.50–4.20) and 2.33 (1.52–3.83) h for R and S‐KP, respectively, but were not significantly different from each other. The mean residence time followed a similar pattern. All pharmacokinetic parameters for R‐ and S‐KP in female camels were significantly different from their corresponding values in male camels.

The extent of protein binding for R‐ and S‐KP was evaluated in vitro by ultrafiltration. The extents of protein binding for R‐ and S‐KP were not significantly different from each other when each enantiomer was supplemented separately. However, when the enantiomers were supplemented together, protein binding of R‐KP was significantly higher than that of S‐KP in female but not in male camels.
Pharmacokinetics of ricobendazole in calves
Authors: Formentini E. A. , Mestorino O. N. , Mariño E. L. , Errecalde J. O.
Published: March 2003

The pharmacokinetics of ricobendazole (RBZ) and its major metabolite albendazole sulphone (ABZSO2) were studied in six calves, after administration of RBZ (7.5 mg/kg), using a 10% experimental solution by the intravenous (i.v.) route, a 10% commercial solution by the subcutaneous (s.c.) route, and a 10% experimental suspension by the intraruminal (i.r.) route. Blood samples were drawn during a 60‐h period. Plasma drug and metabolite concentrations were determined by HPLC. The pharmacokinetic evaluation in each case was prepared by weighted least‐squares nonlinear regression analysis. Ricobendazole i.v. data were best fitted by a two‐compartment model. The best pharmacokinetic exponents and coefficients were estimated, and the pharmacokinetic variables for RBZ and ABZSO2 were calculated from them. Similar patterns of plasma disposition were found for RBZ after i.r. and s.c. administration, suggesting delayed release from the s.c. site resembling the slow release of the drug from the rumen.
Some factors influencing the level of clinical sedation induced by medetomidine in rabbits
Authors: RAEKALLIO M. , ANSAH O. B. , KUUSELA E. , VAINIO O.
Published: March 2003

Rabbits (n=23) received intravenous bolus medetomidine at 100 μg/kg. Prior to medetomidine administration, heart and respiratory rates were measured, arterial blood was collected and analysed for plasma cortisol, glucose and albumin concentrations. Fifteen minutes after medetomidine administration, heart and respiratory rates were measured again and sedation was scored. The rabbit was afterwards anaesthetized with 20 mg/kg ketamine administered intravenously to enable spinal tap and heart puncture. Cerebrospinal fluid (CSF) was collected (this occurred 20 min post medetomidine administration) and analysed for medetomidine concentration. Blood was collected by heart puncture immediately after the spinal tap and analysed for serum medetomidine concentration. Cerebrospinal fluid medetomidine concentration correlated negatively with sedation. Serum medetomidine correlated positively with CSF medetomidine concentration. Cerebro‐spinal fluid medetomidine was 17 ± 13% of serum medetomidine concentration. Plasma cortisol and glucose concentrations correlated negatively with serum medetomidine. We conclude that after an intravenous bolus administration of a low sedative dose of medetomidine to rabbits; CSF concentration of the drug correlate negatively with sedation and that this may be because of the fact that only the free and unbound medetomidine may be available for detection in the CSF, the concentration of medetomidine detected in the CSF was much lower than that in blood and a positive correlation exists between CSF and serum medetomidine concentrations. Stress may have some effect on the distribution or metabolism of medetomidine in rabbits.
Evolution of hepatitis G virus infection and antibody response to envelope protein in patients with transfusion‐associated non‐A, non‐B hepatitis
Authors: Tanaka E. , Kiyosawa K. , Shimoda K. , Hino K. , Tacke M. , Schmolke S. , Engel A. M. , Hess G.
Published: March 2003

