Properties of the static NMR response of a confined thin nematic film of 5CB-d2 under crossed electric and magnetic fields: theory and experiments.

This work describes an investigation of the static (or quasistatic) nuclear magnetic resonance (NMR) response in a nematic liquid crystal confined between two planar conducting plates and subject to a magnetic field and an electric field produced by a difference of voltage applied on the plates. Deuterium NMR spectroscopy of 4-pentyl-d(2)-4'-cyanobiphenyl (5CB-d(2)) under these conditions has revealed a voltage dependent inhomogeneous director distribution for a particular narrow range of voltages and for a fixed magnetic field (that of the spectrometer). In the ideal setup the two plates are assumed to be rigorously parallel, so that a difference of voltage applied on the plates leads to a constant electric field normal to them. When the magnetic field is parallel to the plates (orthogonal geometry) there exists a threshold value of the electric field for which the effect of both fields exactly compensate; moreover, for stronger electric field the director aligns with the electric field while for weaker electric field the director aligns with the magnetic field. If there is a lack of parallelism between the two plates, the electric field becomes inhomogeneous so that it may be larger than the threshold value in some region of the sample and smaller in the remaining part of the sample. In that case the director will adopt essentially two orientations within the sample, namely, parallel or perpendicular to the magnetic field, and the position of the frontier between the two domains depends on the voltage. This feature is clearly shown by deuterium NMR spectra that exhibit a transfer of intensity between two quadrupolar doublets with increase in the applied voltage. The coexistence of two director populations occurs for a range of voltages that depends on the degree of nonparallelism; accordingly, an estimation of this range by NMR yields an experimental estimation of the lack of parallelism. A tiny tilt of the magnetic field (nonorthogonal geometry) entrains a notably different behavior since a single doublet with voltage dependent splitting is observed in this case. In a first stage (simple model) of this work, the main features observed for the orthogonal and nonorthogonal geometries are interpreted within the framework of Leslie-Ericksen theory by employing the concept of a single effective field replacing the two real fields. However, the spectra reveal an additional director distribution, especially for the orthogonal geometry, that cannot be interpreted by this simple approach. In a second stage (advanced model), these less clear features have been investigated by numerical simulations of a two-dimensional model which includes the effects of inversion walls and of the high relative dielectric anisotropy of 5CB.

Field-induced periodic distortions in a nematic liquid crystal: deuterium NMR study and theoretical analysis.

The peculiarities in the dynamic of the director reorientation in a liquid crystal (LC) film under the influence of the electric E field directed at an angle α to the magnetic B field have been investigated both experimentally and theoretically. Time-resolved deuterium NMR spectroscopy is employed to investigate the field-induced director dynamics. Analysis of the experimental results, based on the predictions of hydrodynamic theory including both the director motion and fluid flow, provides an evidence for the appearance of the spatially periodic patterns in 4-n-pentyl-4'-cyanobiphenyl LC film, at the angles α>60∘, in response to the suddenly applied E. These periodic distortions produce a lower effective rotational viscosity. This gives a faster response of the director rotation than for a uniform mode, as observed in our NMR experiment.

Field-induced alignment of the nematic director: studies of nuclear magnetic resonance spectral oscillations in the limit of fast director rotation.

Time-resolved NMR spectroscopy is a powerful method to investigate field-induced rotation of the director in a nematic liquid crystal. The method requires that the director does not rotate significantly during the acquisition of the free induction decay and hence the NMR spectrum. We have extended the method to systems where this is not the case and the observed NMR spectra are now found to contain novel oscillatory features. To understand these oscillations, we have developed a model combining both director and spin dynamics. In addition to increasing the information content of the time-resolved NMR spectra, it also proves possible to determine the field-induced relaxation time from a single spectrum.

Director alignment by crossed electric and magnetic fields: a deuterium NMR study.

