Biomedical subjects

Allan S Hoffman

Publications and source records attributed to Allan S Hoffman.

At least 19 recordsLinked to original sources

PubMed2006

Switchable surface traps for injectable bead-based chromatography in PDMS microfluidic channels.

We report here a reversible microchannel surface capture system for stimuli-responsive grafted bioanalytical beads. Poly(N-isopropylacrylamide) (PNIPAAm) was grafted onto polydimethylsiloxane (PDMS) surfaces by a UV-mediated graft polymerization from a photoinitiator that was preadsorbed in the channel wall. The surface grafting density and resulting switchable hydrophilic/hydrophobic properties were controlled by varying the photo-illumination times and/or the initiator concentration. At limiting PNIPAAm-graft densities, the surfaces demonstrated minimal contact angles of 35 degrees below the lower critical solution temperature (LCST) and maximal contact angles of 82 degrees above it. These contact angles could be varied depending on the graft density. The surface grafts are spatially limited to the photo-illuminated region to define where the trap is constructed. The surface traps capture PNIPAAm-grafted nanobeads uniformly above the LCST and facilitate their rapid release as the temperature is reversed to below the LCST. This dual surface trap and injectable chromatography system could be useful in many applications, such as affinity separations, immunoassays, and enzyme bioprocesses, by providing for the controlled capture and release of chromatography beads.

Acrylic Resins↗

PubMed2006

Preparation and characterization of biocompatible polyelectrolyte complex multilayer of hyaluronic acid and poly-L-lysine.

A novel biocompatible polyelectrolyte complex multilayer (PECML) was successfully prepared using hyaluronic acid (HA) and poly-l-lysine (PLL). The formation of PECML through the electrostatic interaction of HA as a polyanion and PLL as a polycation was confirmed by contact angle measurement, ESCA analysis and HA content analysis. According to the Carbazole assay, HA content increased rapidly up to eight cycles for HA/PLL deposition and then slightly increased with an increasing number of deposition cycle. In vitro release of PLL from the PECML continued up to 4 days exhibiting different release profiles depending on the outer layer of PECML. This result provides evidence for PLL diffusion throughout PECML of HA and PLL during the multilayer buildup. About 25% of HA remained on the cover glass after the in vitro release test for 7 days. From the results, we confirmed that PECML of HA and PLL could remain at least partially on the chitosan-coated cover glass for 7 days. The surface modification with PECML resulted in drastically reduced peripheral blood mononuclear cell (PBMC) attachment according to the lactate dehydrogenase assay for cell counting. This nano-scale control of material deposition may be successfully applied for surface modification of various biomaterials.

Cell Adhesion↗

PubMed2006

Poly(N-isopropylacrylamide-co-propylacrylic acid) copolymers that respond sharply to temperature and pH.

Temperature- and pH-sensitive random copolymers of N-isopropylacrylamide (NIPAAm) and propylacrylic acid (PAA) were prepared using the reversible addition fragmentation chain transfer (RAFT) polymerization method. The lower critical solution temperatures (LCSTs) (or phase separation temperatures) of the NIPAAm-co-PAA copolymer solutions were measured by the cloud-point method. At slightly acidic conditions, the LCST decreased with increase in PAA content, which suggests that the hydrophobic propyl group of PAA has a greater influence on the LCST than the polar carboxylic acid group at those conditions. An increase of pH led to a significant increase in LCST of the copolymers due to the ionization of the -COOH group. The LCSTs were studied as a function of copolymer composition over the pH range from 5.0 to 7.0. Because the pK(a) of the polymers can be tuned to fall close to neutral pH, these polymer compositions can be designed to have phase transitions triggered near physiological pH or at slightly acidic pH values that fall within acidic gradients found in biology. The NIPAAm-co-PAA copolymers thus display tunable properties that could make them useful in a variety of molecular switching and drug delivery applications where responses to small pH changes are relevant.

Acrylamides↗

PubMed2006

pH-responsive poly(styrene-alt-maleic anhydride) alkylamide copolymers for intracellular drug delivery.

