Understanding of cell death induced by the constituents of Taxus yunnanensis wood.

The ethanol extract from the wood of Taxus Yunnanensis (TY) induced apoptosis in all cancer cell lines tested, which was mainly due to activation of an extrinsic pathway in human colon cancer DLD-1 cells. The extrinsic pathway was activated by the upregulation of the expression levels of Fas and TRAIL/DR5, which led to the activation of caspase-8. Of note, the machinery of this increase in expression was promoted by the upregulation of MIR32a expression, which silenced MIR34a-targeting E2F3 transcription factor. Furthermore, ectopic expression of MIR32a or siR-E2F3 silencing E2F3 increased Fas and TRAIL/DR5 expression. Thus, the extract activated the extrinsic pathway through the MIR34a/E2F3 axis, resulting in the autocrine and paracrine release of TRAIL, and upregulated expression of death receptors Fas and DR5 in the treated DLD-1 cells, which were functionally validated by Fas immunocytochemistry, and using anti-Fas and anti-TRAIL antibodies, respectively. In vivo, TY showed significant anti-tumor effects on xenografted and syngeneic model mice. The extract may also aid in chemoprevention by selectively making marked tumor cells susceptible to the tumor immunosurveillance system.

Plant hvu-MIR168-3p enhances expression of glucose transporter 1 (SLC2A1) in human cells by silencing genes related to mitochondrial electron transport chain complex I.

Diet is a crucial factor for preventing most diseases. Edible plant extracts are known to contain exosome-like nanoparticles, in which food-derived plant microRNAs are included and may serve as a novel functional component in human health. Here, we demonstrated that hvu-MIR168-3p included in the nanoparticles of rice aleurone cells down-regulated the expression of the genes related to mitochondrial electron transport chain complex I in human cells. Subsequently, hvu-MIR168-3p enhanced protein and RNA expression levels of glucose transporter I and caused a decrease in the blood glucose level, which findings were obtained by in vitro and in vivo experiments, respectively. These findings suggest that a cross-kingdom relationship between plants and humans with respect to hvu-MIR168-3p exists and may contribute to preventive medicine for GLUT1-related dysfunctions including glucose metabolism, aging, and tumor immunology.

Dehydroepiandrosterone sulfate and cytochrome P450 inducers alleviate fatty liver in male rats fed an orotic acid-supplemented diet.

The effects of the peroxisome proliferator, dehydroepiandrosterone sulfate (DHEAS), and the typical cytochrome P450 (CYP) inducers phenobarbital (PB) and 3-methylcholanthrene (3-MC) on fatty liver were examined in rats. Treating rats with orotic acid caused marked accumulation of lipid droplets in the liver. This effect of orotic acid was almost eradicated by co-treatment with DHEAS and PB. While DHEAS or PB alone also alleviated fatty liver, treatment with 3-MC caused little effect on a reduction in lipid droplets. Histopathological examinations revealed numerous peroxisomes in the liver of rats treated with DHEAS. In addition, a significant increase in the expression on hepatic CYPs was observed in rats the fatty liver of which was attenuated. Regarding other enzymes associated with hepatic fatty acid oxidation, the expression levels of sirtuin 1, sirtuin 6, and carnitine palmitoyltransferase 1 were also upregulated most markedly by treatment with DHEAS alone. Thus, the attenuation in fatty liver observed in the present study is likely due to peroxisome proliferation and the induction of fatty acid-metabolizing enzymes by DHEAS and typical CYP inducers.

Effects of tea catechins on the gastrointestinal mucosa in rats.

Although tea catechins are known to exert a potent antiulcer effect on the alimentary tract, there is scant information concerning their effects on normal mucus cell functions. Using original anti-mucin monoclonal antibodies, we studied the influences of long-term administration of catechins on the quantity and quality of mucin in rat gastrointestinal mucosa. Administration of 0.5% tea catechins significantly increased the mucin content of the ileum, but not the stomach. An enzyme-linked immunosorbent assay (ELISA) showed no remarkable qualitative changes in gastric mucin, but a selective increase and decrease in sulfo- and sialomucins, respectively, in the ileum of rats administered catechins. The ELISA results were consistent with both the immunohistochemical findings and the high-iron diamine-alcian blue staining pattern. These findings indicate that tea catechins modulate ileal mucin metabolism in the ileal mucosa, suggesting that further studies focusing on the ileal epithelium will assist in further elucidation of the mechanism of catechin effects.

