Dietary diacetylene falcarindiol induces phase 2 drug-metabolizing enzymes and blocks carbon tetrachloride-induced hepatotoxicity in mice through suppression of lipid peroxidation

Falcarindiol is a diacetylenic natural product containing unique carbon-carbon triple bonds. Mice were orally administrated falcarindiol (100 mg/kg), and drug-metabolizing and antioxidant enzymes were monitored in several tissues of mice. Treatment with falcarindiol was found to increase glutathione S-transferase (GST) and NAD(P)H: quinone oxidoreductase 1 activities in liver, small intestine, kidney, and lung. No changes were observed in cytochrome P450 (CYP) 1A known to activate procarcinogens. Western blot analysis revealed that various GST subunits including GSTA4, which plays an important role in the detoxification of alkenals produced from lipid peroxides, were induced in liver, small intestine, and kidney of falcarindiol-treated mice. Additionally, we investigated the protective effects of falcarindiol against hepatotoxicity induced by carbon tetrachloride (CCl(4)) and the mechanism of its hepatoprotective effect. Pretreatment with falcarindiol prior to the administration of CCl(4) significantly suppressed both an increase in serum alanine transaminase/aspartate transaminase (ALT/AST) activity and an increase in hepatic thiobarbituric acid reactive substance levels without affecting CCl(4)-mediated degradation of CYP2E1. Formation of hexanoyl-lysine and 4-hydroxy-2(E)-nonenal-histidine adducts, lipid peroxidation biomarkers, in homogenates from the liver of CCl(4)-treated mice was decreased in the group of mice pretreated with falcarindiol. These results suggest that the protective effects of falcarindiol against CCl(4) toxicity might, in part, be explained by anti-lipid peroxidation activity associated with the induction of the GSTs including GSTA4.     

In vitro combination effects of doripenem with aminoglycoside or ciprofloxacin against Pseudomonas aeruginosa

This study evaluated the in vitro activity of combinations of doripenem (DRPM) with aminoglycosides (tobramycin or amikacin) or fluoroquinolone (ciprofloxacin) against 92 isolates of Pseudomonas aeruginosa from 16 clinical facilities in 2004 in Japan. We also tested combination effect of other carbapenems (imipenem (IPM), meropenem, biapenem) with aminoglycosides or fluoroquinolone by checkerboard dilution methods. DRPM showed synergistic or additive effects with the aminoglycosides or the fluoroquinolone against 90% of the isolates. The combination of DRPM and aminoglycosides showed the strongest synergistic effects against IPM-intermediate resistant and IPM resistant strains among the tested combinations. These results suggested that combination of DRPM with aminoglycosides would be useful for the treatment of infections caused by P aeruginosa including IPM-resistant strains.     

Effect of the acute and chronic estrogen on anxiety in the elevated T-maze

Despite the extensive studies on the influences of estrogen (E(2)) on anxiety-like behaviors, there is still conflicting evidence regarding the specific effects of E(2) on anxiety. These discrepancies may be a result of different replacement regimens. The goals of this study were to evaluate anxiety-like behavior in ovariectomized rats (Ovx) using the elevated T-maze (ETM) test for the following variables: (1) the effects of acute versus chronic E(2) dosing, (2) the effects of chronic E(2) at different doses and, (3) the effects of Tamoxifen (Tam) co-administered with E(2). Rats in the acute E(2) dosing group (aE(2)) showed reduced inhibitory avoidance responses with prolong escape latencies compared to Ovx; while rats in the chronic E(2) dosing group (cE(2)) showed reduced inhibitory avoidance responses only. These results suggest that E(2) contains anxiolytic effects when given once or repeatedly. Moreover, when various doses of E(2) (1-100 μg/kg) were chronically given to the Ovx rats, all doses produced impaired inhibitory avoidance responses compared to Ovx, suggesting that chronic replacement of E(2) had no dose-dependent effect on anxiety-like behavior. Interestingly, in the 3-week delay replacement regimen, the low dose E(2) (1 μg/kg, s.c.) group displayed no anxiolytic effects as their inhibitory avoidance responses in the ETM were not different from their Ovx counterparts. On the contrary, the Ovx group that received Tam+E(2) (Tam 1 mg/kg, PO and E(2) 1 μg/kg, s.c.) had reduced inhibitory avoidance responses compared to other groups. These findings indicate that when Tam is co-administered with chronic low dose estrogen, it can act as an estrogen receptor agonist and result in anti-anxiety effects. Therefore, it is likely that the anxiolytic-like behavior relative to generalized anxiety disorder can be conserved when estrogen is given acutely or chronically; while the anxiolytic-like behavior relative to panic disorder can be conserved only when estrogen is given acutely.     

