bluuebunny/crossref_metadata_2025 huggingface dataset:
Putative protective effect of inspiratory threshold loading against exercise-induced supraspinal diaphragm fatigue
Authors: Jonville S. , Jutand L. , Similowski T. , Denjean A. , Delpech N.
Published: October 2004

The present investigation was intended to assess the consequences of an inspiratory load on the diaphragm central component of fatigue during exercise. We recorded the motor potential evoked (MEP) by transcranial magnetic stimulation of the motor cortex in 10 subjects. The diaphragm and rectus femoris were studied before and 10, 20, and 40 min after two 16-min cycling exercise (E) trials requiring 55% of maximal oxygen uptake: 1) one with an inspiratory threshold load (E + ITL), corresponding to 10% of maximal inspiratory pressure; and 2) the other without the load (E). Dyspnea, heart rate, electromyographic activity of the sternocleidomastoid, and diaphragm work were significantly higher in E + ITL than in E. Neither trial affected the response to phrenic magnetic stimulation, which was performed 15 and 25 min postexercise, or the maximal inspiratory pressure (116 and 120 cmH2O before E and E + ITL, respectively, and 110 and 114 cmH2O at 30 min postexercise). Whereas the amplitude of the diaphragm MEP was unaffected by E + ITL (+2.1 ± 29.4%), a significant decrease was observed 10 min after E compared with baseline (−37.1 ± 22.3%) and compared with E + ITL. The MEP amplitude of rectus femoris remained unchanged with E and E + ITL. The recruitment of synergistic agonists during E + ITL may have normalized the major ventilatory stress and reset up the excitability of the diaphragm pathway.
Rapid natriuretic action of aldosterone in the rat
Authors: Rad Abolfazl K. , Balment Richard J. , Ashton Nick
Published: October 2004

Rapid, nongenomic actions of aldosterone have been demonstrated in a number of cell types in vitro, including renal cell lines, but there remains little direct evidence that it is able to exert rapid effects on the kidney in the whole animal. Accordingly, the aim of this study was to determine whether aldosterone induces rapid changes in the renal handling of electrolytes or acid-base balance in the anesthetized rat. With the use of a servo-controlled fluid replacement system, spontaneous urine output by anesthetized male Sprague-Dawley rats was replaced with 2.5% dextrose. After a 3-h equilibration and a 1-h control period, rats were infused with aldosterone (42 pmol/min) or vehicle for 1 h. Aldosterone infusion induced a rapid (within 15 min) increase in sodium excretion that peaked at 0.24 ± 0.08 compared with 0.04 ± 0.01 μmol·min−1 100·body weight−1 ( P = 0.041) in the vehicle-infused rats. This natriuresis was not associated with changes in glomerular filtration rate; urine flow rate; potassium, chloride, or bicarbonate excretion; or urine pH. The mechanisms involved are unclear, but because we have previously shown that aldosterone stimulates a rapid (4 min) increase in cAMP generation in the rat inner medullary collecting duct (IMCD) (Sheader EA, Wargent ET, Ashton N, and Balment RJ. J Endocrinol 175: 343–347, 2002), they could involve cAMP-mediated activation of the cystic fibrosis transmembrane conductance regulator chloride channel, which drives sodium secretion in the IMCD.
Similar Kinematic Properties for Ocular Following and Smooth Pursuit Eye Movements
Authors: Adeyemo Babatunde , Angelaki Dora E.
Published: October 2004

Ocular following (OFR) is a short-latency visual stabilization response to the optic flow experienced during self-motion. It has been proposed that it represents the early component of optokinetic nystagmus (OKN) and that it is functionally linked to the vestibularly driven stabilization reflex during translation (translational vestibuloocular reflex, TVOR). Because no single eye movement can eliminate slip from the whole retina during translation, the OFR and the TVOR appear to be functionally related to maintaining visual acuity on the fovea. Other foveal-specific eye movements, like smooth pursuit and saccades, exhibit an eye-position-dependent torsional component, as dictated by what is known as the “half-angle rule” of Listing's law. In contrast, eye movements that stabilize images on the whole retina, such as the rotational vestibuloocular reflex (RVOR) and steady-state OKN do not. Consistent with the foveal stabilization hypothesis, it was recently shown that the TVOR is indeed characterized by an eye-position-dependent torsion, similar to pursuit eye movements. Here we have investigated whether the OFR exhibits three-dimensional kinematic properties consistent with a foveal response (i.e., similar to the TVOR and smooth pursuit eye movements) or with a whole-field stabilization function (similar to steady-state OKN). The OFR was elicited using 100-ms ramp motion of a full-field random dot pattern that moved horizontally at 20, 62, or 83°/s. To study if an eye-position-dependent torsion is generated during the OFR, we varied the initial fixation position vertically within a range of ±20°. As a control, horizontal smooth pursuit eye movements were also elicited using step-ramp target motion (10, 20, or 30°/s) at similar eccentric positions. We found that the OFR followed kinematic properties similar to those seen in pursuit and the TVOR with the eye-position-dependent torsional tilt of eye velocity having slopes that averaged 0.73 ± 0.16 for OFR and 0.57 ± 0.12 (means ± SD) for pursuit. These findings support the notion that the OFR, like the TVOR and pursuit, are foveal image stabilization systems.
Rapid evolution and diversification of mammalian α-defensins as revealed by comparative analysis of rodent and primate genes
Authors: Patil Amar , Hughes Austin L. , Zhang Guolong
Published: October 2004

