Release Of Macromolecules Research Articles

An explanation for the controlled release of macromolecules from polymers

Controlled release systems composed of hydrophobic polymers such as ethylene—vinyl acetate copolymer have proven useful for releasing various polypeptides and other macromolecules for over 100 days. However, the release mechanism has never been elucidated. Evidence by microscopy is presented suggesting that release occurs through interconnecting pores formed by the macromolecules themselves. A mathematical model has been developed and used to predict the release rates of different proteins.

Response of Legionella pneumophila to beta-lactam antibiotics

Legionella pneumophila Philadelphia strain 1 grown in vitro contained five penicillin-binding proteins that were accessible to the antibiotic in membrane preparations and in live cells as well. The bacterium had reasonably low MICs of several beta-lactam antibiotics and was susceptible to both the bactericidal and the lytic activity of these drugs. An unusual feature of the response of this bacterium to penicillin treatment was that cell lysis as determined by decrease in culture turbidity and release of intracellular macromolecules was not accompanied by degradation of the peptidoglycan.

Human interleukin 1 mediates cartilage matrix degradation

Human monocyte factors mediate cartilage matrix degradation by activation of the resident chondrocytes. In the present work, the cartilage matrix-degrading activity of partially purified human monocyte-derived interleukin 1 has been investigated. Human monocyte or blood mononuclear cell culture supernatants were sequentially purified by phenyl-Sepharose and gel filtration chromatography, or by gel filtration chromatography, isoelectric focusing, or chromatofocusing. Column fractions were simultaneously tested by the standard method that defines interleukin 1 activity—costimulation of mouse thymocyte proliferation—and for matrix macromolecule release from living bovine cartilage expiants in organ culture. The two activities showed identical profiles in purification steps that would discriminate according to molecular size, hydrophobicity, and net charge. Moreover, the thermal denaturation profiles of the material purified by chromatofocusing could not distinguish between thymocyte proliferating and matrix degrading activities. These results suggest that interleukin 1 which is present in inflammatory synovial fluids may play an important role in the mediation of cartilage damage in chronic inflammatory arthritides.

30] Controlled release and magnetically modulated release systems for macromolecules

This chapter presents the methodology of formulating of polymeric delivery systems. The methods of preparing polymeric delivery systems, methods of regulating the release kinetics of these systems, and a discussion of the advantages and limitations of the systems as well as potential directions for future research in this area are presented. The basis for the slow release of macromolecules through these polymers appears to be diffusion through a series of interconnecting macrochannels in the polymer matrix. These channels do not normally exist within the polymer but are caused by the incorporation of solid or liquid in the matrix during the casting procedure. Thus, factors that increase the size of these channels or provide simpler pathways (lower tortuosity) for diffusion out of the matrix would be expected to increase release rates. Modulated delivery systems controlled by external means may ultimately improve the release pattern of certain drugs such as insulin. Modulated release systems triggered by magnetism or other mean may also be useful for delivering hormones for birth control and other therapies.

Ethylene-Vinyl Acetate Copolymer Microspheres for Controlled Release of Macromolecules

A simple technique has been devised for preparing ethylene-vinyl acetate copolymer microspheres containing macromolecular drugs. Beads with good sphericity were formed by a simple extrusion process that can easily be repeated by any laboratory without special equipment. The extruded droplets gelled on immediate contact with cold ethanol in a dry ice-ethanol bath. The ethanol also served to remove the solvent and harden the microspheres as they warmed to room temperature. Release of bovine serum albumin followed the expected trends, i.e., initial linearity with the square root of time and faster release rates with greater drug loading.

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.

Controlled release of polypeptides and other macromolecules.

The use of polymeric matrices for the controlled release of polypeptides and other macromolecular drugs is reviewed. Three principal mechanisms of release include diffusion of the polypeptide through the polymer, erosion of the polymer matrix, and the application of magnetic fields to force more drug out of the matrix. The diffusion controlled systems generally utilize ethylene-vinyl acetate copolymer. The advantage of these systems is facile manipulation of the pore structure to obtain desired release kinetics. Release of many different polypeptides from these systems for periods of months has been demonstrated. Bioerosion provides the advantage that the polymer system does not need to be retrieved. Magnetism provides a mechanism whereby desired increases and decreases in polypeptide release rates can be achieved on demand.

