High-Fructose Diet-Induced Hypertriglyceridemia Is Associated With Enhanced Hepatic Expression of ACAT2 in Rats.

High levels of fructose induce hypertriglyceridemia, characterized by excessive levels of triglyceride-rich lipoproteins such as very low-density lipoprotein (VLDL); however, the underlying mechanisms are poorly understood. The aim of this short communication was to examine hepatic changes in the expression of genes related to cholesterol metabolism in rats with hypertriglyceridemia induced by high-fructose or high-glucose diets. Rats were fed a 65 % (w/w) glucose diet or a 65 % (w/w) fructose diet for 12 days. Serum levels of triglycerides, total cholesterol, and VLDL+LDL-cholesterol, hepatic levels of triglycerides and cholesterol, and ACAT2 expression at the gene and protein levels were significantly higher in the fructose diet group compared to the glucose diet group. The hepatic levels of Abcg5/8 were lower in the fructose group than in the glucose group. Serum high-density lipoprotein (HDL)-cholesterol and hepatic expression levels of Hmgcr, Ldlr, Acat1, Mttp, Apob, and Cyp7a1 did not differ significantly between groups. These findings suggest that high-fructose diet-induced hypertriglyceridemia is associated with increased hepatic ACAT2 expression.

[Critical role for cell cycle regulators in androgen receptor function].

Androgen plays an important role in the growth of prostate cancer, but the molecular mechanism that underlies development of resistance to antiandrogen therapy remains unknown. Cyclin E has now been shown to increase the transactivation activity of the human androgen receptor (AR) in the presence of its ligand dihydrotestosterone. The enhancement of AR activity by cyclin E was resistant to inhibition by the antiandrogen 5-hydroxyflutamide. Cyclin E was shown to bind directly to the AB domain of the AR, and to enhance its AF-1 transactivation function. These results suggest that cyclin E functions as a coactivator of the AR, and that aberrant expression of cyclin E in tumors may contribute persistent activation of AR function, even during androgen ablation therapy.

First synthesis of a alpha-trifluoromethyl allenol ether via the Julia-Lythgoe process.

Alpha-trifluoromethyl allenol ethers 9a-e were prepared in moderate to good yields by the Julia-Lythgoe process using beta-ethoxy-beta-trifluoromethyl vinylic sulfone 3. Several reactions of 9c were examined to give alpha,beta-unsaturated trifluoromethyl ketone derivatives 11 and 12.

Hologram simulation for off-axis electron holography

Hologram simulation for electron holography using an electron biprism is described. An electron hologram is superimposed by Fresnel fringes originating from the electron biprism, which affects both the amplitude and the phase of the object wave and the reference wave. In this simulation, we consider the effects of Fresnel diffraction as well as the electron-wave phase shift due to the electromagnetic field produced by the specimen. We also take into account the phase shift due to the inner potential of the specimen, the amplitude modulation due to the absorption of the incident electrons by the specimen, reference-wave distortion caused by the electromagnetic fields, coherency of the electron wave, and quantum noise of the detected electrons. Simulated and experimentally obtained holograms and reconstructed images are compared for the cases of a charged latex spherical particle and a single magnetic-domain spherical particle placed on a carbon film.

Improvement of light collection efficiency of lens-coupled YAG screen TV system for a high-voltage electron microscope.

A new lens coupling television (TV) system using a YAG (Yttrium Aluminum Garnet: Y(3)Al(5)O(12) : Ce(3+)) single crystal screen has been developed for a high-voltage electron microscope (HVEM), and its performance is examined. The system, using a combination of YAG and lenses, is less damaged by high-energy electron irradiation and reduces the influence of X-rays on the image. YAG screens have not been used for lens-coupling systems, because the high refractive index (n = 1.84) of YAG results in a low light collection efficiency for emitted light. This disadvantage is overcome by combining a thin YAG disk screen (thickness; 100 microm) with a glass hemisphere whose refractive index is 1.81. We found that the light intensity is almost the same as that obtained with a conventional P22 powder screen and lenses system. The resolution is about 55 microm on the YAG screen, and this value is 1.3 times higher than that measured by the conventional system. Shading and distortion do not affect TV observation. Detection quantum efficiency, obtained after correction of the channel mixing effect, is about 0.1.