The static director distribution in thin nematic liquid crystal cells, subject to both electric and magnetic fields, has been investigated using a combination of deuterium nuclear magnetic resonance (NMR) spectroscopy and continuum theory in terms of the director distribution function, which gives the probability density for finding the director at a given orientation. A series of deuterium NMR spectra for the nematic liquid crystal, 4-pentyl-d(2)-4'-cyanobiphenyl deuteriated in the α position of the pentyl chain were acquired as a function of the applied electric field. This powerful experimental technique allowed us to observe uniform and nonuniform director alignment depending on the angle between the two fields and their relative strength. On the basis of the detailed experimental results, we have explored the factors that influence the nature of both the uniform and the nonuniform director distributions. We have discussed the questions that are raised by our attempt to understand the static director distribution as a function of the angle between the two fields. We have discovered that the alignment of the director at the surface of the Teflon spacers is essential in addition to the random variation in the cell thickness in order to account for the static director distribution determined from the NMR spectra.

Electric-field driven director oscillations in a nematic liquid crystal: a NMR investigation.

We have investigated the oscillatory behavior of the nematic director for 4-pentyl-4'-cyanobiphenyl (5CB) when it is subjected to a static magnetic field and a sinusoidal electric field. In these experiments the two fields were inclined at about 50 degrees and the frequency of the electric field was varied from several hertz to approximately 1000 Hz. The director orientation was measured using time-resolved deuterium NMR spectroscopy since this has the advantage of being able to determine the state of director alignment in the sample. In fact, for all of the frequencies studied the director is found to remain uniformly aligned. Since the diamagnetic and dielectric anisotropies are both positive the director oscillates in the plane formed by the two fields. These oscillations were observed to continue for many cycles, indicating that the coherence in the director orientation was not lost during this motion. The maximum and minimum angles made by the director with the magnetic field were determined, as a function of frequency, from the NMR spectrum averaged over many thousand cycles of the oscillations. At low frequencies (several hertz) these limiting angles are essentially independent of frequency but as the frequency increases the two angles approach each other and become equal at high frequencies, typically 1000 Hz. Our results are well explained by a hydrodynamic theory in which the sinusoidal time dependence of the electric field is included in the torque-balance equation. This analysis also shows that, for a range of frequencies between the high and low limits, these NMR experiments can give dynamic as well as static information concerning the nematic phase.

Effect of diadenosine tetraphosphate (AP4A) on coronary arterial microvessels in the beating canine heart.

Diadenosine tetraphosphate (AP4A) can be released from activated platelets and the present study examined its effect on coronary arterial microvessels. The role of purinoceptors in the coronary microcirculation in vivo was also investigated. In open chest dogs, coronary arterioles were observed using a microscope with a floating objective. In Protocol 1, AP4A (1, 10, 100 and 1,000 micromol/L) was superfused onto the heart surface before and during the superfusion of 10 micromol/L of 8-phenyltheophylline (8-PT), a P1 purinoceptor blocker. In Protocol 2, AP4A (0.1, 1, 10, and 100 nmol x kg(-1) x min(-1)) was infused into the left anterior descending coronary artery before and during the superfusion of 10 micromol/L of 8-PT. In addition to 8-PT, 30 micromol/L of pyridoxalphosphate-6-azophenyl 2',4'-disulphonic acid (PPADS), a P2X purinoceptor blocker in Protocol 3, or 300 micromol/L of N(omega)-nitro-L-arginine (LNNA) in Protocol 4, was continuously superfused, and 4 doses of AP4A were cumulatively superfused as in Protocol 1. In Protocol 5, 10 micromol/L of alpha,beta-methylene ATP, an agonist of P2X purinoceptors, was superfused for 60 min. Superfused AP4A dilated arterioles in a dose-dependent manner. The magnitude of dilatation was greater in smaller arterioles (small vessel < or = 150 microm: 24.5+/-2.2% vs large vessel > 150 microm: 10.6+/-1.5% at a dose of 1,000 micromol/L, p<0.001). On the other hand, intraluminally applied AP4A also dilated arterioles, but no size dependency was shown. In the presence of 8-PT, vasodilatory responses to superfused and intraluminally applied AP4A were attenuated and the lower doses of AP4A constricted arterioles. This vasoconstrictor effect was not affected by PPADS. The vasodilatory effect of the higher doses of AP4A was almost abolished in the presence of LNNA. Alpha,beta-methylene ATP had no effect on coronary microvascular diameters. AP4A has bidirectional effects on coronary arterial microvessels: vasodilatory effects mediated by P1 purinoceptors and NO, which might be mediated by P2Y purinoceptors, and a vasoconstrictor effect, which is not mediated by P2X purinoceptors.

