Elevated 1-h plasma glucose following 75-g oral glucose load is a predictor of arterial stiffness in subjects with normal glucose tolerance.

AIMS: The study aimed to investigate arterial stiffness in subjects with normal glucose tolerance. METHODS: BMI, systolic blood pressure, fasting plasma glucose, lipid variables, ankle-brachial pressure index and brachial-ankle pulse wave velocity were measured in 2059 subjects from Takasaki city, located approximately 100 km north of Tokyo in Japan. Following a 75-g oral glucose tolerance test, only subjects with normal glucose tolerance were selected. RESULTS: One-hour post-challenge plasma glucose levels were correlated with brachial-ankle pulse wave velocity values (r = 0.340, P < 0.0001). When subjects with normal glucose tolerance were divided into three groups-group 1 (1-h plasma glucose < 8.56 mmol/l, n = 1595), group 2 (1-h plasma glucose ≥ 8.56 and < 10.17 mmol/l, n = 334) and group 3 (1-h plasma glucose ≥ 10.17 mmol/l, n = 130)-the brachial-ankle pulse wave velocity of group 3 (1473 ± 322 cm/s) was significantly higher than that of group 2 (1355 ± 252 cm/s) and brachial-ankle pulse wave velocity of group 2 was also significantly higher than that of group 1 (1275 ± 212 cm/s). CONCLUSIONS: We have identified that, in normal glucose tolerance, arterial stiffness is advanced in subjects with higher 1-h post-challenge plasma glucose in spite of the normal range for BMI, systolic blood pressure, fasting plasma glucose and lipid variables. Higher 1-h plasma glucose level is a risk factor for arterial stiffness in normal glucose tolerance.

Impaired blood rheology in critically ill patients in an intensive care unit.

Critically ill patients are at increased risk of thromboembolic complications. Japanese patients admitted to the intensive care unit of Gunma University Hospital were divided into critically ill (high score) and moderately ill (low score) groups according to mean Acute Physiology and Chronic Health Evaluation (APACHE) II score. White blood cell count, potassium, creatinine, immunoglobulin G and blood passage time, measured using the microchannel method, were significantly higher and the platelet aggregation score and platelet count were significantly lower in the high-score group than in the low-score group, but other haemorheological parameters did not differ significantly between the two groups. White blood cell count, potassium, creatinine, APACHE II score and levels of immunoglobulins G, A and M were positively correlated with blood passage time in all patients. Critically ill patients had impaired blood rheology, which could result from increased white blood cell count, potassium, creatinine and immunoglobulins and may be associated with the pathophysiology of the thromboembolic process.

A simple and reproducible cerebral thrombosis model in rats induced by a photochemical reaction and the effect of a plasminogen-plasminogen activator chimera in this model.

In this study a new model of cerebral ischemia, based on a middle cerebral artery (MCA) thrombosis in rats is described. Furthermore, the effect of the novel plasminogen activator (SUN9216), a plasminogen-plasminogen activator chimera, comprising the fibrin kringle 1 domain of a plasminogen, and the two kringles, and the serine protease domains of wild-type tissue plasminogen activator (t-PA), including a modification of the mannose glycosylation site on the kringle 1 of t-PA (PK1de1FE1X), was studied in this model. In the newly described model of thrombotic cerebral ischemia, an occlusive thrombus occurred usually within 8 min in the MCA as a consequence of an endothelial injury subsequent to a photochemical reaction between a systemically administered photosensitive dye (rose bengal) and a transillumination of the MCA with a high-intensity green light with a wavelength of 540 nm. The study was quantitated by means of pathological examination of the MCA and the brain. A platelet-rich thrombus was observed in the MCA using electron microscopical analysis based on ion beam bombardment. At 24 hr after induction of the thrombus, the brain was removed from 13 control animals, nine coronal sections were stained from each brain with triphenyltetrazoliumchloride (TTC), and the ischemic area was quantitated. A constant area of infarction was observed in the cortex and the lateral part of the basal ganglia. In a second group (n = 8), at 1 or 8 weeks after induction of the thrombosis in the MCA, the coronal sections were stained with hematoxylin and eosin.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardioprotective profile of MET-88, an inhibitor of carnitine synthesis, and insulin during hypoxia in isolated perfused rat hearts.