High precision phase-shifting electron holography

Today's information-oriented society requires high density and high quality magnetic recording media. The quantitative observation of fine magnetic structures by electron holography is greatly anticipated in the development of such new recording materials. However, the magnetic fields around particles <50 nm have not been observed, because the fields are too weak to observe in the usual way. Here we present a highly precise phase measurement technique: improved phase-shifting electron holography. Using this method, the electric field around a charged polystyrene latex particle (100 nm in diameter) and the magnetic field around iron particles (30 nm in diameter) are observed precisely. A precision of the reconstructed phase image of 2pi/300 rad is achieved in the image of the latex particle.

Facile synthesis of stable and lectin-recognizable DNA-carbohydrate conjugates via diazo coupling.

Stable and lectin-recognizable DNA-carbohydrate conjugates were prepared by diazo coupling of lactose and cellobiose derivatives to fragmented salmon testes DNA. The diazo coupling is suggested to take place selectively to guanine bases since the amount of lactose moiety introduced was directly proportional to the G content of various DNAs with different G contents. According to the CD spectra, the conjugates bearing carbohydrate less than 25% content kept a typical B-type conformation similar to native DNA. The conjugates possessed higher melting temperature and stronger nuclease resistance both to exo- and endonucleases than native DNA. Gel shift assay and fluorescence binding assay showed that the DNA-lactose conjugates were specifically bound to galactose-specific lectin RCA(120) with strong binding affinity (Ka = 10(4)-10(5) M(-1)) due to glycoside cluster effect. This facile method will be a useful protocol of molecular design for cell-targeted gene therapy.

[Clinical role of cell cycle regulators in androgen-dependent cancer cell growth].

The functional and quantitative alterations in cell cycle regulators after androgen depletion in an androgen-dependent cancer cell and the interaction between androgen receptor and cell cycle regulators were examined in order to clarify the initial response of cancer cells to anti-androgen therapy. Fluorescence activated cell sorter analysis (FACS) of androgen-dependent cancer cell line (SC-3) cells cultured with or without 1 nM dihydrotestosterone (DHT) revealed that suppression of cell growth after androgen withdrawal was due to G1 arrest. The protein level of cyclin D1 decreased without any apparent change in the amounts of Cdk2, Cdk4, cyclin E or cyclin A. Among various Cdk inhibitors (CKIs) examined, p27 was upregulated at both mRNA and protein levels 24 h after androgen depletion. On the other hand, cyclin E has been shown to increase the transactivation activity of the human androgen receptor (AR) in the presence of DHT. These results suggest that cell cycle regulators are critical targets in the initial response of androgen-dependent cancer cells to androgen depletion and play a key role in the transcriptional activity of AR.

Oxygen-derived free radical-induced vasoconstriction by thromboxane A2 in aorta of the spontaneously hypertensive rat.

This study was performed to clarify the mechanism of vasoconstriction induced by oxygen-derived free radicals in spontaneously hypertensive rats. The isometric tension of aortic rings from spontaneously hypertensive rats and Wistar-Kyoto rats was measured in Krebs-Henseleit solution. Oxygen-derived free radicals were generated by mixing xanthine and xanthine oxidase. The removal of endothelium enhanced the contractions induced by oxygen-derived free radicals. The inhibition of nitric oxide production with NG-nitro-L-arginine methyl ester (10(-4) M) enhanced the contractions. Treatment with the thromboxane A2 (TXA2) synthetase inhibitor OKY-046 (10(-4) M) or RS-5186 (10(-4) M) markedly reduced the contractions. Treatment with the cyclooxygenase inhibitor indomethacin (10(-5) M) and a TXA2/prostaglandin H2 (PGH2) receptor antagonist, ONO-3708 (10(-6) M), completely abolished the oxygen-derived free radical-induced contractions. In contrast, treatment with the PGI2 synthetase inhibitor tranylcypromine (10(-4) M) did not attenuate the oxygen-derived free radical-induced contractions. Whether endothelium was present or not, the release of TXB2, PGE2, and 6-keto-PGF1alpha, but not PGF2alpha, was increased by the production of oxygen-derived free radicals. Catalase and the hydroxyl radical scavenger deferoxamine plus mannitol markedly inhibited the oxygen-derived free radical-induced contractions. These results suggest that oxygen-derived free radical-induced vasoconstriction in spontaneously hypertensive rat aorta is caused by TXA2 and PGH2 released in smooth muscle.

