Investigation on injectable, thermally and physically gelable poly(ethylene glycol)/poly(octadecanedioic anhydride) amphiphilic triblock co-polymer nanoparticles

A family of injectable, biodegradable and thermosensitive co-polymer nanoparticle (NP) hydrogels based on mPEG-b-POA-b-mPEG, which was synthesized from mono-methoxy poly(ethylene glycol) (mPEG) and poly(octadecanedioic anhydride) (POA), was investigated in this paper. It was found that the aqueous dispersions of these NPs underwent a reversible gel-sol transition upon temperature change. By using paclitaxel and Bovine serum albumin (BSA) as model drugs, we noticed that the in vitro releases of both in situ gel-forming formulations were sustained and no initial burst releases were observed for 7 days. In vitro cytotoxicity tests via MTT assay indicate that mPEG-b-POA-b-mPEG NPs are non-toxic to normal mouse lung fibroblast cells (L929). The in vivo hydrogel formation and in vivo biocompatibility of co-polymer NP hydrogel were also investigated and the results further validate the biocompatible nature of co-polymer NP hydrogel. In conclusion, our mPEG-b-POA-b-mPEG NP hydrogel is able to control the release of incorporated drug for longer duration.     

Mechanical and microstructural properties of hybrid poly(ethylene glycol)-soy protein hydrogels for wound dressing applications

Biomimetic hydrogel made of poly(ethylene glycol) and soy protein with a water content of 96% has been developed for moist wound dressing applications. In this study, such hybrid hydrogels were investigated by both tensile and unconfined compression measurements in order to understand the relationships between structural parameters of the network, its mechanical properties and protein absorption in vitro. Elastic moduli were found to vary from 1 to 17 kPa depending on the composition, while the Poisson's ratio (approximately 0.18) and deformation at break (approximately 300%) showed no dependence on this parameter. Further calculations yielded the crosslinking concentration, the average molecular weight between crosslinks (M(C)) and the mesh size. The results show that reactions between PEG and protein create polymeric chains comprising molecules of PEG and protein fragments between crosslinks. M(C) is three times higher than that expected for a "theoretical network." On the basis of this data, we propose a model for the 3D network of the hydrogel, which is found to be useful for understanding drug release properties and biomedical potential of the studied material.     

Polyethylenimine-grafted copolymer of poly(l-lysine) and poly(ethylene glycol) for gene delivery

A major challenge in gene therapy is the development of effective gene delivery vectors with low toxicity. In the present study, linear poly(ethylenimine) (lPEI) with low molecular weight was grafted onto the block copolymer (PPL) of poly(l-lysine) (PLL) and poly(ethylene glycol)(PEG), yielding a ternary copolymer PEG-b-PLL-g-lPEI (PPI) for gene delivery. In such molecular design, PLL, lPEI and PEG blocks were expected to render the vector biodegradability, proton buffering capacity, low cationic toxicity and potentially long circulation in vivo, respectively. Given proper control of molecular composition, the copolymers demonstrated lower cytotoxicity, proton buffering capacity, ability to condense pDNA and mediate effective gene transfection in various cell lines. With folate as an exemplary targeting ligand, the FA-PPI/pDNA complex showed much higher transgene activity than its nontargeting counterpart for both reporter and therapeutic genes in folate receptor(FR)-positive cells. FA-PPI mediated effective transfection of the TNF-related apoptosis-inducing ligand gene (TRAIL) in human hepatoma Bel 7402 cells, leading to cell apoptosis and great suppression of cell viability. Our results indicate that the copolymers might be a promising vector combining low cytotoxicity, biodegradability, and high gene transfection efficiency.     

Bilayered biodegradable poly(ethylene glycol)/poly(butylene terephthalate) copolymer (Polyactive) as substrate for human fibroblasts and keratinocytes.

The purpose of this study was to find an optimal polymer matrix and to optimize the culture conditions for human keratinocytes and fibroblasts for the development of a human skin substitute. For this purpose porous, dense bilayers made of a block copolymer of poly(ethylene glycol terephthalate) (PEGT) and poly(butylene terephthalate) (PBT; Polyactivetrade mark) with a PEGT/PBT weight ratio of 55/45 and a PEG molecular weight (MW) of 300, 600, 1000, or 4000 Da were used. The best performance was achieved with PEGT/PBT copolymer with MW of PEG 300 D (300PEG55PBT45). When fibroblasts were seeded into the porous underlayer and cultured for 3 weeks in medium supplemented with 100 microg/mL ascorbic acid, all pores were filled with fibroblasts and with extracellular matrix, which was judged from the presence of collagen types I, III, and IV, and laminin. When seeded onto the dense top layer of the bilayered (cell free or fibroblast populated) copolymer matrix, human keratinocytes grew out into confluent sheets. After subsequent lifting to the air-liquid interface, a multilayered epithelium with a morphology corresponding to that of the native epidermis was formed. Some differences could still be observed: the expression and localization of some differentiation specific proteins was different and close to that seen in hyperproliferative epidermis; a basal lamina and anchoring zone were absent.     

