Adriamycin-loaded albumin-heparin conjugate microspheres for intraperitoneal chemotherapy

Adriamycin-loaded albumin-heparin conjugate microspheres (ADR-AHCMS) were evaluated as possible intraperitoneal (i.p.) delivery systems for site-specific cytotoxic action. The biocompatibility of the microspheres after intraperitoneal injection was tested first. 1 day after i.p. administration of empty as well as drug-loaded AHCMS to male Balb/c mice, only a moderate increase in i.p. neutrophils was measured. 3 days after injection neutrophil levels were comparable with the controls. No significant increases in the numbers of other cell types were observed, indicating an acute inflammatory response which can be considered to be mild. Antitumour efficacy was tested in an L1210 tumour-bearing mouse model and in a CC531 tumour-bearing rat model. The use of ADR-AHCMS leads to longer survival times of mice and improved tumour growth delay in rats, as compared with untreated controls and free drug treated animals. In both animal models higher adriamycin doses were initially tolerated if the drug was formulated in microspheres, although long-term adriamycin toxicity effects were evident in all treated groups. Doses and dosage schedules may be optimized to further reduce the toxic effects of the drug

D* Production in Deep Inelastic Scattering at HERA

This paper presents measurements of D^{*\pm} production in deep inelastic scattering from collisions between 27.5 GeV positrons and 820 GeV protons. The data have been taken with the ZEUS detector at HERA. The decay channel $D^{*+}\to (D^0 \to K^- \pi^+) \pi^+$ (+ c.c.) has been used in the study. The $e^+p$ cross section for inclusive D^{*\pm} production with $5<Q^2<100 GeV^2$ and $y<0.7$ is 5.3 \pms 1.0 \pms 0.8 nb in the kinematic region {$1.3<p_T(D^{*\pm})<9.0$ GeV and $| \eta(D^{*\pm}) |<1.5$}. Differential cross sections as functions of p_T(D^{*\pm}), $\eta(D^{*\pm}), W$ and $Q^2$ are compared with next-to-leading order QCD calculations based on the photon-gluon fusion production mechanism. After an extrapolation of the cross section to the full kinematic region in p_T(D^{*\pm}) and $\eta$(D^{*\pm}), the charm contribution $F_2^{c\bar{c}}(x,Q^2)$ to the proton structure function is determined for Bjorken $x$ between 2 $\cdot$ 10$^{-4}$ and 5 $\cdot$ 10$^{-3}$.Comment: 17 pages including 4 figure

Measurement of the Probability of Gluon Splitting into Charmed Quarks in Hadronic Z Decays

We have measured the probability, n(g->cc~), of a gluon splitting into a charm-quark pair using 1.7 million hadronic Z decays collected by the L3 detector. Two independent methods have been applied to events with a three-jet topology. One method relies on tagging charmed hadrons by identifying a lepton in the lowest energy jet. The other method uses a neural network based on global event shape parameters. Combining both methods, we measure n(g->cc~)= [2.45 +/- 0.29 +/- 0.53]%

Inclusive Jet Production in Two-Photon Collisions at LEP

Inclusive jet production, e+e- -> e+e- \ee$ jet X, is studied using 560/pb of data collected at LEP with the L3 detector at centre-of-mass energies between 189 and 209 GeV. The inclusive differential cross section is measured using a k_t jet algorithm as a function of the jet transverse momentum, pt, in the range 3<pt<50 GeV for a pseudorapidity, eta, in the range -1<eta<1. This cross section is well represented by a power law. For high pt, the measured cross section is significantly higher than the NLO QCD predictions, as already observed for inclusive charged and neutral pion production

Targeting of polyelectrolyte RNA complexes to cell surface integrins as an efficient cytoplasmic transfection mechanism

This is the first demonstration of receptor-mediated delivery of mRNA and establishes a new approach to gene therapy. Messenger RNA (mRNA) provides a promising alternative to plasmid DNA as a genetic material for delivery in non-viral gene therapy strategies. Since it does not require access to the nucleus and is less dependent on the cell cycle for expression, mRNA delivered using cationic lipids or short cationic polymers can be effectively translated within target cells. In this study, mRNA formed discrete nanoparticles following self assembly with a range of cationic polymers. Based on transfection activities, the low molecular weight polycations were more efficient than high molecular weight, while protamine and polyethylenimine were far more efficient than poly(L-lysine). Receptor-mediated delivery of mRNA was demonstrated using the synthetic polyamino acid (K),_16,GACDCRGDCFCA designed to promote cell entry following interaction with cell surface α_υ integrins. RGD-bearing mRNA complexes showed very high levels of expression, reaching over 60% transduction of B16F10 cells

Laterally stabilized complexes of DNA with linear reducible polycations: strategy for triggered intracellular activation of DNA delivery vectors

