Mass spectrometric studies of underivatized polyphenolic glycosides

Flavonoid and xanthone glycosides were studied by mass spectrometry using soft ionization techniques such as desorption chemical ionization, fast atom bombardment and field desorption. In all cases a preliminary derivatization was not required. Information on the M_r, the sugar sequence and structure of the aglycone could be obtained.     

Effects of neck exposure to 5.5 mT static magnetic field on pharmacologically modulated blood pressure in conscious rabbits

Static magnetic fields (SMF) in the millitesla (mT) range have been reported to modulate microcirculatory hemodynamics and/or blood pressure (BP) under pharmacologically modified state in mammals. This study was designed to investigate the acute effects of local application of a SMF to neck or pelvic region under pharmacologically modulated BP; norepinephrine (NE)-induced hypertension as well as an L-type voltage-gated Ca(2+) channel blocker, nicardipine (NIC)-induced hypotension in conscious rabbits. Magnetic flux densities were up to 5.5 mT and the spatial magnetic gradient peaked in neck (carotid sinus baroreceptor) region at the level of approximately 0.06 mT/mm. The duration of exposure was 30 min (including 10 min of pretreatment) and the effects on BP were investigated up to 100 min postexposure. Baroreflex sensitivity (BRS) was estimated from invasive recordings of systolic BP and pulse interval. Neck exposure to 5.5 mT significantly attenuated the pharmacologically induced vasoconstriction or vasodilation, and subsequently suppressed the increase or decrease in BP compared with sham exposure. In contrast, pelvic exposure to 5.5 mT did not significantly antagonized NE-elevated BP or NIC-reduced BP. The neck exposure to 5.5 mT has a biphasic and restorative effect on vascular tone and BP acting to normalize the tone and BP. The neck exposure to 5.5 mT caused a significant increase in BRS in NE-elevated BP compared with sham exposure. The buffering effects of the SMF on increased hemodynamic variability under NE-induced high vascular tone and NIC-induced low vascular tone might be, in part, dependent on baroreflex pathways, which could modulate NE-mediated response in conjunction with Ca(2+) dynamics.     

Multiple glycerol shocks increase the calcium phosphate transfection of non-synchronized CHO cells

The exposure of CHO DG44 cells to an osmotic shock, after DNA uptake, results in a cellular volume decrease of approx. 55%. Repetitive osmotic shocks targeted different sub-populations of cells as was demonstrated using two different fluorescent reporter genes. Also the exposure of a calcium phosphate–DNA coprecipitate to high osmolarity in vitro caused the release of the DNA from the precipitate. The results demonstrate the importance of the osmotic shock on the efficient delivery of plasmid DNA to the nucleus of CHO cells following calcium phosphate-mediated transfection.     

Measurement of CTL-induced cytotoxicity: The caspase 3 assay

Cytotoxic T lymphocytes (CTL) are critical effector cells of the immune system. Measurement of target cell damage has historically been an important measure of CTL function. CTL kill their target cells predominantly by inducing programmed cell death, or apoptosis. The gold standard for CTL-mediated cytotoxicity has been the ⁵¹Cr release assay. However, measurement of target cell lysis by ⁵¹Cr release does not provide mechanistic information on the fate of target cells, especially at the single cell level. Given the recent advances in our understanding of programmed cell death, newer assays are required which evaluate the status of the apoptotic pathways in target cells. We have developed a flow cytometry-based assay for CTL-mediated cytotoxicity based on specific binding of antibody to activated caspase 3 in target cells. Our assay is convenient and more sensitive than the ⁵¹Cr release assay. The use of this assay should allow mechanistic studies of the intracellular events resulting from CTL attack.     