Polylactones, 22 ABA triblock copolymers of L-lactide and poly(ethylene glycol)

Triblock copolymers were prepared from L-lactide and a poly(ethylene glycol) of M?_n = 1020. With GeO₂ and SnO₂ only low conversions were obtained, regardless of the reaction temperature. Sb₂O₃ caused partial racemization, and only SnO gave satisfactory results. When other samples of poly(ethylene glycol) were used (e.g., with M?_n = 600 or 3250), partial racemization occurred even with SnO. However, with Sn(II) 2-ethylhexanoate, racemization-free triblock copolymers were obtained under the reaction conditions used. The results of the characterization indicate that the triblock copolymers possess unimodal molecular weight distribution. The lactide blocks crystallize in the α-modification and suppress in most, but not all, cases the crystallization of the poly(ethylene glycol) blocks. The crystallization of long poly(ethylene glycol) blocks observed in several cases indicates a triphasic character of these samples.     

Synthesis and in vitro drug release behavior of amphiphilic triblock copolymer nanoparticles based on poly (ethylene glycol) and polycaprolactone

Novel BAB type amphiphilic triblock copolymers consisting of poly (ethylene glycol) (PEG) (B) as hydrophilic segment and poly (epsilon-caprolactone) (PCL) (A) as hydrophobic block were prepared by coupling reaction using L-lysine methyl ester diisocyanate (LDI) as the chain extender. The triblock copolymers obtained were characterized by FT-IR, 1H NMR, GPC, and DSC. Core-shell type nanoparticles were prepared by nanoprecipitation method and below 100 nm nanoparticles were obtained due to their specific structure. Transmission electron microscopy image demonstrated that these nanoparticles were spherical in shape. Stability of the nanoparticles in biological media was evaluated. Poorly water-soluble anticancer drug 4'-demethyl-epipodophyllotoxin (DMEP) was chosen for controlled drug release because it was easily encapsulated into polymeric nanoparticles via hydrophobic interaction. In vitro release behavior of DMEP from polymeric nanoparticles was investigated, the results showed that the drug release rate can be modulated by the variation of the copolymer composition.     

Biocompatibility of poly(epsilon-caprolactone)/poly(ethylene glycol) diblock copolymers with nanophase separation

In this study, we prepared diblock copolymers of poly(epsilon-caprolactone) (PCL) and poly(ethylene glycol) (PEG) by aluminum alkoxide catalysts. The biological responses to the spin cast surface of different PCL/PEG diblock copolymers were investigated in vitro. Our results showed that surface hydrophilicity improved with the increased PEG segments in diblock copolymers and that bacteria adhesion was inhibited by increased PEG contents. PCL-PEG 23:77 showed nanotopography on the surface. The number of adhered endothelial cells, platelets and monocytes on diblock copolymer surfaces was inhibited in PCL-PEG 77:23 and enhanced in PCL-PEG 23:77. Nevertheless, the platelet and monocyte activation on PCL-PEG 23:77 was reduced. PCL-PEG 23:77 had better cellular response as well as lower degree of platelet and monocyte activation. The current study was the first one to demonstrate that surface nanotopography could influence not only cell adhesion and growth but also platelet and monocyte activation.     

Synthesis and characterization of a polyrotaxane consisting of β-cyclodextrins and a poly(ethylene glycol)-poly(propylene glycol) triblock copolymer

A polyrotaxane in which many β-cyclodextrins (β-CyDs) are threaded onto a triblock copolymer of poly(ethylene glycol) (PEG) and poly(propylene glycol) (PPG) capped with fluorescein-4-isothiocyanate (FITC) was synthesized as a model of stimuli-responsive molecular assemblies for nanoscale devices. β-CyDs threaded onto the triblock copolymer enhance the solubility of the polyrotaxane and presumably contribute to the prevention of aggregation between PPG segments. The interaction of β-CyDs with a terminal FITC moiety was observed to be significant at 10°C, however, with increasing temperature, the interaction of β-CyDs with a PPG segment becomes prominent. From these results, it is concluded that the majority of β-CyDs move toward the PPG segment with increasing temperature although some β-CyDs may reside on PEG segments.     