Mammalian α-defensins constitute a family of cysteine-rich, cationic antimicrobial peptides produced by phagocytes and intestinal Paneth cells, playing an important role in innate host defense. Following comprehensive computational searches, here we report the discovery of complete repertoires of the α-defensin gene family in the human, chimpanzee, rat, and mouse with new genes identified in each species. The human genome was found to encode a cluster of 10 distinct α-defensin genes and pseudogenes expanding 132 kb continuously on chromosome 8p23. Such α-defensin loci are also conserved in the syntenic chromosomal regions of chimpanzee, rat, and mouse. Phylogenetic analyses showed formation of two distinct clusters with primate α-defensins forming one cluster and rodent enteric α-defensins forming the other cluster. Species-specific clustering of genes is evident in nonprimate species but not in the primates. Phylogenetically distinct subsets of α-defensins also exist in each species, with most subsets containing multiple members. In addition, natural selection appears to have acted to diversify the functionally active mature defensin region but not signal or prosegment sequences. We concluded that mammalian α-defensin genes may have evolved from two separate ancestors originated from β-defensins. The current repertoires of the α-defensin gene family in each species are primarily a result of repeated gene duplication and positive diversifying selection after divergence of mammalian species from each other, except for the primate genes, which were evolved prior to the separation of the primate species. We argue that the presence of multiple, divergent subsets of α-defensins in each species may help animals to better cope with different microbial challenges in the ecological niches which they inhabit.
Low-Grade Inflammation and Microalbuminuria in Hypertension
Authors: Pedrinelli Roberto , Dell’Omo Giulia , Di Bello Vitantonio , Pellegrini Giovanni , Pucci Laura , Del Prato Stefano , Penno Giuseppe
Published: October 2004

Background—

Albuminuria and C-reactive protein (CRP), a marker of systemic low-grade inflammation, are frequently elevated in essential hypertension and predict cardiovascular prognosis independent of conventional risk factors. However, in spite of their potentially important links, the interrelationships between the 2 parameters have not been explored in depth in hypertensive patients.

Methods and Results—

Albuminuria (the mean of 3 overnight urine collections), high-sensitive CRP (hs-CRP), 24-hour blood pressure (BP), weight, lipids, poststimulative (75 g PO) plasma glucose, insulin, and insulin sensitivity by the homeostasis model assessment model were evaluated in 220 never treated, nondiabetic, uncomplicated essential hypertensive men. Albuminuria ≥15 μg/min was defined as microalbuminuria and hs-CRP values above and below median (2.3 mg/L) as high and low, respectively. Concentric left ventricular hypertrophy was diagnosed by echocardiography, and a full-blown metabolic syndrome was identified in presence of hypertension and at least 3 of following: obesity, subclinical hyperglycemia, low high-density lipoprotein (HDL) and high triglycerides. Microalbuminuria was present in 54 patients, 29 with high hs-CRP characterized by higher 24-hour systolic BP, postload glucose, body mass index, lower HDL cholesterol, more frequent metabolic syndrome, concentric LVH, and active smoking than those with either isolated microalbuminuria (n=27) or normoalbuminuria.

Conclusions—

Microalbuminuria accompanied by evidence of subclinical inflammation is a strong correlate of metabolic abnormalities in essential hypertension and identifies a patient subset at very high cardiovascular risk. In contrast, isolated microalbuminuria may represent a distinct pathophysiological condition characterized by a more benign profile and possibly a better prognosis.
Induction of Angiogenesis by Heat Shock Protein 90 Mediated by Protein Kinase Akt and Endothelial Nitric Oxide Synthase
Authors: Sun Jianxin , Liao James K.
Published: October 2004

Objective—

A specific inhibitor of heat shock protein 90 (Hsp90), 17-AAG, has been shown to inhibit tumor growth through cell cycle arrest, differentiation, or apoptosis. Because angiogenesis is important for tumor growth, we hypothesize that inhibition of angiogenesis by 17-AAG may mediate some of its antitumor effects.

Methods and Results—

Because protein kinase Akt and endothelial nitric oxide synthase (eNOS) are critical for angiogenesis, we studied the effects of 17-AAG on the phosphorylation and expression of Akt and eNOS in human umbilical vein endothelial cells. In a concentration- and time-dependent manner, inhibition of Hsp90 by 17-AAG decreased Akt and eNOS expression by 74% and 81%, respectively. Inhibition of eNOS expression by 17-AAG occurred at the transcriptional level as determined by eNOS promoter activity and nuclear run-on assay. Furthermore, treatment with 17-AAG decreased basal and vascular endothelial growth factor-stimulated Akt and eNOS phosphorylation. This corresponded with decreased NO production and inhibition of endothelial cell migration and angiogenesis. The anti-angiogenic effect of 17-AAG was partially reversed by the NO donor, SNAP.

Conclusions—

These findings indicate that Hsp90 is important not only for Akt and eNOS phosphorylation but also for eNOS gene transcription and suggests that Hsp90 may be a novel target for anti-angiogenic therapy.
Cardiovascular Risk in Type 2 Diabetes Is Associated With Variation at the
PPARG
Locus
Authors: Doney Alex S.F. , Fischer Bettina , Leese Graham , Morris Andrew D. , Palmer Colin N.A.
Published: October 2004

Objective—

The Pro12Ala polymorphism of
PPARG
modulates risk of developing type 2 diabetes. The Ala allele has also been associated with a reduced risk of cardiovascular events. We have shown previously that the linked T allele of the C1431T polymorphism influences Ala12-associated diabetes risk and that the 2 polymorphisms have opposing associations with body weight. We therefore investigated the association of these 2 variants with cardiovascular events in people with type 2 diabetes.

