bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased protein level of PEPT1 intestinal H+-peptide cotransporter upregulates absorption of glycylsarcosine and ceftibuten in 5/6 nephrectomized rats
Authors: Shimizu Yuriko , Masuda Satohiro , Nishihara Kumiko , Ji Lin , Okuda Masahiro , Inui Ken-ichi
Published: November 2004

In chronic renal failure (CRF), dietary protein is one of the factors that deteriorates residual renal functions. Numerous studies have indicated that the products of protein digestion are mainly absorbed as small peptides. However, how small peptides are absorbed in CRF remains poorly understood. H+-coupled peptide transporter (PEPT1/ SLC15A1) plays an important role in the absorption of small peptides and peptide-like drugs in the small intestine. Because dietary protein intake is one of the risk factors for renal failure, the alteration of intestinal PEPT1 might have implications in the progression of renal disease as well as the pharmacokinetics of peptide-like drugs. In this study, we examined the alteration of intestinal PEPT1 in 5/6 nephrectomized (5/6 NR) rats, extensively used as a model of chronic renal failure. Absorption of [14C]glycylsarcosine and ceftibuten was significantly increased in 5/6 NR rats compared with sham-operated rats, without a change in intestinal protease activity. Western blot analysis indicated that the amount of intestinal PEPT1 protein in 5/6 NR rats was increased mainly at the upper region. On the other hand, the amount of intestinal PEPT1 mRNA was not significantly different from that of sham-operated rats. These findings indicate that the increase in absorption of small peptides and peptide-like drugs, caused by the upregulation of intestinal PEPT1 protein, might contribute to the progression of renal failure as well as the alteration of drug pharmacokinetics.
Interleukin-4- and -13-induced hypercontractility of human intestinal muscle cells-implication for motility changes in Crohn's disease
Authors: Akiho Hirotada , Lovato Paola , Deng Yikang , Ceponis Peter J. M. , Blennerhassett Patricia , Collins Stephen M.
Published: November 2004

Crohn's disease is an idiopathic inflammatory condition. However, little is known about the changes that occur in the muscularis externa, despite the fact that this tissue contributes to motility changes and stricture formation. We characterized immune activity in the muscularis externa from intestinal segments of Crohn's disease patients and evaluated the role of IL-4 and -13 as well as signal transducer and activator of transcription (STAT)6 in the contractility of the cultured human intestinal smooth muscle cells. CD3+ve cells ( P < 0.01) and IL-4 protein ( P < 0.01) were significantly increased in the muscularis externa of Crohn's disease patients compared with noninflamed controls. Preincubation of human cultured smooth muscle cells with IL-4 ( P < 0.001) or IL-13 ( P < 0.05) significantly enhanced carbachol-induced contraction, and this was significantly inhibited by the STAT6 inhibitor leflunomide ( P < 0.0001). A similar profile was observed in muscle cells isolated from Crohn's disease patients. Both IL-4 and IL-13 increased specific STAT6-DNA binding in control cells, and this was inhibited by anti-STAT6 Ab ( P < 0.05) or leflunomide ( P < 0.05). IL-4 and IL-13 mediate the hypercontractility of intestinal muscle via a STAT6 pathway at the level of the smooth muscle cell. The STAT6 pathway may contribute to the hypercontractility of intestinal muscle in Crohn's disease.
Inwardly rectifying chloride channel activity in intestinal pacemaker cells
Authors: Zhu Yaohui , Mucci Andrea , Huizinga Jan D.
Published: November 2004

Cl− channels are proposed to play a role in gut pacemaker activity, but little is known about the characteristics of Cl− channels in interstitial cells of Cajal (ICC), the intestinal pacemaker cells. The objective of the present study was to identify whole cell Cl− currents in ICC associated with previously observed single-channel activity and to characterize its inward rectification. Whole cell patch-clamp studies showed that ICC express an inwardly rectifying Cl− current that was not sensitive to changes in cation composition of the extracellular solutions. Currents were not affected by replacing all cations with N-methyl-d-glucamine (NMDG+). Whole cell currents followed the Cl− equilibrium potential and were inhibited by DIDS and 9-anthracene carboxylic acid. Ramp protocols of single-channel activity showed that inward rectification was due to reduction in single-channel open probability, not a reduction in single-channel conductance. Single-channel data led to the hypothesis that strong cooperation exists between 30-pS channels that show less cooperation at potentials positive to the reversal potential. Hence, an inwardly rectifying Cl− channel plays a prominent role in determining pacemaker activity in the gut.
Purinergic regulation of cholangiocyte secretion: identification of a novel role for P2X receptors
Authors: Doctor R. Brian , Matzakos Thomas , McWilliams Ryan , Johnson Sylene , Feranchak Andrew P. , Fitz J. Gregory
Published: November 2004

The P2X family of ligand-gated cation channels is comprised of seven distinct isoforms activated by binding of extracellular purines. Although originally identified in neurons, there is increasing evidence for expression of P2X receptors in epithelia as well. Because ATP is released by both hepatocytes and cholangiocytes, these studies were performed to evaluate whether P2X receptors are present in cholangiocytes and contribute to local regulation of biliary secretion and bile formation. RT-PCR of cDNA from cultured normal rat cholangiocytes detected transcripts for P2X receptors 2, 3, 4, and 6; products from P2X3 and P2X4 were robust and always detectable. In cholangiocyte lysates, P2X4 protein was readily detected, and immunohistochemical staining of intact rat liver revealed P2X4 protein concentrated in intrahepatic bile ducts. To assess the functional significance of P2X4, isolated Mz-ChA-1 cells were exposed to the P2X4-preferring agonist 2′,3′-O-(4-benzoyl-benzoyl)-ATP (BzATP), which activated inward currents of −18.2 + 3.0 pA/pF. In cholangiocyte monolayers, BzATP but not P2X3 agonists elicited robust Cl−secretory responses (short-circuit current) when applied to either the apical (Δ Isc22.1 ± 3.3 μA) or basolateral (18.5 ± 1.6 μA) chamber, with half-maximal stimulation at ∼10 μM and ∼1 μM, respectively. The response to BzATP was unaffected by suramin (not significant) and was inhibited by Cu2+( P < 0.01). These studies provide molecular and biochemical evidence for the presence of P2X receptors in cholangiocytes. Functional studies indicate that P2X4 is likely the primary isoform involved, representing a novel and functionally important component of the purinergic signaling complex modulating biliary secretion.
Inhibition of β-adrenergic receptor trafficking in adult cardiocytes by MAP4 decoration of microtubules
Authors: Cheng Guangmao , Qiao Fei , Gallien Thomas N. , Kuppuswamy Dhandapani , Cooper George
Published: November 2004