Functionalization of poly(ethylene glycol) and monomethoxy-poly(ethylene glycol)

Simple methods are described for the substitution of poly(ethylene glycol) and monomethoxy-poly(ethylene glycol) substitution. Affinity ligands, coenzymes, or enzymes can be covalently attached to the substitution product or they can be used as liquid ionexchangers.     

Stereocomplex formation in ABA triblock copolymers of poly(lactide) (A) and poly(ethylene glycol) (B)

Two series of triblock copolymers of poly(ethylene glycol) (PEG, number-average molecular weight M _n = 6000) and poly(L -lactide) (PLLA) or poly(D -lactide) (PDLA) were prepared by ring-opening polymerization of lactide initiated by PEG end groups using stannous octoate as a catalyst, either in refluxing toluene or in the melt at 175°C. The weight percentage of PLA in the polymers varied between 15 and 75 wt.-%. Blends of polymers containing blocks of opposite chirality were prepared by co-precipitation from homogeneous solutions. The melting temperatures of the crystalline PEG and PLA phases strongly depended on the composition of the polymers. The melting temperature of the PLA phase in the blends was approximately 40°C higher than that of the single block copolymers. Stereocomplex formation between blocks of enantiomeric poly(lactides) in PEG/PLA block copolymers was established for the first time. Water uptake of polymeric films prepared by solution casting was solely determined by the PEG content of the film.     

Phase behavior and miscibility in blends of poly(sebacic anhydride)/poly(ethylene glycol)

In this research, poly(sebacic anhydride) was synthesized via melt-condensation. The polymer was then blended with poly(ethylene glycol) in various ratios by solvent casting, to obtain the desired polymer blends. A differential scanning calorimeter was employed to investigate the crystalline behavior of the blends. Blends with under 10% poly(ethylene glycol) were found to consist of two partially miscible polymers in the amorphous phase. This compatibility of the two polymers may induce the crystallization of the poly(sebacic anhydride) component from decreasing the glass transition temperature of the poly(sebacic anhydride) component in the blending system due to the presence of poly(ethylene glycol) chain segments. Furthermore, phase separation occurred and the crystallinity of poly(sebacic anhydride) diminished when at least 20% poly(ethylene glycol) was present in the blends. These results were verified by the variation in the absorptions of carbonyl groups in infrared spectra; these spectra exhibit the changes in crystallinity of poly(sebacic anhydride) in the blends.     

Preparation and characterization of ibuprofen-loaded poly(lactide-co-glycolide)/poly(ethylene glycol)-g-chitosan electrospun membranes

Ibuprofen-loaded composite membranes composed of poly(lactide-co-glycolide) (PLGA) and poly(ethylene glycol)-g-chitosan (PEG-g-CHN) were prepared by electrospinning. The electrospun membranes were characterized by scanning electron microscopy (SEM), differential scanning calorimetry (DSC), mechanical evaluation and contact angle measurements. Shrinkage behavior of the membrane in buffer at 37 degrees C was also evaluated. It was found that PLGA glass transition temperature (Tg) decreased with increasing PEG-g-CHN content in the composite membranes, which results in a decrease in tensile stress at break but an increase in tensile strain of the membranes. The degree of shrinkage of these composite membranes decreased from 76 to only 3% when the PEG-g-CHN content in the membranes increased from 10 to 30%. The presence of PEG-g-CHN significantly moderated the burst release rate of ibuprofen from the electrospun PLGA membranes. Moreover, ibuprofen could be conjugated to the side chains of PEG-g-CHN to prolong its release for more than two weeks. The sustained release capacity of the PLGA/PEG-g-CHN composite membranes, together with their compliant and stable mechanical properties, renders them ideal matrices for atrial fibrillation.     