Many macromolecular therapeutics such as peptides, proteins, antisense oligodeoxynucleotides (ASODN), and short interfering RNA (siRNA) are active only in the cytoplasm or nucleus of targeted cells. Endocytosis is the primary route for cellular uptake of these molecules, which results in their accumulation in the endosomal-lysosomal trafficking pathway and loss of therapeutic activity. In this article, we describe the synthesis and pH-dependent membrane-destabilizing activity of a new "smart" polymer family that can be utilized to enhance the intracellular delivery of therapeutic macromolecules through the endosomal membrane barrier into the cytoplasm of targeted cells. These polymers are propylamine, butylamine, and pentylamine derivatives of poly(styrene-alt-maleic anhydride) (PSMA) copolymers. The PSMA-alkylamide derivatives are hydrophilic and membrane-inactive at physiological pH; however, they become hydrophobic and membrane-disruptive in response to endosomal pH values as measured by their hemolytic activity. Results show that the pH-dependent membrane-destabilizing activity of PSMA derivatives can be controlled by varying the length of the alkylamine group, the degree of modification of the copolymer, and the molecular weight of the PSMA copolymer backbone. Butylamine and pentylamine derivatives of PSMA copolymers exhibited more than 80% hemolysis at endosomal pH values, which suggests their potential as a platform of "smart" polymeric carriers for enhanced cytoplasmic delivery of a variety of therapeutic macromolecules.

Animals↗

PubMed2006

Controlling the aggregation of conjugates of streptavidin with smart block copolymers prepared via the RAFT copolymerization technique.

Block copolymers containing stimuli-responsive segments provide important new opportunities for controlling the activity and aggregation properties of protein-polymer conjugates. We have prepared a RAFT block copolymer of a biotin-terminated poly(N-isopropylacrylamide) (PNIPAAm)-b-poly(acrylic acid) (PAA). The number-average molecular weight (M(n)) of the (PNIPAAm)-b-(PAA) copolymer was determined to be 17.4 kDa (M(w)/M(n) = 1.09). The PNIPAAm block had an M(n) of 9.5 kDa and the poly(acrylic acid) (PAA) block had an M(n) of 7.9 kDa. We conjugated this block copolymer to streptavidin (SA) via the terminal biotin on the PNIPAAm block. We found that the usual aggregation and phase separation of PNIPAAm-SA conjugates that follow the thermally induced collapse and dehydration of PNIPAAm (the lower critical solution temperature (LCST) of PNIPAAm is 32 degrees C in water) is prevented through the shielding action of the PAA block. In addition, we show that the cloud point and aggregation properties (as measured by loss in light transmission) of the [(PNIPAAm)-b-(PAA)]-SA conjugate also depended on pH. At pH 7.0 and at temperatures above the LCST, the block copolymer alone was found to form particles of ca. 60 nm in diameter, while the bioconjugate exhibited very little aggregation. At pH 5.5 and 20 degrees C, the copolymer alone was found to form large aggregates (ca. 218 nm), presumably driven by hydrogen bonding between the -COOH groups of PAA with other -COOH groups and also with the -CONH- groups of PNIPAAm. In comparison, the conjugate formed much smaller particles (ca. 27 nm) at these conditions. At pH 4.0, however, large particles were formed from the conjugate both above and below the LCST (ca. 700 and 540 nm, respectively). These results demonstrate that the aggregation properties of the block copolymer-SA conjugate are very different from those of the free block copolymer, and that the outer-oriented hydrophilic block of PAA shields the intermolecular aggregation of the block copolymer-SA bioconjugate at pH values where the -COOH groups of PAA are significantly ionized.

Acrylamides↗

PubMed2006

PEG-cross-linked heparin is an affinity hydrogel for sustained release of vascular endothelial growth factor.