Controlled release of water-soluble macromolecules from bioerodible hydrogels

A bioerodible hydrogel based on water-soluble unsaturated polyesters crosslinked through the double bonds and capable of immobilizing water-soluble macromolecules has been developed. As the ester linkages cleave, the entrapped macromolecule is gradually released to the surrounding aqueous environment. In vitro rate of hydrolysis and concommitant macromolecule release can be controlled by constructing unsaturated polyesters containing varying proportions of esters activated by electron-withdrawing substituents vicinal to the ester function and/or by varying crosslink density. Polyesters containing unsaturation either in the polymer backbone or pendant have been prepared. Macromolecule-containing hydrogels have been fabricated as dessicated microspheres that can be resuspended in water and passed through a 22-gauge hypodermic needle.

In Vivo and In Vitro Release of Macromolecules from Polymeric Drug Delivery Systems

□ In vivo release rates of a macromolecule from an ethylene–vinyl acetate copolymer have been shown to be indistinguishable from those of identical implants tested in vitro. The studies were conducted for ~2months, and two different techniques were used to assess release rates. One of these techniques, using [3H]inulin as a marker, may be particularly useful in future studies assessing in vivo release rates from drug delivery systems. The appearance of [3H]inulin in the urine of rats bearing implants allowed continuous monitoring of release. A histological evaluation of tissue sections surrounding polymer implanted for 7months showed no inflammatory cell reaction.

Controlled release of macromolecules and pharmaceuticals from synthetic polypeptides based on glutamic acid.

AbstractBiocompatible, biodegradable copolymers of glutamic acid and ethyl glutamate were evaluated for their permeability to proteins ranging in molecular weight from 12,300 to 69,000. The results showed that the copolymers were sufficiently permeable that they could be used for the preparation of implantable, controlled‐release systems capable of releasing therapeutic amounts of high‐molecular‐weight drugs.

Differential Effects of Calcium Ions on Glycoconiugate Secretion by Canine Tracheal Explants

Secretion of macromolecules from different exocrine glands is known to be dependent upon calcium ions. 1 Selinger Z Naim E. The effect of calcium on amylase secretion by rat parotid slices.. Biochim Biophys Acta. 1970; 203: 335-337 Crossref PubMed Scopus (40) Google Scholar , 2 Hokin LE. Effects of calcium omission on acetylcholinestimulated amylase secretion and phospholipid synthesis in pigeon pancreas slices.. Biochim Biophys Acta. 1966; 115: 219-221 Crossref PubMed Scopus (85) Google Scholar In these studies, macromolecule release, induced by various secretagogues, is accompanied by a rise in intracellular calcium originating either from the extra cellular space or from intracellular organelles. 3 Case RM Clausen T. The relationship between calcium exchange and enzyme secretion in the isolated rat pancreas.. J Physiol. 1973; 235: 75-102 Crossref PubMed Scopus (147) Google Scholar It is not known whether the secretion of mucus by the airway is dependent on calcium ions although, in a short study of the chicken airway, Balfre 4 Balfre K. The effects of calcium and calcium ionophore A23187 on mucin secretion and potential difference in the isolated chicken trachea.. J Physiol. 1978; 275: 80-P-81P Google Scholar demonstrated both Ca2 + dependent increases and decreases in mucus secretion. In this study, we have used canine tracheal explants to investigate the effect of calcium ions on both the release of 14C-glucosamine-labeled glycoconjugates and the activity of airway secretory cells, particularly of the submucosal glands.

Root canal-simulating experimental model for evaluation of tissue responses to chronic influx of foreign fluid substances.

Searching for an experimental model simulating the root canal, a two-stage procedure was evolved using polyethylene tubes implanted subcutaneously in rabbits. The reaction to the model was histologically evaluated, and the rate of release of macromolecules from the tubes was studied using radio-labeled endotoxin and albumin. The model proved suitable for evaluating an animal's tissue response to the slow and prolonged influx of foreign fluids.

Magnetic modulation of release of macromolecules from polymers.

Sustained-release systems were made by incorporating bovine serum albumin and magnetic steel beads in an ethylene-vinyl acetate copolymer matrix. When exposed to aqueous medium, the polymer matrix released the albumin slowly and continuously. Application of an oscillating magnetic field increased the release rate by as much as 100%. Intervals of 6-hr periods of magnetic exposure and nonexposure were alternated over a 5-day period, resulting in corresponding increases and decreases in release and establishing a pattern of modulated sustained release.

Direct measurement of the increase in intracellular free calcium ion concentration in response to the action of complement.

