bluuebunny/crossref_metadata_2025 huggingface dataset:
The Evolution of an Energy Analyst: Some Personal Reflections
Authors: Reddy Amulya K. N.
Published: October 2002

[Figure: see text]

▪ Abstract  The evolution from an electrochemist was motivated by a growing conviction that Indian science and technology should be reoriented. A cell was created in the Indian Institute of Science in 1974 to initiate and promote work of rural relevance as a weapon against poverty. Surveys led to a detailed empirical study of energy consumption patterns in villages and to the design and construction of rural energy centers. The lessons from this village work are described.

The principal outcome of the collaboration with J. Goldemberg (Brazil), T.B. Johansson (Sweden), and R.H. Williams (United States) was the book Energy for a Sustainable World that contributed significantly to the new paradigm for energy. The application of this paradigm resulted in a detailed electricity demand scenario for the South Indian state of Karnataka. Following mandatory retirement from the Indian Institute of Science, the International Energy Initiative (IEI) was set up in 1991 as a Southern-conceived, Southern-led, Southern-located South-North partnership. Persisting personal concerns about the ethical implications of science resurfaced through opposition to India's nuclear tests in 1998 and a visit to the concentration camps at Auschwitz. The associated human dimensions of energy were emphasized in the acceptance speech at Göteborg of the Volvo Environment Prize 2000. The penultimate endgame involved retirement.
Long-Term Adrenomedullin Administration in Experimental Heart Failure
Authors: Rademaker Miriam T. , Charles Chris J. , Espiner Eric A. , Nicholls M. Gary , Richards A. Mark
Published: October 2002

Short-term administration of adrenomedullin, a recently discovered peptide with potent vasodilator, natriuretic, and aldosterone-inhibitory actions, has beneficial effects in experimental and clinical heart failure. The effects of prolonged adrenomedullin administration have not previously been assessed in this setting. Consequently, in 16 sheep with pacing-induced heart failure, we infused either adrenomedullin (10 ng/kg per minute; n=8) or a vehicle control (Hemaccel; n=8) for 4 days. Compared with control data, infusion of adrenomedullin persistently increased circulating levels of the peptide (by ≈9.5 pmol/L;
P
<0.001), in association with prompt (15 minutes) and sustained (4 days) increases in cardiac output (day 4, 27%), and reductions in peripheral resistance (30%), mean arterial pressure (13%), and left atrial pressure (24%; all,
P
<0.001). Adrenomedullin also significantly enhanced urinary sodium excretion (day 4, 3-fold;
P
<0.05), creatinine excretion (1.2-fold;
P
<0.001), and creatinine clearance (1.4-fold;
P
<0.001) over the 4 days of treatment, whereas urine volume and cAMP excretion tended to be elevated (both, 0.1>
P
>0.05). Plasma renin activity was increased (
P
<0.05), whereas aldosterone levels were reduced in a sustained fashion (
P
<0.01). Plasma endothelin rose transiently (hours 1 to 6) after initiation of treatment (
P
<0.05). Despite substantial cardiac unloading, plasma concentrations of the natriuretic peptides were not significantly different from control. In conclusion, long-term administration of adrenomedullin induces pronounced and sustained cardiovascular and renal effects in experimental heart failure, including reductions in cardiac preload and afterload, as well as augmentation of cardiac output, sodium excretion, and glomerular filtration. These findings support the concept of adrenomedullin as a protective hormone during hemodynamic compromise with therapeutic potential in heart failure.
Comparison of Ion-Channel Subunit Expression in Canine Cardiac Purkinje Fibers and Ventricular Muscle
Authors: Han Wei , Bao Weisheng , Wang Zhiguo , Nattel Stanley
Published: October 2002

Although Purkinje fibers (PFs) play an important role in cardiac electrophysiology, almost nothing is known about the expression of ion-channel subunits in PFs. We applied competitive reverse transcription–polymerase chain reaction, Western blotting, and immunocytochemistry to compare the expression of ion-channel subunit mRNA and protein in canine PFs versus ventricular muscle (VM). For transient outward current–related subunits, Kv4.2 was not detected, and Kv1.4 expression was extremely low. Kv4.3 expression was of the same order for VM and PFs. The tetraethylammonium chloride–sensitive subunit Kv3.4 was expressed much more strongly in PFs than in VM, and Kv channel–interacting protein transcript expression was 25-fold stronger in VM than in PFs. For delayed rectifiers, ERG and KvLQT1 expression was lower in PFs at both mRNA and protein levels. Although minK transcripts were more numerous in PFs, minK protein was significantly more strongly expressed in VM. L-type Ca
2+
current α-subunit (Ca
V
1.2) and Na
+
-Ca
2+
exchanger proteins were more strongly expressed in VM than in PFs. For T-type Ca
2+
current, Ca
V
3.1, Ca
V
3.2, and Ca
V
3.3 transcripts were all more strongly expressed in PFs. For the nonselective cation current, hyperpolarization-activated cation channel 1 (HCN1) expression was subquantifiable, HCN2 transcript expression was comparable in PFs and VM, and HCN4 mRNA expression was strong in PFs but below the detection threshold in VM. HCN2 and HCN4 protein expression was much stronger in PFs than in VM. We conclude that ion-channel subunit expression in PFs differs from that in VM in ways that are consistent with, and shed light on the molecular basis of, well-recognized fundamental PF ionic properties.
Targeted Disruption of the Matrix Metalloproteinase-9 Gene Impairs Smooth Muscle Cell Migration and Geometrical Arterial Remodeling
Authors: Galis Zorina S. , Johnson Chad , Godin Denis , Magid Richard , Shipley J. Michael , Senior Robert M. , Ivan Eugen
Published: October 2002