Methods and Results—

We performed a cohort study of 2016 individuals and used Cox proportional hazards to analyze risk of myocardial infarction or death by
PPARG
Pro12Ala and C1431T genotypes, adjusting for age, sex, and smoking status. In individuals enrolled <70 years of age, the hazard for a first nonfatal event associated with the Ala12 allele was 0.21 (CI, 0.06 to 0.69;
P
=0.01) and the T1431 allele 9.9 (CI, 1.90 to 51.29;
P
=0.007). These opposing associations remained significant after correction for other conventional risk factors. The T1431 allele was also associated with all-cause mortality.

Conclusions—

This study confirms the association of the Ala12 allele with reduced risk of myocardial infarction in a type 2 diabetic population and demonstrates that the T allele independently associates with an increased risk.
Silent Information Regulator 2α, a Longevity Factor and Class III Histone Deacetylase, Is an Essential Endogenous Apoptosis Inhibitor in Cardiac Myocytes
Authors: Alcendor Ralph R. , Kirshenbaum Lorrie A. , Imai Shin-ichiro , Vatner Stephen F. , Sadoshima Junichi
Published: October 2004

Yeast silent information regulator 2 (Sir2), a nicotinamide adenine dinucleotide–dependent histone deacetylase (HDAC) and founding member of the HDAC class III family, functions in a wide array of cellular processes, including gene silencing, longevity, and DNA damage repair. We examined whether or not the mammalian ortholog Sir2 affects growth and death of cardiac myocytes. Cardiac myocytes express Sir2α predominantly in the nucleus. Neonatal rat cardiac myocytes were treated with 20 mmol/L nicotinamide (NAM), a Sir2 inhibitor, or 50 nmol/L Trichostatin A (TSA), a class I and II HDAC inhibitor. NAM induced a significant increase in nuclear fragmentation (2.2-fold) and cleaved caspase-3, as did sirtinol, a specific Sir2 inhibitor, and expression of dominant-negative Sir2α. TSA also modestly increased cell death (1.5-fold) but without accompanying caspase-3 activation. Although TSA induced a 1.5-fold increase in cardiac myocyte size and protein content, NAM reduced both. In addition, NAM caused acetylation and increases in the transcriptional activity of p53, whereas TSA did not. NAM-induced cardiac myocyte apoptosis was inhibited in the presence of dominant-negative p53, suggesting that Sir2α inhibition causes apoptosis through p53. Overexpression of Sir2α protected cardiac myocytes from apoptosis in response to serum starvation and significantly increased the size of cardiac myocytes. Furthermore, Sir2 expression was increased significantly in hearts from dogs with heart failure induced by rapid pacing superimposed on stable, severe hypertrophy. These results suggest that endogenous Sir2α plays an essential role in mediating cell survival, whereas Sir2α overexpression protects myocytes from apoptosis and causes modest hypertrophy. In contrast, inhibition of endogenous class I and II HDACs primarily causes cardiac myocyte hypertrophy and also induces modest cell death. An increase in Sir2 expression during heart failure suggests that Sir2 may play a cardioprotective role in pathologic hearts in vivo.
Plasma Cholesteryl Esters Provided by Lecithin:Cholesterol Acyltransferase and Acyl-Coenzyme A:Cholesterol Acyltransferase 2 Have Opposite Atherosclerotic Potential
Authors: Lee Richard G. , Kelley Kathryn L. , Sawyer Janet K. , Farese Robert V. , Parks John S. , Rudel Lawrence L.
Published: October 2004

Evidence suggests that ACAT2 is a proatherogenic enzyme that contributes cholesteryl esters (CEs) to apoB-containing lipoproteins, whereas LCAT is an antiatherogenic enzyme that facilitates reverse cholesterol transport by esterifying free cholesterol on HDL particles. We hypothesized that deletion of LCAT and ACAT2 would lead to absence of plasma CEs and reduced atherosclerosis. To test this hypothesis, ACAT2
−/−
LCAT
−/−
LDLr
−/−
, ACAT2
−/−
LDLr
−/−
, and LCAT
−/−
LDLr
−/−
mice were fed a 0.15% cholesterol diet for 20 weeks. In comparison to LDLr
−/−
mice, the total plasma cholesterol (TPC) of ACAT2
−/−
LCAT
−/−
LDLr
−/−
mice was 67% lower because of the complete absence of plasma CEs, leading to 94% less CE accumulation in the aorta. In the LCAT
−/−
LDLr
−/−
mice, TPC and atherosclerosis were significantly higher because of increased accumulations of ACAT2-derived CE. In ACAT2
−/−
LDLr
−/−
mice, again compared with LDLr
−/−
mice, TPC was 19% lower, whereas atherosclerosis was 88% lower. Therefore, the absence of ACAT2 led to a significant reduction in TPC although benefits in reduction of atherosclerosis were much more pronounced. Overall, the data suggest that ACAT2-derived CE is the predominant atherogenic lipid in blood, and that an important goal for prevention of atherosclerosis is to limit ACAT2-derived CE accumulation in lipoproteins.
Transient Ischemic Attacks Are More Than “Ministrokes”
Authors: Daffertshofer Michael , Mielke Orell , Pullwitt Arne , Felsenstein Matthias , Hennerici Michael
Published: October 2004

Background and Purpose—

Transient ischemic attacks (TIAs) are warning signs of stroke. Recently, the hypothesis was raised that TIA bears a significant risk for death and dependence and requires the same complex diagnostic workup as a complete stroke.

