Study of the electron-doping mechanism in single-walled carbon nanotubes using dimethylbenzimidazole.

Single-walled carbon nanotubes (SWCNTs) exhibit p-type properties in air, necessitating electron doping using n-dopants (e.g., reducing agents) for the development of SWCNT-based electronic devices. Dimethylbenzimidazole (DMBI-H) derivatives serve as effective electron dopants, not only for SWCNTs, but also for various organic semiconducting materials. However, the doping reaction is still a subject of debate. In this study, the electron-doping reactions of ortho-methoxy-substituted DMBI-H for SWCNTs were analyzed in protic and aprotic solvents in the presence and absence of dioxygen (O2). The presence of O2 was found to cause the reduction of O2 on the SWCNT surface in the protic solvent, resulting in the production of DMBI cations and water through proton-coupled electron transfer (PCET) from the n-doped SWCNT and ethanol. This work elucidates the mechanism behind the air-stability of n-type SWCNTs.

Can radiological technologists serve as primary screeners of low-dose computed tomography for the diagnosis of lung cancer?

BACKGROUND: The Accreditation Council for Lung Cancer CT Screening of Japan established guidelines for the certification of Radiological Technologists in 2009. OBJECTIVE: To analyze the trends in examination pass rates of the Radiological Technologists and discuss the reasons. METHODS: The cohort comprised 1593 Radiological Technologists (as examinees) based on 10-year of data (with a total of 17 examination runs). First, the examinees' written test results were analyzed. Second, an abnormal finding detection test was conducted using >100 client PCs connected to a dedicated server containing low-dose lung cancer CT screening images of 60 cases. The passing scores were correct answer rate >60% and sensitivity (TP) of >90%, respectively. RESULTS: Overall, 1243 examinees passed with an overall rate of 78%. The average pass rate for the written test was 91%, whereas that for the abnormal findings detection test was 85%. There was a moderate correlation between the test pass rate and average years of clinical experience of the examinees for the abnormal findings detection test (R = 0.558), whereas no such correlation existed for the written test (R = 0.105). CONCLUSIONS: In order for accredited Radiological Technologists to serve as primary screeners of low-dose computed tomography, it is important to revise the educational system according to current standard practices.

Cerebellar ataxia in a patient receiving calcineurin inhibitors after living donor liver transplantation: a case report.

Neurological complications of calcineurin inhibitors are frequent problems after transplantation. Cerebellar ataxia with other neurological findings and an abnormal density area in the subcortical white matter are found by MRI in the brains of most patients with central nervous system complications caused by calcineurin inhibitors. Such neurological complications are not life-threatening, but have a negative impact on the quality of life. We describe a 58-year-old woman who developed cerebellar ataxia at 4 days after living donor liver transplantation. She walked with a swaying gait, and after walking for 5 minutes she was unable to stand. Her symptoms persisted after a change from tacrolimus to cyclosporine, but dose reduction of cyclosporine and addition of mycophenolate mofetil cured the ataxia. We diagnosed a case of cerebellar ataxia without leukoencephalopathy or other neurological symptoms, as a new complication of calcineurin inhibitor treatment. We concluded that careful attention should be paid to neurological complications of calcineurin inhibitors.

Cloning and characterization of six highly similar endo-1,3-beta-glucanase genes in hexaploid wheat.

Pathogenesis-related (PR) proteins are often used as a marker of plant defense reactions. Some endo-1,3-beta-glucanase (Gns) genes encode proteins that belong to the PR protein family 2 (PR-2). Although the number of homologous family member genes is significantly greater in hexaploid wheat (Triticum aestivum L.) compared to other model plants, earlier studies did not evaluate the possible contribution of their homologs to hybridization signals in Northern blot analysis. In this study, we have examined whether routine transcriptional analyses of a PR gene is of high reliability or not by isolating six highly similar Gns genes (TaGlb2a, TaGlb2b, TaGlb2c, TaGlb2d, TaGlb2e, and TaGlb2f) and characterizing their expression patterns in detail. While TaGlb2b was shown to be a PR-2 gene, transcription of TaGlb2c and TaGlb2d was not induced upon infection with either powdery mildew (Erysiphe graminis) or head blight (Fusarium graminearum) pathogens; their transcripts were most abundant in healthy spikes (lemmas and in particular paleae). Therefore, in some cases, the conventional analyses do not necessarily provide accurate information on expression pattern of a PR gene in hexaploid wheat. This is also the first report of wheat genes that are specifically expressed in lemma/palea tissues of flowering spikelets.