The clinical significance and course of acute hepatitis G virus (HGV) infection were studied by measuring HGV RNA and antibody to HGV envelope protein E2 (HGV‐E2 antibody). A total of 59 patients with transfusion‐associated non‐A, non‐B hepatitis, who were followed‐up for more than 1 year, were selected retrospectively. HGV RNA was measured by reverse transcriptase (RT) and nested polymerase chain reaction (PCR) was performed, using primer sets, in the 5′‐non‐coding region of the HGV genome. HGV‐E2 antibody was measured by enzyme‐linked immunosorbent assay (ELISA) using recombinant E2 protein. Of the 59 patients, 51 (86%) were infected with hepatitis C virus (HCV) and 12 (20%) were infected with HGV; 11 of the 12 with HGV infection were also infected with HCV. HGV viraemia was cleared during the follow‐up period in seven of the 12 patients with HGV infection. All these seven patients seroconverted for HGV‐E2 antibody just before or just after the clearance of HGV viraemia. In contrast, all five patients without clearance of HGV viraemia were negative for HGV‐E2 antibody (P=0.0013). Of seven patients with continuous HGV viraemia at 1 year from the onset of acute hepatitis, four with HCV RNA showed chronic elevation of alanine aminotransferase (ALT) but three without HCV RNA did not. The severity of acute hepatitis was similar between patients with both HGV and HCV infections and in those with HCV infection alone. The majority of patients with HGV infection cleared the virus during long‐term follow‐up. Appearance of HGV‐E2 antibody was associated with the clearance of HGV viraemia. An abnormal ALT level was noted to depend on HCV infection but not on HGV infection in both the acute and chronic phases of transfusion‐associated hepatitis.
Hepatitis C virus infection: virus/host interactions
Authors: Pawlotsky J. ‐M.
Published: March 2003

Infection with the hepatitis C virus (HCV) is a leading cause of chronic liver disease world‐wide. This paper examines our current understanding of the complex relationship between HCV and its host, especially potential mechanisms of viral persistence and resistance to interferon therapy, and the pathogenesis of liver injury in chronic HCV infection. HCV infection leads to viral persistence and chronic disease in a very high proportion of cases, despite broad humoral and cellular immunological responses to viral proteins. These responses may be thwarted by the high rate of mutation, which leads to the generation of a highly variable mixture of closely related genomes, referred to as a quasispecies, that persists and continuously evolves in infected individuals. Understanding this, and other mechanisms of viral persistence and immune escape, will be essential in developing effective future therapeutic and preventive strategies. As far as the pathogenesis of chronic hepatitis C is concerned, two non‐mutually exclusive hypotheses have been raised: first, that HCV can be cytopathic and induce liver lesions by replicating in infected hepatocytes, and second, that liver lesions could be the result of specific or non‐specific immune responses. In the absence of a cell‐culture model, the direct cytopathogenicity of the virus cannot be assessed confidently. Recent data suggest that cytotoxic T cells and cytokines produced by both CD4 + (T helper) and cytotoxic T cells may be responsible for much of the damage that occurs in the livers of infected patients.
Evidence for sequence selection within the non‐structural 5A gene of hepatitis C virus type 1b during unsuccessful treatment with interferon‐α
Authors: Gerotto M. , Dal Pero F. , Sullivan D. G. , Chemello L. , Cavalletto L. , Polyak S. J. , Pontisso P. , Gretch D. R. , Alberti A.
Published: March 2003

Resistance of the hepatitis C virus (HCV) to interferon‐α (IFN‐α) therapy in patients with hepatitis C may be genetically controlled by an IFN sensitivity‐determining region (ISDR) within the non‐structural 5A (NS5A) gene. To assess whether HCV 1b strains carrying a ‘resistant’ type of ISDR are selected during unsuccessful IFN therapy, we analysed the evolution of the NS5A quasispecies, as detected by the clonal frequency analysis technique, and of the ISDR sequence by nucleotide sequence determination, in 11 patients showing no virological response during two consecutive cycles of IFN‐α therapy. IFN‐resistant patients had a homogeneous ISDR quasispecies with sequences identical to those described as ‘resistant‐’ or ‘intermediate‐’ type ISDR. After retreatment with IFN, further selection towards a homogeneous viral population was observed and 10 out of 11 patients had only one variant of HCV with no or just one single amino acid mutation within the ISDR sequence. Treatment and retreatment with IFN was associated in our non‐responder patients with evolution of the ISDR quasispecies towards a rather homogeneous viral population carrying a conserved or minimally mutated ISDR motif, supporting the idea that this motif may be relevant for IFN resistance in HCV 1b‐infected individuals.
The non‐structural 5A protein of hepatitis C virus
Authors: Pawlotsky J. M. , Germanidis G.
Published: March 2003