Increased oxysterol contents in diabetic rat hearts: their involvement in diabetic cardiomyopathy.

BACKGROUND: Abnormal lipid metabolism associated with diabetes mellitus has been proposed to be involved in the pathogenesis of diabetic cardiomyopathy. Oxysterols, oxidation derivatives of cholesterol, are known to be highly cytotoxic. OBJECTIVE: To monitor changes in myocardial oxysterols and to assess the effect of probucol, a lipid lowering agent, on myocardial lipids and oxysterols in diabetic rats. METHODS AND RESULTS: Streptozotocin-induced diabetic rats were divided into two groups; one group was put on a standard diet, and the other a diet containing 1% (weight/weight) probucol for eight weeks. Two oxysterols, 7 beta-hydroxycholesterol and 7-ketocholesterol, were identified in myocardium by capillary gas chromatography. Both 7 beta-hydroxycholesterol and 7-ketocholesterol were significantly increased in diabetic rats (49.9 +/- 9.4 ng/mg dry weight versus 5.8 +/- 1.2 in controls, P < 0.05; and 5.3 +/- 1.2 ng/mg dry weight versus 1.7 +/- 0.1 in controls, P < 0.01, respectively). Probucol reduced not only plasma lipids but also myocardial lipids except for cholesterol and sphingomyelin fractions. However, probucol did not improve insulin deficiency, glucose metabolism or myocardial oxysterol contents. CONCLUSIONS: This study demonstrated an increase in oxysterols in the myocardium of diabetic rats, suggesting that oxysterols may play a role in the development of diabetic cardiomyopathy. Probucol did not decrease the myocardial oxysterol content at the dose used in this study, suggesting that the increase in oxysterols may not be attributed to high circulating concentrations of lipids, but rather to disturbed myocardial metabolism due to insulin deficiency.

Improved insulin sensitivity by bezafibrate in rats: relationship to fatty acid composition of skeletal-muscle triglycerides.

We investigated the effect of the lipid-lowering agent, bezafibrate, on insulin sensitivity in a dietary model of insulin resistance. Male Sprague-Dawley rats were divided into four groups: control group, administered a standard diet; high-fructose group, given a 40% fructose diet; high-fructose plus lard group, given a 40% fructose diet with 7% lard; and bezafibrate group, given a 40% fructose plus 7% lard diet with 10 mg x kg-1 x day-1 of oral bezafibrate. Insulin action was assessed after 2 weeks with a steady-state plasma glucose (SSPG) level. The fatty acid (FA) composition of skeletal-muscle triglycerides was also determined. A higher SSPG level (20.9 +/- 0.9 vs. 16.5 +/- 1.1 mmol/l in the control group, P < 0.05) as well as a higher systolic blood pressure (120 +/- 2 vs. 101 +/- 2 mmHg, P < 0.01) was observed in the high-fructose plus lard group, but not in the high-fructose group. These changes were prevented by bezafibrate administration. The FA composition of skeletal-muscle triglycerides demonstrated a higher percentage of saturated and monounsaturated FAs (P < 0.01) and a lower percentage of polyunsaturated FAs (P <0.01) in the high-fructose plus lard group versus the control group. These changes were consistent with differences in the dietary intake of FAs. Bezafibrate virtually normalized the FA composition in the high-fructose plus lard group. The ratio of C20:4 to C20:3, an index of delta5 desaturase activity, was significantly higher in the bezafibrate group versus the high-fructose plus lard group (8.60 +/- 0.76 vs. 2.04 +/- 0.27, P < 0.01). In conclusion, the dietary FA composition was closely related to insulin resistance in rats fed 40% fructose. Bezafibrate increased delta5 desaturase activity. Such action may contribute to the improvement of insulin sensitivity.