Decreased β-adrenergic receptor (β-AR) number occurs both in animal models of cardiac hypertrophy and failure and in patients. β-AR recycling is an important mechanism for the β-AR resensitization that maintains a normal complement of cell surface β-ARs. We have shown that 1) in severe pressure overload cardiac hypertrophy, there is extensive microtubule-associated protein 4 (MAP4) decoration of a dense microtubule network; and 2) MAP4 microtubule decoration inhibits muscarinic acetylcholine receptor recycling in neuroblastoma cells. We asked here whether MAP4 microtubule decoration inhibits β-AR recycling in adult cardiocytes. [3H]CGP-12177 was used as a β-AR ligand, and feline cardiocytes were isolated and infected with adenovirus containing MAP4 (AdMAP4) or β-galactosidase (Adβ-gal) cDNA. MAP4 decorated the microtubules extensively only in AdMAP4 cardiocytes. β-AR agonist exposure reduced cell surface β-AR number comparably in AdMAP4 and Adβ-gal cardiocytes; however, after agonist withdrawal, the cell surface β-AR number recovered to 78.4 ± 2.9% of the pretreatment value in Adβ-gal cardiocytes but only to 56.8 ± 1.4% in AdMAP4 cardiocytes ( P < 0.01). This result was confirmed in cardiocytes isolated from transgenic mice having cardiac-restricted MAP4 overexpression. In functional terms of cAMP generation, β-AR agonist responsiveness of AdMAP4 cells was 47% less than that of Adβ-gal cells. We conclude that MAP4 microtubule decoration interferes with β-AR recycling and that this may be one mechanism for β-AR downregulation in heart failure.
Endothelium-derived 2-arachidonylglycerol: an intermediate in vasodilatory eicosanoid release in bovine coronary arteries
Authors: Gauthier Kathryn M. , Baewer David V. , Hittner Sarah , Hillard Cecilia J. , Nithipatikom Kasem , Reddy D. Sudarshan , Falck J. R. , Campbell William B.
Published: November 2004

Acetylcholine stimulates the release of endothelium-derived arachidonic acid (AA) metabolites including prostacyclin and epoxyeicosatrienoic acids (EETs), which relax coronary arteries. However, mechanisms of endothelial cell (EC) AA activation remain undefined. We propose that 2-arachidonylglycerol (2-AG) plays an important role in this pathway. An AA metabolite isolated from bovine coronary ECs was identified as 2-AG by mass spectrometry. In ECs pretreated with the fatty acid amidohydrolase inhibitor diazomethylarachidonyl ketone (DAK; 20 μmol/l), methacholine (10 μmol/l)-stimulated 2-AG release was blocked by the phospholipase C inhibitor U-73122 (10 μmol/l) or the diacylglycerol lipase inhibitor RHC-80267 (40 μmol/l). In U-46619-preconstricted bovine coronary arterial rings, 2-AG relaxations averaging 100% at 10 μmol/l were inhibited by endothelium removal, by DAK, by the hydrolase inhibitor methyl arachidonylfluorophosphate (10 μmol/l), by the cyclooxygenase inhibitor indomethacin (10 μmol/l), but not by the CB1 cannabinoid receptor antagonist SR-141716 (1 μmol/l). The cytochrome P-450 inhibitor SKF-525a (10 μmol/l) and the 14,15-epoxyeicosa-5 Z-enoic acid EET antagonist (14,15-EEZE; 10 μmol/l) further attenuated the indomethacin-resistant relaxations. The nonhydrolyzable 2-AG analogs noladin ether, 2-AG amide, and 14,15-EET glycerol amide did not induce relaxation. N-nitro-l-arginine-resistant relaxations to methacholine were also inhibited by U-73122, RHC-80267, and DAK. 14,15-EET glycerol ester increased opening of large-conductance K+channels 12-fold in cell-attached patches of isolated smooth muscle cells and induced relaxations averaging 95%. These results suggest that methacholine stimulates EC 2-AG production through phospholipase C and diacylglycerol lipase activation. 2-AG is further hydrolyzed to AA, which is metabolized to vasoactive eicosanoids. These studies reveal a role for 2-AG in EC AA release and the regulation of coronary tone.
PPAR-γ activation fails to provide myocardial protection in ischemia and reperfusion in pigs
Authors: Xu Ya , Gen Michael , Lu Li , Fox Jennifer , Weiss Sara O. , Brown R. Dale , Perlov Daniel , Ahmad Hasan , Zhu Peili , Greyson Clifford , Long Carlin S. , Schwartz Gregory G.
Published: November 2004