The non‐structural (NS)5A protein of hepatitis C virus (HCV) is cleaved, after translation, by the NS3‐encoded zinc‐dependent serine proteinase, from the NS4B protein upstream and the NS5B protein downstream. The released, mature NS5A protein is a 56 000 MW phosphoprotein (p56), which also exists within infected cells in a hyperphosphorylated form (p58). The NS5A gene has a quasispecies distribution, meaning that various NS5A sequences co‐exist, in various proportions, in infected individuals. HCV NS5A appears to be located in cytoplasmic membranes surrounding the nucleus. Its precise functions are not known. HCV non‐structural proteins, including NS5A, form a large multiprotein replication complex, which probably directs the replication of the HCV genome. HCV NS5A lacking the 146 N‐terminal amino acids is a potent transcriptional activator in vitro. NS5A can also bind to single‐strand RNA‐dependent protein kinase (PKR) and inhibit its antiviral function. An ‘interferon (IFN) sensitivity‐determining region’ has recently been postulated in the NS5A protein central region in hepatitis C virus (HCV) genotype 1b, but strongly conflicting evidence has been published. In fact, there would seem to be no such region in the NS5A protein, even though NS5A plays an important and complex role in HCV resistance to IFN. Structure–function studies are required to identify precisely how NS5A and IFN interact.
Biochemical and virological response to interferon therapy in patients with chronic hepatitis C, co‐infected with hepatitis G virus
Authors: Shibahara  , Moriyama  , Abe  , Tanaka  , Arakawa
Published: March 2003

It is still unclear whether hepatitis G virus (HGV) and hepatitis C virus (HCV) co‐infection influences the natural course of chronic HCV infection and the response to interferon (IFN) therapy. In this study, we investigated the HGV RNA changes and responses to IFN therapy of HGV RNA‐positive hepatitis C patients. The presence of HGV RNA in 264 patients who received IFN therapy at our institute was examined using the nested reverse transcription–polymerase chain reaction with primers deduced from the 5′ non‐coding region (5′‐NCR) of the HGV. We also analysed the nucleotide sequences of the 5′‐NCR and HGV before and after IFN therapy. Twelve (4.5%) of the 264 patients with chronic hepatitis C were HGV RNA positive. Of these 12 patients, full information was available in nine. Of those nine, seven became HGV RNA negative during IFN therapy but in six of the seven a relapse occurred. HCV RNA was not detected 1 year after IFN therapy in six patients. The serum alanine aminotransferase (ALT) levels of all the HCV RNA‐negative patients post‐treatment was within the normal range, irrespective of their HGV RNA status. Nucleotide sequences did not change after IFN therapy in any patient. Hence, we confirm that HGV is sensitive to IFN therapy but the liver damage caused by HGV infection is very mild or does not modify the liver function tests.
Inhibition of the replication of the DNA polymerase M550V mutation variant of human hepatitis B virus by adefovir, tenofovir, L‐FMAU, DAPD, penciclovir and lobucavir
Authors: Ying  , De Clercq  , Nicholson  , Furman  , Neyts
Published: March 2003