Blasticidin S deaminase gene from Aspergillus terreus (BSD): a new drug resistance gene for transfection of mammalian cells.

Blasticidin S deaminase (BSD) is a drug inactivating enzyme produced by Aspergillus terreus, which convert blasticidin S (BS) to a non-toxic deamino-hydroxy derivative. The BSD gene was fused to SV 40 transcriptional regulatory elements and the resulting vector was used to transfect FM3A cells. Expression of BSD conferred resistance to BS and allowed efficient isolation of integrative transfectants which have stably maintained the BS-resistance phenotype after repeated transfer to fresh selective medium. The frequency of transfection was comparable to that with neo and about 80-times greater than with bsr, a BS-resistance gene of bacterial origin which can be used to isolate efficiently transfectant HeLa cells. Using BSD as a selectable marker, we obtained several stable cell lines expressing the firefly luciferase gene. Four independent transfectants among the randomly selected 5 BS-resistance colonies exhibited detectable luciferase activity under the control of dexamethasone-inducible promoter in the expression vector. The successful application of BSD strongly suggests the usefulness of BS as a versatile selective reagent for introduction of cloned DNA sequences into mammalian cells.

Long-term endothelin receptor antagonist administration improves alterations in expression of various cardiac genes in failing myocardium of rats with heart failure.

BACKGROUND: We reported that long-term (3-month) treatment with the endothelin (ET) type A (ET(A)) receptor antagonist BQ-123 markedly improved survival in rats with chronic heart failure (CHF). However, it is not known whether long-term treatment with an ET receptor antagonist improves alterations in the expression of cardiac genes in failing hearts. METHODS AND RESULTS: CHF rats and control sham-operated rats were treated with BQ-123, SB209670 (ET(A/B) dual receptor antagonist), or saline (vehicle) for 3 months. The survival of CHF rats was markedly higher in the BQ-123 or SB209670 treatment group than in the saline treatment group. The changes in the gene expression of classic molecular markers for failing hearts (mRNA levels of atrial natriuretic peptide and beta-myosin heavy chain) were greatly inhibited by BQ-123 or SB209670 treatment in CHF rats. Long-term BQ-123 treatment also normalized the alterations in the expression of functional molecular markers in failing hearts (eg, mRNA levels of ryanodine receptor, sarcoplasmic reticulum Ca(2+)-ATPase, angiotensin-converting enzyme, angiotensin II type 1 receptor, and prepro-ET-1). CONCLUSIONS: We demonstrated for the first time that long-term (3-month) treatment with an ET receptor antagonist improves the alterations in the expression of various cardiac genes of classic molecular markers (eg, mRNA in atrial natriuretic peptide and beta-myosin heavy chain) and of functional molecular markers (eg, mRNA levels of ryanodine receptor, sarcoplasmic reticulum Ca(2+)-ATPase, angiotensin-converting enzyme, angiotensin II type 1 receptor, and prepro-ET-1) in the failing hearts of CHF rats, suggesting that the great improvement of survival in CHF rats by an ET blocker is partly attributed to the prevention of molecular changes in failing hearts.

Novel molecular mechanism of increased myocardial endothelin-1 expression in the failing heart involving the transcriptional factor hypoxia-inducible factor-1alpha induced for impaired myocardial energy metabolism.

