Genotoxic and Nongenotoxic Effects of Glycidyl Methacrylate on Human Lung Fibroblast Cells

Objective To evaluate the genotoxic and nongenotoxic effects of short-term exposure to glycidyl mathacrylate (GMA) on human lung fibroblast cells (2BS cells) in vitro. Methods DNA strand breakage was determined by single cell gel electrophoresis, and DNA ladder formation assay and flow cytometric analysis were carried out to detect apoptic responses of cells to GMA exposure. The HPRT gene mutation assay was used to evaluate the mutagenicity, and the effect of GMA on gap junctional intercellular communication (GJIC) in the exposed cells was examined with the scrape loading/dye transfer technique. The ability of GMA to transform 2BS cells was also tested by an in vitro cell transformation assay. Results Exposure to GMA resulted in a dose-dependent increase in DNA strand breaks but not apoptic responses. GMA was also shown to significantly induce HPRT gene mutations and morphological transformation in 2BS cells in vitro. In contrast, GMA produced a concentration-dependent inhibition of GJIC. Conclusions GMA elicits both genotoxic and nongenotoxic effects on 2BS cells in vitro. The induction of DNA damage and gene mutations and inhibition of GJIC by GMA may casually contribute to GMA-induced cell transformation.

MONOMER REACTIVITY RATIO AND THERMAL PERFORMANCE OFα-METHYL STYRENE AND GLYCIDYL METHACRYLATE COPOLYMERS

Synthesis and characterization of the copolymers （PAG） of α-methyl styrene （AMS） and glycidyl methacrylate （GMA） are presented. The copolymers of PAG were characterized by gel permeation chromatography （GPC）, Fourier transform infrared spectroscopy （FTIR）, nuclear magnetic resonance （^1H-NMR） and thermogravimetery （TG）. Based on the copolymer compositions determined by ^1H-NMR, the reactivity ratios of AMS and GMA were found to be 0.105 ± 0.012 and 0.883 ± 0.046 respectively by Kelen-Tudos method. TG revealed that thermal stability of the copolymers decreased with increasing the AMS content in the copolymers, which indicated that the degradation was mainly caused by the chain scission of AMS-containing structures. Under heating, the copolymers depolymerize at their weak bonds and form chain radicals, which could further initiate other chemical reactions.

Studies of the Genotoxicity of Glycidyl Methacrylate (GMA)

The following experiments were conducted to evaluate the genotoxic effects of GMA (glycidyl methacrylale) on mammalian and human cells.(1) Using the absorption spectrum shift method in vitro, we observed that the maximums of calf thymus DNA and GMA were shifted toward longer wavelengths (a change of more than 15nm) and the absorbance decreased after incubation at room temperature for 15min or more.The result indicates that binding of DNA and GMA had occurred.The binding force is strong, not affected by the addition of concentrated sodium chloride solution, and only slightly decreased by the addition of 8 M urea solution.Therefore the bond between DNA and GMA might be covalent.(2) In cell cultures, unscheduled DNA synthesis (UDS) in human and/or rat lymphocyte was induced and DNA semiconserva-tive replication was inhibited by GMA at concentrations of less than 5.2 mM.(3) Sperm abnormality tests and assays of UDS in germ cells of male mice were conducted to study the in vivo genotoxicity of GMA.The results revealed that GMA could damage DNA, increase sperm abnormality frequency, and reduce the number of sperm cells, 1990 Academic Press.Inc.

Experimental Study on Malignant Transformation of Human Bronchial Epithelial Cells Induced by Glycidyl Methacrylate and Analysis on its Methylation

Objective To establish the model of human bronchial epithelial cells （16HBE） malignant transformation induced by glycidyl methacrylate （GMA） and define the different methylation genes at different stages. Methods DNA was extracted at different 16HBE malignant phases and methylation at different stages were detected using Methylation chip of Promoter Microarray Methylation＇. Methylation-specific PCR （MSP） was methylation status of some genes, and then compared with the control groups. changes of genes DNA ＇NimbleGen HG18 CpG used to observe the Results The result showed that GMA induced 16HBE morphorlogical transformation at the dose of 8 I^g/mL, and cell exposed to GMA had 1 374 genes in protophase, 825 genes in metaphase, 1 149 genes in anaphase, respectively; 30 genes are all methylation in the 3 stages; 318 genes in protophase but not in metaphase and anaphase; 272 genes in metaphase but not in protophase and anaphase; 683 genes in anaphase but not in metaphase and protophase; 73 genes in protophase and metaphase but not in anaphase; 67 genes in protophase and anaphase but not in metaphase; 59 genes in metaphase and anaphase but not in protophase. Conclusion The pattern of DNA methylation could change in the process of 16HBE induced by GMA.