Matrix remodeling plays an important role in the physiological and pathological remodeling of blood vessels. We specifically investigated the role of matrix metalloproteinase (MMP)-9, an MMP induced during arterial remodeling, by assessing the effects of genetic MMP-9 deficiency on major parameters of arterial remodeling using the mouse carotid artery flow cessation model. Compared with remodeling of matched wild-type (WT) arteries, MMP-9 deficiency decreased intimal hyperplasia, reduced the late lumen loss, eliminated the correlation between intimal hyperplasia and geometric remodeling, and led to significant accumulation of interstitial collagen. Biochemical analysis of MMP-9 knockout (KO) arterial tissue and isolated smooth muscle cells (SMCs) confirmed the lack of MMP-9 expression or compensation by other gelatinases. To investigate potential mechanisms for the in vivo observations, we analyzed in vitro effects of MMP-9 deficiency on the migration, proliferation, and collagen gel contracting capacity of aortic SMCs isolated from MMP-9 KO and WT mice. Although proliferation was comparable, we found that MMP-9-deficient cells had not only decreased migratory activity, but they also had decreased capacity to contract collagen compared with WT cells. Thus, MMP-9 appears to be involved not only in degradation, but also in reorganization of a collagenous matrix, both facets being essential for the outcome of arterial remodeling. Our results also establish MMP-9 as an attractive therapeutic target for limiting the effects of pathological arterial remodeling in restenosis and atherosclerosis.
Neural Correlates of British Sign Language Comprehension: Spatial Processing Demands of Topographic Language
Authors: MacSweeney Mairéad , Woll Bencie , Campbell Ruth , Calvert Gemma A. , McGuire Philip K. , David Anthony S. , Simmons Andrew , Brammer Michael J.
Published: October 2002

In all signed languages used by deaf people, signs are executed in “sign space” in front of the body. Some signed sentences use this space to map detailed “real-world” spatial relationships directly. Such sentences can be considered to exploit sign space “topographically.” Using functional magnetic resonance imaging, we explored the extent to which increasing the topographic processing demands of signed sentences was reflected in the differential recruitment of brain regions in deaf and hearing native signers of the British Sign Language. When BSL signers performed a sentence anomaly judgement task, the occipito-temporal junction was activated bilaterally to a greater extent for topographic than nontopo-graphic processing. The differential role of movement in the processing of the two sentence types may account for this finding. In addition, enhanced activation was observed in the left inferior and superior parietal lobules during processing of topographic BSL sentences. We argue that the left parietal lobe is specifically involved in processing the precise configuration and location of hands in space to represent objects, agents, and actions. Importantly, no differences in these regions were observed when hearing people heard and saw English translations of these sentences. Despite the high degree of similarity in the neural systems underlying signed and spoken languages, exploring the linguistic features which are unique to each of these broadens our understanding of the systems involved in language comprehension.
Testing for Dual Brain Processing Routes in Reading: A Direct Contrast of Chinese Character and Pinyin Reading Using fMRI
Authors: Chen Yiping , Fu Shimin , Iversen Susan D. , Smith Steve M. , Matthews Paul M.
Published: October 2002

Chinese offers a unique tool for testing the effects of word form on language processing during reading. The processes of letter-mediated grapheme-to-phoneme translation and phonemic assembly (assembled phonology) critical for reading and spelling in any alphabetic orthography are largely absent when reading nonalphabetic Chinese characters. In contrast, script-to-sound translation based on the script as a whole (addressed phonology) is absent when reading the Chinese alphabetic sound symbols known as pinyin, for which the script-to-sound translation is based exclusively on assembled phonology. The present study aims to contrast patterns of brain activity associated with the different cognitive mechanisms needed for reading the two scripts. fMRI was used with a block design involving a phonological and lexical task in which subjects were asked to decide whether visually presented, paired Chinese characters or pinyin “sounded like” a word. Results demonstrate that reading Chinese characters and pinyin activate a common brain network including the inferior frontal, middle, and inferior temporal gyri, the inferior and superior parietal lobules, and the extrastriate areas. However, some regions show relatively greater activation for either pinyin or Chinese reading. Reading pinyin led to a greater activation in the inferior parietal cortex bilaterally, the precuneus, and the anterior middle temporal gyrus. In contrast, activation in the left fusiform gyrus, the bilateral cuneus, the posterior middle temporal, the right inferior frontal gyrus, and the bilateral superior frontal gyrus were greater for nonalphabetic Chinese reading. We conclude that both alphabetic and nonalphabetic scripts activate a common brain network for reading. Overall, there are no differences in terms of hemispheric specialization between alphabetic and nonalphabetic scripts. However, differences in language surface form appear to determine relative activation in other regions. Some of these regions (e.g., the inferior parietal cortex for pinyin and fusiform gyrus for Chinese characters) are candidate regions for specialized processes associated with reading via predominantly assembled (pinyin) or addressed (Chinese character) procedures.
A survey of violent and threatening behaviours within an in-patient learning disability unit
Authors: Hakeem Az , Fitzgerald Brian
Published: October 2002

Aims and Method

To analyse violent and threatening behaviour occurring within an in-patient service. We surveyed recorded incidents over a 6-month period.

Results

Ninety-six incidents were recorded. The patients involved were assessed as being aware of their actions. Police were contacted in five cases. No charges were pressed.

Clinical Implications

We believe that violence and threatening behaviours are excessively tolerated in learning disability units. Reasons for this include a staff culture of accepting offending behaviour and an unwillingness or inability to involve the police.
Effect of levothyroxine administration on intestinal P‐glycoprotein expression: Consequences for drug disposition*
Authors: Siegmund Werner , Altmannsberger Stephan , Paneitz Andrea , Hecker Ute , Zschiesche Michael , Franke Gerd , Meng Wieland , Warzok Rolf , Schroeder Eike , Sperker Bernhard , Terhaag Bernd , Cascorbi Ingolf , Kroemer Heyo K.
Published: October 2002

Objective

Thyroid function alters the pharmacokinetics of many drugs; one example is the cardiac glycoside digoxin. Because digoxin disposition is affected by intestinal expression of P‐glycoprotein, we hypothesized that thyroid hormones may regulate P‐glycoprotein and influence disposition of P‐glycoprotein substrates.