Methods—

We prospectively collected pre- and in-hospital procedures, symptoms, outcome, complications, and therapies from a representative sample of all stroke-treating hospitals (n=82) in southwest Germany. Follow-up was attempted 6 months after discharge. End points were death or dependence in activities of daily living (Barthel Index <95, modified Rankin Scale (mRS) of 3 to 6, or institutionalization in a nursing home).

Results—

1380 TIA patients and 3855 stroke patients entered the database. During hospital stay, stroke incidence was 8% for TIA patients and another 5% within the first half-year. Similarly, for ischemic stroke (IS) patients these figures were 7% and 6% (
P
>0.05), respectively. Two percent of TIA patients died in hospital (5% afterward) compared with 9% of stroke patients (10% afterward,
P
<0.001). Seventeen percent TIA compared with 38% IS patients (
P
<0.05) were dependent at follow-up. Whereas an estimated preexisting deficit (mRS >2) was the strongest predictor for death or disability (baseline mRS odds ratio, 4.1; 95% CI, 2.3 to 7.2), admission to a stroke unit was a valid predictor for survival and independence (odds ratio, 0.4; 95% CI, 0.2 to 0.9).

Conclusions—

These data from a large, multicenter, nonselected, observational study underscore the “not so benign” prognosis for TIA patients. There is a relevant individual risk of early stroke, death, or disability in TIA patients. Management and treatment strategies are similar for both TIA and acute stroke.
Treatment of Constipation and Fecal Incontinence in Stroke Patients
Authors: Harari Danielle , Norton Christine , Lockwood Linda , Swift Cameron
Published: October 2004

Background and Purpose—

Despite its high prevalence in stroke survivors, there is little clinical research on bowel dysfunction in this population. This is the first randomized controlled trial to evaluate treatment of constipation and fecal incontinence in stroke survivors.

Methods—

Stroke patients with constipation or fecal incontinence were identified by screening questionnaire (122 community, 24 stroke rehabilitation inpatients) and randomized to intervention or routine care (73 per group). The intervention consisted of a 1-off structured nurse assessment (history and rectal examination), leading to targeted patient/carer education with booklet and provision of diagnostic summary and treatment recommendations (after consultation with geriatrician) to patient’s general practitioner (GP)±ward physician.

Results—

Percentage of bowel movements (BMs) per week graded as “normal” by participants in a prospective 1-week stool diary was significantly higher in intervention versus control patients at 6 months (72% versus 55%;
P
=0.027), as was mean number of BMs per week (5.2 versus 3.6;
P
=0.005). There was no significant reduction in fecal incontinence, although numbers were small. At 12 months, intervention patients were more likely to be modifying their diets (odds ratio [OR], 3.1 [1.2 to 8.0]) and fluid intake (OR, 4.2 [1.4 to 12.2]) to control their bowels and to have visited their GP for their bowel problem (OR, 5.0 [1.4 to 17.5]). GP prescribing of laxatives and suppositories was significantly influenced at 12 months.

Conclusions—

A single clinical/educational nurse intervention in stroke patients effectively improved symptoms of bowel dysfunction up to 6 months later, changed bowel-modifying lifestyle behaviors up to 12 months later, and influenced patient–GP interaction and physician prescribing patterns.
Sexual selection and predator avoidance in an acoustic moth: discriminating females take fewer risks
Authors: Greenfield Michael , Greig Emma
Published: October 2004

Sexual activities in both males and females are expected to incur energetic costs and risk. We tested the expectation that females who exhibit a higher level of discrimination of potential mates' signals incur greater risk than females who are relatively indiscriminate by assaying mate choice and predator evasion in an acoustic moth, Achroia grisella. Female A. grisella evaluate males on the basis of their song, only orienting toward males whose ultrasonic songs are delivered at pulse rates above a lower threshold value. Non-flying females also respond negatively, by ceasing all movement, upon hearing ultrasonic pulses delivered at rates below an upper threshold value; this response is assumed to represent evasion of predatory bats, specifically gleaning species. We found that both the lower and upper pulse-rate thresholds varied considerably among individual females and that the upper pulse-rate threshold for defensive response was always equal to or slightly lower than the lower pulse-rate threshold for a mating response. Thus, females who exhibit heightened discrimination of potential mates' signals experience less, not more, predatory risk. This relationship may reflect the evolutionary origin of acoustic sexual communication in A. grisella as a coöption of the ancestral function of hearing in evasion of predatory bats.
Social play in kaka (Nestor meridionalis) with comparisons to kea (Nestor notabilis)
Authors: Bond Alan , Diamond Judy
Published: October 2004