Role of pertussis toxin-sensitive G protein in metabolic vasodilation of coronary microcirculation.

We have previously demonstrated that pertussis toxin (PTX)-sensitive G protein (G(PTX)) plays a major role in coronary microvascular vasomotion during hypoperfusion. We aimed to elucidate the role of G(PTX) during increasing metabolic demand. In 18 mongrel dogs, coronary arteriolar diameters were measured by fluorescence microangiography using a floating objective. Myocardial oxygen consumption (MVO(2)) was increased by rapid left atrial pacing. In six dogs, PTX (300 ng/ml) was superfused onto the heart surface for 2 h to locally block G(PTX). In eight dogs, the vehicle (Krebs solution) was superfused in the same way. Before and after each treatment, the diameters were measured during control (130 beats/min) and rapid pacing (260 beats/min) in each group. Metabolic stimulation before and after the vehicle treatment caused 8.6 +/- 1. 8 and 16.1 +/- 3.6% dilation of coronary arterioles <100 microm in diameter (57 +/- 8 microm at control, n = 10), respectively. PTX treatment clearly abolished the dilation of arterioles (12.8 +/- 2. 5% before and 0.9 +/- 1.6% after the treatment, P < 0.001 vs. vehicle; 66 +/- 8 microm at control, n = 11) in response to metabolic stimulation. The increases in MVO(2) and coronary flow velocity were comparable between the vehicle and PTX groups. In four dogs, 8-phenyltheophylline (10 microM, superfusion for 30 min) did not affect the metabolic dilation of arterioles (15.3 +/- 2.0% before and 16.4 +/- 3.8% after treatment; 84.3 +/- 11.0 microm at control, n = 8). Thus we conclude that G(PTX) plays a major role in regulating the coronary microvascular tone during active hyperemia, and adenosine does not contribute to metabolic vasodilation via G(PTX) activation.

Effects of low doses of endothelin-1 on basal vascular tone and autoregulatory vasodilation in canine coronary microcirculation in vivo.

The plasma level of endothelin-1 (ET-1) increases in several cardiovascular disorders. The present study examined whether threshold doses of ET-1 affect vascular tone and autoregulatory vasodilation during a reduction in perfusion pressure in the coronary microcirculation in vivo. In anesthetized open-chest dogs, arterial microvessels in the epimyocardium were observed through a microscope equipped with a floating objective. In 6 dogs, ET-1 (10(-13) to 10(-8)mol/L) was superfused onto the epimyocardium in a cumulative fashion. In another set of dogs (n= 16), the perfusion pressure of the observed vascular bed was reduced to 60 mmHg (mild stenosis) and to 40 mmHg (severe stenosis) by a hydraulic occluder, and the microvascular responses were observed in the presence (n=9) or absence (n=7) of ET-1 (10(-12) or 10(-11) mol/L). ET-1 > or =10(-11) mol/L constricted coronary arterioles (< or =100 microm in diameter) and small arteries (>100 microm in diameter) in a dose-dependent fashion. ET-1 of 10(-12) mol/L affected neither the basal diameters nor the dilation of vessels during the pressure reduction. ET-1 of 10(-11) mol/L decreased the diameters of arterioles and small arteries before and during the mild and severe stenosis. However, ET-1 did not attenuate the percentage dilation of arterioles from the baseline in response to the mild and severe stenosis. The data indicates the following: (1) ET-1 at doses > or =10(-11) mol/L similarly constricts coronary arterioles and small arteries; (2) ET-1 at 10(-11) mol/L, which is slightly higher than the pathophysiological plasma level, increases the basal vascular tone, but does not attenuate the autoregulatory vasodilation of the coronary microcirculation.

Troglitazone suppresses cell growth of myeloid leukemia cell lines by induction of p21WAF1/CIP1 cyclin-dependent kinase inhibitor.