Inactivation of basolateral amygdala specifically eliminates palatability-related information in cortical sensory responses

Evidence indirectly implicates the amygdala as the primary processor of emotional information used by cortex to drive appropriate behavioral responses to stimuli. Taste provides an ideal system with which to test this hypothesis directly, as neurons in both basolateral amygdala (BLA) and gustatory cortex (GC)-anatomically interconnected nodes of the gustatory system-code the emotional valence of taste stimuli (i.e., palatability), in firing rate responses that progress similarly through "epochs." The fact that palatability-related firing appears one epoch earlier in BLA than GC is broadly consistent with the hypothesis that such information may propagate from the former to the latter. Here, we provide evidence supporting this hypothesis, assaying taste responses in small GC single-neuron ensembles before, during, and after temporarily inactivating BLA in awake rats. BLA inactivation (BLAx) changed responses in 98% of taste-responsive GC neurons, altering the entirety of every taste response in many neurons. Most changes involved reductions in firing rate, but regardless of the direction of change, the effect of BLAx was epoch-specific: while firing rates were changed, the taste specificity of responses remained stable; information about taste palatability, however, which normally resides in the "Late" epoch, was reduced in magnitude across the entire GC sample and outright eliminated in most neurons. Only in the specific minority of neurons for which BLAx enhanced responses did palatability specificity survive undiminished. Our data therefore provide direct evidence that BLA is a necessary component of GC gustatory processing, and that cortical palatability processing in particular is, in part, a function of BLA activity.     

A promising culture model for analyzing the interaction between adipose tissue and cardiomyocytes

The heart has epicardial adipose tissue that produces adipokines and mesenchymal stem cells. Systemic adipose tissue is involved in the pathophysiology of obesity-related heart diseases. However, the method for analyzing the direct interaction between adipose tissue and cardiomyocytes has not been established. Here we show the novel model, using collagen gel coculture of adipose tissue fragments (ATFs) and HL-1 cardiomyocytes, and electron microscopy, immunohistochemistry, real-time RT-PCR, and ELISA. HL-1 cells formed a stratified layer on ATF-nonembedded gel, whereas they formed almost a monolayer on ATF-embedded gel. ATFs promoted the apoptosis, lipid accumulation, and fatty acid transport protein (FATP) expression of FATP4 and CD36 in HL-1 cells, whereas ATFs inhibited the growth and mRNA expression of myosin, troponin T, and atrial natriuretic peptide. Treatment of leptin (100 ng/ml) and adiponectin (10 μg/ml) neither replicated nor abolished the ATF-induced morphology of HL-1 cells, whereas that of FATP4 and CD36 antibodies (25 μg/ml) never abolished it. HL-1 cells prohibited the development of CD44+/CD105+ mesenchymal stem cell-like cells and lipid-laden preadipocytes from ATFs. HL-1 cells increased the production of adiponectin in ATFs, whereas they decreased that of leptin. The data indicate that our model actively creates adipose tissue-HL-1 cardiomyocyte interaction, suggesting first that ATFs may be related to the lipotoxiciy of HL-1 cells via unknown factors plus FATP4 and CD36 and second that HL-1 cells may help to retain the static state of ATFs, affecting adipokine secretion. Our model will serve to study adipose tissue-cardiomyocyte interaction and mechanisms of obesity-related lipotoxicity and heart diseases.     