An affinity-based controlled release system for growth factors having heparin-binding domains was prepared using a cross-linked heparin gel. The heparin gel was made by reacting hydrazide-functionalized heparin (Hep-ADH) with the N-hydroxysuccinimidyl ester of poly(ethylene glycol)-bis-butanoic acid (SBA-PEG-SBA). The degree of cross-linking could be controlled by defining the stoichiometry of hydrazide modification and the PEG cross-linker addition. The release of vascular endothelial growth factor (VEGF) was characterized as a heparin-binding growth factor. VEGF was directly injected into the heparin gel and the loaded VEGF displayed a slow, controlled release over 3 weeks with little initial burst phase. The biological activity of the released VEGF was measured with a proliferation assay utilizing human umbilical vein endothelial cells. The released VEGF maintained its biological activity at all time points investigated. The heparin gel with loaded VEGF was implanted sub-cutaneously in the dorsal region of mice. A significantly increased density of the endothelial cell marker platelet endothelial adhesion molecule (PECAM-1) was observed in histological specimens of the tissues surrounding the implanted gel.

Animals↗

PubMed2005

Rational design of composition and activity correlations for pH-responsive and glutathione-reactive polymer therapeutics.

Limited cytoplasmic delivery of enzyme-susceptible drugs remains a significant challenge facing the development of protein and nucleic acid therapies that act in intracellular compartments. "Smart" pH-responsive, membrane-destabilizing polymers present a new approach to shuttling therapeutic molecules past the endosomal membrane and into the cytoplasm of targeted cells. This report describes the use of a functionalized monomer, pyridyl disulfide acrylate (PDSA), to develop pH-responsive, membrane-destabilizing, and glutathione-reactive polymers by copolymerization with several pH-responsive and hydrophobic monomers. The activity of the carriers is described as a function of (a) increasing the length of the hydrophobic alkyl group substituted onto the pH-responsive monomer and (b) the incorporation of a hydrophobic monomer such as butyl acrylate (BA) on the pH sensitivity and membrane-destabilizing activity of new polymer compositions. The membrane-destabilizing activity of different polymer compositions was evaluated as a function of pH and polymer concentration using the red blood cell (RBC) hemolysis assay. Hemolysis results show that the increase in the hydrophobic character of the polymer backbone results in a shift in the pH sensitivities and an increase in the membrane-destabilizing activity. Results show that the observed hemolytic activities and pH sensitivity profiles could be designed across a range that matches the properties needed for enhancing the cytoplasmic delivery of macromolecular therapeutic.

Acrylates↗

PubMed2005

Rational design of composition and activity correlations for pH-sensitive and glutathione-reactive polymer therapeutics.

Limited cytoplasmic delivery of enzyme-susceptible drugs remains a significant challenge facing the development of protein and nucleic acid therapies that act in intracellular compartments. "Smart" pH-sensitive, membrane-destabilizing polymers present an attractive approach to shuttle therapeutic molecules past the endosomal membrane and into the cytoplasm of targeted cells. This report describes the use of a new functionalized monomer, pyridyl disulfide acrylate (PDSA), to develop pH-sensitive, membrane-destabilizing, and glutathione-reactive polymers by copolymerization with several pH-sensitive and hydrophobic monomers. The activity of the carriers is described as a function of (a) the influence of increasing the length of the hydrophobic alkyl group substituted onto the pH-sensitive monomer and (b) of the effect of incorporating a hydrophobic monomer such as butyl acrylate (BA) on the pH sensitivity and membrane-destabilizing activity of new polymer compositions. The membrane-destabilizing activity of different polymer compositions was evaluated as a function of pH and polymer concentration using the red blood cells (RBC) hemolysis assay. Hemolysis results show that the increase in the hydrophobic character of polymer backbone results in a shift in the pH sensitivity profile and an increase in the membrane-destabilizing activity. Results show that the observed hemolytic activities and pH sensitivity profiles could be designed across a range that matches the properties needed for drug carriers to enhance the cytoplasmic delivery of therapeutic cargos.

Acrylates↗

PubMed2005

Design and development of polymers for gene delivery.

The lack of safe and efficient gene-delivery methods is a limiting obstacle to human gene therapy. Synthetic gene-delivery agents, although safer than recombinant viruses, generally do not possess the required efficacy. In recent years, a variety of effective polymers have been designed specifically for gene delivery, and much has been learned about their structure-function relationships. With the growing understanding of polymer gene-delivery mechanisms and continued efforts of creative polymer chemists, it is likely that polymer-based gene-delivery systems will become an important tool for human gene therapy.