Genes Differentially Expressed in Human Lung Fibroblast Cells Transformed by Glycidyl Methacrylate

Objective To define the differences in gene expression patterns between glycidyl methacrylate (GMA)-transformed human lung fibroblast cells (2BS cells) and controls. Methods The mRNA differential display polymerase chain reaction (DD-PCR) technique was used. cDNAs were synthesized by reverse transcription and amplified by PCR using 30 primer combinations. After being screened by dot blot analysis, differentially expressed cDNAs were cloned, sequenced and confirmed by Northern blot analysis. Results Eighteen differentially expressed cDNAs were cloned and sequenced, of which 17 were highly homologous to known genes (homology = 89%-100%) and one was an unknown gene. Northern blot analysis confirmed that eight genes encoding human zinc finger protein 217 (ZNF217), mixed-lineage kinase 3 (MLK-3), ribosomal protein (RP) L15, RPL41, RPS16, TBX3, stanniocalcin 2 (STC2) and mouse ubiquitin conjugating enzyme (UBC), respectively, were up-regulated, and three genes including human transforming growth factor b inducible gene (Betaig-h3), a-1,2-mannosidase 1A2 (MAN 1A2) gene and an unknown gene were down-regulated in the GMA-transformed cells. Conclusion Analysis of the potential function of these genes suggest that they may be possibly linked to a variety of cellular processes such as transcription, signal transduction, protein synthesis and growth, and that their differential expression could contribute to the GMA-induced neoplastic transformation.

Analysis of the Phenotype and the Restriction Enzyme Mapping Level of Mutations Induced by the New Mutagen Glycidyl Methacrylate

Glycidyl methaerylate (GMA) is a recently recognized chemical mutagen. In order to explore the mutagenicity and mutagenic process of GMA, plasmid pBR322 was used for in vitro binding, mutant screening, and restriction enzyme mapping. The binding between GMA and DNA in vitro has been verified by means of a spectrophotometric method. When pBR322 and GMAbound pBR322 were used to transform Eschenchia coli HB101, the following results were obtained: (1) The transformation efficiency of GMA-bound pBR322 was much lower than that of pBR322 alone. (2) GMA-bound pBR322 induced phenotype changes in competent cells (i.e., tetracycline-resistance inactivation or ampicillin-resistance inactivation). There were two mutants of pBR322, Ap~RTc~S and Ap~STc~R, in the transformants and a deductive mutant Ap~STc~S in the nontranstormants. (3) All of the selected mutants were stable and heritable. (4) When restriction enzyme maps were used to analyze the mutant Ap~RTc~S, four of seven maps were changed. some sites were shifted to other resistant gene regions, for example, sites of Bgll, EcoRl, Ilindlll. Hinclll, etc., and there was a new recognition site for Hindi (252). We did not observe any DNA fragment insertion or deletion on any maps. Our results suggest that when GMA is covalently linked to the plasmid DNA, it gives rise to a premutagenic lesion of DNA that is converted in vivo into a point mutation. (C)1990 Academic Press, Inc.

PREPARATION OF LOW MOLECULAR WEIGHT POLY(GLYCIDYL METHACRYLATE) BY PHOTOPOLYMERIZATION

Low molecular weight poly（glycidyl methacrylate）s （PGMAs） were prepared by photopolymerization in ethyl acetate, with benzophenone （BP） as photoinitiator, and triethylamine （TEA） as hydrogen donor. The existence of semipinacol dormant end groups in PGMA was confirmed by FT-IR and ^1H-NMR, and the content of the semipinacol dormant groups was determined quantitatively by ^1H-NMR measurement. The effects of various thctors, such as reaction time, BP concentration and monomer concentration on the synthesis of the polymers were investigated systematically. The molecular weights of the polymers were also investigated with GPC. It is shown that increasing BP concentration and decreasing irradiation time and monomer concentration led to a significant decrease of the molecular weights.

