Encapsulating chondrocytes in copolymer gels: bimodal degradation kinetics influence cell phenotype and extracellular matrix development

Hydrogels provide an ideal environment for encapsulating chondrocytes and facilitating the production of cartilaginous tissue. However, the deposition of extracellular matrix (ECM) and ultimate tissue function are significantly affected by degradation of gel scaffolds. It was hypothesized that a bimodal degradation process would capture the critical features necessary for neotissue development. Specifically, most of the initial crosslinks would degrade quickly and enable ECM deposition, whereas a critical amount would remain or degrade much more slowly to provide structural integrity over a longer time period. In this study, chondrocytes were encapsulated in copolymer gels of nondegradable [poly(ethylene glycol) dimethacrylate] and degradable [poly(lactic acid)-b-poly(ethylene glycol)-b-poly(lactic acid) dimethacrylate] macromers to investigate the effects of gel degradation on ECM evolution. All gels were synthesized from 10 wt % total macromer solutions consisting of 0, 19, 21, 23, 25, or 100 mol % nondegradable units. The copolymer constructs were found to have lower DNA content than completely degradable constructs after 8 weeks. However, total biochemical content was very similar among the various copolymer constructs. Histological analysis gave more interesting insight, showing a more uniform spatial distribution of ECM components in copolymer samples than in constructs with 100 mol % nondegradable units. In addition, a number of major structural defects were present in constructs with 0 mol % nondegradable units that became less apparent as the amount of nondegradable units was increased. Overall, the copolymer gels had a higher compressive modulus during neotissue development and also showed no evidence of chondrocyte dedifferentiation. With their bimodal degradation profile, copolymer gels with carefully selected ratios of degrading to slow or nondegrading crosslinks provide distinct advantages for ECM development in tissue-engineered cartilage.     

Hydrophilic molecularly imprinted poly(hydroxyethyl-methacrylate) polymers

Highly cross-linked 2-hydroxyethyl methacrylate (HEMA) and poly(ethylene glycol) dimethacrylate with poly(ethylene glycol) of molecular weight 600 (PEG600DMA) were molecularly imprinted with hydrophilic templates glucose and proxyphylline using water as a solvent. Glucose-imprinted polymers showed increased recognitive capacity compared to nonimprinted polymers as well as increased glucose uptake compared to structurally similar galactose and methylglucopyranoside. Increasing glucose concentration in the imprinting mixture resulted in higher capacity and selective binding. Similar results were obtained for proxyphylline-imprinted P(HEMA-co-PEG600DMA) polymers, where the proxyphylline uptake was higher than structurally similar theophylline. Glucose-imprinted networks also showed diffusion coefficients on the order of 10(-6) cm2/s, conducive to applications in drug delivery and tissue engineering. This work showed that using pairs of hydrogen-bonding monomers and templates, selective, high-affinity sites could be created despite nonspecific binding.     

High-strength hydrogels based on N-vinyl pyrrolidinone and 4-t-butyl-2-hydroxycyclohexylmethacrylate.

A nonionic, high-water-content, high-strength hydrogel based on N-vinyl pyrrolidinone (NVP) and a novel hydrophilic-bulky monomer, 4-t-butyl-2-hydroxycyclohexylmethacrylate (TBCM), was developed. The TBCM was prepared in three relatively simple, high-yield steps. The copolymerization of NVP with varying concentrations of TBCM resulted in transparent hydrogel films possessing a wide range in mechanical and physical properties. A copolymer composition of 91.7 parts NVP, 8.0 parts TBCM, and 0.3 parts ethylene glycol dimethacrylate (EGDMA) gave a transparent, flexible film possessing a water content of 86%, a modulus of 130 g/mm2, and a tear strength of 3.4 g/mm. In contrast, a copolymer composition of 49.7 parts NVP, 50 parts TBCM, and 0.3 parts EGDMA gave a transparent, hard hydrogel film possessing a water content of 26% and a modulus of 86,000 g/mm2. All of the high-water copolymer compositions developed resulted in lysozyme uptake typical of nonionic high-water-content hydrogels and oxygen permeability levels greater than 50 (cm3-O2(STP).cm)/(s.cm2.mm Hg) x 10(-11).     