3-(2,2,2-trimethylhydrazinium) propionate (MET-88) is an inhibitor of carnitine synthesis. This study was carried out to investigate whether or not reduction of carnitine content could attenuate hypoxic damage in isolated perfused rat hearts. Rats were divided into four groups: 1) vehicle control; 2) pretreatment with MET-88 (MET-88); 3) application of insulin (500 muU/mL) in the perfusate (insulin); and 4) pretreatment with MET-88 and application of insulin (MET-88 + insulin). MET-88 (100 mg/kg) was orally administered once a day for 10 days until the day before the experiments. Hearts were initially perfused for a 10 min period under normoxia, followed by a 30 min period under hypoxia. Hearts were frozen at the end of hypoxia for the measurement of high-energy phosphates, carnitine derivatives, and glycolysis intermediates. In a separate series of untreated and MET-88 treated hearts, exogenous glucose and palmitate oxidation was measured. MET-88 decreased the extent of the depression of cardiac contractility (+dP/dt), and aortic flow during the hypoxic state. Insulin also improved cardiac function, and co-treatment of MET-88 and insulin additionally improved cardiac function during hypoxia. MET-88 prevented the decrease of high-energy phosphate and the increase of long-chain acylcarnitine after 30 min of hypoxic perfusion. In addition, MET-88 increased the steady state of glucose oxidation in hypoxic perfused rat hearts. These results indicate that MET-88 has cardioprotective effects on contractile function and energy metabolism of isolated perfused rat hearts in a hypoxic condition. Preventing the accumulation of long-chain acylcarnitine may serve to protect hypoxic hearts.

Normalization of reproductive function in germfree mice following bacterial contamination.

The capacity for reproduction in germfree mice remain inferior to their conventional counterparts even after improvement of feed and other such rearing conditions. The authors provide evidence of increased reproductive capacity in germfree mice following association with bacteria. Estrous cycles were normalized in female mice accidentally contaminated with bacteria, and in mice given fecal suspensions of the accidentally contaminated mice per os. Significant rises were seen in their copulation and implantation rates, reaching levels comparable to values in conventional mice. In male mice, bacterial contamination induced significant increase in sperm motility. Bacteria were identified in the feces of the contaminated mice, and reproductive capacity was examined in mice associated with the identified bacteria. As a result, normalization of the estrous cycle, and rises in copulation and implantation rates were noted in B. distasonis and C. perfringens di-associated mice. Values from B. subtilis mono-associated mice were comparable to those in germfree mice. These results from our accidental contamination indicate that B. distasonis and C. perfringens are capable of normalizing estrous cycles in female germfree mice, and in increasing their reproductive capacity by raising their rates of copulation and implantation.

Development of a CAI program entitled 'Introduction to Nursing Process'. Requirement for nursing education in Japan.

A new teaching strategy is required in nursing education in Japan. A multimedia CAI (Computer Assisted Instruction) program entitled "Introduction to Nursing Process." was developed. The aim of this study was to examine whether CAI could assist students' knowledge acquisition, and increase their motivation and readiness to learn clinical nursing practice. This study involved the process of developing CAI and a pilot study for measuring and evaluating the CAI program. Results demonstrate that CAI can increase students' understanding about the nursing process, enhance their motivation to learn, and provide realistic situations describing the concept of nursing the process.

Uncoated observation and etching of non-conductive materials by ion beam bombardment in scanning electron microscopy.

In a scanning electron microscope, techniques of uncoated observation and subsequent observation of internal and intracellular structures in nonconductive materials removed by ion beam etching have been successfully obtained. An apparatus in which a beam of argon ions is collimated and focused by an electrostatic lens onto an appropriate target and in addition, the specimens are both cooled and rotated during processing to avoid artifacts and thermal damages has been used. Organic and biological specimens can be directly observed without coating of metallic films by pre-bombardment of argon ions. High resolution images of structural details eroded by ion beam etching and coupled with tungsten sputter coating have been clearly observed. The role of angle of incidence of ion beam and accelerating voltage in determining the optimum etching condition without artifacts are experimentally obtained for soft biological specimens as well as hard materials. Step and general views of internal structures in soft biological specimens as well as ceramics and polyvinyl chlorides eroded by cleaning and subsequent etching are revealed without artifacts and thermal damages.