Peroxisome proliferator-activated receptor (PPAR)-γ modulates substrate metabolism and inflammatory responses. In experimental rats subjected to myocardial ischemia-reperfusion (I/R), thiazolidinedione PPAR-γ activators reduce infarct size and preserve left ventricular function. Troglitazone is the only PPAR-γ activator that has been shown to be protective in I/R in large animals. However, because troglitazone contains both α-tocopherol and thiazolidinedione moieties, whether PPAR-γ activation per se is protective in myocardial I/R in large animals remains uncertain. To address this question, 56 pigs were treated orally for 8 wk with troglitazone (75 mg·kg−1·day−1), rosiglitazone (3 mg·kg−1·day−1), or α-tocopherol (73 mg·kg−1·day−1, equimolar to troglitazone dose) or received no treatment. Pigs were then anesthetized and subjected to 90 min of low-flow regional myocardial ischemia and 90 min of reperfusion. Myocardial expression of PPAR-γ, determined by ribonuclease protection assay, increased with troglitazone and rosiglitazone compared with no treatment. Rosiglitazone had no significant effect on myocardial contractile function (Frank-Starling relations), substrate uptake, or expression of proinflammatory cytokines during I/R compared with untreated pigs. In contrast, preservation of myocardial contractile function and lactate uptake were greater and cytokine expression was attenuated in pigs treated with troglitazone or α-tocopherol compared with untreated pigs. Multivariate analysis indicated that presence of an α-tocopherol, but not a thiazolidinedione, moiety in the test compound was significantly related to greater contractile function and lactate uptake and lower cytokine expression during I/R. We conclude that PPAR-γ activation is not protective in a porcine model of myocardial I/R. Protective effects of troglitazone are attributable to its α-tocopherol moiety. These findings, in conjunction with prior rat studies, suggest interspecies differences in the response to PPAR-γ activation in the heart.
Regulation of actin dynamics is critical for endothelial barrier functions
Authors: Waschke J. , Curry F. E. , Adamson R. H. , Drenckhahn D.
Published: November 2004

We tested the hypothesis that the equilibrium between F- and G-actin in endothelial cells modulates the integrity of the actin cytoskeleton and is important for the maintenance of endothelial barrier functions in vivo and in vitro. We used the actin-depolymerizing agent cytochalasin D and jasplakinolide, an actin filament (F-actin) stabilizing and promoting substance, to modulate the actin cytoskeleton. Low doses of jasplakinolide (0.1 μM), which we have previously shown to reduce the permeability-increasing effect of cytochalasin D, had no influence on resting permeability of single-perfused mesenteric microvessels in vivo as well as on monolayer integrity. The F-actin content of cultured endothelial cells remained unchanged. In contrast, higher doses (10 μM) of jasplakinolide increased permeability (hydraulic conductivity) to the same extent as cytochalasin D and induced formation of intercellular gaps in cultured myocardial endothelial (MyEnd) cell monolayers. This was accompanied by a 34% increase of F-actin and pronounced disorganization of the actin cytoskeleton in MyEnd cells. Furthermore, we tested whether an increase of cAMP by forskolin and rolipram would prevent the cytochalasin D-induced barrier breakdown. Conditions that increase intracellular cAMP failed to block the cytochalasin D-induced permeability increase in vivo and the reduction of vascular endothelial cadherin-mediated adhesion in vitro. Taken together, these data support the hypothesis that the state of polymerization of the actin cytoskeleton is critical for maintenance of endothelial barrier functions and that both depolymerization by cytochalasin D and hyperpolymerization of actin by jasplakinolide resulted in an increase of microvessel permeability in vivo. However, cAMP, which is known to support endothelial barrier functions, seems to work by mechanisms other than stabilizing F-actin.
Diabetic dyslipidemia and exercise affect coronary tone and differential regulation of conduit and microvessel K+ current
Authors: Mokelke E. A. , Dietz N. J. , Eckman D. M. , Nelson M. T. , Sturek M.
Published: November 2004

Spontaneous transient outward K+ currents (STOCs) elicited by Ca2+ sparks and steady-state K+ currents modulate vascular reactivity, but effects of artery size, diabetic dyslipidemia, and exercise on these differentially regulated K+ currents are unclear. We studied the conduit arteries and microvessels of male Yucatan swine assigned to one of three groups for 20 wk: control (C, n = 7), diabetic dyslipidemic (DD, n = 6), or treadmill-trained DD animals (DDX, n = 7). Circumflex artery blood flow velocity obtained with intracoronary Doppler and lumen diameters obtained by intravascular ultrasound enabled calculation of absolute coronary blood flow (CBF). Ca2+ sparks were determined in pressurized microvessels, and perforated patch clamp assessed K+ current in smooth muscle cells isolated from conduits and microvessels. Baseline CBF in DD was decreased versus C. In pressurized microvessels, Ca2+ spark activity was significantly lower in DD versus C and DDX ( P < 0.05 vs. DDX). STOCs were pronounced in microvessel (∼35 STOCs/min) in sharp contrast to conduit cells (∼2 STOCs/min). STOCs were decreased by 86% in DD versus C and DDX in microvessels; in contrast, there was no difference in STOCs across groups in conduit cells. Steady-state K+ current in microvessels was decreased in DD and DDX versus C; in contrast, steady-state K+ current in conduit cells was decreased in DDX versus DD and C. We conclude that steady-state K+ current and STOCs are differentially regulated in conduit versus microvessels in health and diabetic dyslipidemia. Exercise prevented diabetic dyslipidemia-induced decreases in baseline CBF, possibly via STOC-regulated basal microvascular tone.
Endothelin-1 decreases endothelial NOS expression and activity through ETA receptor-mediated generation of hydrogen peroxide
Authors: Wedgwood Stephen , Black Stephen M.
Published: November 2004