Role of K+ channels in EDHF-dependent relaxation induced by acetylcholine in canine coronary artery.

To identify the K+ channels responsible for endothelium-derived hyperpolarizing factor (EDHF)-dependent relaxation, we studied the effects of various K+ channel blockers on acetylcholine-induced relaxation, which persists even in the presence of both an inhibitor of nitric oxide synthase and that of cyclooxygenase, in canine coronary artery rings. A nonselective K+ channel blocker, tetrabutylammonium (TBA), a large and intermediate conductance Ca2+-activated K+ channel blocker, charybdotoxin (CTX), and a voltage-dependent K+ channel blocker, 4-aminopyridine (4-AP), significantly inhibited this residual relaxation. A combined treatment with CTX and 4-AP almost completely blocked the relaxation. Neither a large (iberiotoxin) nor a small (apamin) conductance Ca2+-activated K+ channel blocker blocked the relaxation. We also investigated effects of K+ channel blockers on basal tone to determine whether or not EDHF is involved in regulating basal tone. TBA and CTX substantially raised basal tone to a greater degree in endothelium-intact preparations than in endothelium-denuded preparations. These results indicate that EDHF may exert its relaxing action through intermediate conductance Ca2+-activated and voltage-dependent K+ channels in canine coronary arteries. In addition, EDHF may play a role in maintaining basal vascular tone.

XTEM sample preparation technique for n-type compound semiconductors using photochemical etching.

A sample preparation technique based on photochemical etching (PCE) is described for cross-sectional transmission electron microscopy (XTEM) of n-type compound semiconductors. XTEM samples of an InGaAsP/InP single-layer structure, prepared by using a moderately focused laser beam and Br2-methanol solution, gave high quality, damage-free XTEM images. The PCE technique is applicable to other n-type compound semiconductors.

Time courses of cell electroporation as revealed by submicrosecond imaging of transmembrane potential.

Changes in the membrane conductance of sea urchin eggs, during the course of electroporation, were investigated over the time range of 0.5 microsecond to 1 ms by imaging the transmembrane potential at a submicrosecond resolution with the voltage-sensitive fluorescent dye RH292. When a rectangular electric pulse of moderate intensity was applied across an egg, a position-dependent potential developed synchronously with the pulse, as theory predicts for a cell with an insulating membrane. From the rise and fall times, the membrane capacitance of unfertilized eggs was estimated to be 0.95 microF/cm2 and the intracellular conductance 220 omega.cm. Under an electric pulse of much higher intensity, the rise of the induced potential stopped at a certain level and then slowly decreased on the microsecond time scale. This saturation and subsequent reversal of the potential development was ascribed to the introduction of finite membrane conductance, or permeabilization of the membrane, by the action of the intense pulse (electroporation). Detailed analysis indicated the following: already at 0.5 microsecond in the rectangular electric pulse, the two sides of the egg facing the positive and negative electrodes were porated and gave a high membrane conductance in the order of 1 S/cm2; the conductance on the positive side appeared higher. Thereafter, the conductance increased steadily, reaching the order of 10 S/cm2 by 1 ms. This increase was faster on the negative-electrode side; by 1 ms the conductance on the negative side was more than twice that on the positive side. The recovery of the porated membrane after the pulse treatment was assessed from the membrane conductance estimated in a second electric pulse of a small amplitude. At least two recovery processes were distinguished, one with a time constant of 7 microseconds and the other 0.5 ms, at the end of which the membrane conductance was already < 0.1 S/cm2.

Generation of mutagenicity by ozonation of humic substances' components.

Components of humic substances, such as vanillin, syringaldehyde, vanillic acid and di-n-butylphtalate, were ozonated and subjected to the mutagenicity assay using Salmonella typhimurium TA 98 and 100 with and without S9 mix. The strong mutagenic activity was found on all components except di-n-butylphtalate by strain TA 100 with and without S9 mix. Substances with strong mutagenic activity in ozonated vanillin were water-soluble and were slightly extracted with benzene, dichloromethane and ethyl acetate. Following gel chromatography on Sephadex G-10, the strong mutagens generated by ozonation were found with molecular weights greater than 300.