BACKGROUND: Hypoxia-inducible factor (HIF)-1alpha is an important transcriptional factor that activates the gene expression of glycolytic enzymes, which are activated as compensation for impaired beta-oxidation of fatty acid in the failing heart. We reported that cardiac endothelin (ET)-1 expression is markedly increased in heart failure. The mechanism, however, is unknown. Because we found an HIF-1alpha binding site in the 5'-promoter region of the ET-1 gene, we hypothesized that HIF-1alpha is involved in this mechanism. METHODS AND RESULTS: In rat cardiomyocytes, luciferase assay and electrophoretic mobility shift assay showed that HIF-1alpha transcriptionally activates ET-1 gene expression by direct interaction with the predicted DNA binding site in the 5'-promoter region. HIF-1alpha mRNA and ET-1 mRNA in the failing heart increased during the aggravation of heart failure in vivo in animal models, ie, rats with myocardial infarction and hamsters with cardiomyopathy. In cultured cardiomyocytes treated with a mitochondrial inhibitor, HIF-1alpha mRNA and ET-1 mRNA were markedly increased with activated glycolysis, and antisense oligonucleotide for HIF-1alpha largely inhibited the increased gene expression of ET-1. CONCLUSIONS: The present study revealed a novel molecular mechanism of upregulation of myocardial ET-1 in heart failure, indicating that induction of HIF-1alpha to stimulate glycolysis as an adaptation in heart failure against impaired energy metabolism alternatively causes an elevation of cardiac ET-1 gene expression as a maladaptation.

Pulmonary hypertension caused by congestive heart failure is ameliorated by long-term application of an endothelin receptor antagonist. Increased expression of endothelin-1 messenger ribonucleic acid and endothelin-1-like immunoreactivity in the lung in congestive heart failure in rats.

OBJECTIVES: The purpose of this study was to investigate whether 1) endothelin-1, a potent vasoconstrictor peptide, is involved in progression of pulmonary hypertension caused by congestive heart failure (CHF); and 2) whether long-term treatment with BQ-123, an endothelin receptor antagonist, ameliorates pulmonary hypertension caused by CHF. BACKGROUND: Congestive heart failure accompanies pulmonary hypertension, and the severity of pulmonary hypertension is an important determinant of prognosis. Although we reported that production of endothelin-1 is increased in the failing heart in rats with CHF, it is not known whether production of endothelin-1 in the lung is altered by CHF. METHODS: Congestive heart failure was induced by coronary artery ligation in rats. Expression of preproendothelin-1 messenger ribonucleic acid (mRNA) in the lung and kidney was determined. Endothelin-1 staining (immunoreactivity) in the lung was studied by immunohistochemical analysis. Effects of long-term BQ-123 treatment on the rats were studied. RESULTS: Two weeks postoperatively, CHF accompanied by pulmonary hypertension developed in the rats (CHF rats). Expression of preproendothelin-1 mRNA in the lung was markedly higher in the CHF rats than in the sham-operated rats, whereas that in the kidney did not differ between the two groups. Endothelin-1 staining on the pulmonary vascular endothelial cells was more intense in the CHF rats. BQ-123 treatment over a 2-week period in the CHF rats greatly reduced right ventricular systolic pressure and central venous pressure, but it did not affect blood pressure or left ventricular contractility (peak positive first derivative of left ventricular pressure) in these rats. CONCLUSIONS: Long-term BQ-123 treatment greatly ameliorated pulmonary hypertension in the CHF rats. The present study suggests that endothelin-1 plays an important role in the progression of pulmonary hypertension caused by CHF and that an endothelin receptor antagonist may be a new therapeutic agent for CHF-induced pulmonary hypertension.

Response to isoproterenol as a prognostic indicator of evolution from hypertrophic cardiomyopathy to a phase resembling dilated cardiomyopathy.

OBJECTIVES: We sought to assess whether isoproterenol stress echocardiography could detect in advance in which patients hypertrophic cardiomyopathy would progress to a phase resembling dilated cardiomyopathy. BACKGROUND: In a few patients, hypertrophic cardiomyopathy has been reported to progress to a phase characterized by systolic dysfunction and left ventricular dilation, resembling dilated cardiomyopathy. METHODS: Echocardiograms were recorded before and immediately after intravenous infusion of isoproterenol (0.02 microgram/kg body weight per min) for 5 min in 18 patients with typical hypertrophic cardiomyopathy (i.e., hypertrophied, hyperdynamic and nondilated) to determine the difference in fractional shortening. The patients were categorized into those with a good response (difference in fractional shortening > 7%, 14 patients) and those with a poor response (difference < or = 7%, 4 patients). Changes in left ventricular end-diastolic diameter and fractional shortening were evaluated by using serial echocardiography over an average follow-up period of 5.4 years. RESULTS: In the good response group, neither end-diastolic diameter nor fractional shortening changed significantly during the follow-up period. In the poor response group, end-diastolic diameter significantly increased from a mean +/- SD of 41 +/- 5 to 53 +/- 5 mm (p < 0.05), and fractional shortening significantly decreased from 40 +/- 12% to 29 +/- 10% (p < 0.05). All patients in the poor response group showed a substantial decrease (> or = 5%) in fractional shortening and an increase (> or = 5 mm) in end-diastolic diameter. One patient developed congestive heart failure due to systolic dysfunction during the observation period. CONCLUSIONS: The present study confirmed that impaired responses to isoproterenol infusion are related to future deterioration of left ventricular performance in patients with typical hypertrophic cardiomyopathy.