Radical concentrations,environments, and reactivities during crosslinking polymerizations

Radical environments and concentrations during the crosslinking polymerizations of multiethylene glycol dimethacrylates were characterized with electron spin resonance spectroscopy. The relative concentrations of free and trapped radicals were used as a measure of certain structural features in the evolving polymer, especially with respect to microgel formation. The influence of monomer structure and crosslinking density on the polymer structure was studied by examining the radical behavior during the polymerization of diethylene glycol dimethacrylate, poly(ethylene glycol 200) dimethacrylate, and poly(ethylene glycol 600) dimethacrylate. The extent of radical trapping and microgel formation was found to increase with increasing crosslinking density. The influence of the rate of initiation on radical trapping was also investigated, and slower rates of polymerization led to a higher fraction of trapped radicals with respect to the total radical population. Finally, the termination mechanism of radicals was characterized (long-term and short-term) and the corresponding kinetic constants quantified.     

玻璃微珠改性双酚A甲基丙烯酸缩水甘油酯复合的树脂充填材料

背景：口腔复合树脂修复材料存在聚合收缩率大和机械强度低等缺点,不能很好地满足临床使用的要求。 目的：观察双酚A甲基丙烯酸缩水甘油酯和二甲基丙烯酸三甘醇酯的比例及改性实心玻璃微珠添加量对口腔复合树脂弯曲强度与聚合体积收缩率的影响。 方法：以硅烷偶联剂KH-550对实心玻璃微珠进行硅化处理。向双酚A甲基丙烯酸缩水甘油酯与二甲基丙烯酸三甘醇酯配比分别为7/3、6/4、5/5的口腔复合树脂中再分别添加质量分数10%,30%,50%,70%,90%的实心玻璃微珠,机械搅拌均匀后,快速填入模具中固化,设置不添加实心玻璃微珠的树脂作为对照参考,分析口腔复合树脂的机械性能和收缩率。 结果与结论：随着实心玻璃微珠添加量的增加,口腔复合树脂的弯曲强度呈先增加后减小的趋势,体积收缩率呈降低趋势：当双酚A甲基丙烯酸缩水甘油酯与二甲基丙烯酸三甘醇酯的比例为5/5、实心玻璃微珠的添加量为70%时,口腔复合树脂的弯曲强度最高,为(88.29±0.66) MPa(P < 0.05);当双酚A甲基丙烯酸缩水甘油酯与二甲基丙烯酸三甘醇酯的配比为7/3、玻璃微珠添加量为70%时,聚合体积收缩率最小,为0.898%。     

Elution study of unreacted Bis-GMA, TEGDMA, UDMA, and Bis-EMA from light-cured dental resins and resin composites using HPLC

In the present work the elution of residual monomers from light-cured dental resins and resin composites into a 75% ethanol:water solution was studied using High-Performance Liquid Chromatography (HPLC). The resins studied were made by light-curing of bisphenol A glycol dimethacrylate (Bis-GMA), triethylene glycol dimethacrylate (TEGDMA), urethane dimethacrylate (UDMA), ethoxylated bisphenol A glycol dimethacrylate [Bis-EMA(4)] and mixtures of these monomers. The resin composites were made from two commercial light-cured restorative materials (Z100 MP and Filtek Z250), the resin matrix of which is based on copolymers of these monomers. The effect of the curing time on the amount of monomers eluted was investigated. The concentration of the extractable monomers was determined at several immersion periods from 3 h to 30 days. For all the materials studied, it was observed that the chemical structure of the monomers used for the preparation of the resins, which defines the chemical and physical structure of the corresponding resin, directly affects the amount of eluted monomers, as well as the time needed for the elution of this amount. In the case of composites, it seems that the elution process it is not influenced by the presence of filler.     