Similar to infants born with persistent pulmonary hypertension of the newborn (PPHN), there is an increase in circulating endothelin-1 (ET-1) and decreased endothelial nitric oxide synthase (eNOS) gene expression in an ovine model of PPHN. These abnormalities lead to vasoconstriction and vascular remodeling. Our previous studies have demonstrated that reactive oxygen species (ROS) levels are elevated in the pulmonary arteries from PPHN lambs and that ET-1 increases ROS production in pulmonary arterial smooth muscle cells (PASMC) in culture. Thus the objective of this study was to determine whether there was a feedback mechanism between the ET-1-mediated increase in ROS in fetal PASMC (FPASMC) and a decrease in eNOS gene expression in fetal pulmonary arterial endothelial cells (FPAEC). Our results indicate that ET-1 increased H2O2 levels in FPASMC in an endothelin A receptor-dependent fashion. This was observed in both FPASMC monoculture and in cocultures of FPASMC and FPAEC. Conversely, ET-1 decreased H2O2 levels in FPAEC monoculture in an endothelin B receptor-dependent fashion. Furthermore, ET-1 decreased eNOS promoter activity by 40% in FPAEC in coculture with FPASMC. Promoter activity was restored in the presence of catalase. In FPAEC in monoculture treated with 0–100 μM H2O2, 12 μM had no effect on eNOS promoter activity, but it increased eNOS protein levels by 50%. However, at 100 μM, H2O2 decreased eNOS promoter activity and protein levels in FPAEC by 79 and 40%, respectively. These data suggest a role for smooth muscle cell-derived H2O2 in ET-1-mediated downregulation of eNOS expression in children born with PPHN.
Heme oxygenase-derived carbon monoxide promotes arteriolar endothelial dysfunction and contributes to salt-induced hypertension in Dahl salt-sensitive rats
Authors: Teran Federico J. , Johnson Robert A. , Stevenson Blake K. , Peyton Kelly J. , Jackson Keith E. , Appleton Scott D. , Durante William , Johnson Fruzsina K.
Published: November 2004

Vascular tissues express heme oxygenase (HO), which metabolizes heme to form carbon monoxide (CO). Heme-derived CO inhibits nitric oxide synthase and promotes endothelium-dependent vasoconstriction. After 4 wk of high-salt diet, Dahl salt-sensitive (Dahl-S) rats display hypertension, increased vascular HO-1 expression, and attenuated vasodilator responses to ACh that can be completely restored by acute treatment with an inhibitor of HO. In this study, we examined the temporal development of HO-mediated endothelial dysfunction in isolated pressurized first-order gracilis muscle arterioles, identified the HO product responsible, and studied the blood pressure effects of HO inhibition in Dahl-S rats on a high-salt diet. Male Dahl-S rats (5–6 wk) were placed on high-salt (8% NaCl) or low-salt (0.3% NaCl) diets for 0–4 wk. Blood pressure increased gradually, and responses to an endothelium-dependent vasodilator, ACh, decreased gradually with the length of high-salt diet. Flow-induced dilation was abolished in hypertensive Dahl-S rats. Acute in vitro pretreatment with an inhibitor of HO, chromium mesoporphyrin (CrMP), restored endothelium-dependent vasodilation and abolished the differences between groups. The HO product CO prevented the restoration of endothelium-dependent dilation by CrMP. Furthermore, administration of an HO inhibitor lowered blood pressure in Dahl-S rats with salt-induced hypertension but did not do so in low-salt control rats. These results suggest that hypertension and HO-mediated endothelial dysfunction develop gradually and simultaneously in Dahl-S rats on high-salt diets. They also suggest that HO-derived CO underlies the impaired endothelial dysfunction and contributes to hypertension in Dahl-S rats on high-salt diets.
Differential effects of maternal hypoxia or nutrient restriction on carotid and femoral vascular function in neonatal rats
Authors: Williams Sarah J. , Campbell Morag E. , McMillen I. Caroline , Davidge Sandra T.
Published: November 2004

In response to reduced oxygen or nutrient supply, the fetus may redistribute cardiac output to conserve brain and heart growth, at the expense of the peripheral tissues; however, it is not known whether alterations in vascular function are maintained after birth or whether reduced fetal oxygen versus nutrient supply produces distinct effects. Using a pressure myograph, we examined isolated carotid and femoral artery responses to phenylephrine and endothelin-1 in neonatal rats, after either reduced maternal oxygen or global nutrient restriction during late gestation. Timed-pregnant Sprague-Dawley rats were randomly assigned to control ( n = 10), hypoxia (12% O2, n = 9), or nutrient restriction (NR, 40% of control diet, n = 7) protocol and treated from day 15–21 of pregnancy. Pups were collected 3–12 h after birth. Neonatal weights ( P < 0.001) and relative liver weights ( P < 0.001) were lower in hypoxia and nutrient restriction treatments compared with control, while relative heart weights were greater in the hypoxia than in the control or nutrient restriction groups ( P < 0.01). Constriction to phenylephrine was reduced in carotid arteries from the hypoxia and nutrient restriction groups compared with control ( P < 0.001), while the femoral artery response was greater in hypoxia-treated neonates compared with control or nutrient-restricted neonates ( P < 0.01). Only the hypoxia reduced carotid responses to endothelin-1, while no differences were observed in the endothelin-1 responses in femoral arteries. Maternal hypoxia and maternal nutrient restriction produced distinct effects on heart growth and neonatal vascular function, suggesting that regional changes in cardiovascular function after poor fetal growth are dependent on the nature of the insult in utero.
Photoperiod-dependent modulation of cardiac excitation contraction coupling in the Siberian hamster
Authors: Dibb K. M. , Hagarty C. L. , Loudon A. S. I. , Trafford A. W.
Published: November 2004