Animals↗

PubMed2005

Poly(propylacrylic acid)-mediated serum stabilization of cationic lipoplexes.

The serum instability associated with cationic lipoplexes represents one of the major obstacles for the in vivo delivery of nonviral gene therapy vectors. Recently, we have shown that poly(propylacrylic acid) (PPAA), a pH-sensitive polyanionic polymer, can significantly improve the in vitro serum stability of DOTAP lipoplexes and enhance transfection (Cheung et al., Bioconjug. Chem. 12, 906 (2001)). We investigated this serum-stabilizing effect provided by PPAA using methods to identify the specific serum proteins that interact with DOTAP/DNA and DOTAP/DNA/PPAA lipoplexes and determined their modes of interaction with these lipoplexes. Studies showed that only low-density lipoprotein (LDL) caused significant decondensation of DNA from lipoplexes lacking PPAA, but that fully condensed DNA was retained within lipoplexes incorporating PPAA. Another major factor in the loss of transfection activity was due to the reduced cellular uptake of DOTAP lipoplexes upon exposure to serum, with bovine serum albumin (BSA) and high-density lipoprotein (HDL) acting as major contributors to this reduction in vector internalization. In contrast, lipoplexes containing PPAA maintained high levels of uptake into cells in the presence of these proteins. Transfection results generally concurred with the mechanistic studies, suggesting that maintaining effective cellular delivery of intact lipoplexes in the presence of serum proteins is important to retain high transfection efficiencies. These results indicate that the addition of PPAA as a ternary component in DOTAP lipoplexes can overcome some of the serum-related deficiencies encountered with these lipoplexes to provide efficient transfection.

Acrylates↗

PubMed2005

Immobilization of streptavidin-horseradish peroxidase onto a biotinylated poly(acrylic acid) backbone that had been radiation-grafted to a PTFE film.

Films of polytetrafluoroethylene (PTFE) were modified by radiation graft polymerization of acrylic acid (AAc). Optimal conditions for efficient AAc grafting were studied, including pre-irradiation dose in air, monomer concentration, temperature and time of the grafting process. Carboxylic groups of the grafted polyAAc were activated with carbodiimide (EDC) for biotinylation by reaction with 5-(biotinamido) pentylamine. Streptavidin-horseradish peroxidase (SA-HRP) was immobilized by affinity complexation of the SA with the biotin groups on the PTFE surface. The amount of active HRP immobilized on the PTFE films was determined as a function of the extent of polyAAc grafting. This study has demonstrated the utility of combining the processes of (a) radiation grafting of polymers with reactive groups onto inert polymers such as PTFE, (b) biotinylation of the graft polymer reactive groups, (c) immobilization of streptavidin on the biotinylated surface sites, followed by (d) immobilization of biotinylated, biologically active molecules via complexation of their conjugates with streptavidin. In this study, the last two steps were combined by immobilizing the complex of streptavidin and biotinylated HRP onto the biotinylated surface sites. The unique nature of this process is the ability to immobilize biotinylated molecules on an inert surface as PTFE.

Acrylic Resins↗

PubMed2005

Semi-interpenetrating network of poly(ethylene glycol) and poly(D,L-lactide) for the controlled delivery of protein drugs.