Methods

Duodenal expression of P‐glycoprotein measured by reverse transcriptase‐polymerase chain reaction of MDR1 messenger ribonucleic acid (mRNA) and by immunohistochemical examination was studied in 8 healthy volunteers (4 men and 4 women; age range, 22‐29 years; body weight, 59–89 kg) before and after coadministration with levothyroxine (200 μg orally for 17 days), which resulted in suppression of thyroid‐stimulating hormone. The pharmacokinetics of the P‐glycoprotein substrate talinolol was assessed after intravenous (30 mg) and oral (100 mg) administration.

Results

Duodenal MDR1 mRNA expression and immunoreactive P‐glycoprotein were increased 1.4‐fold (not significant; P = .078) and 3.8‐fold (P < .01), respectively, after administration of levothyroxine. The changes in P‐glycoprotein expression were associated with minor alterations in talinolol half‐life after both oral and intravenous administration.

Conclusions

Expression of intestinal P‐glycoprotein in humans appears to be influenced by thyroid hormones. The functional consequences need to be addressed in patients with hyperthyroidism.

Clinical Pharmacology & Therapeutics (2002) 72, 256–264; doi: 10.1067/mcp.2002.126706
Increased genotype frequency of N‐acetyltransferase 2 slow acetylation in patients with rheumatoid arthritis
Authors: Pawlik Andrzej , Ostanek Lidia , Brzosko Iwona , Gawroska‐Szklarz Barbara , Brzosko Marek , Dąbrowska‐Żamojcin Ewa
Published: October 2002

Objectives

Acetylation polymorphism can alter therapeutic responses and toxicity to certain xenobiotics and may also be a factor that influences a patient's susceptibility to certain diseases. We investigated whether patients with rheumatoid arthritis (RA) differed from healthy individuals with regard to genotype of the polymorphic enzyme N‐acetyltransferase 2 (NAT2).

Methods

NAT2 polymorphism was compared in 118 healthy subjects and 82 patients with RA. NAT2 alleles (*4, *5, *6, and *7) were determined by polymerase chain reaction‐restriction fragment length polymorphism methods with deoxyribonucleic acid extracted from peripheral blood.

Results

A statistically significant increase in the proportion of homozygous slow acetylators with 2 mutated alleles (84.1%) was found among patients with RA in comparison with healthy subjects (52.5%; P < .0001). The risk of development of RA was almost 5‐fold greater in slow acetylators than in fast acetylators (odds ratio, 4.79; 95% confidence interval, 2.28–10.21). There was no correlation between NAT2 polymorphism and presence of rheumatoid factor, extra‐articular manifestations, and age at first occurrence of disease symptoms.

Conclusions

NAT2 slow acetylation genotype may be a risk factor of individual susceptibility to RA.

Clinical Pharmacology & Therapeutics (2002) 72, 319–325; doi: 10.1067/mcp.2002.126740
Atovaquone plus chloroguanide versus mefloquine for malaria prophylaxis: A focus on neuropsychiatric adverse events*
Authors: van Riemsdijk M. M. , Sturkenboom M. C. J. M. , Ditters J. M. , Ligthelm R. J. , Overbosch D. , Stricker B. H. Ch.
Published: October 2002

Objectives

We performed a prospective, double‐blind, randomized study to compare the occurrence of neuropsychiatric adverse events and concentration impairment during prophylactic use of either mefloquine or atovaquone plus chloroguanide (INN, proguanil).

Methods

Our potential study population consisted of all persons who were included in the MAL30010 trial at the Travel Clinic, Rotterdam, The Netherlands. All subjects were randomized to receive either active atovaquone (250 mg) plus chloroguanide (100 mg) daily plus a placebo for mefloquine weekly or active mefloquine (250 mg) weekly plus a placebo for atovaquone plus chloroguanide daily. Each subject was followed up from a baseline screening visit up to the index date, 7 days after he or she left the malaria‐endemic area. We measured the interindividual and intraindividual changes in mood disturbance by means of the Dutch shortened Profile of Mood States and 3 domains of the Neurobehavioral Evaluation System, which included sustained attention, coding speed, and visuomotor accuracy between baseline and follow‐up visit.

Results

The cohort consisted of 119 subjects with a mean age of 35 years. A significant deterioration in depression, anger, fatigue, vigor, and total mood disturbance domains occurred during use of mefloquine but not during use of atovaquone plus chloroguanide. Stratification for sex showed between‐treatment differences in female patients but not in male patients. In both treatment groups, sustained attention deteriorated after travel, especially with increased duration of stay.

Conclusions

Prophylactic use of mefloquine was associated with significantly higher scores on scales for depression, anger, and fatigue and lower scores for vigor than prophylactic use of atovaquone plus chloroguanide.

Clinical Pharmacology & Therapeutics (2002) 72, 294–301; doi: 10.1067/mcp.2002.127113
The pharmacokinetics and pharmacodynamics of enoxaparin in obese volunteers
Authors: Sanderink Ger‐Jan , Le Liboux Aimé , Jariwala Navin , Harding Neasa , Ozoux Marie‐Laure , Shukla Umesh , Montay Guy , Boutouyrie Bruno , Miro Adelaida
Published: October 2002

Objectives

The objective of this study was to compare the pharmacokinetics of the low‐molecular‐weight heparin enoxaparin in obese and nonobese volunteers, by means of two administration regimens.