Social play in the kaka (Nestor meridionalis), a New Zealand parrot, is described and contrasted with that of its closest relative, the kea (Nestor notabilis), in one of the first comparative studies of social play in closely related birds. Most play action patterns were clearly homologous in these two species, though some contrasts in the form of specific play behaviors, such as kicking or biting, could be attributed to morphological differences. Social play in kakas is briefer, more predictable, and less sequentially diverse than that shown by keas. Kaka play also appears to be restricted to fledglings and juveniles, while the behavior is more broadly distributed among age groups in keas. Play initiation behaviors were relatively more frequent in kakas and more tightly intercorrelated in occurrence. A primary grouping of action patterns in kakas consisted of arboreal play, which was rare in keas. The most striking species difference was exhibited in social object play, which is pervasive among keas, but which was not observed in kakas. Although the two species are morphologically similar, they differ strikingly in several aspects of their ecology and social behavior, including the duration of the association between juveniles and adults, the degree of exploratory behavior, and the flexibility of their foraging strategies. The observed species differences in play behavior are discussed in relation to the contrasting life histories in the two species, suggesting that many features of social play may reflect evolutionary responses to particular ontogenetic and ecological constraints.
Oxcarbazepine Pharmacokinetics and Tolerability in Children With Inadequately Controlled Epilepsy
Authors: Rey Elisabeth , Bulteau Christine , Motte Jacques , Tran Agnes , Sturm Yvonne , D'Souza Joseph , Markabi Sabri , Pons Gérard , Dulac Olivier
Published: October 2004

This two‐part, open‐label study evaluated the pharmacokinetics, safety, and tolerability of oxcarbazepine as combination therapy in 112 children 2 to 12 years old with inadequately controlled epilepsy. Part I was a pharmacokinetic study in children stratified by age (2–5 years and 6–12 years) and randomized to receive a single oxcarbazepine dose of 5 mg/kg or 15 mg/kg. Mean specific AUC and t1/2 values of the active metabolite (MHD) were approximately 30% lower in younger children compared with older children, regardless of dose. Part II was a 4‐month safety, tolerability, and pharmacokinetic study in which children received oxcarbazepine doses of 11 to 68 mg/kg/day. The mean specific oxcarbazepine daily dose was 38% higher in younger children compared with older children. Similarly, mean trough plasma MHD concentrations were 34% lower in younger children. Six (5%) children discontinued due to adverse events. Oxcarbazepine was safe and well tolerated. Younger children require higher oxcarbazepine doses because of rapid clearance.
Induction of CYP3A4 by Efavirenz in Primary Human Hepatocytes: Comparison With Rifampin and Phenobarbital
Authors: Hariparsad Niresh , Nallani Srikanth C. , Sane Rucha S. , Buckley Donna J. , Buckley Arthur R. , Desai Pankaj B.
Published: October 2004

The antiretroviral agent efavirenz enhances the systemic clearance of coadministered drugs that are cytochrome P450 (CYP) 3A4 substrates. The mechanism of the apparent increase in CYP3A4 activity by efavirenz and the magnitude of change relative to other known inducers are not known. The authors tested the hypothesis that increased enzymatic activity by efavirenz entails CYP3A4 induction and activation of the human pregnane X receptor (hPXR), a key transcriptional regulator of CYP3A4. Employing primary cultures of human hepatocytes, they compared the CYP3A4 inductive effects of efavirenz (1–10 μM) to rifampin (10 μM) and phenobarbital (2 mM). A cell‐based reporter assay was employed to assess hPXR activation. The authors observed that efavirenz caused a concentration‐dependent CYP3A4 induction and hPXR activation. Based on the CYP3A4 activity assay, the average magnitude of induction by efavirenz (5–10 μM) was approximately 3‐to 4‐fold. In comparison, phenobarbital (2 mM) and rifampin (10 μM) caused a 5‐ and 6‐fold induction, respectively.
Induction of T-cell development from human cord blood hematopoietic stem cells by Delta-like 1 in vitro
Authors: La Motte-Mohs Ross N. , Herer Elaine , Zúñiga-Pflücker Juan Carlos
Published: October 2004

The Notch signaling pathway plays a key role at several stages of T-lymphocyte differentiation. However, it remained unclear whether signals induced by the Notch ligand Delta-like 1 could support full T-cell differentiation from a defined source of human hematopoietic stem cells (HSCs) in vitro. Here, we show that human cord blood–derived HSCs cultured on Delta-like 1–expressing OP9 stromal cells undergo efficient T-cell lineage commitment and sustained T-cell differentiation. A normal stage-specific program of T-cell development was observed, including the generation of CD4 and CD8 αβ–T-cell receptor (TCR)–bearing cells. Induction of T-cell differentiation was dependent on the expression of Delta-like 1 by the OP9 cells. Stimulation of the in vitro–differentiated T cells by TCR engagement induced the expression of T-cell activation markers and costimulatory receptors. These results establish an efficient in vitro coculture system for the generation of T cells from human HSCs, providing a new avenue for the study of early T-cell differentiation and function.
Extracellular nucleotide signaling by P2 receptors inhibits IL-12 and enhances IL-23 expression in human dendritic cells: a novel role for the cAMP pathway
Authors: Schnurr Max , Toy Tracey , Shin Amanda , Wagner Moritz , Cebon Jonathan , Maraskovsky Eugene
Published: October 2004