Poly(Glycidyl Methacrylates)-grafted Zinc Oxide Nanowire by Surface-initiated Atom Transfer Radical Polymerization

Poly(glycidyl methacrylates)(PGMA) was grafted from zinc oxide(ZnO) nanowires via surface-initiated atom transfer radical polymerization(SI-ATRP) technique.Firstly,the ZnO nanowires were synthesized by the one-pot hydrothermal technique.Subsequently,the ZnO was functionalized with 3-aminopropyl triethoxysilane,which was converted to macroinitiator by the esterification of them with 2-bromopropionyl bromide.PGMA grafted ZnO nanowires(PGMA-ZnO) were then synthesized in an ATRP of the GMA with CuCl/2,2`-bipyridine as the catalyst system.Kinetics studies revealed an approximate linear increase in weight of polymer with reaction time,indicating that the polymerization process owned some "living" character.The structure and composition of PGMA-ZnO were characterized with scanning electron microscope(SEM),energy-dispersive X-ray(EDX) spectrometer,fourier transform infrared spectroscopy(FT-IR) and thermogravimetric analysis(TGA).

MOLECULAR MUTAGENESIS INDUCED BY GLYCIDYL METHACRYLATE

Glycidyl methacrylate (GMA)is a recently recognized mutagen. In order to explore the mutagenicity and mechanism of GMA, plasmid PBR322 was used for in vitro binding, mutant screening, restriction enzyme mapping,and DNA sequencing. To explore the mechamism by which an initial premutational event is converted into a stable heritable mutation, pBR322 and GMA-bound pBR322 were transformed into E. coli HB101 , and the following results were obtained : 1) GMA-bound PBR322 induced phenotype changes in competent cells. Two stable and heritable mutants were isolated (Ap￣RTc￣S and Ap￣STc￣R). 2) When restriction enzyme mapping was used to analyze the mutant Ap￣RTc￣S , four of seven maps showed changes, but no large DNA insertion or deletion were observed.3) The frequency of deletion and insertion forms counted about 10%. Sequence specificity and hot spot regions were evident in the sequence analysis of mutated plasmid. The above results indicate that the premutagenic lesions of plasmid induced by GMA can be converted into point mutations in vivo.

Glycidyl methacrylate-compatibilized poly(lactic acid)/hemp hurd biocomposites： Processing, crystallization, and thermo-mechanical response

Poly（lactic acid）-based biocomposites were developed with hemp hurd （Cannabis sativa L.） with grafting-based interfacial compatibilization. Poly（lactic acid） was extruded with hemp hurd and glycidyl methacrylate as the polymer/hurd interfacial compatibilizer, and injection molded. Interfacial compatibility between poly（lactic acid） and hemp hurd increased with grafted glycidyl methacrylate in comparison to the non-compatibilized control, as corroborated by scanning electron microscopy fractog- raphy and mechanical analysis, which showed increases in the glycidyl methacrylate-grafted 20% （w/w） hemp hurd/poly（lactic acid） biocomposite, retaining 94% of the neat polymer strength, with increases in crystallinity, and showing a range of thermo-mechanical properties desirable for rigid biocomposite aoolications.

RAFT Copolymerization of Glycidyl Methacrylate and N,N-Dimethylaminoethyl Methacrylate

In this work, copolymerization of two functional monomers, glycidyl methacrylate （GMA） and N,N-dimethylaminoethyl methacrylate （DMAEMA）, was firstly carried out via reversible addition-fragmentation chain transfer （RAFT） polymerization successfully. The copolymerization kinetics was investigated under the molar ratio of n[GMA＋DMAEMA]o/n[AIBN]o/n[CPDN]o=300/1/3 at 60℃. The copolymerization showed typical ＂living＂ features such as first-order polymerization kinetics, linear increase of molecular weight with monomer conversion and narrow molecular weight distribution. The reactivity ratios of GMA and DMAEMA were calculated by the extended Kelen-Tudos linearization methods. The epoxy group of the copolymer PGMA-co-PDMAEMA remained intact under the conditions of RAFT copolymerization and could easily be post-modified by ethylenedia- mine. Moreover, the modified copolymer could be used as a gene carrier.