In mammals, changes in photoperiod regulate a diverse array of physiological and behavioral processes, an example of which in the Siberian hamster ( Phodopus sungorus) is the expression of bouts of daily torpor following prolonged exposure to a short photoperiod. During torpor, body temperature drops dramatically; however, unlike in nonhibernating or nontorpid species, the myocardium retains the ability to contract and is resistant to the development of arrhythmias. In the present study, we sought to determine whether exposure to a short photoperiod results in alterations to cardiac excitation-contraction coupling, thus potentially enabling the heart to survive periods of low temperature during torpor. Experiments were performed on single ventricular myocytes freshly isolated from the hearts of Siberian hamsters that had been exposed to either 12 wk of short-day lengths (SD) or 12 wk of long-day lengths (LD). In SD-acclimated animals, the amplitude of the systolic Ca2+ transient was increased (e.g., from 142 ± 17 nmol/l in LD to 229 ± 31 nmol/l in SD at 4 Hz; P < 0.001). The increased Ca2+ transient amplitude in the SD-acclimated animals was not associated with any change in the shape or duration of the action potential. However, sarcoplasmic reticulum Ca2+ content measured after current-clamp stimulation was increased in the SD-acclimated animals (at 4 Hz, 110 ± 5 vs. 141 ± 15 μmol/l, P < 0.05). We propose that short photoperiods reprogram the function of the cardiac sarcoplasmic reticulum, resulting in an increased Ca2+ content, and that this may be a necessary precursor for maintenance of cardiac function during winter torpor.
Test of the principle of initial value in rat genetic models of exercise capacity
Authors: Koch Lauren Gerard , Green Cheryl L. , Lee Abraham D. , Hornyak Joseph E. , Cicila George T. , Britton Steven L.
Published: November 2004

An inverse relationship between initial level of physical capacity and the magnitude of response to training is termed the principle of initial value. We tested the operation of this principle under experimental conditions of minimal genetic and environmental variation. Inbred rat strains previously identified as genetic models of low [Copenhagen (COP)] and high [Dark Agouti (DA)] intrinsic (untrained) exercise capacity were trained for 8 wk on a treadmill using two disparate protocols: 1) a relative mode where each rat exercised daily according to its initial capacity, and 2) an absolute mode where both strains received the same amount of training independent of initial capacity. Response to exercise was the change in running capacity as estimated by meters run to exhaustion before and after training. When trained with the relative mode, COP rats gained 88 m (+21%; NS) whereas DA rats increased distance run by 228 m (+36%; P < 0.001). When each strain trained with the same absolute amount of training, the COP strain showed essentially no change (−6 m, −2%) and the DA strain gained 325 m (+49%; P < 0.009). Differences in response to exercise between the COP and DA could not be explained by body mass differences, oxidative enzyme activity (citrate synthase or ATP), or spontaneous behavioral activity. Our data demonstrate that genetic factors causative of high response to exercise are not uniquely associated with genetic factors for low intrinsic capacity and thus are not in accord with the principle of initial value.
Acute ventilator-induced vascular permeability and cytokine responses in isolated and in situ mouse lungs
Authors: Yoshikawa S. , King J. A. , Lausch R. N. , Penton A. M. , Eyal F. G. , Parker J. C.
Published: November 2004

To determine the influence of experimental model and strain differences on the relationship of vascular permeability to inflammatory cytokine production after high peak inflation pressure (PIP) ventilation, we used isolated perfused mouse lung and intact mouse preparations of Balb/c and B6/129 mice ventilated at high and low PIP. Filtration coefficients in isolated lungs and bronchoalveolar lavage (BAL) albumin in intact mice increased within 20–30 min after initiation of high PIP in isolated Balb/c lungs and intact Balb/c, B6/129 wild-type, and p55 and p75 tumor necrosis factor (TNF) dual-receptor null mice. In contrast, the cytokine response was delayed and variable compared with the permeability response. In isolated Balb/c lungs ventilated with 25–27 cmH2O PIP, TNF-α, interleukin (IL)-1β, IL-1α, macrophage inflammatory protein (MIP)-2, and IL-6 concentrations in perfusate were markedly increased in perfusate at 2 and 4 h, but only MIP-2 was detectable in intact Balb/c mice using the same PIP. In intact wild-type and TNF dual-receptor null mice with ventilation at 45 cmH2O PIP, the MIP-2 and IL-6 levels in BAL were significantly increased after 2 h in both groups, but there were no differences between groups in the BAL albumin and cytokine concentrations or in lung wet-to-dry weight ratios. TNF-α was not be detected in BAL fluids in any group of intact mice. These results suggest that the alveolar hyperpermeability induced by high PIP ventilation occurs very rapidly and is initially independent of TNF-α participation and unlikely to depend on MIP-2 or IL-6.
Association of interleukin-15 protein and interleukin-15 receptor genetic variation with resistance exercise training responses
Authors: Riechman Steven E. , Balasekaran G. , Roth Stephen M. , Ferrell Robert E.
Published: November 2004

Interleukin-15 (IL-15) is an anabolic cytokine that is produced in skeletal muscle and directly affects muscle anabolism in animal and in vitro models. The contribution of IL-15 variability in muscle responses to 10 wk of resistance exercise training in young men and women was examined by measuring acute and chronic changes in IL-15 protein in plasma and characterizing genetic variation in the IL-15 receptor-α gene (IL15RA). Participants trained 3 days a week at 75% of one repetition maximum, performing three sets (6–10 repetitions) of 13 resistance exercises. Plasma IL-15 protein was significantly increased ( P < 0.05) immediately after acute resistance exercise but did not change with training and was not associated with variability in muscle responses with training. A single nucleotide polymorphism in exon 7 of IL15RA was strongly associated with muscle hypertrophy and accounted for 7.1% of the variation in regression modeling. A polymorphism in exon 4 was also independently associated with muscle hypertrophy and accounted for an additional 3.5% of the variation in hypertrophy. These results suggest that IL-15 is an important mediator of muscle mass response to resistance exercise training in humans and that genetic variation in IL15RA accounts for a significant proportion of the variability in this response.
Timing of cortical excitability changes during the reaction time of movements superimposed on tonic motor activity
Authors: Schneider Cyril , Lavoie Brigitte A. , Barbeau Hugues , Capaday Charles
Published: November 2004