Dissociation of photoreceptor cells from the pineal organ of the lamprey, Lampetra japonica.

Photoreceptor cells, nerve cells and supporting cells were dissociated from the pineal organ of the river lamprey, Lampetra japonica, by the use of 10 U/ml papain solution at 28 degrees C for 20 min, followed by repeated trituration. With the aid of Nomarski interference-contrast optics, photoreceptor cells, nerve cells and supporting cells were readily identified. Electron-microscopic examination revealed that isolated photoreceptor cells display an outer segment endowed with a few lamellar disks and connected to the inner segment (ellipsoid) via a connecting cilium. The structural features of the dissociated photoreceptor and supporting cells strongly resemble the morphology of the respective cellular elements in situ. We succeeded in culturing dissociated cells for time periods up to 48 h when the procedure described in detail was applied.

Dynamic reaction in a homogeneous HDL-cholesterol assay visualized by electron microscopy.

BACKGROUND: Measurement of HDL-cholesterol (HDL-C) by homogeneous assays with automated analyzers is replacing precipitation methods. However, in this reaction-type assay, interactions between the reagents and lipoproteins remain unknown. METHODS: Electron microscopy was used to investigate the reactions in a homogeneous HDL-C assay. Negative staining with 10 g/L uranyl acetate was performed for lipoprotein visualization by electron microscopy. Observations of the interactions between lipoproteins and the reagents of a polyanion-polymer/detergent assay were achieved by cooling the reaction mixture in ice water. This treatment also allowed observation of the time course of the reaction. RESULTS: In the first-reagent reaction (polyanion-polymer), every lipoprotein aggregated almost completely. In the second-reagent reaction (enzymes and detergent), only HDL in the lipoprotein aggregates was selectively resolved and reacted enzymatically. Reagent 1 contains two important substances: polyanion and synthetic polymer. Using x-ray microanalysis, we confirmed that aggregation of lipoproteins in the first reaction occurred through interaction with the phosphotungstate of the polyanion. CONCLUSION: Electron microscopy morphologically revealed the dynamic reaction in a homogeneous HDL-C assay.

Alterations of glomerular basement membrane relevant to haematuria.

To elucidate the morphological basis of glomerular haematuria, morphometric analysis of the glomerular basement membrane (GBM) and lamina densa (LD) was performed on silver impregnated samples for electron microscopy. The cases studied consisted of 3 groups: group A, normal controls, being from donors for kidney transplantation; group B, haematuric; and group C, non-haematuric cases with isolated proteinuria. Qualitative analysis revealed that gap formation, splitting, segmental and diffuse thinning of the GBM occur preferentially in haematuric cases. The morphometry of the GBM and LD yielded increased mean values of the GBM and of LD thickness in groups B and C. The coefficient of variation (CV, SD/mean) for the GBM and LD, however, was the highest in group B among the 3 groups, suggesting the most irregular GBM and LD in group B. In addition, CV was significantly higher in cases with splitting, segmental attenuation and gap of the GBM than cases without. The findings suggest that the irregularity of the GBM rather than its mean thickness is clearly associated with splitting and ultimately with haematuria via the gaps produced.

Digital processing methods using scanning densitometer and microcomputer for the structural analysis of a scanning electron micrograph.

The fundamental equation of image contrast with practical resolving power is shown taking account of the optical and specimen characteristics. Then, the structural factor which is used to indicate the scattering from the unit of structure to form the periodic object in a scanning electron microscopy can be derived straight-forwardly by digital processing. Based on the Fourier analysis, the reconstruction and subsequent interpretation of structural images such as biological specimens of rat microvilli have been successful. These microvilli must exhibit regularity of a hexagonal lattice consisting of rod-like crystalline arrays. Lattice defects are analysed by means of contrast enhancement, level slicing and histogram plotting techniques. The resolution of SEM can be tested from the granularity distance of coated films by the image sharpening by differentiation and subsequent superimposition of reconstructed images. Furthermore, the structural details in high brightness owing to reducing of the brightness level and filtering the high frequency component have been analysed.