We have prepared a semi-interpenetrating network (IPN) of poly(ethylene glycol) dimethacrylate (PEGDMA) with entrapped poly(D,L-lactide) (PLA) using photochemical techniques. These IPNs were developed for the controlled delivery of protein drugs such as growth factors. The PEG component draws water into the network, forming a hydrogel within the PLA matrix, controlling and facilitating release of the protein drug, while the PLA component both strengthens the PEG hydrogel and enhances the degradation and elimination of the network after the protein drug is released. The rate and extent of swelling and the resultant protein release kinetics could be controlled by varying the PEG/PLA ratio and total PLA content. These IPNs were prepared using a biocompatible benzyl benzoate/benzyl alcohol solvent system that yields a uniform, fine dispersion of the protein throughout the PEG/PLA IPN matrix. IPNs composed of high molecular mass PLA and lower PEG/PLA ratios exhibited lower equilibrium swelling ratios. The release of bovine serum albumin (BSA), a model protein, from these IPNs was characterized by a large initial burst, regardless of the PEG/PLA ratio, due to the entrapment of residual solvent within the network. Microparticles of the PEG/PLA IPNs were also prepared using a modified Prolease strategy. Residual solvent removal was significantly enhanced using this process. The microparticles also exhibited a significant reduction in the initial burst release of protein. Mixtures of different compositions of PEG/PLA microparticles should be useful for the delivery of a variety of protein drugs with different release kinetics from any tissue-engineering matrix.

Acrylates↗

PubMed2005

Monocyte activation on polyelectrolyte multilayers.

The adherence and activation of primary human monocytes was investigated on a polyelectrolyte multilayer film containing hyaluronic acid (HA) and poly-L-lysine (PLL). The sequential layer-by-layer deposition of the multilayer film was characterized by surface plasmon resonance. Eight alternating bilayers displayed an effective thickness of 16.15 nm with a total polymer coverage of 2.10 microg/cm2. For cell studies, HA-PLL multilayers were constructed on tissue culture polystyrene (TCPS) substrates and characterized by time of flight second ion mass spectrometry (ToF-SIMS) analysis. Principal component analysis of the ToF-SIMS spectra resolved no significant difference in surface chemistry between PLL-terminated and HA-terminated multilayer surfaces. Monocyte adhesion on PLL- and HA-terminated surfaces was measured by the lactate dehydrogenase assay and showed a significant decrease in cell adhesion after 24 h incubation. Cell viability measured by Live/Dead fluorescent staining showed significant cell death in the adherent cell population over these 24 h. Tumor necrosis factor-alpha (TNF-alpha) production, a measure of monocyte activation, was quantified by ELISA and normalized to the number of adherent monocytes. The activation of monocytes on PLL-terminated and HA-terminated surfaces was nearly identical, and both surfaces had TNF-alpha levels that were 8-fold higher than TCPS. These results demonstrate that sufficient PLL had diffused into the surface layer to direct monocyte adherence and to induce cytokine activation and cell death on the HA-terminated multilayer films. The diffusion of the second multilayer component to the coating surface should, thus, be taken into account in the design of polyelectrolyte-based biomaterial coating strategies.

Cell Adhesion↗

PubMed2005

Smart polymeric carriers for enhanced intracellular delivery of therapeutic macromolecules.

Limited cytoplasmic delivery of enzyme-susceptible drugs remains a significant challenge facing the development of protein and nucleic acid therapies that act in intracellular compartments. Researchers have examined several approaches, including fusogenic proteins and protein transduction domains, to enhance the intracellular delivery of the therapeutic cargo. This review summarises efforts to develop 'smart' pH-sensitive and membrane-destabilising polymers that can shuttle therapeutic peptide, protein and nucleic acid molecules past the endosomal membrane into the cytoplasm of targeted cells. Several classes of 'smart' non-degradable polymeric carriers have been developed that have proved effective both in vitro and in vivo in enhancing the cytoplasmic delivery of a variety of therapeutic molecules.

Animals↗

PubMed2004

Hyaluronic acid grafting mitigates calcification of glutaraldehyde-fixed bovine pericardium.

Pathologic calcification is the leading cause of the clinical failure of glutaraldehyde-fixed bovine pericardium used in bioprosthetic valves. A novel surface modification of glutaraldehyde fixed bovine pericardium was carried out with high molecular weight hyaluronic acid (HA). HA was chemically modified with adipic dihydrazide (ADH) to introduce hydrazide functional groups onto the HA backbone. Glutaraldehyde-fixed bovine pericardium (GFBP) was modified by grafting this HA to the free aldehyde groups on the tissue via the hydrazide groups. Following a 2-week subcutaneous implantation in osteopontin (OPN)-null mice, the calcification of HA-modified bovine pericardium was drastically reduced (by 84.5%) compared to positive controls (tissue without HA-modification) (p = 0.005). The calcification-mitigating effect of HA surface modification was also confirmed by microscopic analysis of explanted tissue stained with Alizarin Red S for calcium.