Intrasynaptosomal Sequestration of [3H]Amphetamine and [3H]Methylenedioxyamphetamine: Characterization Suggests the Presence of a Factor Responsible for Maintaining Sequestration

We examined the incorporation of [3H]methylenedioxyamphetamine ([3H]MDA) and [3H]amphetamine into rat brain synaptosomes. Saturation studies, using increasing concentrations of nonradioactive ligand, revealed that [3H]-MDA interacted with two saturable sites that were sensitive to boiling of the tissue. Eadee-Scatchard plots of [3H]MDA saturation data were curvilinear; nonlinear curve-fitting analysis of these data suggested the presence of high- and low-affinity [3H]MDA sites of association: KD high = 295 nM, Bmax high = 32 pmol/mg of protein; KD low = 45 microM, Bmax low = 5.2 nmol/mg of protein. Association of [3H]MDA to the low-affinity site was dependent on the presence of isotonic sucrose in the incubation medium. The high capacities of these sites argue against a bimolecular interaction of [3H]MDA with monovalent protein binding sites. [3H]MDA incorporation was reduced under conditions that disrupt the integrity of plasma membranes, such as sonication, incubation in hypotonic media, and incubation in the presence of the detergent digitonin. These data indicate that [3H]MDA incorporation into synaptosomes may represent an internalization and sequestration phenomenon. [3H]MDA incorporation was also reduced by preincubation of the synaptosomal preparation at 37 degrees C or in hypotonic buffer at 4 degrees C, a result suggesting that this sequestration is maintained by an intrasynaptosomal component that is lost under the preincubation conditions described above. [3H]MDA incorporation was pH dependent (maximal at pH 7.5) and temperature sensitive (maximal incorporation occurred at 21 degrees C and was substantially reduced at 37 degrees C). [3H]Amphetamine was also incorporated into synaptosomes, and this incorporation was sensitive to the same physical manipulations of the tissue preparation as [3H]MDA incorporation. The synaptosomal sequestration of both [3H]MDA and [3H]amphetamine was inhibited by permeant cations, such as sodium and potassium, a result suggesting that the proposed intrasynaptosomal component that maintains the sequestration is anionic. Preliminary pharmacological profiles of [3H]MDA and [3H]amphetamine sequestration were identical. The rank order of inhibitor potencies for the incorporation of both ligands was desipramine greater than amphetamine greater than MDA greater than methylphenidate. This order of potency does not correspond to the lipophilicity of the test drugs. The synaptosomal incorporation and sequestration of [3H]MDA, [3H]methylenedioxymethamphetamine, and [3H]amphetamine described in the present report may be important in the molecular mechanism of action of monoamine release induced by the amphetamines.     

A four coil exposure system (tetracoil) producing a highly uniform magnetic field

We propose a magnetic field exposure system (tetracoil) for in vitro and in vivo experiments, composed of two couples of circular coils satisfying a spherical constraint, whose characteristics are chosen in order to maximize the uniformity region of the magnetic field. Analytical calculations and computer simulations show that our system, as compared to the other most largely used magnetic field exposure systems, represents an optimal compromise in terms of field uniformity, accessibility for biological experiments, and ratio between overall dimension and uniformity region.     

Genotypic characterization to identify markers associated with putative hypervirulence in Swedish Escherichia coli O157:H7 cattle strains

Aims: To establish whether investigated subtyping methods could identify any specific characteristics that distinguish Swedish VTEC O157:H7 strains isolated from cattle farms associated with human enterohaemorrhagic Escherichia coli (EHEC) cases from cattle strains isolated in prevalence studies. Methods and Results: Strains (n = 32) isolated in a dairy herd prevalence study and strains isolated from farms associated with human cases (n = 13) were subjected to typing. Partial sequencing of the vtx₂ genes could not identify any unique variants of vtx₂ or vtx_2c in strains associated with human cases. A specific variant of VTEC O157:H7, which was overrepresented among farms associated with human cases (P = 0·01), was by two different single‐nucleotide‐polymorphism (SNP) assays identified as clade 8, a subgroup of VTEC O157:H7 strains considered to be putatively hypervirulent. Multi‐locus variable number tandem repeat analysis (MLVA) typing of all strains produced similar results as pulsed‐field gel electrophoresis (PFGE) typing regarding clustering of the strains, but MLVA distinguished slightly better among strains than PFGE. Conclusion: In Sweden, VTEC O157:H7 strains from the putatively hypervirulent clade 8 are overrepresented among isolates from cattle farms associated with human cases compared with VTEC O157:H7 strains isolated in prevalence studies. Significance and Impact of the Study: Real‐time PCR SNP typing for clade 8 can be used to identify cattle farms that are at higher risk of causing EHEC infections in humans.     