In a human eosinophilic leukemia cell line, EoL-1, cell proliferation was suppressed by 2-day treatment with troglitazone. EoL-1 cells treated with troglitazone were arrested and maintained in the G0/G1 phase in the cell cycle. This suppression correlated with the up-regulation of mRNA for p21WAF1/CIP1 cyclin-dependent kinase (Cdk) inhibitor. The inhibitory effects of troglitazone on cell proliferation and expression of p21 mRNA were observed in a human myelomonocytic cell line, U937, and a human myelomonoblastic cell line, KPB-M15. In addition, in EoL-1 cells, p21 protein was induced by troglitazone treatment and the induction was inhibited by protein synthesis inhibitor, cycloheximide. These data suggest that troglitazone inhibits cell proliferation in myeloid leukemia cell lines at least in part by induction of p21 Cdk inhibitor.

A human myeloid cell line producing stem cell growth factor, KPB-M15, secretes another growth factor active on murine hematopoietic progenitor cells.

Human stem cell growth factor (SCGF) produced by a myeloid cell line, KPB-M15, exhibits species-specific hematopoietic activities. However, KPB-M15-conditioned medium induced colony formation of mouse bone marrow cells. KPB-M15-derived colony-stimulating activity (CSA) was purified through Butyl-Toyopearl 650c and Cu2+ chelating-Sepharose 6B chromatography. TSK-G3000SW gel filtration of the purified preparation presented 3 distinct peaks around Vo, 150 kD and 85 kD. Gel fractions extracted from SDS-PAGE had macrophage colony-stimulating factor (M-CSF)-specific amino acid sequences. PCR, Northern hybridization and ELISA demonstrated that KPB-M15 cells secreted a significant amount of M-CSF and IL-6. Anti-M-CSF but not anti-IL-6 antibody abrogated CSA in KPB-M15-CM. IL-6 hardly synergized with M-CSF to enhance colony formation. Collectively, M-CSF is a sole CSA for murine hematopoietic progenitor cells in KPB-M15-CM. This is the first report of a human myeloid cell line, KPB-M15, constitutively producing M-CSF in addition to SCGF and IL-6. It can be useful in investigating the mechanism of production of M-CSF.

Cloning, expression, and characterization of a cDNA encoding a novel human growth factor for primitive hematopoietic progenitor cells.

Multiple growth factors synergistically stimulate proliferation of primitive hematopoietic progenitor cells. A human myeloid cell line, KPB-M15, constitutively produces a novel hematopoietic cytokine, termed stem cell growth factor (SCGF), possessing species-specific proliferative activities. Here we report the molecular cloning, expression, and characterization of a cDNA encoding human SCGF using a newly developed lambdaSHDM vector that is more efficient for differential and expression cloning. cDNA for SCGF encodes a 29-kDa polypeptide without N-linked glycosylation. SCGF transiently produced by COS-1 cells supports growth of hematopoietic progenitor cells through a short-term liquid culture of bone marrow cells and exhibits promoting activities on erythroid and granulocyte/macrophage progenitor cells in primary semisolid culture with erythropoietin and granulocyte/macrophage colony-stimulating factor, respectively. Expression of SCGF mRNA is restricted to myeloid cells and fibroblasts, suggesting that SCGF is a growth factor functioning within the hematopoietic microenvironment. SCGF could disclose some human-specific mechanisms as yet unidentified from studies on the murine hematopoietic system.

Mechanisms of coronary microvascular dilation induced by the activation of pertussis toxin-sensitive G proteins are vessel-size dependent. Heterogeneous involvement of nitric oxide pathway and ATP-sensitive K+ channels.