Methods

Enoxaparin was administered subcutaneously (1.5 mg/kg once daily for 4 days) and in a single 6‐hour infusion (1.5 mg/kg) to 24 obese volunteers and 24 age‐, sex‐, and height‐matched nonobese volunteers in a randomized, open‐label, 2‐way crossover design. Blood plasma was assessed for anti‐Xa and anti‐IIa activity and activated partial thromboplastin time.

Results

After subcutaneous administration, steady‐state exposure was achieved after the second dose in nonobese volunteers and after the third dose in obese volunteers. Time to maximum anti‐Xa activity was 1 hour longer in obese volunteers, but maximum anti‐Xa activity was similar in both groups. For anti‐Xa activity, exposure at steady‐state was 16% higher in obese volunteers than in nonobese volunteers (90% confidence interval, 108%‐125%). After intravenous infusion, total body clearance and volume of distribution at steady state were higher in obese volunteers than in nonobese volunteers, but when adjusted for weight, these values were about 10% lower in obese volunteers. Anti‐IIa activity after subcutaneous administration did not differ significantly between obese and nonobese volunteers. Pharmacodynamic analysis of activated partial thromboplastin time showed similar results in obese and nonobese volunteers after both intravenous and subcutaneous administration. No deaths or serious adverse events occurred during the study.

Conclusions

Enoxaparin was well tolerated when administered subcutaneously or intravenously, and there appears to be no need to modify the currently recommended dose for obese volunteers.

Clinical Pharmacology & Therapeutics (2002) 72, 308–318; doi: 10.1067/mcp.2002.127114
Pharmacokinetics of diltiazem and its metabolites in relation to CYP2D6 genotype
Authors: Molden Espen , Johansen Per Wiik , Bøe Grethe Helen , Bergan Stein , Christensen Hege , Rugstad Hans Erik , Rootwelt Helge , Reubsaet Leon , Lehne Gustav
Published: October 2002

Objectives

Recently, it was shown in vitro that the polymorphic enzyme cytochrome P450 (CYP) 2D6 mediates O‐demethylation of diltiazem. The aim of this study was to compare the pharmacokinetics of diltiazem and its major metabolites in healthy human volunteers representing different CYP2D6 genotypes.

Methods

Norwegians of Caucasian origin were screened for their CYP2D6 genotype on the LightCycler (Roche Diagnostics, Mannheim, Germany) by melting‐curve analysis of allele‐specific fluorescence resonance energy transfer probes hybridized to polymerase chain reaction‐amplified deoxyribonucleic acid. The first 5 individuals identified with genotypes corresponding to a homozygous extensive, heterozygous extensive, or homozygous poor CYP2D6‐metabolizing phenotype, respectively, were voluntarily enrolled in the pharmacokinetic study. The participants received diltiazem, 120 mg, as a single oral dose, and plasma samples were collected up to 24 hours after administration. Plasma samples were purified by solid phase extraction. Diltiazem and 7 phase I metabolites were analyzed by liquid chromatography‐mass spectrometry.

Results

The pharmacokinetics of diltiazem was not significantly different between the subgroups. However, the systemic exposure of the pharmacologically active metabolites desacetyl diltiazem and N‐demethyldesacetyl diltiazem was ≥5 times higher in poor CYP2D6 metabolizers than in extensive CYP2D6 metabolizers (P < .01).

Conclusions

CYP2D6 activity does not have a major impact on the disposition of diltiazem. In contrast, desacetyl diltiazem and N‐demethyldesacetyl diltiazem are markedly accumulated in individuals expressing a deficient CYP2D6 phenotype. Because these metabolites exhibit pharmacologic properties of possible importance, individual CYP2D6 activity might be an aspect to consider in the clinical use of diltiazem.

Clinical Pharmacology & Therapeutics (2002) 72, 333–342; doi: 10.1067/mcp.2002.127396
Cytochrome P450 2E1 and 3A activities do not differ between Mexicans and European Americans*
Authors: Poland Russell A. , Lin Keh‐Ming , Nuccio Cenci , Wilkinson Grant R.
Published: October 2002

Objectives

Population differences in the activity of various cytochrome P450 (CYP) enzymes have been demonstrated on the basis of either genetic or environmental determinants. Hispanics are a large demographic group both worldwide and within the United States; hence the possibility of differences in metabolism between one such group—Mexicans—and a European‐derived population was determined with respect to CYP2E1 and CYP3A.

Methods

Young healthy Mexican immigrants living in Los Angeles, Calif, who had maintained a traditional diet were compared with previously and identically studied groups of age‐, sex‐, and weight‐matched European Americans who resided in middle Tennessee and ate a “western” diet (15 men and 15 women). In one study carried out in 15 women, the disposition of chlorzoxazone after an oral dose (250 mg) was compared. In the other investigation, all of the 15 subjects were men and received intravenous [15N3]‐labeled midazolam (1 mg) and oral midazolam (2 mg) simultaneously to characterize the disposition of benzodiazepine in the two populations.

Results

Plasma concentration‐time profiles of chlorzoxazone and its 6‐hydroxy metabolite and the 0‐ to 24‐hour urinary recovery of the latter were not different between Mexicans and European Americans. This indicates that CYP2E1 activity is similar in the two populations. Similarly, no significant intergroup differences were noted in the plasma concentration‐time profiles of midazolam after either intravenous or oral administration. Accordingly, CYP3A does not appear to be different between Mexicans and European Americans.

Conclusions

Similarity in the metabolism of chlorzoxazone between Mexicans and European Americans suggests that the risk associated with CYP2E1‐mediated activation of procarcinogens is not different between these two populations. Likewise, the absence of any difference in the disposition of midazolam indicates that, from a pharmacokinetic standpoint, dosages of drugs metabolized by CYP3A need not be different between Mexicans and European Americans.