A novel biocompatible adhesive incorporating plant-derived monomers

We describe a new class of biomaterials with potential for a variety of applications in tissue engineering, wound healing, and transdermal drug delivery. These materials are based on oleic methyl ester (OME), which is derived from various plant oils including soybean oil. The OME was acrylated (AOME) and subsequently copolymerized with methyl methacrylate (MMA) and ethylene glycol dimethacrylate (EGDMA) to form pressure sensitive adhesives (PSAs). We assessed the cytocompatibility of each PSA product using Alamar Blue and Live/Dead assays. It was found that after 2 h, human fibroblast cells attached on all four of the PSA polymers tested. After 24 h, cell spreading was seen on all materials with the exception of the polymerized AOME product (PAOME). Cells attached to the copolymer PSA products continued to proliferate for up to 2 weeks, as shown by fluorescent confocal microscopy imaging. Finally, a mechanical analysis of each of the copolymers is presented demonstrating that they have a range of mechanical properties and cell adhesiveness depending on the formulation, making them attractive candidates for use as bioactive adhesives.     

Hydrogels in endovascular embolization. V. Antitumour agent methotrexate-containing p(HEMA).

The aminohexyl derivative of the copolymer of 2-hydroxyethyl methacrylate and ethylene dimethacrylate (p(HEMA)-Hex) in the form of regular spherical particles was used as a polymeric carrier for chemically sorbed methotrexate (MTX). The effect of the washed p(HEMA)-Hex-MTX carrier on the morphological structure of donor blood, on blood coagulation indicators and on responses of the living tissue surrounding the material which has been in the blood vessel of the rabbit for various periods of time was evaluated. MTX is capable of diffusion for several days from the embolic material.     

Pathomorphological study of the pancreas during phospholipase A2-induced experimental pancreatic necrosis

The pathomorphogenesis of experimental acute pancreatic necrosis induced by administration of phospholipase A2 into the pancreatic tissue is characterized by pathomorphological signs of hemorrhagic pancreatic necrosis (total necrosis and purulent fusion of some acini and massive hemorrhages in the interlobular and intralobular interstitial tissue) and fatty pancreatic necrosis (necrobiotic and necrotic changes in acinar cells that spread from the peripheral area and form the demarcation line).     

Tissue anti-adhesion potential of ibuprofen-loaded PLLA-PEG diblock copolymer films

This study was designed to evaluate the effect of polyethylene glycol (PEG) and nonsteroidal anti-inflammatory drug (ibuprofen) on the prevention of postsurgical tissue adhesion. For this, poly(L-lactic acid) (PLLA)-PEG diblock copolymers were synthesized by ring opening polymerization of L-lactide and methoxy polyethylene glycol (Mw 5000) of different compositions. The synthesized copolymers were characterized by gel permeation chromatography and 1H-nuclear magnetic resonance spectroscopy. PLLA-PEG copolymer films were prepared by solvent casting. The prepared copolymer films were more flexible and hydrophilic than the control PLLA film, as investigated by the measurements of glass transition temperature, water absorption content, and water contact angle. The drug release behavior from the ibuprofen (10 wt%)-loaded copolymer films was examined by high performance liquid chromatography. It was observed that the drug was released gradually up to about 40% of total loading amount after 20 days, depending on PEG composition; more drug release from the films with higher PEG compositions. In vitro cell adhesions on the copolymer films with/without drug were compared by the culture of NIH/3T3 mouse embryo fibroblasts on the surfaces. For in vivo evaluation of tissue anti-adhesion potential, the copolymer films with/without drug were implanted between the cecum and peritoneal wall defects of rats and their tissue adhesion extents were compared. It was observed that the ibuprofen-containing PLLA-PEG films with high PEG composition (particularly PLLA113-PEG113 film with PEG composition, 50 mol%) were very effective in preventing cell or tissue adhesion on the film surfaces, probably owing to the synergistic effects of highly mobile, hydrophilic PEG and anti-inflammatory drug, ibuprofen.     