Electrophysiological actions of mexiletine on isolated rabbit atria and canine ventricular muscle and purkinje fibres.

The electrophysiological effects of varying concentrations of mexiletine on isolated rabbit sinus node and atrial preparations. Purkinje fibres, Purkinje-fibre-ventricular muscle junction and distal conduction system ('gate') were studied in oxygenated Tyrode solution at 35 degrees C using standard microelectrode techniques. 'Therapeutic' concentrations of mexiletine (0.5 to 2.0 micrograms.cm-3) had little effect on the atrial action potentials or on sinus node automaticity but sino-atrial conduction was delayed. In Purkinje fibres, these concentrations of the drug depressed the dV/dtmax of phase 0, conduction velocity and membrane responsiveness accompanied by significant shortening of the action potential duration and the effective refractory period, the change in the former always being greater than that in the latter. Mexiletine decreased antegrade as well as retrograde conduction at the Purkinje-fibre-ventricular muscle junction, while producing a uniform shortening of the action potential duration in the distal conduction system without a preferential effect at the 'gate'. The mechanisms of antiarrhythmic actions of mexiletine are thus complex but resemble those of lignocaine in superfused isolated cardiac muscle.

Electrophysiological effects of verapamil.

Previous studies have demonstrated that verapamil possess potent anti-arrhythmic effects. The present study has been designed to define the cardiovascular effects of this drug. Isolated tissue studies performed in rabbit right atrium demonstrated that prompt and prominent slowing of the sinus rate even at a dose of 1 X 10(-7) mol . litre-1. This dose produced significant decrease in action potential amplitude and phase 4 slope, shifted the 'threshold potential' to a less negative value, prolonged action potential duration but did not change maximum diastolic potential. At this dose of verapamil, sinoatrial conduction time prolonged significantly (control: 40.0 +/- 4.8 ms; 1 X 10(-7) mol . litre-1 verapamil: 50.0 +/- 6.4 ms). Purkinje fibre studies demonstrated decreases in dV/dt, resting potential, total amplitude, action potential duration at 75, 95% of recovery and effective refractory period only after exposure to greater than or equal to 1 X 10(-5) mol . litre-1 verapamil. Electrophysiological studies in conscious dogs demonstrated, after bolus administration of verapamil, progressive increases in the A-H interval and heart rate, but no changes in H-V and QRS intervals. Anaesthetised dog studies showed the lack of significant effect on A-H and H-V intervals or QRS duration regardless of the bolus dose of verapamil. However, verapamil produced statistically significant increases in heart rate after 0.025 mg . kg-1. Verapamil administration did not produce a statistically significant change in escape pacemaker rate in vagal stimulation experiments or with spontaneously beating isolated Purkinje fibres. Finally, the effect of increasing intravenous bolus does of verapamil on ischaemic arrhythmias was studied in five conscious dogs 24 h following LAD ligation. Only one dog with ventricular tachycardia and another dog with junctional escape rhythm were converted to sinus rhythm after the 0.05 mg . kg-1 and 0.2 mg . kg-1 doses, respectively. In conclusion, these studies demonstrated that administration of verapamil specifically depresses tissue with electrophysiological dependence on slow channel current. Therefore, sinus and A-V nodal events would be suppressed and slow-channel mediated events in ischaemic ventricle also would be inhibited. Clinically, acute administration of verapamil would lead to depression of sinus and A-V nodal function as well as potentially eliminate slow current mechanisms in ischaemic arrhythmias.

Mating-type genes from asexual phytopathogenic ascomycetes Fusarium oxysporum and Alternaria alternata.