Clinical Pharmacology & Therapeutics (2002) 72, 288–293; doi: 10.1067/mcp.2002.127398
Glyburide and glimepiride pharmacokinetics in subjects with different CYP2C9 genotypes
Authors: Niemi Mikko , Cascorbi Ingolf , Timm Ramona , Kroemer Heyo K. , Neuvonen Pertti J. , Kivistö Kari T.
Published: October 2002

Objectives

Our objective was to investigate the effects of genetic polymorphisms of cytochrome P450 (CYP) 2C9 on the pharmacokinetics of glyburide (INN, glibenclamide) and glimepiride, two widely used sulfonylurea antidiabetic drugs.

Methods

We conducted CYP2C9 genotyping for 29 healthy volunteers who had participated in our previous pharmacokinetic studies on glyburide or glimepiride.

Results

There were 17 subjects (59%) with the CYP2C9*1/*1 (wild‐type) genotype, 8 (28%) with the CYP2C9*1/*2 genotype, 3 (10%) with the CYP2C9*1/*3 genotype, and 1 (3%) with the CYP2C9*2/*3 genotype. The pharmacokinetics of glyburide or glimepiride were not significantly changed among subjects with the CYP2C9*1/*2 genotype. However, in individuals heterozygous for the CYP2C9*3 allele, the median total area under the plasma concentration‐time curve of glyburide (n = 2) was 280% (P ≤ .05) and that of glimepiride (n = 3) was 267% (P ≤ .01) of the respective values in subjects with the CYP2C9*1/*1 genotype (n = 5 and n = 12, respectively). Blood glucose responses to glyburide and glimepiride were not significantly affected by the CYP2C9 genotype.

Conclusions

Genetic polymorphisms of CYP2C9 markedly affect the pharmacokinetics of both glyburide and glimepiride. The influence of the CYP2C9*3 variant allele on glyburide and glimepiride pharmacokinetics may be clinically significant.

Clinical Pharmacology & Therapeutics (2002) 72, 326–332; doi: 10.1067/mcp.2002.127495
Evaluation of potential losartan‐phenytoin drug interactions in healthy volunteers*
Authors: Fischer Tracy L. , Pieper John A. , Graff Donald W. , Rodgers Jo E. , Fischer Jeffrey D. , Parnell Kimberly J. , Goldstein Joyce A. , Greenwood Robert , Patterson J. Herbert
Published: October 2002

Background

Phenytoin, a cytochrome P450 (CYP) 2C9 substrate, has a narrow therapeutic index and nonlinear pharmacokinetics. Therefore there is the potential for significant concentration‐related adverse effects when phenytoin is coadministered with other CYP2C9 substrates. Losartan, an antihypertensive agent, is also a substrate for CYP2C9.

Objective

Our objective was to assess the effects of losartan on the pharmacokinetics of phenytoin and the effects of phenytoin on the pharmacokinetics of losartan in a healthy population of volunteers.

Methods

A prospective, randomized, 3‐period crossover study was conducted in 16 healthy volunteers with phenytoin alone, phenytoin in combination with losartan, and losartan alone. Each treatment was given for 10 days with a 3‐week washout period between treatments. On day 10, plasma concentrations of phenytoin and plasma and urine concentrations of losartan and its active carboxylic‐acid metabolite E3174 were measured to determine steady‐state pharmacokinetic parameters.

Results

Coadministration of losartan had no effect on the pharmacokinetics of phenytoin. Coadministration of phenytoin increased the mean area under the concentration‐time curve from time zero to 24 hours [AUC(0–24)] of losartan by 17% (355 ± 220 ng · h/mL versus 427 ± 177 ng · h/mL; P = .1), but this difference was not statistically significant. In the 14 CYP2C9*1/*1 subjects, the mean AUC(0–24) of losartan was increased by 29% (284 ± 84 ng · h/mL versus 402 ± 128 ng · h/mL; P = .008). Coadministration of phenytoin significantly reduced the AUC(0–24) of E3174 by 63% (1254 ± 256 ng · h/mL versus 466 ± 174 ng · h/mL; P = .0001) and the formation clearance of losartan to E3174 (1.91 ± 0.8 mL/h per kilogram versus 0.62 ± 0.4 mL/h per kilogram; P = .0001).

Conclusions

Losartan, a CYP2C9 substrate, had no effect on the pharmacokinetics of phenytoin. However, phenytoin inhibited the CYP2C9‐mediated conversion of losartan to E3174.

Clinical Pharmacology & Therapeutics (2002) 72, 238–246; doi: 10.1067/mcp.2002.127945
Nucleic Acid-Independent Retrovirus Assembly Can Be Driven by Dimerization
Authors: Johnson Marc C. , Scobie Heather M. , Ma Yu May , Vogt Volker M.
Published: October 2002

The Gag protein of retroviruses alone can polymerize into regular virus-like particles (VLPs) both in vitro and in vivo. In most circumstances the capsid (CA) and nucleocapsid (NC) domains of Gag as well as some form of nucleic acid are required for this process. The mechanism by which NC-nucleic acid interaction promotes assembly has remained obscure. We show here that while deletion of the NC domain of Rous sarcoma virus Gag abolishes formation and budding of VLPs at the plasma membranes of baculovirus-infected insect cells
,
replacement of NC with a dimer-forming leucine zipper domain restores budding of spherical particles morphologically similar to wild-type VLPs. The positioning of the dimerization domain appears to be critical for proper assembly, as the insertion of a 5-amino-acid flexible linker upstream of the zipper domain leads to budding of tubular rather than spherical particles. Similar tubular particles are formed when the same linker is inserted upstream of NC. The tubes are morphologically distinct from tubes formed when the p10 domain upstream of CA is deleted. The fact that a foreign dimerization domain can functionally mimic NC suggests that the role of nucleic acid in retroviral assembly is not to serve as a scaffold but rather to promote the formation of Gag dimers, which are critical intermediates in the polymerization of the Gag shell.
Disruption of Epstein-Barr Virus Latency in the Absence of Phosphorylation of ZEBRA by Protein Kinase C
Authors: El-Guindy Ayman S. , Heston Lee , Endo Yoshimi , Cho Myung-Sam , Miller George
Published: October 2002