Several nucleoside analogues (penciclovir, lobucavir, dioxalane guanine [DXG], 1‐β‐2,6‐diaminopurine dioxalane [DAPD], L‐FMAU, lamivudine) and acyclic nucleoside phosphonate analogues (adefovir, tenofovir) that are in clinical use, in clinical trials or under preclinical development for the treatment of hepatitis B virus (HBV) infections, were evaluated for their inhibitory effect on the replication of a la‐ mivudine‐resistant HBV variant containing the methionine → valine substitution (M550V) in the polymerase nucleoside‐binding domain. The antiviral activity was determined in the tetracycline‐responsive HepAD38 and HepAD79 cells, which are stably transfected with either a cDNA copy of the wild‐type pregenomic RNA or with cDNA containing the M550V mutation. As expected, lamivudine was much less (≈ 200‐fold) effective at inhibiting replication of the M550V mutant virus than the wild‐type virus. In contrast, adefovir, tenofovir, lobucavir, L‐FMAU, DXG and DAPD proved almost equally effective against both viruses. A second objective of this study was to directly compare the antiviral potency of the anti‐HBV agents in HepG2 2.2.15 cells (which are routinely used for anti‐HBV drug‐screening purposes) with that in HepAD38 cells. HepAD38 cells produce much larger quantities of HBV than HepG2 2.2.15 cells, and thus allow drug screening in a multiwell plate format. All compounds were found to be almost equally effective at inhibiting HBV replication in HepAD38 cells (as in HepG2 2.2.15 cells), except for penciclovir, which was clearly less effective in HepAD38 cells.
Methodological issues in papers on IFN therapy: time for reappraisal
Authors: Kondili  , Taliani  , Tosti  , De Bac  , Pasquazzi  , Mele
Published: March 2003

We conducted an analytical review of 194 full papers on interferon (IFN) therapy for chronic hepatitis C to evaluate current methodology (i.e. study design, criteria for evaluating the efficacy of therapy and predictors of response). Of the papers evaluated, 64 were randomized controlled trials (RCT), 40 were non‐randomized controlled trials (NRCT) and 90 were observational studies (OS). The methodological analysis was focused mainly on clinical trials. The number of patients enrolled in RCT was higher compared with the number enrolled in NRCT. Uniform enrolment criteria were used in less than 50% of the trials. Only 20% of RCT and 2.5% of NRCT used criteria for defining sample size. The response rate was calculated on an intention‐to‐treat basis in 36 of the RCT and in 14 of the NRCT. The outcome of treatment and the criteria employed to define the response to treatment were found to be far from standardized. In 51.5% of the RCT and 42.5% of the NRCT, normalization of alanine aminotransferase (ALT) level at the end of follow‐up was the only marker of response studied. Only 57.6% of the trials considered histological evidence as an important outcome. Among the clinical trials, 71.1% evaluated predictors of good response to IFN therapy. In 51% of the OS, ALT normalization by the end of follow‐up was the only criterion for defining response. In conclusion, to ensure a high level of reliability in comparing or combining the results of different studies, some basic general requirements must be followed when planning trials on antiviral therapy.
How can the cellular immune response control hepatitis B virus replication?
Authors: Maini  , Bertoletti
Published: March 2003

In this review we focus on aspects of the virus‐specific cellular immune response, although we should point out that all the components of the innate and adaptive immune response are likely to play a role in successful control of hepatitis B virus (HBV) infection. We concentrate particularly on the relevance of the polyclonality and multispecificity of the HBV‐specific cytotoxic T cell response to its antiviral activity. In this context, we discuss the possible role of viral escape mutations and highlight evidence from other models of the benefit of multispecificity in antiviral responses. We stress the contribution of CD4 help for effective CD8 responses and raise the possibility that HBV may produce factors inhibiting the antiviral response.
Evaluation of an enzyme‐linked immunosorbent assay for detection and quantification of hepatitis B virus PreS1 envelope antigen in serum samples: comparison with two commercial assays for monitoring hepatitis B virus DNA
Authors: Guillou D. Buffello‐Le , Duclos‐Vallée  , Eberle  , Capel  , Petit
Published: March 2003