Effect of tea catechins on body fat accumulation in rats fed a normal diet.

Although it is known that tea catechins exert potent effects in obese subjects, there is scant information concerning these effects on body weight gain and body fat accumulation in the non-obese. We studied normal rats fed a normal diet and water containing either 0.1% or 0.5% tea catechins to examine the effects on body fat content and serum cholesterol levels, as well as evaluating whether the effect is related to bile acids, which in recent years have emerged as an inducer of energy expenditure. The administration of 0.5% catechins decreased the accumulation of body fat and the serum levels of cholesterol and bile acids. These results indicate that tea catechins modulate lipid metabolism not only in obese subjects, but also in the non-obese.

Rapid determination of carbamate pesticides in food using dual counter-current chromatography directly interfaced with mass spectrometry.

Dual counter-current chromatography (dual CCC)-tandem mass spectrometry (MS/MS) was successfully performed with a newly designed spiral column for dual CCC. The small column capacity required for directly coupling with electrospray MS/MS was accomplished by forming a rectangular spiral groove on a plastic disk and sealing it with a PTFE sheet. This novel dual CCC-MS/MS technique was successfully applied for the rapid determination of methomyl, fenobucarb and carbaryl pesticides in food. A two-phase solvent system of n-hexane-acetonitrile-0.1% formic acid (45:45:10) was suitable for both good dual CCC separation and sufficient ionization of pesticides. Recoveries of these three pesticides from mandarin orange and spinach samples fortified at 0.05 mg/kg were in the range of 93-107% with relative standard deviations of 2.4-3.8%.

Eupalinin A isolated from Eupatorium chinense L. induces autophagocytosis in human leukemia HL60 cells.

Eupalinin A, a natural phytoalexin included in Eupatorium chinense L., exhibited a marked inhibitory effect on cell growth in HL60 cells. The morphological aspects of eupalinin A-treated cells evaluated by Hoechst 33342 nuclear staining indicated cell death, only a small part of which showed a typical apoptosis with nuclear fragmentation and condensation. To determine what type of cell death is caused by eupalinin A, we examined the contribution of caspases, Bcl-2 family proteins, MAP kinase, and PI3K/Akt, and mitochondrial membrane potential to this cell death. As a result, most part of the cell death was not associated with apoptosis because of caspase independence and no death factor released from mitochondria. Electron microscopic study indicated a characteristic finding of autophagy such as the formation of autophagosomes. Furthermore, the level of microctubule-associated-protein light chain 3 (LC3) II protein and monodancylcanaverin (MDC) incorporation were gradually increased with reduction of mitochondrial membrane potential by the accumulation of intracellular ROS after eupalinin A treatment. From these results, we can conclude that eupalinin A-induced cell death was mainly due to autophagy, which was initiated by increased ROS, resulting in the perturbation of mitochondrial membrane potential. Since the class III PI3K inhibitor such as 3-MA or LY294002 did not inhibit the eupalinin A-induced type II programmed cell death (PCD II), it was suggested that the PCD II was executed by Beclin-1 independent pathway of damage-induced mitochondrial autophagy (mitophagy).

Application of dual counter-current chromatography for rapid sample preparation of N-methylcarbamate pesticides in vegetable oil and citrus fruit.