Synchronous Synthesis and Immobilization of Metal Phthalocyanine for Aerobic Oxidation of Styrene

In this study,the precursor 4-(4-carboxy-phenoxy)phthalonitrile(CPPN)was first bonded onto the silica gel surface modified with poly(glycidyl methacrylate)(PGMA)(PGMA/SiO2)to prepare CPPN-PGMA/SiO2,and metal phthalocyanine(MPc;M=Co,Fe,Cu,Mn)was supported on the PGMA/SiO2 surface to prepare MPc-PGMA/SiO2 by synchronous synthesis and immobilization with phthalonitrile and metal salt in the solution.The chemical composition and surface morphology were characterized by the Fourier transform infrared(FTIR)spectroscopy,UV-Vis spectroscopy,scanning electron microscopy(SEM),X-ray photoelectron spectroscopy(XPS),and thermogravimetry analysis(TGA).The catalytic performance of MPc-PGMA/SiO2 in epoxidation of styrene was also investigated with molecular oxygen acting as the oxidant.The results show that MPc-PGMA/SiO2 can efficiently and selectively catalyze molecular oxygen for oxidation of styrene to styrene oxide under mild conditions.However,the catalytic activity differs substantially depending on the central metal,and a highest catalytic activity is achieved by CoPc-PGMA/SiO2.The CoPc-PGMA/SiO2 amount and temperature can also affect the catalytic oxidation of styrene,and at normal atmospheric pressure,a maximum conversion rate of styrene(99%)and selectivity of styrene oxide(53%)are obtained using 0.1 g of CoPc-PGMA/SiO2(22.61μmol of CoPc)at 100℃ for 6 h.CoPc-PGMA/SiO2 also has excellent reusability,and the conversion rate of styrene is still over 90%after 5 cycles.

Acrylate Copolymers as Impact Modifi er for Epoxy Resin

P（BA-GMA）（PBG）, having various molecular weights, was synthesized by in situ polymerization of butyl acrylate（BA） and glycidyl methacrylate（GMA）, and further used as a modifier to improve the comprehensive properties of the epoxy curing system. The copolymers were characterized by gel permeation chromatography（GPC）. The effects of various molecular weights of copolymers on the mechanical properties, thermal performance, and phase behavior of the curing system were carefully evaluated. The experimental results of differential scanning calorimetry（DSC） and dynamic mechanical analysis（DMA） showed that glass transition temperature decreased and the tan δ peak shifted to a lower temperature with decreasing molecular weight of copolymer. Mechanical properties analysis of curing films showed that the impact strength and fracture toughness increased significantly upon the addition of PBG, indicating good toughness of the modified epoxy resins. From scanning electron microscopy（SEM） studies of the fracture surfaces of ER/PBG systems, the fracture behavior of epoxy matrix was changed from brittleness to toughness.

STUDY ON GMA-DNA ADDUCTS

Objective. DNA modification fixed as mutations in the cells may be an essential factor in the initiation step of chemical carcinogenesis. In order to explore the mechanism of gene mutation and cell transformation induced by glycidyl methacrylate (GMA), the current test studied the characteristics of GMA DNA adducts formation in vitro. Methods. In vitro test, dAMP, dCMP, dGMP, dTMP and calf thymus DNA were allowed to react with GMA (Glycidyl Methacrylate). After the reaction, the mixtures were detected by UV and subjected to reversed phase HPLC on ultrasphere ODS reversed phase column, the reaction products were eluted with a linear gradients of methanol (solvent A) and 10mmol/L ammonium formate, pH5 0 (solvent B). The synthesized adducts were then characterized by UV spectroscopy in acid (pH1 0), neutral (pH7 2), alkaline (pH11 0) and by mass spectroscopy. Results. The results showed that GMA could bind with dAMP, dCMP, dGMP and calf thymus DNA by covalent bond, and the binding sites were specific (N 6 of adenine, N 3 of cytosine). Meanwhile, a main GMA DNA adduct in the reaction of GMA with calf thymus DNA was confirmed as N 3 methacrylate 2 hydroxypropy1 dCMP. Conclusions. GMA can react with DNA and /or deoxynucleotide monophosphate and generate some adducts such as N 6 methacrylate 2 hydroxypropyl dAMP and N 3 methacrylate 2 hydroxypropyl dCMP, ets. Formation of GMA DNA adducts is an important molecular event in gene mutation and cell transformation induced by GMA.

Artificial cilia for soft and stable surface covalent immobilization of bone morphogenetic protein-2