Effects of MET-88, a gamma-butyrobetaine hydroxylase inhibitor, on tissue carnitine and lipid levels in rats.

MET-88, 3-(2,2,2-trimethylhydrazinium) propionate, suppresses carnitine synthesis by inhibiting (gamma-butyrobetaine hydroxylase. The purpose of this study was to clarify the effects of suppression of carnitine synthesis on carnitine and lipid contents in tissues. MET-88 (50, 100, 200 or 400 mg/kg/d) was administered orally to male SD rats for 10, 30 or 60 d. Total carnitine and lipid (triglycerides, non-esterified fatty acids) contents were measured in heart and liver. In both tissues, treatment with MET-88 dose-dependently decreased total carnitine levels, and the reduction reached the plateau state after 30 d at each dose. MET-88 had no effect on lipid content in the heart, but increased the lipid content in the liver at the highest doses. Treatment with MET-88 at 400 mg/kg for 60 d resulted in no pathologic findings in the histological study, and also had no effect on parameters of liver function such as glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase as judged from the results of blood biochemical analysis. We concluded that long-term treatment with MET-88 decreased the carnitine content to a constant level in both heart and liver, but had no effect on lipid contents in the heart, although it affected lipid metabolism in the liver.

Beneficial effect of MET-88, a gamma-butyrobetaine hydroxylase inhibitor, on energy metabolism in ischemic dog hearts.

The effect of MET-88 [3-(2, 2, 2-trimethylhydrazinium) propionate], a gamma-butyrobetaine hydroxylase inhibitor, on the ischemic changes of energy metabolism was studied in the anesthetized dog. In the dog pretreated orally with MET-88 (50, 100 or 200 mg/kg/day) or placebo for 10 days, the left anterior descending coronary artery was occluded for 60 min, and the myocardium was taken from the left anterior descending coronary area (ischemic area) and left circumflex area (nonischemic area) for metabolic analysis. In the ischemic area, occlusion of the left anterior descending coronary artery decreased the tissue levels of adenosine triphosphate, adenosine diphosphate and creatine phosphate, increased the tissue levels of adenosine monophosphate and lactate, and decreased the value of the energy charge potential. These metabolic alterations, induced by occlusion of the left anterior descending coronary artery, were dose-dependently attenuated by MET-88. In the nonischemic area, MET-88 did not markedly change either the tissue levels of energy metabolites or the value of the energy charge potential. These results indicate that MET-88 attenuates the derangement of the energy metabolism in the ischemic myocardium, without affecting the energy metabolism in the nonischemic myocardium.

Relationship between triglyceride concentrations and LDL size evaluated by malondialdehyde-modified LDL.

BACKGROUND: Hypertriglyceridemia is associated with decreased HDL-cholesterol (HDL-C) and increased small dense LDL. In addition, small dense LDL is known to be susceptible to oxidation. METHODS: We measured LDL particle size, using gradient gel electrophoresis, and malondialdehyde-modified LDL (MDA-LDL), using an ELISA, and investigated the association between triglyceride (TG) concentrations, LDL size, and MDA-LDL. RESULTS: TG concentrations correlated negatively with the predominant LDL size (r = -0.650) and HDL-C concentration (r = -0.556). The relationship between TG concentration and LDL size, evaluated by measuring MDA-LDL, distinguished subgroups derived from four subfractions of TG concentrations and four distribution ranges of LDL size. These experiments indicated that there is a threshold for oxidation susceptibility at an LDL size of 25.5 nm and a TG concentration of 1500 mg/L. To investigate the relationship between LDL size, MDA-LDL concentration, and other lipids (TGs, HDL-C, apolipoprotein B, and total cholesterol), we evaluated them in control subjects and patients with diabetes mellitus or hypertriglyceridemia. When the size range for normal LDL was postulated to be 25.5 < or = phi (LDL diameter) < 26.5 nm, the MDA-LDL concentration was significantly higher in the subgroups of patients with LDL in the size range 24.5 < or = phi < 25.5 nm compared with patients with normal LDL. This result also suggests that the threshold is at a LDL size of 25.5 nm. CONCLUSION: The threshold for oxidation susceptibility coincided with the point of LDL size separation between the LDL subclass patterns A and B as an atherosclerotic risk.

