Inhibition of angiogenesis- and inflammation-inducing factors in human colon cancer cells <Emphasis Type=Italic>in vitro</Emphasis> and <Emphasis Type=Italic>in ovo</Emphasis> by free and nanoparticle-encapsulated redox dye,DCPIP

Background  

The redox dye, DCPIP, has recently shown to exhibit anti-melanoma activity in vitro and in vivo. On the other hand, there is increasing evidence that synthetic nanoparticles can serve as highly efficient carriers of drugs and vaccines for treatment of various diseases. These nanoparticles have shown to serve as potent tools that can increase the bioavailability of the drug/vaccine by facilitating absorption or conferring sustained and improved release. Here, we describe results on the effects of free- and nanoparticle-enclosed DCPIP as anti-angiogenesis and anti-inflammation agents in a human colon cancer HCT116 cell line in vitro, and in induced angiogenesis in ovo.     

Association between plasma zinc concentration and zinc kinetic parameters in premenopausal women

The objective of this study was to measure relationships between plasma zinc (Zn) concentrations and Zn kinetic parameters and to measure relationships of Zn status with taste acuity, food frequency, and hair Zn in humans. The subjects were 33 premenopausal women not taking oral contraceptives and dietary supplements containing iron and Zn. Main outcomes were plasma Zn concentrations, Zn kinetic parameters based on the three-compartment mammillary model using 67Zn as a tracer, electrical taste detection thresholds, and food frequencies. Lower plasma Zn was significantly (P < 0.01) associated with smaller sizes of the central and the lesser peripheral Zn pools, faster disappearance of tracer from plasma, and higher transfer rate constants from the lesser peripheral pool to the central pool and from the central pool to the greater peripheral pool. The break points in the plasma Zn-Zn kinetics relationship were found between 9.94 and 11.5 micromol/l plasma Zn. Smaller size of the lesser peripheral pool was associated with lower frequency of beef consumption and higher frequency of bran breakfast cereal consumption. Hypozincemic women with plasma Zn <10.7 micromol/l or 700 ng/ml had decreased thresholds of electrical stimulation for gustatory nerves. Our results based on Zn kinetics support the conventional cutoff value of plasma Zn (10.7 micromol/l or 700 ng/ml) between normal and low Zn status.     

A P-loop Mutation in G�� Subunits Prevents Transition to the Active State: Implications for G-protein Signaling in Fungal Pathogenesis

Heterotrimeric G-proteins are molecular switches integral to a panoply of different physiological responses that many organisms make to environmental cues. The switch from inactive to active Gαβγ heterotrimer relies on nucleotide cycling by the Gα subunit: exchange of GTP for GDP activates Gα, whereas its intrinsic enzymatic activity catalyzes GTP hydrolysis to GDP and inorganic phosphate, thereby reverting Gα to its inactive state. In several genetic studies of filamentous fungi, such as the rice blast fungus Magnaporthe oryzae, a G42R mutation in the phosphate-binding loop of Gα subunits is assumed to be GTPase-deficient and thus constitutively active. Here, we demonstrate that Gα(G42R) mutants are not GTPase deficient, but rather incapable of achieving the activated conformation. Two crystal structure models suggest that Arg-42 prevents a typical switch region conformational change upon Gα_i1(G42R) binding to GDP·AlF₄ ⁻ or GTP, but rotameric flexibility at this locus allows for unperturbed GTP hydrolysis. Gα(G42R) mutants do not engage the active state-selective peptide KB-1753 nor RGS domains with high affinity, but instead favor interaction with Gβγ and GoLoco motifs in any nucleotide state. The corresponding Gα_q(G48R) mutant is not constitutively active in cells and responds poorly to aluminum tetrafluoride activation. Comparative analyses of M. oryzae strains harboring either G42R or GTPase-deficient Q/L mutations in the Gα subunits MagA or MagB illustrate functional differences in environmental cue processing and intracellular signaling outcomes between these two Gα mutants, thus demonstrating the in vivo functional divergence of G42R and activating G-protein mutants.     

Photodimerization in pyrimidine-substituted dipeptides.

Ten N(epsilon)-glycylornithineamide derivatives have been synthesized containing various N(alpha)-linked pyrimidine-1-ylacetyl groups which can undergo (2pi + 2pi) photodimerization on irradiation with UV light at 254 nm. The dimerization efficiency of the free and bound pyrimidine groups was compared in aqueous solution: it was dependent on the substitution of the pyrimidine ring. N(alpha),N(alpha')-bis-(uracil-1-ylacetyl)-(N(epsilon)-glycylornithineamide) and N(alpha),N(alpha')-bis-(5-bromouracil-1-ylacetyl)-(N(epsilon)-glycylornithineamide) were identified as possible candidates for optical data storage.     

Pulses of darkness shift the phase of a circadian rhythm in an insectivorous bat

Summary Microspectrophotometric measurements on the retina ofDeilephila elpenor were performed using fresh (cut by razor blade) as well as frozen (cryostate microtome) sections. Difference spectra from various parts of the rhabdom clearly showed that in the distal part of the rhabdom the UV pigment is predominant, while in the medial part only green pigment was found. From the difference spectra, wave-length and relative height of the absorption maxima of rhodopsins and metarhodopsins were computed and were found to be in close agreement with other results reported in the literature. These findings confirm the work of Welsch (1977) who concluded from his electron microscopic analysis that the two distal receptor cells are UV sensitive, while the six median receptor cells are green sensitive. There is also good evidence to assume that the ninth, proximal receptor is violet sensitive.We would like to thank Mrs. E. Brüning for excellent technical assistance. This work was supported by the Deutsche Forschungsgemeinschaft SFB 114 Bionach.     