Controlled release of cytarabine from poly(2-hydroxyethyl methacrylate-co-N-vinyl-2-pyrrolidone) hydrogels

Controlled release of cytarabine (ara-C) from poly(2-hydroxyethyl methacrylate-co-N-vinyl-2-pyrrolidone) [p(HEMA-co-VP)] hydrogels cross-linked with ethylene glycol dimethacrylate (EGDMA) is reported. Three compositions of copolymer, each one with a different cross-linking degree, have been studied: H50/VP50, H75/VP25, and H80/VP20. Ara-C (5-25 mg by disc) was trapped in the gels by including it in the polymerization feed mixture. The ara-C release time was between 1 day from H50/VP50/E0.5 discs and 16 days from H80/VP20/E15 discs. In all cases there is a time period for which the drug release rate is constant.     

In vitro cytotoxicity and in vivo biocompatibility of poly(propylene fumarate-co-ethylene glycol) hydrogels.

The in vitro cytotoxicity and in vivo biocompatibility of poly(propylene fumarate-co-ethylene glycol) [P(PF-co-EG)] hydrogels were assessed in order to investigate the influence of poly(ethylene glycol) molecular weight and copolymer composition. These materials have application as injectable cardiovascular implants; cytotoxicity due to leachable products, as well as inflammation caused by the biomaterial itself, may ultimately affect the biocompatibility of the implant. We utilized a 7-day in vitro cytotoxicity assay to quantify cell density and cellular proliferation in the presence of copolymer films. The copolymer films exhibited slight to moderate cytotoxicity toward cultured endothelial cells, showing 20-86% viability relative to controls. Cell viability increased with an increasing weight percent of PEG or, to a lesser extent, the molecular weight of PEG. In vivo biocompatibility was assessed using a cage implantation model over a 21-day time period. This system was used to characterize the local cellular and humoral inflammatory response in the surrounding exudate, as well as the size and density of macrophages adherent to the material itself. All copolymer formulations exhibited excellent biocompatibility relative to controls with no significant differences in total leukocyte count among the different formulations. The in vivo inflammatory reaction displayed normal wound healing over 21 days as shown by a progressive decrease in both leukocyte concentration and enzymatic activity. The surface coverage of the copolymer films remained relatively constant from 7 to 21 days. There were no cells larger than 0.003 mm2, which was previously shown to be the threshold value for foreign-body giant cells. These data suggest that P(PF-co-EG) hydrogels have potential for use as injectable biomaterials.     

Poly(oligoethylene glycol methacrylate) Star-Shaped Copolymers with Hydroxypropyl Methacrylate Cores

Star-shaped copolymers with crosslinked hyperbranched cores, varying in both number and length of the arms, are prepared by the reversible addition-fragmentation chain transfer (RAFT) polymerization technique. Star synthesis is conducted via a simple, “core-first” method. Hydroxypropyl methacrylate monomer is used for the formation of the hyperbranched/crosslinked core, with ethylene glycol dimethacrylate used as a bifunctional monomer in the role of the cross-linker. Two different cross-linker/chain transfer agent ratios are chosen to manipulate the number of the arms in the final star nanostructure. Oligoethylene glycol methacrylate (OEGMA₅₀₀) is used as a biocompatible and hydrophilic component for the creation and extension of the arms. Arm length is controlled by the amount of OEGMA monomer used in the second polymerization step. Molecular characterization of the four synthesized stars is achieved via size exclusion chromatography and nuclear magnetic resonance, which provides information regarding the molar mass, dispersity, and composition of each star-shaped copolymer. Dynamic light scattering (DLS) studies of copolymers in N,N-dimethyl formamide solutions provide sizes of individual copolymer molecules in the size range of 8–20 nm. Aqueous dispersions of copolymers are investigated by DLS, cryogenic transmission electron microscopy, and atomic force microscopy, which show the formation of spherical aggregates with diameters of 60–350 nm, dependent on the arm number and length of star copolymers. The obtained sizes and size distributions, along with the hydroxyl-group-bearing cores, indicate that the star copolymer can be used as potential nanocarriers for drugs or bioimaging agents.     

U75412E, a lazaroid, prevents progressive burn ischemia in a rat burn model.