Constitutive androstane receptor agonist, TCPOBOP, attenuates steatohepatitis in the methionine choline-deficient diet-fed mouse

AIM： To ascertain whether constitutive androstane receptor （CAR） activation by 1,4-bis-[2-（3,5,- dichloropyridyloxy）] benzene （TCPOBOP） modulates steatohepatitis in the methionine choline-deficient （MCD） diet-fed animal.METHODS： C57/BL6 wild-type mice were fed the MCD or standard diet for 2 wk and were treated with either the CAR agonist, TCPOBOP, or the CAR inverse agonist, androstanol.RESULTS： Expression of CYP2B10 and CYP3A11, known CAR target genes, increased 30-fold and 45-fold, respectively, in TCPOBOP-treated mice fed the MCD diet. TCPOBOP treatment reduced hepatic steatosis （44.6 ＋ 5.4% vs 30.4 ＋ 4.5%, P 〈 0.05） and serum triglyceride levels （48 ＋ 8 vs 20 ＋ 1 mg/dL, P 〈 0.05） in MCD diet- fed mice as compared with the standard diet-fed mice. This reduction in hepatic steatosis was accompanied by an increase in enzymes involved in fatty acid microsomal co-oxidation and peroxisomal p-oxidation, namely CYP4A10, LPBE, and 3-ketoacyI-CoA thiolase. The reduction in steatosis was also accompanied by a reduction in liver cell apoptosis and inflammation. In contrast, androstanol was without effect on any of the above parameters.CONCLUSION： CAR activation stimulates induction of genes involved in fatty acid oxidation, and ameliorates hepatic steatosis, apoptosis and inflammation.     

Accelerated degradation and improved bone-bonding ability of hydroxyapatite ceramics by the addition of glass

Dense hydroxyapatite (HA) ceramics are useful bone substitutes, but they degrade minimally. One solution is to incorporate degradable materials in the HA. In this study, we manufactured glass-containing HA and investigated whether the degradability and bone-bonding ability of the HA were improved. The glass-containing HA was manufactured from a mixture of HA powder and 1.0 wt% glass powder. The control HA was manufactured from pure HA powder. In vitro degradability was evaluated by soaking in physiological saline, and a rabbit model was used to evaluate in vivo degradability and bone-bonding ability. Detaching tests were performed for all removed samples to quantify bone-bonding ability of each type of HA. The glass-containing HA showed higher degradability than the control HA, both in vitro and in vivo. The detaching failure load of the glass-containing HA was rapidly elevated after implantation and was higher than that of the control HA. Our results suggest that the dissolution of the added glass made the glass-containing HA degradable and that the detaching failure load of the glass-containing HA was elevated by reinforcement of the mechanical locking at the roughened interface. Incorporation of glass additives into HA can be concluded to be a good candidate for producing a bone substitute that can partially degrade and bond to bone firmly and rapidly.     

Neuroprotective effects of edaravone,a free radical scavenger,on the rat hippocampus after pilocarpine-induced status epilepticus

PurposeEdaravone (MCI-186) is a newly developed antioxidative radical scavenger for the treatment of acute cerebral infarction, exerting neuroprotective effects against ischemic insult. The neuroprotective effects of edaravone on pilocarpine-induced seizures in rats were investigated.MethodsRats were treated intraperitoneally with saline or edaravone (1–30 mg/kg), applied 30 min before pilocarpine hydrochloride (330 mg/kg). The onset of status epilepticus (SE) and mortality were recorded for a period of at least 3 days. The cell loss and immunoreactivities of nitric oxide synthase (NOS) in the hippocampus from control and the day 3 rats after SE, treated with saline or edaravone, were evaluated.ResultsEdaravone (1 mg/kg) significantly prevented cell loss in the hippocampus after SE while easier inducing SE. The higher dose of drug could not induce SE significantly but tended to increase the rate of mortality. Inducible NOS (iNOS) expression was significantly decreased in the hippocampus from day 3 rats treated with 1 mg/kg edaravone, compared with saline group, while neuronal NOS (nNOS) and iNOS significantly increased in the hippocampus treated with saline, compared with control group. Significant alteration of endothelial NOS (eNOS) expression in the hippocampus among control group, saline group, and edaravone group was not shown.ConclusionsEdaravone may act as a neuroprotector for the hippocampus after SE by reducing at least iNOS although the low dose of drug easier induces SE because of preventing an endogenous antiepileptic effect of NO.     