The interleukin-12 (IL-12) cytokine family plays important roles in the orchestration of innate and adaptive immunity by dendritic cells (DCs). The regulation of IL-12 expression has been thoroughly studied, but little is known about factors governing the expression of IL-23 and IL-27, 2 novel IL-12 family members acting on memory and naive T cells, respectively. We report that the expression of these cytokines by DCs was critically dependent on the mode of activation. DC activation by CD40L predominantly induced IL-12. Ligands of the Toll-like receptor (TLR) 3 and TLR4 induced IL-12 and IL-27, whereas exposure to intact Escherichia coli resulted in high expression of IL-12, IL-27, and IL-23. The nucleotide adenosine triphosphate (ATP) has been shown to inhibit IL-12 production by P2 receptors. We found that ATP also inhibited IL-27 expression but enhanced IL-23 expression. Interestingly, the reciprocal regulation of IL-12/IL-27 and IL-23 by ATP was mediated by 2 distinct P2 receptors and was also induced by prostaglandin E2 by cyclic adenosine monophosphate (cAMP)–elevating EP2/EP4 receptors. As a consequence, DCs were selectively impaired in their ability to induce interferon-γ (IFN-γ) in naive T cells but continued to promote IFN-γ and IL-17 production in memory T cells. These studies identify P2 receptors as promising targets for the design of novel strategies to manipulate specific stages of T-cell responses and to treat IL-12– and IL-23–mediated disorders.
Tumor metastasis but not tumor growth is dependent on Src-mediated vascular permeability
Authors: Criscuoli Michele L. , Nguyen Mai , Eliceiri Brian P.
Published: October 2004

Vascular endothelial growth factor (VEGF)–induced vascular permeability (VP) is a hallmark of tumor growth and metastasis. Previous studies have shown a requirement for Src kinase in VEGF-mediated VP and signaling in blood vessels. In this study, we have examined the effect of Src-mediated reduced VP on tumor growth and metastasis. The growth and spontaneous metastasis of VEGF-expressing tumor cells were determined in Src-knockout (src–/–) or control mice (src+/+ or src+/–). In comparison to control mice, src-null mice had a significant reduction in tumor-induced VP as well as a subsequent reduction in spontaneous metastasis. In contrast, primary tumor weight and vascular density were unchanged between src-null and control mice. Consistent with a role for Src in the extravasation of tumor cells from the circulation, direct intravenous injection of lung carcinoma cells resulted in a more than 2-fold reduction in lung tumor burden in src-null mice compared to control mice. The comparison of the results from the experimental metastasis and the spontaneous metastasis models suggests that there are defects in VP in the primary site of Src-deficient mice and that there may be an essential role for Src and Src-mediated VP in tumor metastasis to the lung.
Induction of tolerance using Fas ligand: a double-edged immunomodulator
Authors: Askenasy Nadir , Yolcu Esma S. , Yaniv Isaac , Shirwan Haval
Published: October 2004

Apoptosis mediated by Fas ligand (FasL) interaction with Fas receptor plays a pivotal regulatory role in immune homeostasis, immune privilege, and self-tolerance. FasL, therefore, has been extensively exploited as an immunomodulatory agent to induce tolerance to both autoimmune and foreign antigens with conflicting results. Difficulties associated with the use of FasL as a tolerogenic factor may arise from (1) its complex posttranslational regulation, (2) the opposing functions of different forms of FasL, (3) different modes of expression, systemic versus localized and transient versus continuous, (4) the level and duration of expression, (5) the sensitivity of target tissues to Fas/FasL-mediated apoptosis and the efficiency of antigen presentation in these tissues, and (6) the types and levels of cytokines, chemokines, and metalloproteinases in the extracellular milieu of the target tissues. Thus, the effective use of FasL as an immunomodulator to achieve durable antigen-specific immune tolerance requires careful consideration of all of these parameters and the design of treatment regimens that maximize tolerogenic efficacy, while minimizing the non-tolerogenic and toxic functions of this molecule. This review summarizes the current status of FasL as a tolerogenic agent, problems associated with its use as an immunomodulator, and new strategies to improve its therapeutic potential.
Dendritic-cell activation by secretory phospholipase A2
Authors: Ramoner Reinhold , Putz Thomas , Gander Hubert , Rahm Andrea , Bartsch Georg , Schaber Claudia , Thurnher Martin
Published: October 2004

Dendritic cells (DCs), also referred to as the sentinels of the immune system, induce and coordinate important functions of immune surveillance. DCs acquire immunity-initiating capacity only after a process of maturation usually induced by ligands that bind to members of the tumor necrosis factor (TNF) or toll-like receptor families. Secretory phospholipase A2 (sPLA2), which hydrolyzes the sn-2 ester bond of glycerophospholipids, regulates a variety of cellular functions including migration of endothelial cells and neurite outgrowth. In the present study we investigated the role of sPLA2 in DC biology. We report that human monocyte-derived DC cultures lack sPLA2 activity but respond to exogenous sPLA2. sPLA2 alone and in cooperation with TNF-α and interleukin 1 β (IL-1β) induced fatty acid release from DC membranes, which was accompanied by upregulation of surface markers and by an increase in the migratory and immunostimulatory capacity of the DCs. Our findings indicate that secreted enzymes such as sPLA2 can contribute to DC maturation and emphasize the role of lipid mediators in the regulation of immune responses. This observation may also have implications for DC-based vaccine development.
Rank Difference Analysis of Microarrays (RDAM), a novel approach to statistical analysis of microarray expression profiling data
Authors: Martin Dietmar E , Demougin Philippe , Hall Michael N , Bellis Michel
Published: October 2004

Background

A key step in the analysis of microarray expression profiling data is the identification of genes that display statistically significant changes in expression signals between two biological conditions.

Results

We describe a new method, Rank Difference Analysis of Microarrays (RDAM), which estimates the total number of truly varying genes and assigns a p-value to each signal variation. Information on a group of differentially expressed genes includes the sensitivity and the false discovery rate. We demonstrate the feasibility and efficiency of our approach by applying it to a large synthetic expression data set and to a biological data set obtained by comparing vegetatively-growing wild type and tor2-mutant yeast strains. In both cases we observed a significant improvement of the power of analysis when our method is compared to another popular nonparametric method.