An in‐house sensitive and easy‐to‐use solid‐phase enzyme‐linked immunoassay (ELISA) was adapted for the detection and quantification of hepatitis B virus (HBV) PreS1 envelope antigen in serum, and compared with the HBV DNA Hybrid Capture™ system from Murex and the polymerase chain reaction (PCR) Amplicor™ HBV Monitor assay from Roche. Twenty‐five patients with chronic hepatitis B after liver transplantation were included in this study. The sensitivity of our ELISA was found to be 50 pg of HBsAg/PreS1Ag ml–1. The linearity was between 0.1 and 100 ng ml–1. Intra‐assay reproducibility was obtained with a standard deviation of <1%. No correlation between the presence of serum PreS1 antigen and viral DNA detected by direct hybridization (Murex) was observed. In contrast, there was a significant 96% correspondence in the presence of PreS1 antigen and viral DNA detected and quantified by the PCR assay (Roche). In conclusion, the most important and reliable markers for monitoring residual HBV replication in serum were HBV DNA by the PCR assay, and virus envelope PreS1Ag by our in‐house ELISA. Thus, PreS1Ag disappearance in serum could be used for evaluating the efficacy of antiviral therapies.
Serum cytokine values and fatigue in chronic hepatitis C infection
Authors: Gershon A. S. , Margulies M. , Gorczynski R. M. , Heathcote E. J.
Published: March 2003

Patients with chronic hepatitis C infection often experience fatigue. In many clinical situations, an association between fatigue and altered serum cytokine levels has been found. Altered cytokine levels in patients with hepatitis C have not shown a correlation with the degree of serum transaminase elevation or pathological change on liver biopsy. The aim of our study was to examine whether there was an association between abnormal serum cytokine levels and fatigue in patients with compensated chronic hepatitis C. Patients referred to a tertiary care hepatology clinic who were hepatitis C antibody positive and who had elevated alanine aminotransferase (ALT) levels were eligible for entry into the study. A control group was also included. Subjects in both groups who had characteristics other than hepatitis C that were known to alter cytokine values and/or cause fatigue were excluded. Patients completed a validated questionnaire to determine their fatigue severity score (FSS). Bioassays were used to measure interleukin (IL)‐1, IL‐6 and tumour necrosis factor‐α (TNF‐α) levels in early morning serum samples taken from patients and controls. Altered cytokine values were defined as those more than two standard deviations above the mean control value. Data was analysed using SPSS, version 8.01. Of the 78 patients with chronic hepatitis C who participated in the study, 19 (24%), 24 (30%) and 45 (56%) had elevated levels of IL‐1, IL‐6 and TNF‐α, respectively, compared with only two (6%) of the control group who had elevation of any of the three cytokines. No correlation was found between the FSS and serum cytokine levels, when analysed singly or in combination, in patients with chronic hepatitis C. Hence, alteration in early morning serum levels of IL‐1, IL‐6 and TNF‐α in patients with chronic hepatitis C infection and elevated ALT levels bear no correlation with the symptom of fatigue.
Sequence, expression and reconstitution of an HCV genome from a British isolate derived from a single blood donation
Authors: Kumar U. , Tuthill T. , Thomas H. C. , Monjardino J.
Published: March 2003

Morphological analysis of hepatitis C virus and development of antiviral drugs to eradicate this agent have been seriously hampered by the low viraemias observed during natural infection and the unavailability of a cell culture system for virus propagation. Recently a low‐grade hepatitis has been reported in chimpanzees after intrahepatic transfection of full‐length synthetic HCV RNA and successful infections shown to be critically dependent on the integrity and genetic homogeneity of the reconstituted clone. In this study we describe and characterize a full HCV RNA sequence derived from a case of chronic sporadic hepatitis. The genotype was shown to be 1a with a low level of intraclonal sequence heterogeneity, and processing of both structural and nonstructural proteins has been documented. The assembly of the full genome has also been achieved.