Thermal energy causes an immediate, irreversible injury at the burn site, followed by a delayed, reversible tissue loss in the area surrounding the burn site due to progressive ischemia. We investigated the role of lipid peroxidation in the pathogenesis of progressive ischemia in a rat burn model. The burn model consisted of a row of four 10 x 20 mm burns separated by three unburned 5 x 20 mm skin bridges (interspaces). The interspaces became ischemic and necrotic by 24 hours, producing a single wound with the merger of the burn sites. U75412E, a lipid peroxidation inhibitor, preserved vascular patency, restored blood flow, prevented a rise in tissue conjugated dienes, and maintained tissue viability in the interspaces. Four separate burn wounds healed between three viable strips of hair-bearing interspaces. The treatment was effective, when given systemically during the period between 2 hours before and 1 hour after the burn. U75412E prevented progressive burn ischemia and the expansion of tissue loss.     

Granulomatous inflammation and monstrous giant cells in response to intraperitoneal hormone implants in channel catfish (Ictalurus punctatus)

Plastic implants (2.7 mm maximum dimension) of an ethyl vinyl acetate copolymer (EVAc) matrix, containing inulin, bovine serum albumin (BSA) and luteinizing hormone releasing hormone (LHRH), were covered with impervious EVAc and then surgically placed into the peritoneal cavity of 1-year-old channel catfish, Ictalurus punctatus. In fish kept in cold water (13 degrees C), 10 per cent of the implants per month were encapsulated by granulation tissue. In fish kept in warm water (27 degrees C), 20 per cent of the implants per month were encapsulated, with a total of 86 per cent encapsulated at 5 months. In addition to fibroblasts and capillaries, the granulation tissue included macrophages, neutrophils, lymphocytes, plasma cells, multinucleated giant cells and a matrix of collagen fibres. The density of the fibrous capsule increased with time. In a separate investigation, it was found that the thickness of the capsule was directly proportional to the degree of exposure of the EVAc matrix to the fish (exposure influenced by the rate of dissolution of the capsule content). Monstrous giant cells with up to 600 nuclei per 5 microns thick section were seen in capsules around implants. On intraperitoneally implanted cover glasses, whole giant cells contained up to 6000 nuclei and were interconnected by cytoplasmic bridges. Signs of neoplasia, implant expulsion or massive adhesions were not seen.     

Hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) copolymers for cell microencapsulation: effect of HEMA purity

Thermoplastic copolymers of 2-hydroxyethyl methacrylate (HEMA) and methyl methacrylate (MMA) (molar ratio: 75/25 HEMA-MMA) were synthesized using HEMA containing different amounts of ethylene glycol dimethacrylate (EGDMA) to investigate their suitability for cell microencapsulation. Pure HEMA (0.0% EGDMA) was obtained with preparative chromatography to prepare a linear copolymer. Microcapsules (with a diameter of 300-400 microm) were readily made with the copolymers by interfacial precipitation. Smaller and more transparent capsules were obtained using the copolymer prepared from purer HEMA. Chinese hamster ovary (CHO) fibroblasts, as model cells, were microencapsulated in the linear copolymer. The CHO cells survived the microencapsulation process and the metabolic activity of the encapsulated cells increased within the 14 days observation period.     

Preparation and characterization of infection-resistant antibiotics-releasing hydrogels rods of poly[hydroxyethyl methacrylate-co-(poly(ethylene glycol)-methacrylate]: biomedical application in a novel rabbit penile prosthesis model

In this work, preparation and characterization of novel three different antibiotic loaded penile prosthesis in the rod form were investigated by copolymerization of 2-hydroxyethylmethacrylate (HEMA) with poly(ethylene glycol)-methacrylate, (PEG-MA). To achieve this goal, a series of novel copolymer hydrogels were prepared in rod form using HEMA and PEG-MA monomers via UV initiated photopolymerization. The thermal stability of the copolymer was found to be lowered by increase in the ratio of PEG-MA in the rod structure. Contact angle measurements on the surface of copolymer hydrogel demonstrated that the copolymer gave rise to a significant hydrophilic surface compared with pure poly(HEMA). The blood protein adsorption and platelet adhesion were significantly reduced on the surface of the copolymer hydrogels compared with control pure poly(HEMA). Poly(HEMA:PEG-MA;1:1)-1 formulation containing different antibiotics (20 mg antibiotic/g polymer) released about 90, 91, and 55% of the total loaded cephtriaxon, vancomycin, and gentamicin in 48 h at pH 7.4, respectively. Finally, antibiotics loaded biocompatible poly(HEMA:PEG-MA;1:1)-1 hydrogel compositions was used as a penile prosthesis in preventing cavernous tissue infections in a rabbit prosthesis model. The efficacy of the three different antibiotics loaded hydrogel system was evaluated in four different groups of rabbits, in which various infectious agents were inoculated. The animals were sacrificed after predetermined time periods, and clinical, histological and microbiological assessment on the implant side were carried out to detect infections. Eventually, we concluded that three different antibiotic loaded penile prostheses (i.e. poly(HEMA:PEG-MA;1:1)-1 hydrogel systems) were as effective as parenteral antibiotics applications.     