1. The effect of rabbit anti-(pigeon erythrocyte) antibodies plus human complement on the concentration of intracellular free Ca2+ in sealed pigeon erythrocyte 'ghosts' was investigated with the photoprotein obelin. 2. The addition of human serum, as a source of complement, to 'ghosts' coated with antibody caused a rapid increase in intracellular free Ca2+ after a lag of 20-40 s, as detected by an increase in obelin luminescence. 3. The increase in obelin luminescence could not be explained by release of obelin into the medium. It was also Ca2+-dependent in that extracellular EGTA abolished the effect and intracellular EGTA inhibited it and required the complete terminal complex (C56789). No effect was seen with C5678. 4. The concentration of intracellular free Ca2+ before addition of complement was approx. 0.3 microM. This increased to a maximum of 5-30 microM after complement addition and then remained constant for at least 1-2 min. 5. Antibody plus complement induced a rapid increase in 42K+ efflux and an inhibition of cyclic AMP formation. 6. When partially purified complement components (C5b-9) were used in 'reactive lysis' it was possible to inhibit the release of macromolecules from pigeon erythrocyte 'ghosts' by extracellular EGTA. 7. It was concluded that the increase in intracellular free Ca2+ concentration caused by anti-cell antibody plus complement occurred before cell lysis and may be involved in the mechanism of complement-induced cell injury.

4] Polymers for the sustained release of macromolecules: Their use in a single-step method of immunization

This chapter describes the methodology for fabricating and utilizing the polymeric systems for the sustained release of macromolecules. Polymeric delivery systems could provide a simple, safe, and effective single-step immunization. Polyvinyl alcohol or ethylene-vinyl acetate copolymer may cause a mild inflammatory response. This can be eliminated for ethylene-vinyl acetate, or reduced for polyvinyl alcohol, by washing with solvents to extract impurities. It is found that to retard the diffusion of the macromolecules from the polymer matrix, the pellets were sometimes coated with pure polymer solution. Coating was accomplished by one of the two procedures. To test the effectiveness of the polymers in inducing an immune response, a method was used to prepare ethylene-vinyl acetate copolymer pellets. The polymer implants were made by mixing 5 mg of dry protein powder with 100 μl of polymer solution. Prolonged stimulation with antigen encapsulated in a polymer increased antibody production with time and did not induce tolerance. The antibodies formed were the IgG types. Mice implanted with polymer pellets showed no early decline in antibody formation, as was observed, with the complete Freund's adjuvant-sponsored primary response in controls.

Polymers for Sustained Macromolecule Release: Procedures to Fabricate Reproducible Delivery Systems and Control Release Kinetics

Matrixes composed of ethylene–vinyl acetate copolymer are useful vehicles for the sustained release of macromolecules such as proteins. A new procedure for fabricating these polymeric delivery systems involved mixing the dry, powdered macromolecule with a polymer solution and casting the mixture at −80°. The resulting matrix was dried in two 48-hr stages, first at −20° and then at 20°. These polymer systems had uniform drug distribution, and their release kinetics were reproducible. Fabrication parameters such as drug particle size, drug loading, and matrix coating all significantly affected release kinetics.

Dopamine beta-hydroxylase in health and disease.

DBH is a copper-containing oxygenase that catalyzes the hydroxylation of the beta carbon of a wide variety of phenylethylamine derivatives using molecular oxygen ascorbate as cofactors. It is a glycoprotein with a molecular weight of 290,000 and consists of four identical subunits, each with a single copper atom and 5% carbohydrate by weight. The enzyme is a constituent of catecholamine storage vesicles in chromaffin cell and adrenergic neurons in the peripheral and central nervous system where it functions to synthesize noradrenaline from dopamine. Although endogenous inhibitors have been isolated, they have not been demonstrated to have a physiological function, and the kinetics of the enzyme in vitro and in vivo suggest that the enzyme is not a rate limiting step in catecholamine synthesis under normal conditions. DBH exists in both a soluble form within vesicles and as a constituent of their membranes with its active site directed inward. The significance of the partition of the enzyme into soluble and membrane forms is not understood, although the soluble form has a fivefold greater homospecific activity. DBH has been one of the most intensively investigated enzymes in neurochemistry for several reasons. It is a readily assayable constitutent of catecholamine storage vesicles and, as such, provides a convenient biochemical marker for subcellular fractionation work and studies of the cellular regulation of catecholamine synthesis, storage, and release. The adrenal medulla is a rich source of the enzyme for purification, and the purified enzyme is highly antigenic, thereby enabling the use of several immunological techniques to study the cellular dynamics of the enzyme and the organelles in which it is located. These include radioimmunoassay, immunohistochemistry, and cytochemistry. This review firstly summarizes the present state of knowledge concerning the molecular properties of DBH. It then describes the tissue, cellular, and subcellular localization of the enzyme and its physiological regulation. The remainder of the review concentrates on those aspects of research on DBH in which the authors have participated that have led to general advances such as the development of the concept of homospecific activity, the introduction of immunohistochemistry for the localization of enzymes involved in transmitter metabolism, the release of macromolecules from synaptic vesicles during the process of exocytosis, the use of antibodies to DBH administered in vivo to study the fate of synaptic vesicle membranes and to produce specific immunological lesions of noradrenergic nerves in the peripheral and central nervous system, the genetic, environmental, and physiological determinants of serum DBH activity as an index of sympathetic function in animals and man, and the question of its diagnostic value in disease.

