Probenecid-induced changes in the clearance of pranoprofen enantiomers

Probenecid is known to inhibit the elimination of several acidic drugs. Its influence on the pharmacokinetics of pranoprofen was investigated in rabbit after a single intravenous injection of racemic mixture (5 mg/kg). Levels of (-)-(R)- and (+)-(S)-pranoprofen and their glucuronide (after hydrolysis with sodium hydroxide) were determined in plasma, urine, and several tissues. The plasma concentration of the (+)-(S)-isomer was higher than that of the (-)-(R)-form. Oral coadministered probenecid (100 mg/kg) resulted in an increased plasma concentration of both enantiomers. Probenecid reduced the apparent total clearance and excretion of pranoprofen enantiomers in urine. It had a slight effect on the tissue distribution of pranoprofen at the dose used, but significantly reduced the formation of glucuronide for both enantiomers to the same extent in kidney microsomes. The differences caused by probenecid were significant with respect to its ability to inhibit glucuronidation in the kidney and subsequent excretion into urine, but enantioselective effects were negligible.     

Inhibition of chromosome separation in fertilized starfish eggs by kalihinol F, a topoisomerase I inhibitor obtained from a marine sponge

Kalihinol F, a naturally occurring diterpene from a marine sponge, Acanthella sp., inhibited chromosome separation in fertilized starfish (Asterina pectinifera) eggs but allows the first cleavage to occur, thereby forming unseparated metaphase chromosomes which were elongated between the two daughter cells. The chromosomes were eventually torn off in the embryonic cells. Most of the cells gradually lost the chromosomes during the cell cycle progression. The embryonic development halted at the morula stage just before the onset of blastulation. The mitotic failure occurred when kalihinol F was applied to a fertilized egg during the second meiotic process, but not after the completion of the second meiotic division. Kalihinol F inhibited topoisomerase I activity in vitro, but had no effects on activities of DNA polymerases alpha, beta, and gamma, and of topoisomerase II. These results suggest that the topoisomerase I plays an essential role in meiosis II in this species.     

Promoting effect of kaempferol on the differentiation and mineralization of murine pre-osteoblastic cell line MC3T3-E1

A number of agents have been reported to influence osteoblastic differentiation and to prevent and treat bone loss. We found that kaempferol, a flavonoid identified in extracts of the medicinal plant, Polygonum tinctorium. Lour, had stimulatory effects on the differentiation and mineralization of the murine pre-osteoblastic cell line, MC3T3-E1. After enhancing the alkaline phosphatase activity, significant augmentation of calcification by kaempferol was observed between concentrations of 10 and 20 microM, without any marked effect on cell proliferation. When kaempferol was combined with ipriflavone, which is clinically applied to treat bone loss, calcification was synergistically augmented, suggesting that these two flavonoids may have different mechanisms of action. These results suggest that kaempferol may be a promising agent for the prevention or treatment of bone loss, especially when combined with ipriflavone.     

Denitrification of nitrate by the fungus Cylindrocarpon tonkinense

The denitrifying fungus Cylindrocarpon tonkinense was thought to be able to denitrify only nitrite (NO2-) but not nitrate (NO3-) to form nitrous oxide (N2O). Here we found, however, that C. tonkinense can denitrify NO3- under certain conditions. Presence of ammonium (NH3+) in addition to NO3- and the use of a fermentable sugar as an electron donor were key conditions for inducing the denitrifying activity. Such induction accompanied a remarkable increase in the intracellular level of the enzyme activities related to NO3- metabolism. These activities contained assimilatory type NADPH (or NADH)-dependent NO3- reductase (aNar), dissimilatory nitrite reductase (dNir), and nitric oxide reductase (P450nor), but did not contain ubiquinol-dependent, dissimilatory NO3- reductase (dNar). The denitrification was inhibited by tungstate, an inhibitor of Nar. These results demonstrated occurrence of a novel type of denitrification in C. tonkinense, in which assimilatory type Nar is possibly involved.     

Properties of rice cooked with commercial water-soluble soybean polysaccharides extracted under weakly acidic conditions from soybean cotyledons

It has been found that commercial water-soluble soybean polysaccharides (SSPS) can make cooked rice and noodles non-sticky and prevent rice grains and noodles from adhering to each other. We studied in detail the phenomenon of rice cooked with SSPS. We assumed that the phenomenon resulted from the interaction between SSPS and starch during cooking, and studied the effects of SSPS on the gelatinizing behavior of rice starch by using a Rapid-Visco-Analyzer. The addition of SSPS reduced the viscosity of the gelatinized starch. This lower final viscosity of the rice starch was more distinct from than that of potato starch. These results imply that the properties of SSPS in forming a non-sticky condition might result from a decrease in the viscosity of the gelatinized starch.     

Inhibitory effects of brown algal phlorotannins on secretory phospholipase A2s, lipoxygenases and cyclooxygenases

The inhibitory effects of brown algal phlorotannins on secretory phospholipase A₂s (sPLA₂s), lipoxygenases (LOXs) and cyclooxygenases (COXs) were determined with an in vitro assay. Oligomers of phloroglucinol; eckol (a trimer), phlorofucofuroeckol A (a pentamer), dieckol (a hexamer) and 8,8-bieckol (a hexamer) isolated from the brown alga Eisenia bicyclis had pronounced inhibitory effects on sPLA₂ from porcine pancreas and bee venom (IC₅₀ 100–200 M). The phlorotannins inhibited LOX activity more effectively than the well-known LOX inhibitors; resveratrol and epigallocatechin gallate. 8,8-Bieckol, the strongest LOX inhibitor in this study, inhibited soybean LOX and 5-LOX with IC₅₀ values of 38 and 24 M, respectively. Negligible or very weak effects of the phlorotannins on COX-1 and COX-2 were found, except for an inhibitory effect of dieckol on COX-1 (74.7%) and of eckol on COX-2 (43.2%) at 100 M.     