Optical Spectral Surveillance of Breast Tissue Landscapes for Detection of Residual Disease in Breast Tumor Margins

We demonstrate a strategy to “sense” the micro-morphology of a breast tumor margin over a wide field of view by creating quantitative hyperspectral maps of the tissue optical properties (absorption and scattering), where each voxel can be deconstructed to provide information on the underlying histology. Information about the underlying tissue histology is encoded in the quantitative spectral information (in the visible wavelength range), and residual carcinoma is detected as a shift in the histological landscape to one with less fat and higher glandular content. To demonstrate this strategy, fully intact, fresh lumpectomy specimens (n = 88) from 70 patients were imaged intra-operatively. The ability of spectral imaging to sense changes in histology over large imaging areas was determined using inter-patient mammographic breast density (MBD) variation in cancer-free tissues as a model system. We discovered that increased MBD was associated with higher baseline β-carotene concentrations (p = 0.066) and higher scattering coefficients (p = 0.007) as measured by spectral imaging, and a trend toward decreased adipocyte size and increased adipocyte density as measured by histological examination in BMI-matched patients. The ability of spectral imaging to detect cancer intra-operatively was demonstrated when MBD-specific breast characteristics were considered. Specifically, the ratio of β-carotene concentration to the light scattering coefficient can report on the relative amount of fat to glandular density at the tissue surface to determine positive margin status, when baseline differences in these parameters between patients with low and high MBD are taken into account by the appropriate selection of threshold values. When MBD was included as a variable a priori, the device was estimated to have a sensitivity of 74% and a specificity of 86% in detecting close or positive margins, regardless of tumor type. Superior performance was demonstrated in high MBD tissue, a population that typically has a higher percentage of involved margins.     

Phenol, 2,4-bis(1,1-dimethylethyl) of marine bacterial origin inhibits quorum sensing mediated biofilm formation in the uropathogen Serratia marcescens

Intercellular communication in bacteria (quorum sensing, QS) is an important phenomenon in disease dissemination and pathogenesis, which controls biofilm formation also. This study reports the anti-QS and anti-biofilm efficacy of seaweed Gracilaria gracilis associated Vibrio alginolyticus G16 against Serratia marcescens. Purification and mass spectrometric analysis revealed the active principle as phenol, 2,4-bis(1,1-dimethylethyl) [PD]. PD affected the QS regulated virulence factor production in S. marcescens and resulted in a significant (p < 0.05) reduction in biofilm (85%), protease (41.9%), haemolysin (69.9%), lipase (84.3%), prodigiosin (84.5%) and extracellular polysaccharide (84.62%) secretion without hampering growth, as evidenced by XTT [2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] assay. qPCR analysis confirmed the down-regulation of the fimA, fimC, flhD and bsmA genes involved in biofilm formation. Apart from biofilm inhibition and disruption, PD increased the susceptibility of S. marcescens to gentamicin when administered synergistically, which opens another avenue for combinatorial therapy where PD can be used to enhance the efficacy of conventional antibiotics.     

Delivery-Corrected Imaging of Fluorescently-Labeled Glucose Reveals Distinct Metabolic Phenotypes in Murine Breast Cancer

When monitoring response to cancer therapy, it is important to differentiate changes in glucose tracer uptake caused by altered delivery versus a true metabolic shift. Here, we propose an optical imaging method to quantify glucose uptake and correct for in vivo delivery effects. Glucose uptake was measured using a fluorescent D-glucose derivative 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-deoxy-D-glucose (2-NBDG) in mice implanted with dorsal skin flap window chambers. Additionally, vascular oxygenation (SO₂) was calculated using only endogenous hemoglobin contrast. Results showed that the delivery factor proposed for correction, “R_D”, reported on red blood cell velocity and injected 2-NBDG dose. Delivery-corrected 2-NBDG uptake (2-NBDG₆₀/R_D) inversely correlated with blood glucose in normal tissue, indicating sensitivity to glucose demand. We further applied our method in metastatic 4T1 and nonmetastatic 4T07 murine mammary adenocarcinomas. The ratio 2-NBDG₆₀/R_D was increased in 4T1 tumors relative to 4T07 tumors yet average SO₂ was comparable, suggesting a shift toward a “Warburgian” (aerobic glycolysis) metabolism in the metastatic 4T1 line. In heterogeneous regions of both 4T1 and 4T07, 2-NBDG₆₀/R_D increased slightly but significantly as vascular oxygenation decreased, indicative of the Pasteur effect in both tumors. These data demonstrate the utility of delivery-corrected 2-NBDG and vascular oxygenation imaging for differentiating metabolic phenotypes in vivo.     

Filament organization of the bacterial actin MreB is dependent on the nucleotide state

MreB, the bacterial ancestor of eukaryotic actin, is responsible for shape in most rod-shaped bacteria. Despite belonging to the actin family, the relevance of nucleotide-driven polymerization dynamics for MreB function is unclear. Here, we provide insights into the effect of nucleotide state on membrane binding of Spiroplasma citri MreB5 (ScMreB5). Filaments of ScMreB5^WT and an ATPase-deficient mutant, ScMreB5^E134A, assemble independently of the nucleotide state. However, capture of the filament dynamics revealed that efficient filament formation and organization through lateral interactions are affected in ScMreB5^E134A. Hence, the catalytic glutamate functions as a switch, (a) by sensing the ATP-bound state for filament assembly and (b) by assisting hydrolysis, thereby potentially triggering disassembly, as observed in other actins. Glu134 mutation and the bound nucleotide exhibit an allosteric effect on membrane binding, as observed from the differential liposome binding. We suggest that the conserved ATP-dependent polymerization and disassembly upon ATP hydrolysis among actins has been repurposed in MreBs for modulating filament organization on the membrane.