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1.
ACS Appl Mater Interfaces ; 8(34): 21939-45, 2016 Aug 31.
Article in English | MEDLINE | ID: mdl-27089518

ABSTRACT

Surface topography has been introduced as a new tool to coordinate cell selection, growth, morphology, and differentiation. The materials explored so far for making such structural surfaces are mostly rigid and impermeable. Hydrogel, on the other hand, was proved a better synthetic media for cell culture because of its biocompatibility, softness, and high permeability. Herein, we fabricated a poly(2-hydroxyethyl methacrylate) (pHEMA) hydrogel substrate with high-aspect-ratio surface microfeatures. Such structural surface could effectively guide the orientation and shape of human mesenchymal stem cells (HMSCs). Notably, on the flat hydrogel surface, cells rounded up, whereas on the microplate patterned hydrogel surface, cells elongated and aligned along the direction parallel to the plates. The microplates were 2 µm thick, 20 µm tall, and 10-50 µm wide. The interplate spacing was 5-15 µm, and the intercolumn spacing was 5 µm. The elongation of cell body was more pronounced on the patterns with narrower interplate spacing and wider plates. The cells behaved like soft solid. The competition between surface energy and elastic energy defined the shape of the cells on the structured surfaces. The soft permeable hydrogel scaffold with surface structures was also demonstrated as being viable for long-term cell culture, and could be used to generate interconnected tissues with finely tuned cell morphology and alignment across a few centimeter sizes.


Subject(s)
Hydrogels/chemistry , Cell Differentiation , Cell Line , Humans , Mesenchymal Stem Cells , Polyhydroxyethyl Methacrylate
2.
Ann Biomed Eng ; 44(4): 895-902, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26101035

ABSTRACT

Leukocyte recruitment plays a key role in chronic inflammatory diseases such as cardiovascular disease, rheumatoid arthritis, and cancer. Leukocyte rolling and arrest are mediated in part by the temporally-regulated surface expression of vascular cell adhesion molecule-1 (VCAM1) on endothelial cells (ECs). In this paper, we engineered a pH-responsive vehicle comprised of 30 mol% dimethylaminoethyl methacrylate (30D) and 70 mol% hydroxyethyl methacrylate (70H) to encapsulate, protect, and deliver VCAM1 small interfering RNA (siRNA). The ability of siRNA to reduce VCAM1 gene expression is in direct opposition to its activation by cytokines. At 12 h post-activation, VCAM1 gene knockdown was 90.1 ± 7.5% when delivered via 30D/70H nanoparticles, which was on par with a leading commercial transfection agent. This translated into a 68.8 ± 6.7% reduction in the surface density of VCAM1 on cytokine-activated ECs. The pH-responsive delivery of VCAM1 siRNA efficiently reduced temporal surface protein expression, which may be used to avert leukocyte recruitment.


Subject(s)
Human Umbilical Vein Endothelial Cells/metabolism , RNA, Small Interfering/administration & dosage , Vascular Cell Adhesion Molecule-1/genetics , Cell Survival/drug effects , Gene Expression , HeLa Cells , Human Umbilical Vein Endothelial Cells/drug effects , Humans , Hydrogen-Ion Concentration , Interleukin-1alpha/pharmacology , Methacrylates/chemistry , Nanoparticles/administration & dosage , Nanoparticles/chemistry , RNA, Small Interfering/chemistry
3.
Biomaterials ; 57: 22-32, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25956194

ABSTRACT

A principal challenge in wound healing is a lack of cell recruitment, cell infiltration, and vascularization, which occurs in the absence of temporal and spatial cues. We hypothesized that a scaffold that expands due to local changes in pH may alter oxygen and nutrient transport and the local cell density, leading to enhanced cell deposition and survival. In this study, we present a pH-responsive scaffold that increases oxygen transport, as confirmed by our finite element model analysis, and cell proliferation relative to a non-responsive scaffold. In vivo, responsive scaffolds induce a pro-healing gene expression profile indicative of enhanced angiogenesis, granulation tissue formation, and tissue remodeling. Scaffolds that stretch in response to their environment may be a hallmark for tissue regeneration.


Subject(s)
Methacrylates/chemistry , Oxygen/metabolism , Tissue Scaffolds/chemistry , Wound Healing , Animals , Cell Proliferation , Cell Survival , Finite Element Analysis , Hydrogen-Ion Concentration , Mice , NIH 3T3 Cells , Porosity , Rats, Wistar , Tissue Engineering
4.
Mol Pharm ; 11(3): 755-65, 2014 Mar 03.
Article in English | MEDLINE | ID: mdl-24467226

ABSTRACT

Because breast cancer patient survival inversely correlates with metastasis, we engineered vehicles to inhibit both the C-X-C chemokine receptor type 4 (CXCR4) and lipocalin-2 (Lcn2) mediated migratory pathways. pH-responsive liposomes were designed to protect and trigger the release of Lcn2 siRNA. Liposomes were modified with anti-CXCR4 antibodies to target metastatic breast cancer (MBC) cells and block migration along the CXCR4-CXCL12 axis. This synergistic approach--coupling the CXCR4 axis blockade with Lcn2 silencing--significantly reduced migration in triple-negative human breast cancer cells (88% for MDA-MB-436 and 92% for MDA-MB-231). The results suggested that drug delivery vehicles engineered to attack multiple migratory pathways may effectively slow progression of MBC.