Seated subjects were instructed to react to an auditory cue by simultaneously contracting the tibialis anterior (TA) muscle of each ankle isometrically. Focal transcranial magnetic stimulation of the leg area of the motor cortex (MCx) was used to determine the time course of changes in motor-evoked potential amplitude (MEP) during the reaction time (RT). In one condition the voluntary contraction was superimposed on tonic EMG activity maintained at 10% of maximal voluntary contraction. In the other condition the voluntary contraction was made starting from rest. MEPs in the TA contralateral to the stimulation coil were evoked at various times during the RT in each condition. These were compared to the control MEPs evoked during tonic voluntary activity or with the subject at rest. The RT was measured trial by trial from the EMG activity of the TA ipsilateral to the magnetic stimulus, taking into account the nearly constant time difference between the two sides. The MEPs became far greater than control MEPs during the RT (mean = 332%, SD = 44 %, of control MEPs, P < 0.001) without any measurable change in the background level of EMG activity. The onset of this facilitation occurred on average 12.80 ms (SD = 7.55 ms) before the RT. There was no difference in the onset of facilitation between the two conditions. Because MEPs were facilitated without a change in the background EMG activity, it is concluded that this facilitation is specifically due to an increase of MCx excitability just before voluntary muscle activation. This conclusion is further reinforced by the observation that MEPs evoked by near-threshold anodal stimuli to the MCx were not facilitated during the RT, in contrast to those evoked by near-threshold transcranial magnetic stimulation. However, several observations in the present and previous studies indicate that MEP amplitude may be more sensitive to α-motoneuron activity than to motor cortical neuron activity, an idea that has important methodological implications.
Monocarboxylate transporters, blood lactate removal after supramaximal exercise, and fatigue indexes in humans
Authors: Thomas C. , Perrey S. , Lambert K. , Hugon G. , Mornet D. , Mercier J.
Published: November 2004

The present study investigated whether muscular monocarboxylate transporter (MCT) 1 and 4 contents are related to the blood lactate removal after supramaximal exercise, fatigue indexes measured during different supramaximal exercises, and muscle oxidative parameters in 15 humans with different training status. Lactate recovery curves were obtained after a 1-min all-out exercise. A biexponential time function was then used to determine the velocity constant of the slow phase (γ2), which denoted the blood lactate removal ability. Fatigue indexes were calculated during 1-min all-out (FIAO) and repeated 10-s (FISprint) cycling sprints. Biopsies were taken from the vastus lateralis muscle. MCT1 and MCT4 contents were quantified by Western blots, and maximal muscle oxidative capacity ( Vmax) was evaluated with pyruvate + malate and glutamate + malate as substrates. The results showed that the blood lactate removal ability (i.e., γ2) after a 1-min all-out test was significantly related to MCT1 content ( r = 0.70, P < 0.01) but not to MCT4 ( r = 0.50, P > 0.05). However, greater MCT1 and MCT4 contents were negatively related with a reduction of blood lactate concentration at the end of 1-min all-out exercise ( r = −0.56, and r = −0.61, P < 0.05, respectively). Among skeletal muscle oxidative indexes, we only found a relationship between MCT1 and glutamate + malate Vmax ( r = 0.63, P < 0.05). Furthermore, MCT1 content, but not MCT4, was inversely related to FIAO ( r = −0.54, P < 0.05) and FISprint ( r = −0.58, P < 0.05). We concluded that skeletal muscle MCT1 expression was associated with the velocity constant of net blood lactate removal after a 1-min all-out test and with the fatigue indexes. It is proposed that MCT1 expression may be important for blood lactate removal after supramaximal exercise based on the existence of lactate shuttles and, in turn, in favor of a better tolerance to muscle fatigue.
Dual Effect of Angiotensin-Converting Enzyme Inhibition on Angiogenesis in Type 1 Diabetic Mice
Authors: Ebrahimian Téni G. , Tamarat Radia , Clergue Michel , Duriez Micheline , Levy Bernard I. , Silvestre Jean-Sébastien
Published: November 2004

Objective—

We analyzed the beneficial therapeutic effect of angiotensin converting enzyme inhibitor (ACEI) on both retinal and hind limb neovascularization in diabetic mice.

Methods and Results—

Diabetic mice (streptozotocin, 40 mg/kg) were treated with or without ACEI (Perindopril, 3 mg/kg per day) or AT1 receptor blocker (Candesartan, 20 mg/kg) for 4 months. Hind limb ischemia was then induced by right femoral artery ligature for 1 additional month. In the ischemic leg, angiographic score, capillary density, and foot perfusion were increased by 2.7, 2.0-fold, and 1.6-fold, respectively, in ACEI-treated diabetic mice compared with untreated diabetic animals (
P
<0.01). ACEI also raised vascular endothelial growth factor (VEGF) protein level by 1.4-fold in ischemic diabetic leg. This ACEI pro-angiogenic effect was totally blunted in diabetic bradykinin B2 receptor-deficient animals, suggesting that it was mediated by the bradykinin pathway. In the diabetic retina, angiotensinogen and ACE mRNA levels were increased by 2.8-fold and 4.1-fold, respectively (
P
<0.01 versus nondiabetic mice), highlighting a local activation of renin-angiotensin system. Diabetes also raised VEGF protein level by 1.5-fold (
P
<0.05 versus nondiabetic mice). Treatments with ACEI and AT1 receptor blocker hampered diabetes-induced VEGF upregulation and retinal neovascularization.

