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At least 775 records · Page 43Linked to original sources
PubMed2004

Dendrimer-based nanosized MRI contrast agents.

Paramagnetic metals can induce T1 shortening by interaction with free water molecules. Two metal ions, Gadolinium and Manganese, are currently available for human use. Gadolinium-based MRI contrast agents (CAs) can operate using a approximately 100-fold lower concentration of Gadolinium ions in comparison to the necessary concentration of Iodine atoms employed in CT imaging in the tissues. Therefore, numerous macromolecular MRI CAs prepared employing relatively simple chemistry are readily available that can provide sufficient enhancement for multiple applications. Herein, we describe the synthesis, characteristics, and potential applications of dendrimer-based macromolecular MRI CAs in our recently reported libraries. This entire series of dendrimer-based macromolecular MRI CAs have a spherical shape and possess similar surface charges. Changes in molecular size altered the route of excretion. Smaller sized contrast agents, of less than 60 kD molecular weight, were excreted through the kidney resulting in these agents being potentially suitable as functional renal contrast agents. Less hydrophilic and larger sized contrast agents were found better suited for use as blood pool contrast agents. Hydrophobic variants of CAs formed with polypropylenimine diaminobutane dendrimer cores quickly accumulated in the liver and can function as liver contrast agents. Larger hydrophilic agents are also useful for lymphatic imaging. Finally, contrast agents conjugated with either monoclonal antibodies or with avidin are able to function as tumor-specific contrast agents and might also be employed as therapeutic drugs for either gadolinium neutron capture therapy or in conjunction with radioimmunotherapy.

Animals↗
PubMed1979

Electroimmunoassay of a subunit protein in a macromolecular complex (apolipoprotein B in human plasma very low density lipoprotein); implications for other electroimmunoassay systems.

With an electroimmunoassay ("rocket") system for the apolipoprotein B component of the plasma very low density lipoprotein complex we obtained results which were similar to those obtained by a colorimetric tetramethylurea extraction method. Results were up to twice as high as those using radioimmunoassay. Low density lipoprotein containing apolipoprotein B as the only demonstrable protein component was used as the standard for these assays. This protein produced larger and higher rockets at pH 8.6 when the negative particle charge was increased by maleylation. The very low density lipoprotein complex has a higher negative charge at pH 8.6 than low density lipoprotein. These findings suggest that some apolipoprotein B in vary low density lipoprotein is not "recognised" by anti-apolipoprotein B antibodies, hence radioimmunoassay results are lower than those obtained with the tetramethylurea extraction method. The higher negative charge on very low density lipoprotein particles (compared with low density lipoprotein), as a factor tending to increase rocket area and height, is counterbalanced by reduced recognition by antiapolipoprotein B antibodies. The net result of these opposing tendencies is that the rocket electroimmunoassay of apolipoprotein B in very low density lipoprotein fortuitously gives valid results, under the specified assay conditions. We conclude that electroimmunoassays of complex proteins are not necessarily valid if protein subunits are used for standards. This has implications for the electroimmunoassay of other apolipoproteins.

Apolipoproteins↗
PubMed2006

Fluorescence correlation spectroscopy simulations of photophysical phenomena and molecular interactions: a molecular dynamics/monte carlo approach.

Fluorescence correlation spectroscopy (FCS) is being applied increasingly to study diffusion and interactions of fluorescently labeled macromolecules in complex biological systems. Fluctuations in detected fluorescence, deltaF(t), are expressed as time-correlation functions, G(tau), and photon-count histograms, P(k;DeltaT). Here, we developed a generalized simulation approach to compute G(tau) and P(k;DeltaT) for complex systems with arbitrary geometry, photophysics, diffusion, and macromolecular interactions. G(tau) and P(k;DeltaT) were computed from deltaF(t) generated by a Brownian dynamics simulation of single-molecule trajectories followed by a Monte Carlo simulation of fluorophore excitation and detection statistics. Simulations were validated by comparing analytical and simulated G(tau) and P(k;DeltaT) for diffusion of noninteracting fluorophores in a three-dimensional Gaussian excitation and detection volume. Inclusion of photobleaching and triplet-state relaxation produced significant changes in G(tau) and P(k;DeltaT). Simulations of macromolecular interactions and complex diffusion were done, including transient fluorophore binding to an immobile matrix, cross-correlation analysis of interacting fluorophores, and anomalous sub- and superdiffusion. The computational method developed here is generally applicable for simulating FCS measurements on systems complicated by fluorophore interactions or molecular crowding, and experimental protocols for which G(tau) and P(k;DeltaT) cannot be computed analytically.