TAS-301, an inhibitor of smooth muscle cell migration and proliferation, inhibits intimal thickening after balloon injury to rat carotid arteries.

The purpose of this study was to determine the efficacy and the possible mechanism of action of a recently synthesized drug, TAS-301 [3-bis (4-methoxyphenyl)methylene-2-indolinone], on intimal formation in comparison with those of tranilast, the clinical efficacy of which was reported earlier. Rat carotid arteries were injured using a balloon catheter. Neointimal thickening, measured 14 days after injury, was reduced by the oral administration of TAS-301 in a dose-dependent fashion (3-100 mg/kg), and the effect of TAS-301 at a dose of 100 mg/kg was significantly greater than that of tranilast (300 mg/kg). Fewer cells were found on the intima of balloon-injured arteries of TAS-301-treated rats than on arteries of tranilast-treated rats. In an in vitro assay, TAS-301 inhibited the migration of smooth muscle cells (SMCs) stimulated by platelet-derived growth factor-BB, insulin-like growth factor-1 or heparin-binding epidermal growth factor-like growth factor. In addition, TAS-301 and tranilast reduced the proliferation of medial and intimal SMCs at 4 and 8 days, respectively, after the injury. In vitro, TAS-301 inhibited basic fibroblast growth factor-induced proliferation of SMCs dose dependently. These findings indicate that TAS-301 shows a higher inhibitory potency on intimal formation than tranilast due to inhibition of both migration of medial SMCs and proliferation of medial and intimal SMCs. Our results suggest that further evaluation of TAS-301 as an inhibitor of postangioplasty intimal thickening is warranted.

Inhibitory effect of TAS-301, a new synthesized constrictive remodeling regulator, on renarrowing after balloon overstretch injury of porcine coronary artery.

The purpose of this study was to determine the efficacy and the possible mechanism of action of a recently synthesized drug, TAS-301 [3-bis(4-methoxyphenyl)methylene-2-indolinone], on stenosis after balloon overstretch injury of porcine arteries. We measured the diameter of vessels by angiography and conducted histological analysis. The oral administration of TAS-301 kept dilated the angiographic luminal diameter of injured segment 4 weeks after overstretch injury and reduced calculated stenosis ratio in a dose-dependent manner, significantly reducing it at doses of 30 and 100 mg/kg. Histopathological analysis showed that TAS-301 significantly reduced the adventitial area at doses of 30 and 100 mg/kg with moderate reduction of the neointimal area, resulting in the larger residual lumen. In an in vitro assay, TAS-301 dose dependently inhibited the proliferation of adventitial fibroblasts stimulated by basic fibroblast growth factor or transforming growth factor-beta(1). In addition, the drug reduced adventitial fibroblast-mediated three-dimensional collagen gel contraction. These findings indicate that TAS-301, the first compound developed for targeting the constrictive remodeling, showed a high inhibitory potency on coronary artery stenosis of micropigs after injury, mainly due to inhibition of adventitial fibroblast proliferation and of the contractile ability of myofibroblasts. Our results suggest the strong possibility that TAS-301 may be efficacious for prevention of restenosis after angioplasty and the need to examine the therapeutic usefulness of this drug in clinical trials.

Inhibitory effects of bovine lactoferrin on the adherence of enterotoxigenic Escherichia coli to host cells.

Adherence is an essential and prerequisite step for the colonization of mucosal surfaces by enterotoxigenic Escherichia coli (ETEC). We studied the effect of bovine lactoferrin (BLF) on the adherence of ETEC to human epithelial cells in vitro, and to intestinal mucosa of ICR germfree mice in vivo. In the in vitro study, BLF was found to inhibit the adherence of ETEC. This adhesion-inhibiting activity of BLF was found to lessen with decreasing BLF concentration, but the data obtained suggest a positive inhibitory effect of BLF against the adhesion of ETEC cells. In the in vivo study, the counts of adherent bacteria in various sections of the intestinal tract (duodenum, jejunoileum, and large intestine) were lower in the BLF group than in the control group, suggesting the possible action of BLF as an intestinal tract adherence-blocking agent with regards to ETEC.