Conclusion—

ACE inhibition improved neovascularization in the diabetic ischemic leg through activation of bradykinin signaling, whereas it reduced vessel growth in the diabetic retina through inhibition of overacting Ang II pathway.
Reduced In Vivo Aortic Uptake of Radiolabeled Oxidation-Specific Antibodies Reflects Changes in Plaque Composition Consistent With Plaque Stabilization
Authors: Torzewski Michael , Shaw Peter X. , Han Kyoo-Rok , Shortal Brian , Lackner Karl J. , Witztum Joseph L. , Palinski Wulf , Tsimikas Sotirios
Published: November 2004

Objective—

Labeled oxidation-specific antibodies (Ox-AB) detect, quantify, and noninvasively image lipid-rich atherosclerotic lesions. However, it is unknown whether Ox-AB detect plaque stabilization.

Methods and Results—

The aortic uptake of intravenously injected
125
I-MDA2 (Ox-AB to malondialdehyde [MDA]–low-density lipoprotein [LDL]) was quantitated in: (1) LDL receptor−/− mice with established atherosclerosis continued on Western diet (Progression) or switched to chow (Regression) or chow+vitamins E and C (Regression-VIT) for 6 months; and (2) Watanabe rabbits (3- to 57-months old) with naturally evolved atherosclerotic lesions. In mice, the Progression group had more extensive atherosclerosis, higher
125
I-MDA2 uptake, high concordance of Sudan (lipid)-staining and
125
I-MDA2 uptake, and stronger oxidized LDL (OxLDL) and macrophage immunostaining than both Regression groups. In contrast, the Regression groups showed Sudan-positive lesions with focally diminished
125
I-MDA2 uptake, which coincided with reduced OxLDL and macrophages but more smooth muscle cells (SMCs) and collagen. In rabbits, areas of increased
125
I-MDA2 uptake were associated with high Sudan concordance and strong immunostaining for OxLDL and macrophages. Interestingly, advanced lesions with focally diminished
125
I-MDA2 uptake showed stronger immunostaining for SMCs and collagen, particularly at the fibrous cap.

Conclusion—

Ox-AB uptake is focally diminished in plaques displaying accepted features of plaque stability. Imaging techniques to detect the presence and depletion of OxLDL may be useful in assessing plaque stabilization.
MRI of Atherothrombosis Associated With Plaque Rupture
Authors: Viereck Jason , Ruberg Frederick L. , Qiao Ye , Perez Alexandra S. , Detwiller Kara , Johnstone Michael , Hamilton James A.
Published: November 2004

Objective—

MRI would be a valuable noninvasive diagnostic tool to study plaque-associated thrombi. We defined the imaging characteristics of these thrombi, composed primarily of platelets and fibrin, and distinguished them clearly from the vessel lumen and underlying atherosclerotic plaque in an animal model of plaque rupture.

Methods and Results—

After triggering plaque rupture in New Zealand White male rabbits, segments of infrarenal aorta containing either red or white thrombi were fixed in formalin. Compared with postmortem red cell–rich thrombi, atherothrombi yielded complex magnetic resonance images with intermediate signal intensity in standard T1- and T2-weighted imaging sequences and were often difficult to distinguish from the aortic wall. Diffusion-weighted imaging sequences revealed restricted diffusion of the atherothrombus relative to the vessel wall and provided excellent contrast. The apparent diffusion coefficient of the thrombus is 1.0×10
−3
mm
2
/s, compared with 1.5×10
−3
mm
2
/s in tissue. Similar results were obtained using purified aggregated platelets.

Conclusions—

We present the first detailed description of the MRI appearance of plaque rupture–associated thrombosis in histologically validated platelet-rich thrombi. Diffusion-weighted imaging provided the best distinction between thrombus and vessel wall and has potential application for the noninvasive in vivo detection of atherothrombosis.
Sulfatides Activate Platelets Through P-Selectin and Enhance Platelet and Platelet–Leukocyte Aggregation
Authors: Merten Michael , Beythien Christian , Gutensohn Kai , Kühnl Peter , Meinertz Thomas , Thiagarajan Perumal
Published: November 2004

Objective—

Sulfatides are sulfated glycosphingolipids present on the surface of a variety of cells; however, their exact physiological function is not known. Recently, we have shown that the inhibition of sulfatide–P-selectin interactions leads to disaggregation of platelet aggregates.

Methods and Results—

In this study, we show that sulfatides activated platelets as they increased activation of GPIIb/IIIa (PAC-1 epitope) and expression of P-selectin on the platelet surface. Furthermore, sulfatides aggregated washed platelets in a dose-dependent manner and enhanced platelet aggregation in platelet-rich plasma. Previous activation of platelets was necessary for this effect. Monoclonal anti–P-selectin antibodies inhibited not only sulfatide-induced PAC-1 binding to platelets but also sulfatide-induced platelet aggregation, suggesting that sulfatides activate platelet GPIIb/IIIa via signaling through P-selectin. The proaggegatory effect of sulfatides was also observed in an ex vivo thrombosis model using whole blood and pulsatile flow at 37°C. In this model, sulfatides significantly enhanced platelet aggregation and the formation of platelet–leukocyte aggregates.

Conclusions—

We show that sulfatide-P-selectin interactions lead to subsequent platelet activation and P-selectin expression, forming a positive feedback loop that can potentiate formation of stable platelet aggregates. In addition, sulfatides enhance the aggregation of platelet–leukocyte aggregates. These mechanisms may play a significant role in hemostasis and thrombosis.
Quantitative Evaluation of Carotid Plaque Composition by In Vivo MRI
Authors: Saam T. , Ferguson M.S. , Yarnykh V.L. , Takaya N. , Xu D. , Polissar N.L. , Hatsukami T.S. , Yuan C.
Published: November 2004

Objective—

This study evaluates the ability of MRI to quantify all major carotid atherosclerotic plaque components in vivo.