The low level of intraclonal variation observed may reflect infection with a single isolate and the fact that cloning was performed on virus obtained from a single blood donation makes this clone a good candidate for future in vivo and in vitro transfection studies.
Failure to infect rhesus monkeys with hepatitis C virus strains of genotypes 1a, 2a or 3a
Authors: Bukh J. , Apgar C. L. , Govindarajan S. , Emerson S. U. , Purcell R. H.
Published: March 2003

The chimpanzee is the only recognized animal model for the study of hepatitis C virus (HCV). However, recently it was reported that rhesus monkeys were susceptible to HCV and developed hepatitis during infection. In the present study, we inoculated two rhesus monkeys each with HCV strain H77 (genotype 1a), strain HC‐J6 (genotype 2a) or strain S52 (genotype 3a). Weekly serum samples were tested for liver enzyme values, HCV antibodies and HCV RNA. We did not find evidence of HCV infection in any of the monkeys during 24 weeks of follow‐up. Our study demonstrates that rhesus monkeys are not readily infected with HCV and apparently do not represent a useful animal model for the study of HCV.
World‐wide epidemiology of HBeAg‐negative chronic hepatitis B and associated precore and core promoter variants
Authors: Funk M. L. , Rosenberg D. M. , Lok A. S. F.
Published: March 2003

Hepatitis B is a serious disease that is endemic in many parts of the world. A significant proportion of patients with chronic hepatitis B (CHB) are infected with a variant form of the hepatitis B virus (HBV) which decreases or abolishes the production of hepatitis B e‐antigen (HBeAg). The purpose of this literature review is to describe the epidemiology of HBeAg‐negative CHB (e‐CHB) worldwide. A literature search was conducted to identify studies pertaining to e‐CHB and underlying variants (precore and core promoter). Fifty studies were included in our analysis. The median prevalence of e‐CHB among patients with chronic HBV infection was 33% in the Mediterranean, 15% in Asia Pacific, and 14% in the USA and Northern Europe. The pre core stop codon variant was detected in a median of 60% (range 0–100%) of HBeAg‐negative patients overall, 92% in the Mediterranean, 50% in Asia Pacific and 24% in the USA and Northern Europe. There were very few data on the prevalence of core promoter variants outside Asia where the median prevalence among HBeAg‐negative patients was 77%. This literature review revealed that e‐CHB is more common than previously suspected and that it is present worldwide with marked variations in the prevalence of associated HBV variants across different geographical regions. Additional research using population based samples of adequate size based on a consensus definition of e‐CHB and using standardized HBV DNA assays is needed to better estimate the true prevalence of e‐CHB and its associated HBV variants.
Kinetics of serum cytokines reflect changes in the severity of chronic hepatitis C presenting minimal fibrosis
Authors: Neuman M. G. , Benhamou J. P. , Malkiewicz I. M. , Ibrahim A. , Valla D. C. , Martinot‐Peignoux M. , Asselah T. , Bourliere M. , Katz G. G. , Shear N. H. , Marcellin P.
Published: March 2003