Conclusions

This study provided a valuable new statistical method to analyze microarray data. We conclude that the good quality of the results obtained by RDAM is mainly due to the quasi-perfect equalization of variation distribution, which is related to the standardization procedure used and to the measurement of variation by rank difference.
Protein kinases of the human malaria parasite Plasmodium falciparum: the kinome of a divergent eukaryote
Authors: Ward Pauline , Equinet Leila , Packer Jeremy , Doerig Christian
Published: October 2004

Background

Malaria, caused by the parasitic protist Plasmodium falciparum, represents a major public health problem in the developing world. The P. falciparum genome has been sequenced, which provides new opportunities for the identification of novel drug targets. Eukaryotic protein kinases (ePKs) form a large family of enzymes with crucial roles in most cellular processes; hence malarial ePKS represent potential drug targets. We report an exhaustive analysis of the P. falciparum genomic database (PlasmoDB) aimed at identifying and classifying all ePKs in this organism.

Results

Using a variety of bioinformatics tools, we identified 65 malarial ePK sequences and constructed a phylogenetic tree to position these sequences relative to the seven established ePK groups. Predominant features of the tree were: (i) that several malarial sequences did not cluster within any of the known ePK groups; (ii) that the CMGC group, whose members are usually involved in the control of cell proliferation, had the highest number of malarial ePKs; and (iii) that no malarial ePK clustered with the tyrosine kinase (TyrK) or STE groups, pointing to the absence of three-component MAPK modules in the parasite. A novel family of 20 ePK-related sequences was identified and called FIKK, on the basis of a conserved amino acid motif. The FIKK family seems restricted to Apicomplexa, with 20 members in P. falciparum and just one member in some other Apicomplexan species.

Conclusion

The considerable phylogenetic distance between Apicomplexa and other Eukaryotes is reflected by profound divergences between the kinome of malaria parasites and that of yeast or mammalian cells.
In vitro pharmacokinetics of anti-psoriatic fumaric acid esters
Authors: Litjens Nicolle HR , van Strijen Elisabeth , van Gulpen Co , Mattie Herman , van Dissel Jaap T , Thio H Bing , Nibbering Peter H
Published: October 2004

Background

Psoriasis is a chronic inflammatory skin disease that can be successfully treated with a mixture of fumaric acid esters (FAE) formulated as enteric-coated tablets for oral use. These tablets consist of dimethylfumarate (DMF) and salts of monoethylfumarate (MEF) and its main bioactive metabolite is monomethylfumarate (MMF). Little is known about the pharmacokinetics of these FAE. The aim of the present study was to investigate the hydrolysis of DMF to MMF and the stability of MMF, DMF and MEF at in vitro conditions representing different body compartments.

Results

DMF is hydrolyzed to MMF in an alkaline environment (pH 8), but not in an acidic environment (pH 1). In these conditions MMF and MEF remained intact during the period of analysis (6 h). Interestingly, DMF was hardly hydrolyzed to MMF in a buffer of pH 7.4, but was rapidly hydrolyzed in human serum having the same pH. Moreover, in whole blood the half-life of DMF was dramatically reduced as compared to serum. The concentrations of MMF and MEF in serum and whole blood decreased with increasing time. These data indicate that the majority of the FAE in the circulation are metabolized by one or more types of blood cells. Additional experiments with purified blood cell fractions resuspended in phosphate buffered saline (pH 7.4) revealed that at concentrations present in whole blood monocytes/lymphocytes, but not granulocytes and erythrocytes, effectively hydrolyzed DMF to MMF. Furthermore, in agreement with the data obtained with the pure components of the tablet, the enteric-coated tablet remained intact at pH 1, but rapidly dissolved at pH 8.

Conclusion

Together, these in vitro data indicate that hydrolysis of DMF to MMF rapidly occurs at pH 8, resembling that within the small intestines, but not at pH 1 resembling the pH in the stomach. At both pHs MMF and MEF remained intact. These data explain the observation that after oral FAE intake MMF and MEF, but not DMF, can be readily detected in the circulation of human healthy volunteers and psoriasis patients.
Length of sick leave – Why not ask the sick-listed? Sick-listed individuals predict their length of sick leave more accurately than professionals
Authors: Fleten Nils , Johnsen Roar , Førde Olav Helge
Published: October 2004

Background

The knowledge of factors accurately predicting the long lasting sick leaves is sparse, but information on medical condition is believed to be necessary to identify persons at risk. Based on the current practice, with identifying sick-listed individuals at risk of long-lasting sick leaves, the objectives of this study were to inquire the diagnostic accuracy of length of sick leaves predicted in the Norwegian National Insurance Offices, and to compare their predictions with the self-predictions of the sick-listed.

Methods

Based on medical certificates, two National Insurance medical consultants and two National Insurance officers predicted, at day 14, the length of sick leave in 993 consecutive cases of sick leave, resulting from musculoskeletal or mental disorders, in this 1-year follow-up study. Two months later they reassessed 322 cases based on extended medical certificates. Self-predictions were obtained in 152 sick-listed subjects when their sick leave passed 14 days. Diagnostic accuracy of the predictions was analysed by ROC area, sensitivity, specificity, likelihood ratio, and positive predictive value was included in the analyses of predictive validity.