Dual counter-current chromatography (dual CCC) has been successfully applied to rapid sample preparation for the simultaneous determination of residual carbaryl, fenobucarb and methomyl in vegetable oil and citrus fruit. The citrus fruit samples were extracted with n-hexane solution containing stable isotopically labeled internal standards (methomyl-d3, fenobucarb-d3 and carbaryl-d9), and applied to dual CCC using a two-phase solvent system of n-hexane-acetonitrile to purify the carbamate pesticides from aliphatic sample matrix. The coiled column was rotated at 420 rpm, the lower mobile phase was introduced through the head toward the tail, and the upper mobile phase in the opposite direction. Due to the high partition efficiency of dual CCC, the lower phase fraction collected from 2 to 5 min after injection could be subjected to flow-injection tandem mass spectrometry directly after concentration. Repetitive sample injection can be performed at high reproducibility without a risk of contamination from the compounds retained in the column.

Lovastatin inhibits tumor growth and lung metastasis in mouse mammary carcinoma model: a p53-independent mitochondrial-mediated apoptotic mechanism.

The effects of lovastatin, a potent inhibitor of hydroxymethylglutaryl-coenzyme A reductase, were studied in a mouse model of metastatic mammary cancer carrying a p53 mutation. Mice bearing mammary tumors, induced by inoculation of syngeneic BALB/c mice with BJMC3879 cells, were treated with lovastatin at 0, 25 and 50 mg/kg three times a week. Tumor volumes were significantly reduced in a dose-dependent manner throughout the 6 week study and were associated with both a decrease in DNA synthesis and an increase in apoptosis. The high dose of lovastatin also inhibited lung metastasis. In a corollary in vitro study, flow cytometric analyses of lovastatin-treated mammary cancer cells additionally showed cell cycle arrest at G1 phase and decreases in S and G2/M phases. Laser scanning cytometric analyses further demonstrated that cancer cells in S and G2/M were particularly susceptible to the effects of lovastatin. Transmission electron microscopic evaluation of TUNEL-confirmed apoptotic bodies in lovastatin-treated mammary carcinoma cells revealed many free 3'-OH ends of DNA in condensed chromatin within fragmented nuclei that occasionally assumed a characteristic half-moon shape. Consistent with initiation of apoptosis, cellular caspase-8, caspase-9 and caspase-3 activities were elevated in lovastatin-treated cells. The mitochondrial membrane potential was also decreased, with subsequent release of cytochrome c. However, lovastatin-induced cell death was significantly reduced by the broad spectrum caspase inhibitor z-VAD-fmk, as well as the caspase-9 inhibitor z-LEHD-fmk and the caspase-3 inhibitor z-DEVD-fmk, but not by the specific caspase-8 inhibitor z-IETD-fmk. Since immunoelectron microscopy showed translocation of Bax to the mitochondria in lovastatin-treated cells, lovastatin-induced apoptosis may, therefore, be ultimately dependent on Bax induction of cytochrome c release. These results suggest that lovastatin may be useful as an adjuvant therapy in breast cancers containing p53 mutations due to its ability to both suppress DNA synthesis and induce p53-independent mitochondria-mediated apoptosis.

Effectiveness of right or left radial approach for coronary angiography.

The transradial approach for catheterization is becoming increasingly more popular. At present, the choice of the right or left radial artery depends on the operator's preference. We examined how the laterality influenced the effectiveness of the approach. Employing Judkins-type catheters, we performed coronary angiography in 232 patients with the left approach and in 205 patients with the right approach. Although access time did not differ between the two groups of patients, the duration of catheter manipulation was shorter in the left- than in the right-approach group (11.7 +/- 5.9 vs. 9.8 +/- 4.4 min; P < 0.001). Because of the shorter duration of catheter manipulation, the total procedural duration was shorter in the left-approach group (13.7 +/- 6.4 vs. 11.4 +/- 4.8 min; P < 0.001). The fluoroscopy time was shorter in the left- than in the right-approach group (3.7 +/- 2.5 vs. 5.0 +/- 3.3 min; P < 0.001). The amount of contrast material did not differ between the groups (79 +/- 27 vs. 83 +/- 25 ml). The rate of guidewire usage to engage the coronary ostium was higher in the right- than in the left-approach group because of the severe tortuosity of the right subclavian artery (20/205 vs. 0/232; P < 0.001). Thus, for operators with significant experience, the left radial approach may provide increased procedural efficacy for coronary angiography compared to the right radial approach.