Food and specific macronutrient limitation in an assemblage of predatory beetles

Habitats vary in food resources with carnivores often being prey limited, but it is unclear whether habitats facilitate a nutritionally balanced diet. Two paradigms in nutritional ecology, ecological stoichiometry and nutritional geometry, predict that carnivores are limited mainly by protein or lipid, respectively. Using the carabid beetle Anchomenus dorsalis and 10 other predatory beetles from agricultural fields, we developed and tested two simple procedures for quantifying macronutrient‐specific habitat conditions without requiring information about the natural prey. Both procedures assume that predators forage for nutrients rather than specific prey. Our results show that 10 of 11 species were food limited. Five species were lipid limited and one species was protein limited in the field. Co‐existing predator species showed considerable segregation of fundamental macronutritional niches. A linear relationship between specific nutrient limitation and the target lipid:protein (L:P) intake ratio indicates that species with high L:P target are more protein limited while species with low L:P target are more lipid limited. The study illustrates how species within a natural assemblage vary in nutritional niche and in specific nutrient limitation.     

The role of the Na+/Ca2+ exchangers in Ca2+ dynamics in ventricular myocytes

The role of the Na⁺/Ca²⁺ exchanger (NCX) as the main pathway for Ca²⁺ extrusion from ventricular myocytes is well established. However, both the role of the Ca²⁺ entry mode of NCX in regulating local Ca²⁺ dynamics and the role of the Ca²⁺ exit mode during the majority of the physiological action potential (AP) are subjects of controversy. The functional significance of NCXs location in T-tubules and potential co-localization with ryanodine receptors was examined using a local Ca²⁺ control model of low computational cost. Our simulations demonstrate that under physiological conditions local Ca²⁺ and Na⁺ gradients are critical in calculating the driving force for NCX and hence in predicting the effect of NCX on AP. Under physiological conditions when 60% of NCXs are located on T-tubules, NCX may be transiently inward within the first 100 ms of an AP and then transiently outward during the AP plateau phase. Thus, during an AP NCX current (I_NCX) has three reversal points rather than just one. This provides a resolution to experimental observations where Ca²⁺ entry via NCX during an AP is inconsistent with the time at which I_NCX is thought to become inward. A more complex than previously believed dynamic regulation of I_NCX during AP under physiological conditions allows us to interpret apparently contradictory experimental data in a consistent conceptual framework. Our modelling results support the claim that NCX regulates the local control of Ca²⁺ and provide a powerful tool for future investigations of the control of sarcoplasmic reticulum (SR) Ca²⁺ release under pathological conditions.     

磁流体肿瘤热疗技术的研究进展

热疗在近年来已经成为肿瘤治疗最为重要的手段之一,但存在一定的局限性。磁流体热疗技术作为新兴发展起来的热疗手段,克服了常规热疗的缺陷,可以辅助治疗,甚至发展成独立治疗手段。本文综述了国内外近年来有关磁流体热疗基础研究及试验领域的最近进展,首先对磁流体特性、常见磁流体材料和交变磁场装置等方面进行了介绍,最后介绍了磁流体肿瘤热疗技术在体外试验、动物试验和临床研究方面的进展状况。虽然磁流体热疗逐步进入临床阶段,但仍存在不足,需要进一步的完善提高治疗效果。