Algorithms↗
PubMed2003

Post-transcriptional regulation of gene expression by degradation of messenger RNAs.

Recent evidence suggests that gene expression may be regulated, at least in part, at post-transcriptional level by factors inducing the extremely rapid degradation of messenger RNAs. These factors include reactions between adenyl-uridyl-rich elements (AREs) of the relevant mRNA and either specific proteins that bind to these elements or exosomes. This review deals with examples of the proteins (AU-rich binding proteins, AUBPs) and exosomes, which have been shown to form complexes with AREs and bring about rapid degradation of the relevant mRNA, and with certain other factors, which protect the RNA from such degradation. The biochemical and physiological factors underlying the stability of messenger RNAs carrying the ARE motifs will be reviewed in the light of their emerging significance for cell physiology, human pathology, and molecular medicine. We also consider the possible application of the results of recent insights into the mechanisms to pharmacological interventions to prevent or cure disorders, especially developmental disorders, which the suppression of gene expression may bring about. Molecular targeting of specific steps in protein degradation by synthetic compounds has already been utilized for the development of pharmacological therapies.

Animals↗
PubMed1984

Sintering technique for the preparation of polymer matrices for the controlled release of macromolecules.

A new method for making polymeric systems for the controlled release of macromolecular drugs is described. The method consists of mixing drug and polymer (ethylene-vinyl acetate copolymer) powders below the glass transition temperature of the polymer and compressing the mixture at a temperature above the glass transition point. The macromolecule is not exposed to organic solvent during the fabrication process. Kinetic studies indicate that there is sustained release, and the bioactivity of macromolecules tested is unchanged throughout the sintering and release processes.

Delayed-Action Preparations↗
PubMed1999

The optical biosensor studies on the role of hydrophobic tails of NADPH-cytochrome P450 reductase and cytochromes P450 2B4 and b5 upon productive complex formation within a monomeric reconstituted system.

The optical biosensor study of interaction between microsomal proteins-NADPH-cytochrome P450 reductase, cytochrome P450 2B4, and cytochrome b5-was carried out in the monomeric reconstituted system in the absence of phospholipids. The formation of individual complexes was kinetically characterized and their association and dissociation rate constants were determined. The association rate constants for the complexes formed were found to be close to the diffusiion limit-(0.5-4) x 10(6) M-1 s-1-while their dissociation rate constants did not exceed 0.5 s-1. It was shown that the interprotein electron transfer can occur both through complex formation and due to random collision. The dominant role of hydrophobic membraneous protein fragments in formation of productive electron transfer complexes was demonstrated.

Animals↗
PubMed1999

AFM study of membrane proteins, cytochrome P450 2B4, and NADPH-cytochrome P450 reductase and their complex formation.

The application of the AFM technique for visualization of membrane proteins and for measuring their dimensions was demonstrated. The AFM images of the microsomal monooxygenase system components-cytochrome P450 2B4 and NADPH-cytochrome P450 reductase-were obtained by using two types of supports-hydrophobic, highly oriented pyrolytic graphite (HOPG) and hydrophilic mica. It was shown that hemo- and flavoprotein monomers and oligomers can be adsorbed to and visualized on HOPG. On the negatively charged mica matrix, flavoprotein oligomers dissociated to monomers while hemoprotein oligomers dissociated into less aggregated particles. The images of cytochrome P450 2B4 and NADPH-cytochrome P450 reductase monomers were about 3 and 5 nm high, respectively, while the images of oligomeric forms of these proteins were about 10 and 8 nm high, respectively. We were able to observe the binary complexes composed of monomeric proteins, cytochrome P450 2B4 and its reductase and to measure the heights of these complexes (7 nm). The method is applicable for visualization of not only individual proteins but also their complexes.

Animals↗
PubMed2001

Crowding and hydration effects on protein conformation: a study with sol-gel encapsulated proteins.