Methods and Results—

Thirty-one subjects scheduled for carotid endarterectomy were imaged with a 1.5T scanner using time-of-flight–, T1-, proton density–, and T2-weighted images. A total of 214 MR imaging locations were matched to corresponding histology sections. For MRI and histology, area measurements of the major plaque components such as lipid-rich/necrotic core (LR/NC), calcification, loose matrix, and dense (fibrous) tissue were recorded as percentages of the total wall area. Intraclass correlation coefficients (ICCs) were computed to determine intrareader and inter-reader reproducibility. MRI measurements of plaque composition were statistically equivalent to those of histology for the LR/NC (23.7 versus 20.3%;
P
=0.1), loose matrix (5.1 versus 6.3%;
P
=0.1), and dense (fibrous) tissue (66.3% versus 64%;
P
=0.4). Calcification differed significantly when measured as a percentage of wall area (9.4 versus 5%;
P
<0.001). Intrareader and inter-reader reproducibility was good to excellent for all tissue components, with ICCs ranging from 0.73 to 0.95.

Conclusions—

MRI-based tissue quantification is accurate and reproducible. This application can be used in therapeutic clinical trials and in prospective longitudinal studies to examine carotid atherosclerotic plaque progression and regression.
Crucial Role of the CCL2/CCR2 Axis in Neointimal Hyperplasia After Arterial Injury in Hyperlipidemic Mice Involves Early Monocyte Recruitment and CCL2 Presentation on Platelets
Authors: Schober Andreas , Zernecke Alma , Liehn Elisa A. , von Hundelshausen Philipp , Knarren Sandra , Kuziel William A. , Weber Christian
Published: November 2004

Monocyte chemoattractant protein-1 (also known as CC chemokine ligand 2 [CCL2]) and its receptor CC chemokine receptor 2 (CCR2) play a central role in the inflammatory response and neointimal formation after vascular injury. In the context of hyperlipidemia, this appears to involve neointimal monocyte infiltration. Hence, we investigated the function of the CCL2/CCR2 axis in early monocyte recruitment to injured arteries. Wire-induced injury of the carotid artery in apoE
−/−
mice caused a rapid increase of JE/CCL2 protein in the vessel wall peaking at 24 hours after injury, whereas serum JE/CCL2 was increased solely at 6 hours and blood cell-associated levels were unaltered, as demonstrated by enzyme-linked immunosorbent assay. Immunohistochemistry revealed intense staining for JE/CCL2 in smooth muscle cells (SMCs) and in association with platelets adherent to the denuded vessel wall 24 hours after injury. In vitro, exogenous or SMC-derived JE/CCL2 binds to the platelet surface and triggers monocyte arrest on adherent platelets but not on SMCs in flow assays. Accordingly, monocyte arrest in ex vivo perfused apoE
−/−
carotid arteries isolated 24 hours after injury was profoundly inhibited by pretreatment with a JE/CCL2 antibody. In CCR2
−/−
/apoE
−/−
mice, neointimal plaque area was reduced by 47% compared with CCR2
+/+
/apoE
−/−
mice. Moreover, CCR2 deletion markedly decreased neointimal macrophage content while expanding SMC content. Vascular JE/CCL2 expressed by SMCs and immobilized by adherent platelets after endothelial denudation is crucial for mediating early monocyte recruitment to injured arteries in hyperlipidemic mice. This mechanism may explain reduced neointimal macrophage infiltration and lesion formation in CCR2-deficient apoE
−/−
mice.
Alterations in Myofilament Function Contribute to Left Ventricular Dysfunction in Pigs Early After Myocardial Infarction
Authors: van der Velden J. , Merkus D. , Klarenbeek B.R. , James A.T. , Boontje N.M. , Dekkers D.H.W. , Stienen G.J.M. , Lamers J.M.J. , Duncker D.J.
Published: November 2004

Myocardial infarction (MI) initiates cardiac remodeling, depresses pump function, and predisposes to heart failure. This study was designed to identify early alterations in Ca
2+
handling and myofilament proteins, which may contribute to contractile dysfunction and reduced β-adrenergic responsiveness in postinfarct remodeled myocardium. Protein composition and contractile function of skinned cardiomyocytes were studied in remote, noninfarcted left ventricular (LV) subendocardium from pigs 3 weeks after MI caused by permanent left circumflex artery (LCx) ligation and in sham-operated pigs. LCx ligation induced a 19% increase in LV weight, a 69% increase in LV end-diastolic area, and a decrease in ejection fraction from 54±5% to 35±4% (all
P
<0.05), whereas cardiac responsiveness to exercise-induced increases in circulating noradrenaline levels was blunted. Endogenous protein kinase A (PKA) was significantly reduced in remote myocardium of MI animals, and a negative correlation (
R
=0.62;
P
<0.05) was found between cAMP levels and LV weight-to-body weight ratio. Furthermore, SERCA2a expression was 23% lower after MI compared with sham. Maximal isometric force generated by isolated skinned myocytes was significantly lower after MI than in sham (15.4±1.5 versus 19.2±0.9 kN/m
2
;
P
<0.05), which might be attributable to a small degree of troponin I (TnI) degradation observed in remodeled postinfarct myocardium. An increase in Ca
2+
sensitivity of force (pCa
50
) was observed after MI compared with sham (ΔpCa
50
=0.17), which was abolished by incubating myocytes with exogenous PKA, indicating that the increased Ca
2+
sensitivity resulted from reduced TnI phosphorylation. In conclusion, remodeling of noninfarcted pig myocardium is associated with decreased SERCA2a and myofilament function, which may contribute to depressed LV function. The full text of this article is available online at http://circres.ahajournals.org.