Preservation of growth factor sensitivity and bioactivity(e.g.,bone morphogenetic protein-2(BMP-2))post-immobilization to tissue engineering scaffolds remains a great challenge.Here,we develop a stable and soft surface modification strategy to address this issue.BMP-2(a model growth factor)is covalently immobilized onto homogeneous poly(glycidyl methacrylate)(PGMA)polymer brushes which are grafted onto substrate surfaces(Au,quartz glass,silica wafer,or common biomaterials)via surface-initiated atom transfer radical polymerization.This surface modification method multiplies the functionalized interfacial area;it is simple,fast,gentle,and has little effect on the loaded protein owing to the cilia motility.The immobilized BMP-2(i-BMP-2)on the surface of homogeneous PGMA polymer brushes exhibits excellent bioactivity(-87%bioactivity of free BMP-2 in vitro and 20%-50%higher than scaffolds with free BMP-2 in vivo),with conformation and secondary structure well-preserved after covalent immobilization and ethanol sterilization.Moreover,the osteogenic activity of i-BMP-2 on the nanoline pattern(PGMA-poly(N-isopropylacrylamide))shows-110%bioactivity of free BMP-2.This is superior compared to conventional protein covalent immobilization strategies in terms of both bioactivity preservation and therapeutic efficacy.PGMA polymer brushes can be used to modify surfaces of different tissue-engineered scaffolds,which facilitates in situ immobilization of growth factors,and accelerates repair of a wide range of tissue types.

Preparation of magnetic anion exchange resin and their adsorption kinetic behavior of reactive blue

Magnetic anion exchange resin （MD-1） was prepared from quaternization of magnetic copolymeric resin （glycidyl methacry- late-co-divinylbenzene）. For comparison, magnetic resin MD-0 without quaternization and non-magnetic resin （D-l） were also synthesized for the adsorption process. It was found that the adsorption was mainly contributed to the chemical interaction between quaternary ammonium groups and reactive blue RXHC. Due to the smaller size, MD- 1 had faster adsorption and desorption kinetics than D-1. Coupled with the advantage of easy separation, the magnetic anion exchange resin was considered to be superior to common anion exchange resin in removal of reactive dye.

SPECTRAL AND CHEMICAL MONITORING OF CYCLO-ADDITION REACTION OF CO_2 WITH POLY(MMA-co-GMA) COPOLYMERS

This paper deals with the monitoring cyclo-addition of CO2 to methyl methacrylate （MMA）-glycidyl methacrylate （GMA） copolymers using spectral （IH-NMR and FTIR） and chemical （elemental analysis and titration） methods. Thus, poly（MMA-co-GMA）, was first prepared via solution polymerization. The copolymer was then treated with CO2 gas flow in the presence of cetyltrimethyl ammoniumbromide as a catalyst. In terms of the carbonation reaction time, the terpolymer poly（MMA-eo-GMA-co-2-oxo-l,3-dioxolane-4-yl-methyl methacrylate） was prepared in various yield of CO2 fixation （〉 90%）. The peak intensity changes in the 1H-NMR and FTIR spectra provided excellent demonstrative techniques to monitor the carbonation reaction progression. In a comparative analytical viewpoint, the NMR and elemental analysis were recognized to be the most accurate ways to follow the cyclo-addition reaction progression. However, titration was recognized to be the most preferred method, because it is a very inexpensive, facile and available method with a reasonable cost- accuracy balance.

Facile synthesis of wormlike quantum dots-encapsulated nanoparticles and their controlled surface functionalization for effective bioapplications

Semiconductor quantum dots (QDs) are considered as ideal fluorescent probes owing to their intrinsic optical properties. It has been demonstrated that the size and shape of nanoparticles significantly influence their behaviors in biological systems. In particular, one-dimensional (1D) nanoparticles with larger aspect ratios are desirable for cellular uptake. Here, we explore a facile and green method to prepare novel 1D wormlike QDs@SiO2nanoparticles with controlled aspect ratios, wherein multiple QDs are arranged in the centerline of the nanoparticles. Then, an excellent cationic gene carrier, ethanolamine-functionalized poly(glycidyl methacrylate) (denoted by BUCT-PGEA), was in-situ produced via atom transfer radical polymerization on the surface of the QDs@SiO2nanoparticles to achieve stable surfaces (QDs@SiO2-PGEA) for effective bioapplications. We found that the wormlike QDs@SiO2-PGEA nanoparticles demonstrated much higher gene transfection performance than ordinary spherical counterparts. In addition, the wormlike nanoparticles with larger aspect ratio performed better than those with smaller ratio. Furthermore, the gene delivery processes including cell entry and plasmid DNA (pDNA) escape and transport were also tracked in real time by the QDs@SiO2-PGEA/pDNA complexes. This work realized the integration of efficient gene delivery and real-time imaging within one controlled 1D nanostructure. These constructs will likely provide useful information regarding the interaction of nanoparticles with biological systems. [Figure not available: see fulltext.] © 2016, Tsinghua University Press and Springer-Verlag Berlin Heidelberg.