Synthesis of chemotactic peptide analogs with a photoaffinity cross-linker as new probes for analysis of receptor-ligand interaction

Formylpeptide receptors are well-characterized receptors which participate in host defense responses of neutrophils. We designed and synthesized chemotactic peptide analog with p-benzoylphenylalanine (Bpa) and biotin to probe structural and mechanistic aspects of peptide-receptor interaction. These peptides possess biological activities which were dependent upon spacer residue length of and Bpa position. The covalent photoaffinity label was detected by Streptavidine-blot, which was inhibited by the parent peptide.     

Prickle 1 regulates cell movements during gastrulation and neuronal migration in zebrafish

During vertebrate gastrulation, mesodermal and ectodermal cells undergo convergent extension, a process characterised by prominent cellular rearrangements in which polarised cells intercalate along the medio-lateral axis leading to elongation of the antero-posterior axis. Recently, it has become evident that a noncanonical Wnt/Frizzled (Fz)/Dishevelled (Dsh) signalling pathway, which is related to the planar-cell-polarity (PCP) pathway in flies, regulates convergent extension during vertebrate gastrulation. Here we isolate and functionally characterise a zebrafish homologue of Drosophila prickle (pk), a gene that is implicated in the regulation of PCP. Zebrafish pk1 is expressed maternally and in moving mesodermal precursors. Abrogation of Pk1 function by morpholino oligonucleotides leads to defective convergent extension movements, enhances the silberblick (slb)/wnt11 and pipetail (Ppt)/wnt5 phenotypes and suppresses the ability of Wnt11 to rescue the slb phenotype. Gain-of-function of Pk1 also inhibits convergent extension movements and enhances the slb phenotype, most likely caused by the ability of Pk1 to block the Fz7-dependent membrane localisation of Dsh by downregulating levels of Dsh protein. Furthermore, we show that pk1 interacts genetically with trilobite (tri)/strabismus to mediate the caudally directed migration of cranial motor neurons and convergent extension. These results indicate that, during zebrafish gastrulation Pk1 acts, in part, through interaction with the noncanonical Wnt11/Wnt5 pathway to regulate convergent extension cell movements, but is unlikely to simply be a linear component of this pathway. In addition, Pk1 interacts with Tri to mediate posterior migration of branchiomotor neurons, probably independent of the noncanonical Wnt pathway.     

The role of Grg4 in tectal laminar formation

Mature chick optic tecta consist of 16 laminae and receive retinal fiber projections in a precise retinotopic manner. Retinal axons arborize in laminae a-f of the SGFS, but do not cross the border between lamina f and g. In order to elucidate molecular mechanisms of tectal laminar formation, we first looked at the migration of tectal postmitotic cells. We found that the migration pattern of postmitotic cells changes around E5 and that late migratory cells intervened laminae that were formed by early migratory cells. The coincident appearance of Grg4 expression in the tectal ventricular layer and the change in migration pattern suggested an important role for Grg4. Clonal misexpression of Grg4 resulted in cells migrating to laminae h-j of the SGFS. Massive misexpression of Grg4 resulted in disruption of laminae that were formed by early migratory cells, in particular lamina g of the SGFS. Application of Grg4 morpholino antisense oligonucleotide or the misexpression of a dominant-negative form of Grg4 exerted the opposite effect. We concluded that Grg4 may direct tectal postmitotic cells to follow a late migratory pathway.     

Noninvasive measurement of chloride concentration in rat olfactory receptor cells with use of a fluorescent dye

Inwardly directed Ca(2+)-dependent chloride currents are thought to prolong and boost the odorant-induced transient receptor currents in olfactory cilia. Cl(-) inward current, of course, requires a sufficiently high intracellular Cl(-) concentration ([Cl(-)](i)). In previous measurements using a fluorescent Cl(-) probe, N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE), [Cl(-)](i) of newt olfactory cells was estimated to be only 40 mM. This low value led us to reexamine the [Cl(-)](i) by an improved procedure. When isolated rat olfactory neurons were bathed in Tyrode's solution (150 mM Cl(-)) at room temperature, the [Cl(-)] was 81.5 +/- 13.5 mM (mean +/- SE) in the tip of the dendrite (olfactory knob) and 81.8 +/- 10.2 mM (mean +/- SE) in the soma. The corresponding Cl(-) equilibrium potentials were -15.4 and -15.3 mV, respectively. Therefore, at resting potentials in the range of -90 to -50 mV, Cl(-) currents are predicted to be inward and capable of contributing to the depolarization induced by odorants. Yet, if the cell was depolarized beyond -15 mV, somal Cl(-) currents would be outward and facilitate repolarization during excitation. The measured [Cl(-)] in soma and knob are of interest, because in the cilia the chloride content may be expected to equilibrate with that of the knob in the resting state. They provide a starting point for the decrease in ciliary [Cl(-)] predicted to occur during transduction.