Subject(s)
Acute-Phase Proteins/antagonists & inhibitors , Antibodies, Monoclonal/pharmacology , Cell Movement , Gene Expression Regulation, Neoplastic , Lipocalins/antagonists & inhibitors , Proto-Oncogene Proteins/antagonists & inhibitors , RNA, Small Interfering/genetics , Receptors, CXCR4/antagonists & inhibitors , Triple Negative Breast Neoplasms/therapy , Acute-Phase Proteins/genetics , Acute-Phase Proteins/metabolism , Apoptosis , Blotting, Western , Cell Proliferation , Combined Modality Therapy , Drug Delivery Systems , Drug Synergism , Female , Fluorescent Antibody Technique , Humans , Hydrogen-Ion Concentration , Lipocalin-2 , Lipocalins/genetics , Lipocalins/metabolism , Liposomes , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , RNA, Small Interfering/administration & dosage , Receptors, CXCR4/immunology , Receptors, CXCR4/metabolism , Signal Transduction , Triple Negative Breast Neoplasms/secondary , Tumor Cells, Cultured
5.
Biomaterials ; 35(9): 3071-9, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24397987

ABSTRACT

An excessive tissue response to prosthetic arterial graft material leads to intimal hyperplasia (IH), the leading cause of late graft failure. Seroma and abnormal capsule formation may also occur after prosthetic material implantation. The matricellular protein Thrombospondin-2 (TSP-2) has shown to be upregulated in response to biomaterial implantation. This study evaluates the uptake and release of small interfering RNA (siRNA) from unmodified and surface functionalized electrospun PET graft materials. ePET graft materials were synthesized using electrospinning technology. Subsets of the ePET materials were then chemically modified to create surface functional groups. Unmodified and surface-modified ePET grafts were dip-coated in siRNAs alone or siRNAs complexed with transfection reagents polyethyleneimine (PEI) or Lipofectamine RNAiMax. Further, control and TSP-2 siRNA-PEI complex treated ePET samples were placed onto a confluent layer of human aortic smooth muscle cells (AoSMCs). Complexation of all siRNAs with PEI led to a significant increase in adsorption to unmodified ePET. TSP-2 siRNA-PEI released from unmodified-ePET silenced TSP-2 in AoSMC. Regardless of the siRNA-PEI complex evaluated, AoSMC migrated into the ePET. siRNA-PEI complexes delivered to AoSMC from dip-coated ePET can result in gene knockdown. This methodology for siRNA delivery may improve the tissue response to vascular and other prosthetics.


Subject(s)
Aorta/cytology , Coated Materials, Biocompatible/pharmacology , Gene Silencing/drug effects , Myocytes, Smooth Muscle/metabolism , Polyethylene Glycols/pharmacology , Polyethyleneimine/pharmacology , RNA, Small Interfering/metabolism , Adsorption , Cell Adhesion/drug effects , Cell Shape/drug effects , Gene Knockdown Techniques , Humans , Myocytes, Smooth Muscle/cytology , Myocytes, Smooth Muscle/drug effects , Myocytes, Smooth Muscle/ultrastructure , Polyethylene Terephthalates , RNA, Messenger/genetics , RNA, Messenger/metabolism , Reproducibility of Results , Transfection
7.
Biomaterials ; 33(32): 8104-10, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22884683

ABSTRACT

The primary cause of mortality in breast cancer is tumor aggressiveness, characterized by metastases to regional lymph nodes, bone marrow, lung, and liver. C-X-C chemokine receptor type 4 (CXCR4) has been shown to mobilize breast cancer cells along chemokine gradients. Quantification of CXCR4 surface expression may predict the efficacy of anti-CXCR4 labeled liposomal therapeutics to target and kill breast cancer cells. We evaluated gene and surface receptor expression of CXCR4 on breast cancer cell lines distinguished as having low and high invasiveness, MDA-MB-175VII and HCC1500, respectively. CXCR4 surface expression did not correlate with invasiveness. MDA-MB-175VII exhibited more binding to anti-CXCR4 labeled liposomes relative to HCC1500. Increased binding correlated with greater cell death relative to IgG labeled liposomes. Quantitative cell characterization may be used to select targeted therapeutics with enhanced efficacy and minimal side effects.