We are developing an experimental system for testing the effects of macromolecular crowding and molecular confinement on protein structure. In the present study, solvent effects on the secondary structure of two proteins were examined by circular dichroism following encapsulation in the hydrated pores of a silica glass matrix by the sol-gel method. Changes in the unfolded conformations of encapsulated apomyoglobin and reduced serum albumin were analyzed after equilibration with aqueous solutions of natural osmolytes, short-chain alcohols, polyethylene glycol, and a complete series of Hofmeister cations. In many instances, the alpha-helical content of the encapsulated protein was increased by addition of solutes at concentrations that have no effect on the protein in the absence of the glass. The results are discussed from the perspective of water structure. We argue that perturbed water at the silica interface causes an increase in the average free energy of the bulk water phase which, consequently, diminishes the strength of the hydrophobic effect inside the glass matrix and destabilizes the conformation of encapsulated proteins. We propose that solutes can increase the strength of the hydrophobic effect and influence folding equilibria without directly interacting with the protein. A hypothesis is provided for the apparent paradox that kosmotropic (strongly water binding) anions favor native protein structure, whereas chaotropic (weakly water binding) cations enhance native protein structure. The encapsulation results suggest that macromolecular crowding and molecular confinement are accompanied by hydration effects that may oppose or potentiate the stabilizing effects of excluded volume on protein structure, depending on the surface chemistry of the crowding agent and its influence on bulk water structure. In the crowded environment of a living cell, excluded volume effects, surface-induced water structure, and compatible solutes are expected to complement the dominant forces in protein folding.

Apoproteins↗
PubMed1976

The role of cytochrome P-450 in the regulation of steroid biosynthesis.

A cytochrome P-450 from bovine adrenocortical mitochondria has been purified to near homogeneity. The protein catalyzes side-chain cleavage of cholesterol (cholesterol leads to pregnenolone) but neither 11beta- nor 18-hydroxylation. It consists of 16 subunits of two species (MW 52,000) and contains 8 heme groups. The enzyme has been used to determine the stoichiometry of side-chain cleavage with the following results: (TPNH and O2 consumed/mole of cleavage), cholesterol 3:3:1, 20S-hydroxycholesterol 2:2:1 and 20S,22R-dihydroxycholesterol 1:1:1. These findings support the occurrence of the proposed pathway for the side-chain cleavage of cholesterol. Cleavage of the diol is inhibited by CO and shows a characteristic P-450 photochemical action spectrum. Evidently the diol is cleaved in a typical monoxygenase reaction. The active form of the enzyme contains 16 subunits (protein 16); forms consisting of 8 (protein 8) and 4 (protein 4) subunits can be isolated and are enzymatically active only by prior conversion to protein 16.

Adrenal Cortex↗
PubMed1988

Biological effects of non-ionizing radiations: cellular properties and interactions.

The Lauriston Taylor lectures honor the founder of the National Committee on Radiation Protection and Measurement, soon to be followed by the corresponding international organization. These standard setting bodies had a vast influence on proper recognition of radiation hazards. The 10th Taylor lecture is the first to deal with nonionizing radiations and may be, therefore, of particular interest to the bioengineer. During early history biophysics and bioengineering were primarily concerned with ionizing radiation bioeffects and electrophysiology. The nonionizing part of the radiation field and electrophysiology are closely related. Biomedical observation, biophysical and bioengineering efforts in the nonionizing radiation field are defined and complement each other. Topics concentrate on the relevant biophysical and bioengineering efforts of the author and his colleagues. They include: electrical properties of biological systems; established electrical field interactions (excitation, macromolecular responses and cellular responses); problems of dosimetry (macroscopic and microscopic considerations); conclusions about relative merits of various research approaches.

Animals↗
PubMed1984

Drug targeting by drug entrapment into ultrafine compartments as carriers.