Mating-type (MAT) loci were cloned from two asexual (mitosporic) phytopathogenic ascomycetes, Fusarium oxysporum (a pyrenomycete) and Alternaria alternata (a loculoascomycete), by a polymerase chain reaction (PCR)-based strategy. The conserved high mobility group (HMG) box domain found in the MAT1-2-1 protein was used as a starting point for cloning and sequencing the entire MAT1-2 idiomorph plus flanking regions. Primer pairs designed to both flanking regions were used to amplify the opposite MAT1-1 idiomorph. The MAT1-1 and MAT1-2 idiomorphs were approximately 4.6 and 3.8 kb in F. oxysporum and approximately 1.9 and 2.2 kb in A. alternata, respectively. In both species, the MAT1-1 idiomorph contains at least one gene that encodes a protein with a putative alpha box domain and the MAT1-2 idiomorph contains one gene that encodes a protein with a putative HMG box domain. MAT-specific primers were used to assess the mating type of F. oxysporum and A. alternata field isolates by PCR. MAT genes from A. alternata were expressed. The A. alternata genes were confirmed to be functional in a close sexual relative, Cochliobolus heterostrophus, by heterologous expression.

Characterization of heparin-induced osteopenia in rats.

Heparin has been known to induce osteopenia, but its precise mechanism of action is unknown. In the present study, we examined the effect of heparin on the rat femur using single photon absorptiometry and characterized the osteopenia biochemically and pharmacologically. Daily heparin injection dose dependently induced osteopenia in rats. Significant bone loss was observed from 2 weeks after starting heparin treatment (2000 U/kg.day) and peaked at 4 weeks. Serum PTH levels were significantly elevated from 1 week onward after starting heparin treatment, whereas no significant changes were seen in serum total calcium or ionized calcium levels. A bone resorption inhibitor, FR78844 (a bisphosphonate compound), significantly attenuated the heparin-induced osteopenia, as did 1 alpha-hydroxyvitamin D3; with the latter, the effective dose was 10 times lower than that needed for a similar effect against immobilization and ovariectomy-induced osteopenia, suggesting an up-regulation of 1 alpha, 25-dihydroxyvitamin D3 receptors in the heparin-treated rats. This speculation was supported by the finding that serum 1 alpha, 25-dihydroxyvitamin D levels were significantly decreased by 54% in the heparin-treated rats compared to those in normal rats. These results suggest that the enhanced bone resorption by high PTH blood levels and the reduction of 1 alpha, 25-dihydroxyvitamin D are involved in the pathogenesis of heparin-induced osteopenia.

Renal dopamine-1 receptors in hypertensive inbred rat strains with and without hyperactivity.

Renal dopamine-1 (DA-1) receptors are involved in the regulation of sodium transport in several nephron segments, including the proximal convoluted tubule (PCT). DA-1 receptors in the PCT and cortical collecting duct of normotensive rats are linked to the stimulation of adenylyl cyclase (AC). We have reported a defect in the DA-1 receptor/AC coupling in the PCT of the spontaneously hypertensive rat (SHR) of the Okamoto-Aoki strain. Hyperactivity and hypertension are both expressed in the SHR. To determine if the DA-1 receptor coupling defect is associated with hyperactivity or hypertension, we studied the DA-1 receptor in the PCT of two new inbred rat strains derived from the SHR: the hyperactive WKHA and the hypertensive WKHT rat. Tail-cuff blood pressures taken at 4 weeks indicated that WKHT rats were not hypertensive (86 +/- 3 mm Hg, n = 6), whereas at 12 weeks systolic pressures in both SHR and WKHT rats exceeded 150 mm Hg. Hyperactivity, however, was noted in WKHA rats even at this early age. Basal AC activity was similar in WKHA and WKHT PCT in either age group. In the older rats, the DA-1 agonist fenoldopam (10(-7) mol/L) stimulated AC activity in WKHA (70.6 +/- 16.1 fmol per 3 mm PCT per 20 minutes, n = 3) but not in WKHT PCT (43.3 +/- 5.3 fmol per 3 mm PCT per 20 minutes, n = 4). Gpp(NH)p (10(-5) mol/L), a nonhydrolyzable GTP analogue, stimulated AC activity to a similar extent in WKHA and WKHT PCT.(ABSTRACT TRUNCATED AT 250 WORDS)

Dopamine D1A receptors and renin release in rat juxtaglomerular cells.