Surface modification using silanated poly(ethylene glycol)s

Surface-grafted poly(ethylene glycol) (PEG) molecules are known to prevent protein adsorption to the surface. The protein-repulsive property of PEG molecules are maximized by covalent grafting. We have synthesized silanated monomethoxy-PEG (m-PEG) for covalent grafting of PEG to surfaces with oxide layers. Two different trialkoxysilylated PEGs were synthesized and characterized. The first trialkoxysilylated PEG was prepared by direct coupling of m-PEG with 3-isocyanatopropyltriethoxysilane through a urethane bond (silanated PEG I). The other silanated PEG (silanated PEG II) containing a long hydrophobic domain between PEG and a silane domain was prepared by reacting m-PEG with 1,6-diisocyanatohexane and 10-undecen-1-ol in sequence before silylation with 3-mercaptopropyl trimethoxysilane. Silanated PEGs I and II were grafted onto glass, a model surface used in our study. The PEG-grafted glass surfaces were characterized by contact angle, X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM). Although contact angle did not change much as the bulk concentration of silanated PEG used for grafting increased from 0.1 to 20 mg/ml for both PEGs I and II, the surface atomic concentrations from XPS measurements showed successful PEG grafting. Surface PEG grafting increased concentration of surface carbon but decreased silicone concentration. The high resolution C1s spectra showed higher ether carbon with lower hydrocarbon compositions for the PEG-grafted surfaces compared to the control surface. AFM images showed that more PEG molecules were grafted onto the surface as the bulk concentration used for grafting was increased. AFM images of the dried surfaces showed that the surfaces were not completely covered by PEG molecules. After hydration, however, the surface appears to be covered completely probably due to the hydration of the grafted PEG chains. Glass surfaces modified with silanated PEGs reduced fibrinogen adsorption by more than 95% as compared with the control surface. Silanated PEGs provides a simple method for PEG grafting to the surface containing oxide layers.     

Photopolymerized poly(ethylene glycol)/poly(L-lysine) hydrogels for the delivery of neural progenitor cells

Neural progenitor cells (NPCs) have shown promise in a number of models of disease and injury, but for these cells to be safe and effective, they must be directed to differentiate appropriately following transplantation. We have developed a photopolymerized hydrogel composed of macromers of poly(ethylene glycol) (PEG) bound to poly(L-lysine) (PLL) that supports NPC survival and directs differentiation. Green fluorescent protein (GFP) positive NPCs were encapsulated in these gels and demonstrated survival up to 17 days. When encapsulated in the gels at a photoinitiator concentration of 5.0 mg/ml, few NPCs (0.5 +/- 0.25%) demonstrated apoptosis. Furthermore, 55 +/- 6% of the NPCs cultured within the gels in epidermal growth factor (EGF) containing media differentiated into a mature neuronal cell type (neurofilament 200 positive) while the remainder 44 +/- 8% were undifferentiated (nestin positive). A small percentage, 1 +/- 0.4%, expressed the astrocytic marker glial acidic fibrilary protein. Photopolymerized PEG/PLL gels promote the survival and direct the differentiation of NPCs, making this system a promising delivery vehicle for NPCs in the treatment of injuries and diseases of the central nervous system.     

Photopolymerized poly(ethylene glycol)/poly(L-lysine) hydrogels for the delivery of neural progenitor cells

Neural progenitor cells (NPCs) have shown promise in a number of models of disease and injury, but for these cells to be safe and effective, they must be directed to differentiate appropriately following transplantation. We have developed a photopolymerized hydrogel composed of macromers of poly(ethylene glycol) (PEG) bound to poly(L-lysine) (PLL) that supports NPC survival and directs differentiation. Green fluorescent protein (GFP) positive NPCs were encapsulated in these gels and demonstrated survival up to 17 days. When encapsulated in the gels at a photoinitiator concentration of 5.0 mg/ml, few NPCs (0.5 ± 0.25%) demonstrated apoptosis. Furthermore, 55 ± 6% of the NPCs cultured within the gels in epidermal growth factor (EGF) containing media differentiated into a mature neuronal cell type (neurofilament 200 positive) while the remainder 44 ± 8% were undifferentiated (nestin positive). A small percentage, 1 ± 0.4%, expressed the astrocytic marker glial acidic fibrilary protein. Photopolymerized PEG/PLL gels promote the survival and direct the differentiation of NPCs, making this system a promising delivery vehicle for NPCs in the treatment of injuries and diseases of the central nervous system.     

Branched poly(ethylene glycol) linkers

Novel types of methoxy poly(ethylene glycol) (PEG) linkers (U-PEG linkers) have been synthesized. These PEG linkers are linear polymers that attach to bioactive agents via a functional group, derived from a 2° alcohol, located in the center of the polymer chain versus the traditional terminal attachment site. These new types of linkers can be prepared with different functional groups (e.g. active ester, succinimidyl carbonate, and carbazate) for selected point of attachment, including ethylene oxide oligomers to provide “stems” when steric factors need to be addressed. Conversion of p-nitrophenyl carbonates to the more desirable succinimidyl carbonates has also been accomplished by a novel nucleophilic displacement procedure. Modification of proteins with these reagents is easily accomplished and is illustrated by the conjugation of a U-PEG linker with L -asparaginase.     