ZEBRA protein converts Epstein-Barr virus (EBV) infection from the latent to the lytic state. The ability of ZEBRA to activate this switch is strictly dependent on the presence of serine or threonine at residue 186 of the protein (A. Francis, T. Ragoczy, L. Gradoville, A. El-Guindy, and G. Miller, J. Virol. 72:4543-4551, 1999). We investigated whether phosphorylation of ZEBRA protein at this site by a serine-threonine protein kinase was required for activation of an early lytic cycle viral gene, BMRF1, as a marker of disruption of latency. Previous studies suggested that phosphorylation of ZEBRA at S186 by protein kinase C (PKC) activated the protein (M. Baumann, H. Mischak, S. Dammeier, W. Kolch, O. Gires, D. Pich, R. Zeidler, H. J. Delecluse, and W. Hammerschmidt, J. Virol 72:8105-8114, 1998). Two residues of ZEBRA, T159 and S186, which fit the consensus for phosphorylation by PKC, were phosphorylated in vitro by this enzyme. Several isoforms of PKC (α, β1, β2, γ, δ, and ε) phosphorylated ZEBRA. All isoforms that phosphorylated ZEBRA in vitro were blocked by bisindolylmaleimide I, a specific inhibitor of PKC. Studies in cell culture showed that phosphorylation of T159 was not required for disruption of latency in vivo, since the T159A mutant was fully functional. Moreover, the PKC inhibitor did not block the ability of ZEBRA expressed from a transfected plasmid to activate the BMRF1 downstream gene. Of greatest importance, in vivo labeling with [32P]orthophosphate showed that the tryptic phosphopeptide maps of wild-type ZEBRA, Z(S186A), and the double mutant Z(T159A/S186A) were identical. Although ZEBRA is a potential target for PKC, in the absence of PKC agonists, ZEBRA is not constitutively phosphorylated in vivo by PKC at T159 or S186. Phosphorylation of ZEBRA by PKC is not essential for the protein to disrupt EBV latency.
A Transactivator Function of Cottontail Rabbit Papillomavirus E2 Is Essential for Tumor Induction in Rabbits
Authors: Jeckel Sonja , Huber Evamaria , Stubenrauch Frank , Iftner Thomas
Published: October 2002

Infection of domestic rabbits with cottontail rabbit papillomavirus (CRPV) causes local papillomas which progress to carcinomas in more than 80% of cases. This animal model system therefore allows the identification of molecular mechanisms required for the induction and progression of epithelial tumors. The viral E2 protein stimulates both viral DNA replication and transcription, and these functions can be genetically separated. We introduced the respective mutations into CRPV E2 and found, in line with published data for other papillomavirus E2 proteins, that mutation of the highly conserved amino acid 37 or 73 resulted in replication-competent but transactivation-deficient E2 proteins, whereas E2 proteins with mutations at residue 39 were replication deficient and transactivation competent. The R37A, I73L, and I73A E2 mutants, showing a loss of transactivation function, and the R37K E2 mutant, which is still transactivation competent, were introduced into the whole genome of CRPV, which was then injected into the skin of rabbits. Strikingly, the ability to induce tumors within 6 weeks was abolished by each of the E2 mutations, in contrast to the tumor induction rate (93%) obtained with wild-type CRPV DNA. Two small papillomas induced by mutant E2 I73A CRPV DNA appeared as late as 12 or 24 weeks postinjection, were significantly smaller, and showed no further extension of growth. These data suggest that functionally conserved amino acids in the transactivation domain of E2 are also required for the induction and growth of epithelial tumors in rabbits infected with CRPV.
Mandelic Acid Condensation Polymer: Novel Candidate Microbicide for Prevention of Human Immunodeficiency Virus and Herpes Simplex Virus Entry
Authors: Herold B. C. , Scordi-Bello I. , Cheshenko N. , Marcellino D. , Dzuzelewski M. , Francois F. , Morin R. , Casullo V. Mas , Anderson R. A. , Chany C. , Waller D. P. , Zaneveld L. J. D. , Klotman M. E.
Published: October 2002

Presently marketed vaginal barrier methods are cytotoxic and damaging to the vaginal epithelium and natural vaginal flora when used frequently. Novel noncytotoxic agents are needed to protect men and women from sexually transmitted diseases. One novel candidate is a mandelic acid condensation polymer, designated SAMMA. The spectrum and mechanism of antiviral activity were explored using clinical isolates and laboratory-adapted strains of human immunodeficiency virus (HIV) and herpes simplex virus (HSV). SAMMA is highly effective against all CCR5 and CXCR4 isolates of HIV in primary human macrophages and peripheral blood mononuclear cells. SAMMA also inhibits infection of cervical epithelial cells by HSV. Moreover, it exhibits little or no cytotoxicity and has an excellent selectivity index. SAMMA, although not a sulfonated or sulfated polymer, blocks the binding of HIV and HSV to cells by targeting the envelope glycoproteins gp120 and gB-2, respectively, and also inhibits HSV entry postattachment. SAMMA is an excellent, structurally novel candidate microbicide that warrants further preclinical evaluation.
Roles of Nonstructural Protein nsP2 and Alpha/Beta Interferons in Determining the Outcome of Sindbis Virus Infection
Authors: Frolova Elena I. , Fayzulin Rafik Z. , Cook Susan H. , Griffin Diane E. , Rice Charles M. , Frolov Ilya
Published: October 2002