The N terminus of PA polymerase of swine-origin influenza virus H1N1 determines its compatibility with PB2 and PB1 subunits through a strain-specific amino acid serine 186

Despite several lines of evidence suggesting possible mechanisms by which the influenza virus polymerase complex, comprising PB2, PB1 and PA, work in concert during virus replication, exactly how they function is not entirely understood. The N terminal region of the PA subunit has been shown to play a key role in various functions through a number of conserved amino acid residues. However, little is known about the role of amino acids reported to be unique for a virus strain. Here, we investigated the functional implication of an amino acid (S186) present uniquely in the N terminus of the PA subunit of the pandemic H1N1 influenza virus and determined the effect of its mutation in terms of polymerase activity as well as virus growth. Using chimeric constructs of PA derived from A/PR/8/34 (H1N1) (PR8) and the swine-origin influenza virus (S-OIV) H1N1, we found that, when complexed with PB2 and PB1 of PR8, the chimeric PA protein containing the N terminus of S-OIV (1-213) with the remaining region from PR8 showed significantly reduced polymerase activity. Recombinant viruses harboring the chimeric PA also grew poorly in MDCK cells and embryonated eggs. Likewise, the chimeric PA in which the N terminus of PA of PR8 (1-213) was assembled with the remaining region of PA of S-OIV showed a similar phenotype when complexed with PB2 and PB1 of S-OIV. Interestingly, when S186 in the N terminus was altered to the residue common in most strains of influenza virus (G186), the chimeric as well as wild-type PA of S-OIV showed severely impaired polymerase activity when assayed with PB2 and PB1 of S-OIV. Collectively, this finding suggests that S186 at the N terminal region of PA of S-OIV is necessary for the protein to function optimally.     

Double mapping with subareolar blue dye and peritumoral green dye injections decreases the false-negative rate of dye-only sentinel node biopsy for early breast cancer: 2-site injection is more accurate than 1-site injection

BACKGROUND: The optimum sentinel node biopsy (SNB) mapping method for breast cancer remains to be determined. No matter which mapping agents are used, 2-site injection may be superior to 1-site injection in limiting the false-negative rate. METHODS: We examined whether a double-mapping method with subareolar injection of blue dye and peritumoral injection of green dye would decrease the false-negative rate of dye-only SNB in 145 patients with early breast cancer. RESULTS: The identification rate for blue-dyed and/or green-dyed (including mixed color-dyed) lymph nodes was 96.6% (140/145). Sensitivity and specificity were 95.1% (39/41) and 100% (99 of 99), respectively. Accuracy was 98.6% (138/140) with a false-negative rate of 4.9% (2/41). There were 4 patients in whom nodes of each color were found, but nodes of only 1 color were shown to be positive. The primary tumors of these 4 patients and of the 2 patients with false-negative results were located in the upper-outer quadrant of the breast. When only blue-dyed or green-dyed nodes (including mixed color-dyed nodes) were counted, the false-negative rates were 10.3% (4/39) for the subareolar mapping technique and 10.0% (4/40) for the peritumoral mapping technique. CONCLUSIONS: The double-mapping method based on subareolar and peritumoral injections decreases the false-negative rate of dye-only SNB for early breast cancer. Variations in lymphatic channels may exist in the lateral half of the breast and thus may influence identification of positive sentinel nodes. This finding should be taken into account in cases of multicentric breast cancer.