Two dopamine D1-like receptors have been cloned from mammals, the D1 and D5 receptors, also known as D1A and D1B receptors, respectively, in rodents. Although D1-like receptors are known to stimulate renin release, the receptor subtype mediating this action has not been determined. We investigated D1 receptor subtype expression in rat juxtaglomerular cells obtained after enzymatic dispersion of kidney cortex and differential centrifugation. Juxtaglomerular cells in primary culture were immunocytochemically 85% to 95% renin positive. These cells expressed the D1A but not the D1B receptor (mRNA and protein). D1-like receptor function was demonstrated by a concentration-dependent stimulation of cAMP production by dopamine (n = 5-9 per group). Fenoldopam, a D1-like receptor agonist, also caused a concentration-dependent increase in cAMP production and renin secretion that was blocked by the selective D1-like receptor antagonist SCH23390 (n = 4-13 per group). Although the D1 ligands do not distinguish between the cloned D1-like receptors, the actions of fenoldopam were due to occupancy of the D1A receptor: (1) the D1B receptor, the only other mammalian D1-like receptor, is not expressed in juxtaglomerular cells; (2) antisense but not sense D1A oligonucleotides completely blocked the stimulatory effect of fenoldopam on cAMP production and renin secretion. We conclude that there is selective dopamine receptor gene expression in juxtaglomerular cells; the dopamine receptor subtype linked to the stimulation of cAMP and renin secretion in juxtaglomerular cells is the D1A subtype.

The mystery of the trichothecene 3-O-acetyltransferase gene. Analysis of the region around Tri101 and characterization of its homologue from Fusarium sporotrichioides.

The trichothecene 3-O-acetyltransferase gene, Tri101, plays a pivotal role for the well-being of the type B trichothecene producer Fusarium graminearum. We have analyzed the cosmids containing Tri101 and found that this resistance gene is not in the biosynthetic gene cluster reported so far. It was located between the UTP-ammonia ligase gene and the phosphate permease gene which are not related to trichothecene biosynthesis. These two 'house-keeping' genes were also linked in Fusarium species that do not produce trichothecenes. The result suggests that the isolated occurrence of Tri101 is attributed to horizontal gene transfer and not to the reciprocal translocation of the chromosome containing the gene cluster. Interestingly, 3-O-acetylation was not always a primary self-defensive strategy for all the t-type trichothecene producers; i.e. the type A trichothecene producer Fusarium sporotrichioides did not acetylate T-2 toxin in vivo although the fungus possessed a functional 3-O-acetyltransferase gene. Thus Tri101 appears to be a defense option which the producers have independently acquired in addition to their original resistance mechanisms.

Highly sensitive hepatitis B surface antigen detection by measuring stable nitroxide radical formation with ESR spectroscopy.

In areas where hepatitis B virus (HBV) is prevalent, HBV carriers negative for hepatitis B surface antigen (HbsAg) by enzyme-linked immunosorbent assay (ELISA) have been reported. Moreover, even after screening donor blood for HbsAg and hepatitis B core antibody (HBcAb), post-transfusion hepatitis B continues to occur, though with a decreasing frequency. Therefore, screening tests far more sensitive for detecting HBsAg than those currently available are needed. We developed a highly sensitive method for HBsAg detection. It is based on the recognition of peroxidase activity through measuring the formation of stable nitroxide radical with electron spin resonance (ESR) spectroscopy in the presence of hydrogen peroxide, p-acetamidophenol (p-AP), and 4-hydrazonomethyl-1-hydroxy-2,2,5,5,-tetramethyl-3-imidazoline-3-o xide (HHTIO). A cut-off value was established by testing of 186 healthy adults and 50 HBsAg-positive individuals. The signal to noise (S/N) ratio of less than 1.488 obtained by ESR spectroscopy was considered to be negative and more than 2.181, positive. The p-AP/HHTIO method was found to be 10 times more sensitive than the standard ELISA and reproducibility was excellent. Additional investigations were made on the HBsAg levels in the serum from 26 healthy subjects, in whom cut-off index levels on ELISA were negative but relatively high (range: 0.6 to 1.0); and on 15 patients with non B non C hepatitis. Three of 26 cases and 3 of 15 with non B non C hepatitis were judged to be HBsAg positive. Of these, 5 were found to be positive for HBV DNA by polymerase chain reaction (PCR). It was shown in this study that the p-AP/HHTIO method is practical and useful in screening HBV carriers because of the sensitivity in HBsAg detection, which is comparable to PCR analysis.