Our aims were to measure the kinetics of serum tumour necrosis alpha (TNF‐α) and transforming growth factor beta (TGF‐β) levels as markers of progression of disease in nontreated chronic hepatitis C virus (HCV)‐infected patients with minimal or no fibrosis and minimal histology activity index (HAI) scores. Our study group consisted of 56 patients diagnosed with minimal (1) or no fibrosis (0) and minimal HAI (0–1) on their first biopsy as defined by Knodell and METAVIR scores. We compared their initial (entry of study) cytokine levels with a group of 103 HCV controls with minimal (0–1) to mild fibrosis (0–3) and mild HAI (5.5). Serum TNF‐α and TGF‐β levels were measured by enzyme‐linked‐immunosorbent‐assay. A significant difference was seen in TNF‐α levels at baseline in the study group vs. controls. Regardless of their HAI, there was a correlation between TGF‐β and degree of fibrosis. As shown by their biopsies, during the 3 years (from entry to follow up), many of the patients that initially had minimal fibrosis progressed to higher degree of fibrosis. This progression is paralleled by an increase in TGF‐β levels when comparing initial and follow‐up levels. In conclusion, serum TNF‐α reflects the progression of inflammation as seen in liver biopsies and TGF‐β reflects the degree of fibrosis in HCV patients.
Interferon retreatment reduces or delays the incidence of hepatocellular carcinoma in patients with chronic hepatitis C
Authors: Hino K. , Kitase A. , Satoh Y. , Fujiwara D. , Yamaguchi Y. , Korenaga M. , Shingai Y. , Konishi T. , Yamashita S. , Uchida K. , Mori K. , Hanada H. , Kodama T. , Nukui K. , Okita K.
Published: March 2003

summary. Inhibition of hepatocarcinogenesis is a crucial issue in treating chronic hepatitis C patients, especially those who do not respond completely to interferon therapy. Interferon has been reported to reduce the incidence of hepatocellular carcinoma (HCC) not only in sustained virological responders but also in transient biochemical responders. However, the incidence of HCC increases in 5 years or more after interferon therapy in transient biochemical responders. The aim of this study is to assess whether interferon retreatment reduces the incidence of HCC in chronic hepatitis C patients in whom hepatitis C virus was not eradicated during initial interferon therapy. We enrolled 309 patients who were not sustained virological responders after initial interferon treatment consisting of a total dose of more than 250 megaunits of interferon and were followed for more than 2 years after treatment. Ninety‐nine patients received interferon retreatment and 210 did not. Two courses of interferon therapy were administered in 84, three courses in 14 and five courses in one. The incidence of HCC was compared between patients with retreatment and those without. In the clinical characteristics, retreated patients were younger and followed up for a longer time period. The cumulative incidence of HCC was significantly lower in retreated patients. In multivariate analysis, patients' age (P=0.018) and the number of courses of interferon therapy (P=0.022) were independently associated with HCC incidence. These results suggest that interferon retreatment reduces or delays the incidence of HCC in chronic hepatitis Cpatients who did not completely respond to initial therapy.
Therapy of chronic hepatitis B: current challenges and opportunities
Authors: Liaw Y.‐F.
Published: March 2003

summary. Better understanding of hepatitis B virus (HBV) replication, natural history and the immunopathogenesis of chronic hepatitis B, together with the introduction of effective agents with different mechanisms of action are the basis for better therapeutic strategies against chronic hepatitis B. Among currently available drugs, interferon‐α and thymosin‐α1 have only modest efficacy (∼ 40%vs 9–20% in controls). In the past decade, lamivudine has dominated in the treatment of chronic HBV infection because it is easy to use, safe, and is effective in terms of hepatitis B e antigen and/or HBV‐DNA loss, ALT normalization, and improvement in histology. The response rate increases with increasing pretherapy alanine aminotransferase (ALT) levels, suggesting that patients with stronger endogenous immune response against HBV have a better response to direct antiviral agents. Lamivudine is also beneficial in decompensated cirrhotics with HBV replication. Hepatitic flares may occur after stopping lamivudine therapy in nonresponders and also in responders. Therefore, prolonged therapy is usually required. However, tyrosine‐methionine‐aspartate‐aspartate (YMDD ) mutations conferring resistance to lamivudine start to emerge after 6–9 months of therapy, and hepatitis flare, even decompensation, may develop after viral breakthrough. Thus the benefits of long‐term lamivudine therapy must be balanced against the concern about YMDD mutations and the durability of treatment response. Adefovir dipivoxil, entecavir, emtricitabine, clevudine and other nucleoside/ nucleotide analogues have shown encouraging results and some agents appear effective in patients with YMDD mutants. Further development of new drugs and new strategies may help to improve treatment in the new century.