Water sorption and mechanical properties of light-cured proprietary composite tooth restorative materials.

Seven light-cured proprietary composite restorative materials, P-50 (3M), P-10 (3M), P-30 (3M), FulFil (Caulk), Herculite (Kerr), Silux Plus (3M) and Silux (3M) were characterized in terms of water uptake at 37 degrees C. For several of the systems, elastic modulus and glass transition temperature were evaluated with a dynamic mechanical analyser (Autovibron DDV-II-C). Model systems such as bis-GMA (75%) + triethylene glycol dimethacrylate (25%), bis-GMA (30%) + triethylene glycol dimethacrylate (10%) + lithium aluminium silicate (60%), bis-GMA + triethylene glycol dimethacrylate (14%) + barium glass (silanated) (86%) and bis-GMA + triethylene glycol dimethacrylate (14%) + zinc glass (silanated) (86%) were also studied with reference to water sorption. It was concluded that the changes in elastic modulus tend to confirm the hypothesis that the matrix-filler interface contains water. Silux and Silux Plus accommodated the greatest amount of water at the interface. Silux Plus displayed a dramatic reduction in elastic modulus c. 0 degrees C, indicating possible melting of water clusters. It appeared that the only available hole for water clusters was at the matrix-filler interface.     

Development of the rectal fascia

The development of the fascia recti was investigated by 300-600 microns thick sections through the pelves of 8-35-week-old human fetuses, of newborn children and adults. The pelves were impregnated with an epoxy resin and cut with a diamond wire-saw. We here report that the fascia recti develops as part of the adventitia recti. In 9-week-old fetuses the latter consists of condensed mesenchyme which in 18-20-week-old fetuses develops to a fibrous connective tissue surrounding the rectum. In newborn children adipose tissue starts spreading between the connective tissue lamellae of the adventitia recti and thereby separates them. The most external of these connective tissue lamellae is rather dense and can be regarded as fascia recti. In the adult we found it not always as strongly marked as in newborn children. The close developmental relations between the fascia recti and the tissue of the adventitia recti might be seen as cause for the temporary stop of tumor spread at the fascia recti.     

Manipulations in hydrogel degradation behavior enhance osteoblast function and mineralized tissue formation

Hydrogels were prepared by copolymerizing a degradable macromer, poly(lactic acid)-b-poly(ethylene glycol)-b-poly(lactic acid) endcapped with methacrylate groups (PEG-LA-DM), with a nondegradable macromer, poly(ethylene glycol) dimethacrylate (PEGDM). Copolymer networks consisted of 100:0, 83:17, 67:33, and 50:50 PEGDM:PEG-LA-DM mass%, essentially creating scaffolds that exhibit 0, 17, 33, and 50% degradation over the time course of the experiment. Osteoblasts were photoencapsulated in these copolymer hydrogels and cultured for 3 weeks in vitro. Metabolic activity, proliferation, and alkaline phosphatase production were enhanced by an increase PEG-LADM content and corresponding degradation. Gene expression of the cultured osteoblasts, normalized to beta-actin, was analyzed, and osteopontin and collagen type I gene expression increased with degradation. Finally, as a measure of mineralized tissue formation, calcium and phosphate deposition was analyzed biochemically and histologically. Mineralization increased with increasing concentration of PEG-LA-DM and biochemically resembled that of hydroxyapatite.