Protective action of a calcium antagonist, nilvadipine, against aortic calcium deposition--a pathogenic factor in atherosclerosis.

Nilvadipine and other calcium antagonists were studied for their effect on 1-alpha-hydroxyvitamin D3 (1 - alpha (OH)D3)-induced aortic calcium deposition in rats. The animals were treated orally with 1-alpha (OH)D3 (10 micrograms/kg) for 2 weeks. Calcium antagonists were given orally twice a day during the same period. The aortic calcium content in 1-alpha (OH)D3-treated rats increased to about 100 times that in the control. Nilvadipine reduced the aortic calcium deposition dose-dependently, with percent inhibition of 6, 43, 72 and 92%, at doses of 0.1, 1, 10 and 100 mg/kg, respectively. Similar activities were obtained for the other calcium antagonists except diltiazem which had no effect even at the largest dose of 100 mg/kg. According to the ED50 values, nilvadipine (2.2 mg/kg) was more potent than nifedipine (23.2 mg/kg), nicardipine (12.4 mg/kg) and verapamil (32.0 mg/kg). Scanning and transmission electron microscopy showed clear-cut degenerative changes in the endothelial cells after 1-alpha (OH)D3 treatment. Nilvadipine exerted a protective effect against these degenerative changes but not against 1-alpha (OH)D3-induced hypercalcemia. Furthermore, the drug had only minimal effect on in vitro calcification of the aorta. Our findings suggest that nilvadipine inhibits aortic calcification by protecting the aortic wall cells.

Smooth muscle cell migration induced by inflammatory cell products and its inhibition by a potent calcium antagonist, nilvadipine.

The chemotactic activities of inflammatory cell products for rat aortic smooth muscle cells (SMC) were examined in modified Boyden chambers. A checker board analysis revealed that interleukin-1 (IL-1), leukotriene B4 (LTB4), platelet-derived growth factor (PDGF) and inflammatory exudate from zymosan-activated air pouches stimulated chemotaxis of SMC. The chemotaxis, irrespective of the attractants used, was strongly inhibited by nilvadipine, a potent calcium antagonist, and the IC50 values were around 1 x 10(-10) M. Removal of extracellular calcium abolished the chemotactic activities of the attractants. These results suggest that inflammatory cells such as macrophages and polymorphonuclear leukocytes (PMN) have an important role in the migration of SMC into the intima during atherogenesis, and that nilvadipine might be useful for preventing and treating atherosclerosis.

Role of inflammatory responses in initiation of atherosclerosis: effects of anti-inflammatory drugs on cuff-induced leukocyte accumulation and intimal thickening of rabbit carotid artery.

Immediately after a cuff-sheathing of rabbit carotid artery, a large number of leukocytes adhered to injured endothelium then infiltrated into the media. These inflammatory responses were followed by an atherosclerotic change, intimal thickening, of the artery. A simultaneous injection of dexamethasone (10 mg/kg i.m.) inhibited the leukocyte accumulation by 74% when evaluated 18 h thereafter. Similarly, 39% inhibition was obtained with the same dose of FR110302, a potent 5-lipoxygenase inhibitor. On the other hand, the same dose of indomethacin, a cyclooxygenase inhibitor, had little effect on the leukocyte accumulation. The intimal thickening which was evaluated 3 weeks after the cuff-treatment was attenuated by a daily dose (10 mg/kg i.m.) of dexamethasone or FR110302 but not by one of indomethacin. The inhibition by the two former drugs were 91 and 58%, respectively. In vitro, the three drugs in concentrations up to 10 microM hardly affected endothelial adhesion of PMN which was induced by LPS or IL-1. Though 10 microM of FR110302 and indomethacin significantly decreased PMN chemotaxis induced by LTB4, the decreases were less than that at 10 microM dexamethasone. These results confirm a possible linkage between inflammation and atherosclerosis, and suggest that 5-lipoxygenase products contribute to the initiation and development of atherosclerosis.