Target-specific DNA delivery requires vectors that combine stability in the biological milieu, receptor-mediated uptake into target cells, and intracellular activation to mediate transgene expression. This is achieved here using polymer-coated vectors based on plasmid DNA complexed with a reductively degradable polycation (RPC), designed for intercellular degradation. The RPC were prepared by oxidation of the terminal cysteinyl thiol groups of Cys(Lys)&lt;sub&gt;10&lt;/sub&gt;Cys. The complexes were coated and surface-cross-linked using multivalent reactive copolymers of N-(2-hydroxypropyl)methacrylamide (PHPMA), providing a unique combination of steric and reversible lateral stabilization, known to promote extended circulation in the bloodstream. Coated complexes containing RPC exhibited lateral stabilization that was reversible by treatment with 2.5 mM dithiothreitol, releasing free DNA after incubation with a polyanion. In contrast, coated complexes containing nonreducible poly(l-lysine) (PLL) were not destabilized by reduction. The biological usefulness of this trigger mechanism was examined by measuring transfection activity in human retinoblast 911 cells of coated complexes, based on PLL or RPC, targeted to cell surface receptors by covalent linkage of basic fibroblast growth factor. The levels of transgene expression observed for RPC-based targeted vectors indicated efficient intracellular activation, authenticating the concept that lateral stabilization introduced by surface coating with PHPMA can be reversed by intracellular reduction

Methodologies for monitoring nanoparticle formation by self-assembly of DNA with poly(L-lysine)

DNA self-assembly with polycations produces nanoparticles suitable for gene delivery, although there is no standard methodology to measure particle formation and stability. Here we have compared three commonly used assays, namely, light scattering, inhibition of ethidium bromide fluorescence, and modified electrophoretic mobility of DNA. Analysis by light scattering and loss of ethidium bromide fluorescence both showed poly(l-lysine) (pLL)/DNA nanoparticles form over the lysine/phosphate ratio range 0.6–1.0, although retardation of DNA electrophoretic mobility commenced at lower lysine/phosphate ratios. This probably indicates that the first two assays monitor DNA collapse into particles, while the electrophoresis assay measures neutralization of the charge on DNA. Gel analysis of the complexes showed disproportionation during nanoparticle formation, probably reflecting cooperative binding of the polycation. The assays were used to examine stability of complexes to dilution in water and physiological salts. Whereas all pLL/DNA nanoparticles were stable to dilution in water, the presence of physiological salts provoked selective disruption of complexes based on low-molecular-weight pLL. Polyelectrolyte complexes for targeted application in vivo should therefore be based on high-molecular-weight polycations, or should be stabilized to prevent their dissociation under physiological salt conditions

Limit dilution analysis of peripheral blood T lymphocytes specific to periodontopathic bacteria

Limit dilution analysis (LDA) was used to determine the presence and frequency of periodontopathic-bacteria-specific T cells in the peripheral blood of patients with chronic inflammatory periodontal disease. Twelve adult periodontitis (AP), 13 marginal gingivitis (MG) and 12 healthy control subjects took part in the study. Bacteroides gingivalis and Actinomyces viscosus were used as test organisms, while tetanus toxoid was used as the control antigen. The median PTL-p frequencies to B. gingivalis were 46.33 x 10(-6), 45.33 x 10(-6) and 58.83 x 10(-6) in the control, gingivitis and AP groups respectively, while the median PTL-p frequencies to A. viscosus were 13.8 x 10(-6), 17.33 x 10(-6) and 11.5 x 10(-6), again in the control, gingivitis and AP groups. There were no statistically significant differences between the groups. All subjects displayed 'single-hit' kinetics with the control tetanus toxoid antigen and, with three exceptions, 'single-hit' kinetics was also found with the two test organisms. One control subject displayed a 'saw-tooth' curve with A. viscosus and a 'suppressor' curve with B. gingivalis, while two MG subjects had a 'saw-tooth' curve with B. gingivalis. These complex curves suggest that, in some subjects, more than one limiting cell type may exist in the cultures. Nevertheless, the results of the present study illustrate that lymphocytes specific to periodontopathic bacteria exist in the peripheral blood of both diseased and non-diseased subjects

Effect of initial treatment of chronic inflammatory periodontal disease on the frequency of peripheral blood T-lymphocytes specific to periodontopathic bacteria.

Limit dilution analysis (LDA) was used to determine the effect of initial treatment of chronic inflammatory periodontal disease on the frequency of periodontopathic bacteria‐specific T‐cells in peripheral blood. Eleven marginal gingivitis (MG) and 8 adult periodontitis (AP) subjects took part in the study. The proliferative T‐lymphocyte precursor (PTL‐P) frequencies to Porphyromonas gingivalis and Actinomyces viscosus were determined using LDA and Poisson statistics both before and after treatment. Tetanus toxoid was used as a control antigen. Treatment resulted in a significant reduction in clinical disease parameters in both groups. The median peak PTL‐P frequency for P. gingivalis was significantly higher in the AP group compared with the MG group before treatment. This was not the case after treatment nor with A. viscosus. In the MG group the median peak PTL‐P frequency with both P. gingivalis and A. viscosus declined as a result of treatment. Although this decline was not statistically significant it may indicate an antigen‐specific response in this group. In the AP group the median peak PTL‐P frequency with P. gingivalis before treatment was 83.76 × 10 (approximately 1 in 12,000) and after treatment it was 36.17 × 10 (approximately 1 in 28,000). Dose‐response relationships showed at each concentration of organisms/ well this trend for a decline in PTL‐P frequency after treatment, suggesting that any increased responsiveness to this organism in this group may be largely antigen‐specific. However, there was no difference in this group in the median peak PTL‐P frequency with A. viscosus before and after treatment. This, together with the low PTL‐P frequency to A. viscosus, suggests that any increased responsiveness to this organism in this group may be non‐specific. Copyrigh