Metabolism of cell surface-associated sulfated glycosaminoglycans in cultured human smooth muscle cells.

Smooth muscle cells grown from human aorta synthesize chondroitin sulfate, dermatan sulfate, and heparan sulfate as sulfated glycosaminoglycans. These polymers are found mainly extracellularly and in association with the cell membrane. Each compartment is characterized by a distinct distribution pattern of sulfated glycosaminoglycans though considerable variability was noted between different cell lines. On incubation of the cells in the presence of [35S]sulfate for up to 72 h no significant change in the distribution pattern of newly synthesized extracellular and membrane-associated glycosaminoglycans was found. Prelabeling experiments revealed that pericellular glycosaminoglycans are metabolically heterogenous. they leave their compartment with half-lives of less than 10 h and 1--3 days respectively. During the initial period of the chase experiment (up to 12 h) about equal proportions of the material disappeared either by shedding into the culture medium or by endocytosis. Thereafter, release of macromolecules into the medium exceeded endocytotic uptake. None of the individual glycosaminoglycans on the cell surface showed a clear preference for its removal either by shedding or by endocytosis. It is concluded that cell surface-associated glycosaminoglycans are neither direct precursors of the extracellular glycosaminoglycans nor do they represent mainly extracellular glycosaminoglycans which are in the process of endocytosis.

Stimulation of Synthesis and Release of Lipids in Tumor Cells Under Attack by Antibody Plus Complement

Abstract Resistance to antibody-complement- (Ab-C) mediated killing of guinea pig hepatoma cells (line-1 and line-10) correlates with the ability of the cells to synthesize lipids, but not DNA, RNA, protein, or complex carbohydrate. We therefore studied the synthesis and release of macromolecules (especially lipids) by tumor cells under attack by Ab plus C. Ab-sensitized tumor cells (TA) incorporated increased amounts of acetate, glycerol, or fatty acids into complex lipids compared to control cells within 5 to 10 min after the addition of guinea pig C (GPC) to the cells (TAC). TA, TC, or TAΔC (Ab plus heat-inactivated GPC) did not show this effect. No enhancement of DNA, RNA, or protein synthesis in TAC as compared with control cells was noted. Line-1 cells, which are killed by GPC when sensitized with excess anti-Forssman Ab (TAxs), demonstrated enhanced lipid synthesis within 1 to 3 min after the addition of GPC to TAxs, before measurable killing of the cells had occurred.

Mechanisms of cellular enzyme release. I. Alteration in membrane fluidity and permeability.

Above-normal plasma enzyme activities resulting from increased release of intracellular macromolecules are an important diagnostic feature of Duchenne muscular dystrophy. These and other biochemical and histological characteristics of this disease are faithfully duplicated when imipramine and serotonin are administered to the rat. Imipramine, but not serotonin alone, causes release of enzymes from rat diaphragms and human lymphocytes and release of hemoglobin from erythrocytes in vitro. Ouabain causes neither. Imipramine-induced enzyme release is decreased by adding ATP to the specimen in vitro or in hypertonic solution. Imipramine inhibits the capping phenomenon (an aggregation of antigen/antibody complexes of the membrane) of human B lymphocytes labeled with fluorescein-conjugated antihuman immunoglobulins. Serotonin alone has no such effect, but, administered together with imipramine, it potentiates the inhibition of capping by imipramine.