Alphaviruses productively infect a variety of vertebrate and insect cell lines. In vertebrate cells, Sindbis virus redirects cellular processes to meet the needs of virus propagation. At the same time, cells respond to virus replication by downregulating virus growth and preventing dissemination of the infection. The balance between these two mechanisms determines the outcome of infection at the cellular and organismal levels. In this report, we demonstrate that a viral nonstructural protein, nsP2, is a significant regulator of Sindbis virus-host cell interactions. This protein not only is a component of the replicative enzyme complex required for replication and transcription of viral RNAs but also plays a role in suppressing the antiviral response in Sindbis virus-infected cells. nsP2 most likely acts by decreasing interferon (IFN) production and minimizing virus visibility. Infection of murine cells with Sindbis virus expressing a mutant nsP2 leads to higher levels of IFN secretion and the activation of 170 cellular genes that are induced by IFN and/or virus replication. Secreted IFN protects naive cells against Sindbis virus infection and also stops viral replication in productively infected cells. Mutations in nsP2 can also attenuate Sindbis virus cytopathogenicity. Such mutants can persist in mammalian cells with defects in the alpha/beta IFN (IFN-α/β) system or when IFN activity is neutralized by anti-IFN-α/β antibodies. These findings provide new insight into the alphavirus-host cell interaction and have implications for the development of improved alphavirus expression systems with better antigen-presenting potential.
Sialic Acid Functions in Enterovirus 70 Binding and Infection
Authors: Alexander David A. , Dimock Kenneth
Published: October 2002

The interaction of viruses with host cell receptors is the initial step in viral infection and is an important determinant of virus host range, tissue tropism, and pathogenesis. The complement regulatory protein decay-accelerating factor (DAF/CD55) is an attachment receptor for enterovirus 70 (EV70), a member of thePicornaviridae, commonly associated with an eye infection in humans known as acute hemorrhagic conjunctivitis. In early work, the EV70 receptor on erythrocytes, responsible for its hemagglutinating activity, was shown to be sensitive to neuraminidase, implying an essential role for sialic acid in virus attachment. Here, we extend these results to show that cell surface sialic acid is required for EV70 binding to nucleated cells susceptible to virus infection and that sialic acid binding is important in productive infection. Through the use of site-directed mutagenesis to eliminate the single N-linked glycosylation site of DAF and of a chimeric receptor protein in which the O-glycosylated domain of DAF was replaced by a region of the HLA-B44 molecule, a role in EV70 binding for the sialic acid residues of DAF was excluded, suggesting the existence of at least one additional, sialylated EV70-binding factor at the cell surface. Treatment of cells with metabolic inhibitors of glycosylation excluded a role for the N-linked oligosaccharides of glycoproteins but suggested that O-linked glycosylation is important for EV70 binding.
Infectious Bursal Disease Virus Capsid Protein VP3 Interacts both with VP1, the RNA-Dependent RNA Polymerase, and with Viral Double-Stranded RNA
Authors: Tacken Mirriam G. J. , Peeters Ben P. H. , Thomas Adri A. M. , Rottier Peter J. M. , Boot Hein J.
Published: October 2002

Infectious bursal disease virus
(IBDV) is a double-stranded RNA (dsRNA) virus of the
Birnaviridae
family. Its two genome segments are encapsidated together with multiple copies of the viral RNA-dependent RNA polymerase, VP1, in a single-shell capsid that is composed of VP2 and VP3. In this study we identified the domains responsible for the interaction between VP3 and VP1. Using the yeast two-hybrid system we found that VP1 binds to VP3 through an internal domain, while VP3 interacts with VP1 solely by its carboxy-terminal 10 amino acids. These results were confirmed by using a reverse-genetics system that allowed us to analyze the interaction of carboxy-terminally truncated VP3 molecules with VP1 in infected cells. Coimmunoprecipitations with VP1- and VP3-specific antibodies revealed that the interaction is extremely sensitive to truncation of VP3. The mere deletion of the C-terminal residue reduced coprecipitation almost completely and also fully abolished production of infectious virions. Surprisingly, these experiments additionally revealed that VP3 also binds to RNA. RNase treatments and reverse transcription-PCR analyses of the immunoprecipitates demonstrated that VP3 interacts with dsRNA of both viral genome segments. This interaction is not mediated by the carboxy-terminal domain of VP3 since C-terminal truncations of 1, 5, or 10 residues did not prevent formation of the VP3-dsRNA complexes. VP3 seems to be the key organizer of birnavirus structure, as it maintains critical interactions with all components of the viral particle: itself, VP2, VP1, and the two genomic dsRNAs.
Role of the Alfalfa Mosaic Virus Methyltransferase-Like Domain in Negative-Strand RNA Synthesis
Authors: Vlot A. Corina , Menard Aymeric , Bol John F.
Published: October 2002

RNAs 1 and 2 of the tripartite genome of alfalfa mosaic virus (AMV) encode the replicase proteins P1 and P2, respectively. P1 contains a methyltransferase-like domain in its N-terminal half, which has a putative role in capping the viral RNAs. Six residues in this domain that are highly conserved in the methyltransferase domains of alphavirus-like viruses were mutated individually in AMV P1. None of the mutants was infectious to plants. Mutant RNA 1 was coexpressed with wild-type (wt) RNAs 2 and 3 from transferred DNA vectors in
Nicotiana benthamiana
by agroinfiltration. Mutation of His-100 or Cys-189 in P1 reduced accumulation of negative- and positive-strand RNA in the infiltrated leaves to virtually undetectable levels. Mutation of Asp-154, Arg-157, Cys-182, or Tyr-266 in P1 reduced negative-strand RNA accumulation to levels ranging from 2 to 38% of those for the wt control, whereas positive-strand RNA accumulation by these mutants was 2% or less. The (transiently) expressed replicases of the six mutants were purified from the agroinfiltrated leaves. Polymerase activities of these preparations in vitro ranged from undetectable to wt levels. The data indicate that, in addition to its putative role in RNA capping, the methyltransferase-like domain of P1 has distinct roles in replication-associated functions required for negative-strand RNA synthesis. The defect in negative-strand RNA synthesis of the His-100 and Cys-189 mutants could be complemented in
trans
by coexpression of wt P1.
Adenovirus RIDβ Subunit Contains a Tyrosine Residue That Is Critical for RID-Mediated Receptor Internalization and Inhibition of Fas- and TRAIL-Induced Apoptosis
Authors: Lichtenstein Drew L. , Krajcsi Peter , Esteban David J. , Tollefson Ann E. , Wold William S. M.
Published: October 2002