Biodegradable nanoparticles of amphiphilic triblock copolymers based on poly(3-hydroxybutyrate) and poly(ethylene glycol) as drug carriers

New amorphous amphiphilic triblock copolymers of poly(3-hydroxybutyrate)-poly(ethylene glycol)-poly(3-hydroxybutyrate) (PHB-PEG-PHB) were synthesized using the ring-opening copolymerization of beta-butyrolactone monomer. They were characterized by fluorescence, SEM and (1)H NMR. These triblock copolymers can form biodegradable nanoparticles with core-shell structure in aqueous solution. Comparing to the poly(ethylene oxide)-PHB-poly(ethylene oxide) (PEO-PHB-PEO) copolymers, these nanoparticles exhibited much smaller critical micelle concentrations and better drug loading properties, which indicated that the nanoparticles were very suitable for delivery carriers of hydrophobic drugs. The drug release profile monitored by fluorescence showed that the release of pyrene from the PHB-PEG-PHB nanoparticles exhibited the second-order exponential decay behavior. The initial biodegradation rate of the PHB-PEG-PHB nanoparticles was related to the enzyme amount, the initial concentrations of nanoparticle dispersions and the PHB block length. The biodegraded products detected by (1)H NMR contained 3HB monomer, dimer and minor trimer, which were safe to the body.     

Control of vitamin B12 release from poly(ethylene glycol)/poly(butylene terephthalate) multiblock copolymers

The release of vitamin B12 (1355 Da) from matrices based on multiblock copolymers was studied. The copolymers were composed of hydrophilic poly(ethylene glycol)-terephthalate (PEGT) blocks and hydrophobic poly(butylene terephthalate) (PBT) blocks. Vitamin B12 loaded films were prepared by using a water-in-oil emulsion method. The copolymer properties, like permeability, could be varied by increasing the PEG-segment length from 300 up to 4,000 g/mol and by changing the wt% of PEGT. From permeation and release experiments. the diffusion coefficient of vitamin B12 through PEGT/PBT films of different compositions was determined. The diffusion coefficient of Vitamin B12 was strongly dependent on the composition of the copolymers. Although an increased wt% of PEGT (at a constant PEG-segment length) resulted in a higher diffusion coefficient, a major effect was observed at increasing PEG-segment length. By varying the copolymer composition, a complete release of vitamin B12 in 1 day up to a constant release for over 12 weeks was obtained. The release rate could be effectively tailored by blending copolymers with different PEG-segment lengths. The swelling and the crystallinity of the matrix could explain the effect of the matrix composition on the release behavior.     

Physical characterization of thin semi-porous poly(L-lactic acid)/poly(ethylene glycol) membranes for tissue engineering

This study examines physical properties of solvent-cast poly(L-lactic acid) (PLLA): poly(ethylene glycol) PEG membranes as a function of PEG molecular weight (MW) and incubation in vitro for 6 weeks. PEGs of MW 400, 1450 and 8000 were used. The morphological, thermal, mechanical and permeability properties of the membranes were studied prior to and after 3 and 6 weeks of incubation in phosphate-buffered saline (PBS) at 37 degrees C. The membranes showed a thickness of about 35+/-5 microm and were found to be semi-porous, with a non-porous surface as well as a porous surface with pore-diameters of 0.5-5 microm. The surface pore size was found to be a function of PEG MW used. All membranes were mechanically strong, with elastic moduli and tensile strength of 150-440 MPa and 7-36 MPa, respectively, all through the 6-week incubation period. The lower-MW PEGs plasticized PLLA based on high initial percent elongation; however, the effect was lost after 3 weeks of incubation in PBS. All membranes except those fabricated with PEG 8000 were impermeable for up to 6 weeks of incubation in PBS. Permeability studies showed that only PLLA:PEG 8000 membranes were permeable to methylene blue after 3 weeks of degradation.     

Molecular calculations of poly(ethylene glycol) transport across a swollen poly (acrylic acid)/mucin interface

The transport of poly(ethylene glycol) chains than can promote mucoadhesion across the interface between lightly cross-linked poly(acrylic acid) and mucin may be analyzed as a function of molecular characteristics using theories of chain penetration in a dilute network. The fracture energy for the ensuing adhesive bond is proportional to the number of polymer chains crossing the interface, which, in turn, is related to the polymer volume fraction, the chain diffusion coefficient, and the degree of polymerization. Relevant calculations were performed for a number of cross-linked poly(acrylic acid) gels and three different types of poly(ethylene glycol) chains.