The incorporation of drugs into vesicles is one of several technological methods for the optimization of targeted drug delivery and controlled drug targeting. The main problems will always remain: To design inert auxiliary accompanying materials in order to overcome side reactions; To use body-friendly and biodegradable macromolecular carrier materials for the therapeutic system; To miniaturize the dosage form dramatically in the submicroscopic size range in order to eliminate foreign body irritations; To develop ultrafine solid and amorphous vesicular compartments (nanocapsules, nanopellets, nanoparticles) to get stable systems with good tissue transfer and organ targeting properties The actual stand of the incorporation of drugs and biologic active material into ultrafine colloidal solid capsules is reviewed here as for instance: Immunoactive material; Fluorescent indicators in body fluids; Controlled and sustained release systems Nonspecific drug targeting of the first-order (passage through endothelial tissues); Second-order targeting (a specific transparenchymal migration), and a highly specific targeting of the third-order (transcellular passage, especially lysosomal transports). Examples for some of these applications are given. It can be shown that such ultrafine vesiculated capsules offer some advantages when applied parenterally, but also partly for oral application. In the future, still more studies are necessary finally to clarify the importance and practical use of such ultrafine targeting carriers.

Animals↗
PubMed2005

Influence of osmolytes on inactivation and aggregation of muscle glycogen phosphorylase b by guanidine hydrochloride. Stimulation of protein aggregation under crowding conditions.

The effects of the osmolytes trimethylamine-N-oxide (TMAO), betaine, proline, and glycine on the kinetics of inactivation and aggregation of rabbit skeletal muscle glycogen phosphorylase b by guanidine hydrochloride (GuHCl) have been studied. It is shown that the osmolytes TMAO and betaine exhibit the highest protective efficacy against phosphorylase b inactivation. A test system for studying the effects of macromolecular crowding induced by osmolytes on aggregation of proteins is proposed. TMAO and glycine increase the rate of phosphorylase b aggregation induced by GuHCl.

Animals↗
PubMed1990

Retroviral-mediated gene transfer of the leukocyte integrin CD18 subunit.

Children with leukocyte adherence deficiency (LAD) exhibit heterogeneous defects in the leukocyte integrin CD18 subunit that prevent surface expression of functional CD11/CD18 leukocyte integrin adherence complexes. We used a retroviral vector, designated LCD18SN, to transfer the CD18 cDNA into K562 human myeloid leukemia cells and into EBV B-cells from a child with LAD. Transfer of the LCD18SN retroviral construct, which expresses the CD18 cDNA from the Moloney Murine leukemia virus (MoMLV) long terminal repeat (LTR), into K562 cells resulted in relatively high levels of CD18 mRNA and intracellular protein. Retroviral-mediated gene transfer of CD18 into LAD EBV B-cells resulted in low, but readily measurable, levels of surface expression of the CD11a/CD18 complex in these previously deficient lymphocytes. The reconstitution of surface expression of the CD11a/CD18 complex by gene transfer of the CD18 cDNA into LAD EBV B-cells indicates that this syndrome represents a candidate disorder for gene therapy.

Antigens, CD↗
PubMed1980

Binding of [125I]insulin to specific receptors and stimulation of nucleotide incorporation in cells cultured from rat brain.

The occurrence of insulin receptors and biological responses to insulin has been investigated in trypsin-dissociated fetal rat brain cells maintained in culture for 8 days. Binding of [125I]insulin to brain cells in culture was time- and pH-dependent and 85--90% specific. Porcine insulin competed for [125I]insulin binding in a dose-dependent manner. Unrelated polypeptides, including angiotensin II, glucagon, bovine growth hormone, and bovine prolactin did not compete for [125I]insulin binding. The half-life of [125I]insulin dissociation from receptors at 24 degrees C was 15 min and a plot of In[B/Bo] vs time suggested two dissociated rate constants of 2.7 X 10(-4) sec-1 and 5.0 X 10(-5) sec-1. Scatchard analysis of the binding data gave a curvilinear plot which may indicate negative cooperativity or the occurrence of both high affinity (Ka = 2 X 10(11) M-1) and low affinity (Ka = 4 X 10(10) M-1) sites. Of the estimated total of 4.9 X 10(4) binding sites per cell, 28--30% appear to be high affinity sites. Incubation of cultures with insulin caused a time- and dose-dependent stimulation of [3H]thymidine and [3H]uridine incorporation into TCA-precipitable material. Maximum stimulation of thymidine incorporation (2--5-fold) occurred 11 h after incubation with 167 nM insulin. The same concentration of insulin caused a 2.2-fold increase in [3H]uridine incorporation in 2 h. These results indicate that cells cultured from rat brain contain specific insulin receptors capable of mediating effects of insulin on macromolecular synthesis in the central nervous system.

Animals↗