The adenovirus-encoded receptor internalization and degradation (RID) protein (previously named E3-10.4K/14.5K), which is composed of RIDα and RIDβ subunits, down-regulates a number of cell surface receptors in the tumor necrosis factor (TNF) receptor superfamily, namely Fas, TRAIL receptor 1, and TRAIL receptor 2. Down-regulation of these “death” receptors protects adenovirus-infected cells from apoptosis induced by the death receptor ligands Fas ligand and TRAIL. RID also down-regulates certain tyrosine kinase cell surface receptors, especially the epidermal growth factor receptor (EGFR). RID-mediated Fas and EGFR down-regulation occurs via endocytosis of the receptors into endosomes followed by transport to and degradation within lysosomes. However, the molecular interactions underlying this function of RID are unknown. To investigate the molecular determinants of RIDβ that are involved in receptor down-regulation, mutations within the cytoplasmic tail of RIDβ were constructed and the mutant proteins were analyzed for their capacity to internalize and degrade Fas and EGFR and to protect cells from death receptor ligand-induced apoptosis. The results demonstrated the critical nature of a tyrosine residue near the RIDβ C terminus; mutation of this residue to alanine abolished RID function. Mutating the tyrosine to phenylalanine did not abolish the function of RID, arguing that phosphorylation of the tyrosine is not required for function. These data suggest that this tyrosine residue forms part of a tyrosine-based sorting signal (Yxxφ). Additional mutations that target another potential sorting motif and several possible protein-protein interaction motifs had no discernible effect on RID function. It was also demonstrated that mutation of serine 116 to alanine eliminated phosphorylation of RIDβ but did not affect any of the functions of RID that were examined. These results suggest a model in which the tyrosine-based sorting signal in RID plays a role in RID's ability to down-regulate receptors.
A Limited Number of Transducible Hepatocytes Restricts a Wide-Range Linear Vector Dose Response in Recombinant Adeno-Associated Virus-Mediated Liver Transduction
Authors: Nakai Hiroyuki , Thomas Clare E. , Storm Theresa A. , Fuess Sally , Powell Sharon , Wright J. Fraser , Kay Mark A.
Published: October 2002

Recombinant adeno-associated virus (rAAV) vectors are promising vehicles for achieving stable liver transduction in vivo. However, the mechanisms of liver transduction are not fully understood, and furthermore, the relationships between rAAV dose and levels of transgene expression, total number of hepatocytes transduced, and proportion of integrated vector genomes have not been well established. To begin to elucidate the liver transduction dose response with rAAV vectors, we injected mice with two different human factor IX or
Escherichia coli lacZ
-expressing AAV serotype 2-based vectors at doses ranging between 4.0 × 10
8
and 1.1 × 10
13
vector genomes (vg)/mouse, in three- to sixfold increments. A 2-log-range linear dose-response curve of transgene expression was obtained from 3.7 × 10
9
to 3.0 × 10
11
vg/mouse. Vector doses above 3.0 × 10
11
vg/mouse resulted in disproportionately smaller increases in both the number of transduced hepatocytes and levels of transgene expression, followed by saturation at doses above 1.8 × 10
12
vg/mouse. In contrast, a linear increase in the number of vector genomes per hepatocyte was observed up to 1.8 × 10
12
vg/mouse concomitantly with enhanced vector genome concatemerization, while the proportion of integrated vector genomes was independent of the vector dose. Thus, the mechanisms that restrict a wide-range linear dose response at high doses likely involve decreased functionality of vector genomes and restriction of transduction to fewer than 10% of total hepatocytes. Such information may be useful to determine appropriate vector doses for in vivo administration and provides further insights into the mechanisms of rAAV transduction in the liver.
Regulation of Bovine Papillomavirus Replicative Helicase E1 by the Ubiquitin-Proteasome Pathway
Authors: Malcles Marie-Helene , Cueille Nathalie , Mechali Francisca , Coux Olivier , Bonne-Andrea Catherine
Published: October 2002

Papillomaviruses maintain their genomes in a relatively constant copy number as stable extrachromosomal plasmids in the nuclei of dividing host cells. The viral initiator of replication, E1, is not detected in papillomavirus-infected cells. Here, we present evidence that E1 encoded by bovine papillomavirus type 1 is an unstable protein that is degraded through the ubiquitin-proteasome pathway. In a cell-free system derived from
Xenopus
egg extracts, E1 degradation is regulated by both cyclin E/Cdk2 binding and E1 replication activity. Free E1 is readily ubiquitinated and degraded by the proteasome, while it becomes resistant to this degradation pathway when bound to cyclin E/Cdk2 complexes before the start of DNA synthesis. This stabilization is reversed in a process involving E1-dependent replication activity. In transiently transfected cells, E1 is also polyubiquitinated and accumulates when proteasome activity is inhibited. Thus, the establishment and maintenance of a stable number of papillomavirus genomes in latently infected cells are in part a function of regulated ubiquitin-mediated degradation of E1.