Animals↗

PubMed2004

"Smart" mobile affinity matrix for microfluidic immunoassays.

There is a current need for simple methods for immobilizing biomolecules within microfluidic channels. Here, a technique is reported for reversibly immobilizing immunoassay components in a channel zone that can be simply controlled by integrated heating elements. Latex beads were modified with the temperature-responsive polymer poly(N-isopropylacrylamide)(PNIPAAm) and co-modified with biotinylated poly(ethylene glycol)(PEG). PNIPAAm undergoes a hydrophilic-to-hydrophobic transition when the temperature is raised above the lower critical solution temperature (LCST)( approximately 28 degrees C in the solutions used here). This reversible transition drives the aggregation and dis-aggregation of the modified beads in heated zones within poly(ethylene terephthalate)(PET) microchannels. Biotinylated monoclonal antibodies for the drug digoxin were bound via streptavidin to the biotin-PEG-coated beads. These antibody-functionalized beads were then reversibly immobilized by aggregation and hydrophobic adhesion to the surface of PET microfluidic channels in response to a thermal stimulus. The antibodies on the beads immobilized in the channel were shown to bind digoxin and a competitor fluorescent ligand from a flow stream in a quantitative competitive assay format that reported the digoxin concentration. The antibodies could be replenished for each immunoassay trial, using the reversible, temperature-controlled immobilization process. This technique allows reagent immobilization immediately prior to an analytical procedure, following the removal of previously utilized beads, guaranteeing fresh and active immobilized biomolecules. Furthermore, it provides a simple approach to multiplexing through the simultaneous or sequential injection of different antibody-coated bead species, potentially at multiple sites in the integrated device channels.

Biotin↗

PubMed2004

Effect of polymer surface activity on cavitation nuclei stability against dissolution.

The persistence of acoustic cavitation in a pulsed wave ultrasound regime depends upon the ability of cavitation nuclei, i.e., bubbles, to survive the off time between pulses. Due to the dependence of bubble dissolution on surface tension, surface-active agents may affect the stability of bubbles against dissolution. In this study, measurements of bubble dissolution rates in solutions of the surface-active polymer poly(propyl acrylic acid) (PPAA) were conducted to test this premise. The surface activity of PPAA varies with solution pH and concentration of dissolved polymer molecules. The surface tension of PPAA solutions (55-72 dynes/cm) that associated with the polymer surface activity was measured using the Wilhelmy plate technique. Samples of these polymer solutions then were exposed to 1.1 MHz high intensity focused ultrasound, and the dissolution of bubbles created by inertial cavitation was monitored using an active cavitation detection scheme. Analysis of the pulse echo data demonstrated that bubble dissolution time was inversely proportional to the surface tension of the solution. Finally, comparison of the experimental results with dissolution times computed from the Epstein-Plesset equation suggests that the radii of residual bubbles from inertial cavitation increase as the surface tension decreases.

Biocompatible Materials↗

PubMed2004

Grafting copolymerization of 2-methacryloyloxyethyl phosphorylcholine (MPC) onto pre-irradiated cellulose films.

Grafting of 2-methacryloyloxyethyl phosphorylcholine (MPC) and poly(ethylene glycol mathacrylate) (PEGMA) onto cellulose films was performed using the pre-irradiation grafting method. The effects of monomer concentration, solvent system and co-solvent composition, and reaction time on the degree of grafting were determined. The grafted samples were confirmed by FT-IR-ATR spectra. The blood compatibilities of the grafted cellulose were evaluated by platelet-rich plasma contact studies and viewed by scanning electron microscopy; non-grafted cellulose film sample was used as references. As a result, MPC and MPC/PEGMA were grafted on the surface of cellulose films. It was found that fewer platelets adhered to the MPC-grafted surfaces and that they showed less shape variation than the ungrafted references.

Biocompatible Materials↗