Results

The sick-listed identified sick leave lasting 12 weeks or longer with an ROC area of 80.9% (95% CI 73.7–86.8), while the corresponding estimates for medical consultants and officers had ROC areas of 55.6% (95% CI 45.6–65.6%) and 56.0% (95% CI 46.6–65.4%), respectively. The predictions of sick-listed males were significantly better than those of female subjects, and older subjects predicted somewhat better than younger subjects. Neither formal medical competence, nor additional medical information, noticeably improved the diagnostic accuracy based on medical certificates.

Conclusion

This study demonstrates that the accuracy of a prognosis based on medical documentation in sickness absence forms, is lower than that of one based on direct communication with the sick-listed themselves.
Prospective study of clinician-entered research data in the Emergency Department using an Internet-based system after the HIPAA Privacy Rule
Authors: Kline Jeffrey A , Johnson Charles L , Webb William B , Runyon Michael S
Published: October 2004

Background

Design and test the reliability of a web-based system for multicenter, real-time collection of data in the emergency department (ED), under waiver of authorization, in compliance with HIPAA.

Methods

This was a phase I, two-hospital study of patients undergoing evaluation for possible pulmonary embolism. Data were collected by on-duty clinicians on an HTML data collection form (prospective e-form), populated using either a personal digital assistant (PDA) or personal computer (PC). Data forms were uploaded to a central, offsite server using secure socket protocol transfer. Each form was assigned a unique identifier, and all PHI data were encrypted, but were password-accessible by authorized research personnel to complete a follow-up e-form.

Results

From April 15, 2003-April 15 2004, 1022 prospective e-forms and 605 follow-up e-forms were uploaded. Complexities of PDA use compelled clinicians to use PCs in the ED for data entry for most forms. No data were lost and server log query revealed no unauthorized entry. Prospectively obtained PHI data, encrypted upon server upload, were successfully decrypted using password-protected access to allow follow-up without difficulty in 605 cases. Non-PHI data from prospective and follow-up forms were available to the study investigators via standard file transfer protocol.

Conclusions

Data can be accurately collected from on-duty clinicians in the ED using real-time, PC-Internet data entry in compliance with the Privacy Rule. Deidentification-reidentification of PHI was successfully accomplished by a password-protected encryption-deencryption mechanism to permit follow-up by approved research personnel.
The evolution of drug-activated nuclear receptors: one ancestral gene diverged into two xenosensor genes in mammals
Authors: Handschin Christoph , Blättler Sharon , Roth Adrian , Looser Renate , Oscarson Mikael , Kaufmann Michel R , Podvinec Michael , Gnerre Carmela , Meyer Urs A
Published: October 2004

Background

Drugs and other xenobiotics alter gene expression of cytochromes P450 (CYP) by activating the pregnane X receptor (PXR) and constitutive androstane receptor (CAR) in mammals. In non-mammalian species, only one xenosensor gene has been found. Using chicken as a model organism, the aim of our study was to elucidate whether non-mammalian species only have one or two xenosensors like mammals.

Results

To explore the evolutionary aspect of this divergence, we tried to identify additional xenobiotic sensing nuclear receptors in chicken using various experimental approaches. However, none of those revealed novel candidates. Ablation of chicken xenobiotic receptor (CXR) function by RNAi or dominant-negative alleles drastically reduced drug-induction in a chicken hepatoma cell line. Subsequently, we functionally and structurally characterized CXR and compared our results to PXR and CAR. Despite the high similarity in their amino acid sequence, PXR and CAR have very distinct modes of activation. Some aspects of CXR function, e.g. direct ligand activation and high promiscuity are very reminiscent of PXR. On the other hand, cellular localization studies revealed common characteristics of CXR and CAR in terms of cytoplasmic-nuclear distribution. Finally, CXR has unique properties regarding its regulation in comparison to PXR and CAR.

Conclusion

Our finding thus strongly suggest that CXR constitutes an ancestral gene which has evolved into PXR and CAR in mammals. Future studies should elucidate the reason for this divergence in mammalian versus non-mammalian species.
Identification of endogenous retroviral reading frames in the human genome
Authors: Villesen Palle , Aagaard Lars , Wiuf Carsten , Pedersen Finn Skou
Published: October 2004

Background

Human endogenous retroviruses (HERVs) comprise a large class of repetitive retroelements. Most HERVs are ancient and invaded our genome at least 25 million years ago, except for the evolutionary young HERV-K group. The far majority of the encoded genes are degenerate due to mutational decay and only a few non-HERV-K loci are known to retain intact reading frames. Additional intact HERV genes may exist, since retroviral reading frames have not been systematically annotated on a genome-wide scale.

Results

By clustering of hits from multiple BLAST searches using known retroviral sequences we have mapped 1.1% of the human genome as retrovirus related. The coding potential of all identified HERV regions were analyzed by annotating viral open reading frames (vORFs) and we report 7836 loci as verified by protein homology criteria. Among 59 intact or almost-intact viral polyproteins scattered around the human genome we have found 29 envelope genes including two novel gammaretroviral types. One encodes a protein similar to a recently discovered zebrafish retrovirus (ZFERV) while another shows partial, C-terminal, homology to Syncytin (HERV-W/FRD).

Conclusions

This compilation of HERV sequences and their coding potential provide a useful tool for pursuing functional analysis such as RNA expression profiling and effects of viral proteins, which may, in turn, reveal a role for HERVs in human health and disease. All data are publicly available through a database at http://www.retrosearch.dk.