G proteins are critically important mediators of many signal transduction systems. In the present study, we investigated the effect of direct activation of pertussis toxin (PTX)-sensitive G protein (GPTX) on coronary arterial microvascular tone in 37 open-chest anesthetized dogs in vivo. Coronary arterial microvessels on the surface of the beating left ventricle were visualized by performing fluorescence coronary microangiography using an intravital microscope with a floating objective system. Microvessels were divided into two groups, small microvessels (inner diameter, < or = 130 microns) and large microvessels (inner diameter, > 130 microns). Topically applied mastoparan (G protein activator, 10, 30, and 100 mumol/L) produced homogeneous microvascular dilation in a concentration-dependent manner (10 mumol/L, 7.9 +/- 2.0%; 30 mumol/L, 10.3 +/- 2.4%; and 100 mumol/L, 16.7 +/- 4.5% in small microvessels; 10 mumol/L, 5.3 +/- 1.2%; 30 mumol/L, 9.8 +/- 2.5%; and 100 mumol/L, 15.5 +/- 3.9% in large microvessels). These dilations were reversed to constriction by pretreatment with PTX (300 ng/mL, 2 hours) in both microvessel groups. Blockade of nitric oxide production by NG-nitro-L-arginine (LNNA, 300 mumol/L) offset the mastoparan-induced dilation in large microvessels but not in small microvessels. Cosuperfusion of glibenclamide (10 mumol/L) with LNNA produced constriction of all sizes of microvessels in response to mastoparan, whereas charybdotoxin (10 nmol/L) did not affect the mastoparan effect. Pretreatment with glibenclamide alone reversed mastoparan dilation to constriction in small microvessels, whereas it only offset the dilation without producing constriction in large microvessels. We conclude that the activation of GPTX produces homogeneous coronary arterial microvascular dilation and that the underlining mechanisms of the dilation are vessel size dependent. The L-arginine-nitric oxide pathway mediates the dilation only in large microvessels, whereas ATP-sensitive K+ channel activation plays a central role in the dilation of small microvessels when GPTX is directly activated. ATP-sensitive K+ channels are also involved in the dilation of large microvessels in a synergistic fashion with nitric oxide production.

Vasodilatory effect of nicorandil on coronary arterial microvessels: its dependency on vessel size and the involvement of the ATP-sensitive potassium channels.

We aimed to clarify the size dependency of nicorandil-induced dilation in coronary microcirculation and the involvement of adenosine triphosphate (ATP)-sensitive potassium channels. Coronary arterial microvessels were observed through a microscope equipped with a floating objective in anesthetized open-chest dogs (n = 29). Heart rate and mean aortic pressure were maintained at control level. In 16 dogs, nicorandil was infused into the coronary in a cumulative fashion (0.1, 1.0, 10, and 100 micrograms/kg/min, for 5 min for each dose). In 13 dogs, glibenclamide (10 microM) was topically applied onto the observed area, and nicorandil was similarly infused. Nicorandil dilated vessels < 100 microns in diameter at all applied doses in a dose-dependent manner. Glibenclamide abolished the dilation of these vessels at the lower two doses. Vessels > 100 microns in diameter dilated only at the two higher doses and the dilation was not affected by glibenclamide. These data suggest that the vessels < 100 microns are more sensitive to this agent than other size vessels, and that ATP-sensitive potassium channels are involved in the nicorandil-induced dilation of vessels smaller than 100 microns, whereas the dilation of other size vessels occurs independently of this channel.

Effect of an ATP sensitive potassium channel opener, levcromakalim, on coronary arterial microvessels in the beating canine heart.

OBJECTIVE: The aim was to clarify the site in the coronary microcirculation that is dilated by an ATP sensitive potassium channel opener, levcromakalim, and to examine whether the magnitude of dilatation is size dependent. METHODS: Coronary arterial microvessels were observed through an intravital microscope equipped with a floating objective in beating canine left ventricles in situ. Flow velocity of the left anterior descending coronary artery was measured with a suction-type Doppler probe. Heart rate and aortic pressure were maintained at control levels throughout the experiments. Three doses of levcromakalim (0.01-1.0 microgram.kg-1.min-1) or a single dose (1.0 microgram.kg-1.min-1) were infused into the coronary artery in groups, with or without intracoronary glibenclamide pretreatment (200 or 400 micrograms.kg-1). The effect of levcromakalim on different sized vessels was assessed by dividing them into three groups according to control diameter (small, internal diameter < 100 microns; medium, > or = 100, < 200 microns; large, > or = 200 microns). RESULTS: The lowest dose of levcromakalim dilated only the small vessels. The two higher doses dilated vessels of all sizes, but the magnitude of dilatation was greater in the small vessel group than in the other two groups. Coronary resistance significantly decreased dose dependently during the infusion of 0.1 and 1.0 microgram.kg-1.min-1 of levcromakalim. Pretreatment with glibenclamide markedly attenuated the levcromakalim induced dilatation of all vessel groups and the reduction in coronary vascular resistance. CONCLUSIONS: Levcromakalim heterogeneously dilates coronary arterial microvessels via the opening of ATP sensitive potassium channels, and small vessels are more sensitive to levcromakalim.