Subject(s)
Antibiotics, Antineoplastic/administration & dosage , Breast Neoplasms/drug therapy , Breast Neoplasms/genetics , Doxorubicin/administration & dosage , Molecular Targeted Therapy , Receptors, CXCR4/genetics , Antibiotics, Antineoplastic/pharmacology , Breast Neoplasms/pathology , Cell Line, Tumor , Cell Survival/drug effects , Doxorubicin/pharmacology , Female , Gene Expression Regulation, Neoplastic , Humans , Liposomes/immunology , Receptors, CXCR4/immunology
8.
Biomaterials ; 33(15): 3880-6, 2012 May.
Article in English | MEDLINE | ID: mdl-22364701

ABSTRACT

Cell adhesion molecules govern leukocyte-endothelial cell (EC) interactions that are essential in regulating leukocyte recruitment, adhesion, and transmigration in areas of inflammation. In this paper, we synthesized hydrogel matrices modified with antibodies against vascular cell adhesion molecule-1 (VCAM1) and endothelial leukocyte adhesion molecule-1 (E-Selectin) to mimic leukocyte-EC interactions. Adhesion of human umbilical vein ECs to polyvinyl alcohol (PVA) hydrogels was examined as a function of the relative antibody ratio (anti-VCAM1:anti-E-Selectin) and substrate elasticity. Variation of PVA backbone methacrylation was used to affect hydrogel matrix stiffness, ranging from 130 to 720 kPa. Greater EC adhesion was observed on hydrogels presenting 1:1 anti-VCAM1:anti-E-Selectin than on gels presenting either arginine-glycine-asparagine (RGD) peptide, anti-VCAM1, or anti-E-Selectin alone. Engineered cell adhesion - based on complementing the EC surface presentation - may be used to increase the strength of EC-matrix interactions. Hydrogels with tunable and synergistic adhesion may be useful in vascular remodeling.


Subject(s)
Human Umbilical Vein Endothelial Cells/cytology , Human Umbilical Vein Endothelial Cells/drug effects , Polyvinyl Alcohol/pharmacology , Amines/chemistry , Antibodies/pharmacology , Cell Adhesion/drug effects , Human Umbilical Vein Endothelial Cells/metabolism , Humans , Interleukin-1alpha/pharmacology , Magnetic Resonance Spectroscopy , Materials Testing , Mechanical Phenomena/drug effects , Methacrylates/chemistry , Microscopy, Confocal , Polymerization/drug effects , Polyvinyl Alcohol/chemistry
9.
Nano Lett ; 11(9): 3643-8, 2011 Sep 14.
Article in English | MEDLINE | ID: mdl-21800912

ABSTRACT

Scaffolds that couple electrical and elastic properties may be valuable for cardiac cell function. However, existing conductive materials do not mimic physiological properties. We prepared and characterized a tunable, hybrid hydrogel scaffold based on Au nanoparticles homogeneously synthesized throughout a polymer templated gel. Conductive gels had Young's moduli more similar to myocardium relative to polyaniline and polypyrrole, by 1-4 orders of magnitude. Neonatal rat cardiomyocytes exhibited increased expression of connexin 43 on hybrid scaffolds relative to HEMA with or without electrical stimulation.


Subject(s)
Connexin 43/biosynthesis , Gene Expression Regulation , Heart/physiology , Myocardium/metabolism , Myocytes, Cardiac/cytology , Nanotechnology/methods , Animals , Animals, Newborn , Anisotropy , Electric Conductivity , Gold/chemistry , Metal Nanoparticles/chemistry , Methacrylates/chemistry , Polymers/chemistry , Rats
10.
Langmuir ; 27(18): 11282-6, 2011 Sep 20.
Article in English | MEDLINE | ID: mdl-21823661

ABSTRACT

Inspired by morphogenesis in biology, we present a strategy for developing functional 3D materials with the capacity to morph based on environmental cues. We utilized local mechanical stresses to cause global shape changes in colloidosomes. Colloidosomes were assembled from pH-sensitive calcium alginate particles (CAPs) with high and low swelling ratios. Colloidosomes were subsequently cross-linked via diamine compounds with varying carbon chain lengths. New colloidosome isoforms were generated from heterogeneous mixtures of CAPs, which resulted in nonuniform stresses. Our study demonstrated that coordinated networks of heterogeneous subunits may be used to design programmable materials.


Subject(s)
Alginates/chemistry , Biomimetics , Carbon/chemistry , Colloids , Diamines/chemistry , Glucuronic Acid/chemistry , Hexuronic Acids/chemistry , Hydrogen-Ion Concentration , Stress, Mechanical
11.
J Biol Eng ; 4: 15, 2010 Nov 29.
Article in English | MEDLINE | ID: mdl-21114841

ABSTRACT

For years, the field of drug delivery has focused on (1) controlling the release of a therapeutic and (2) targeting the therapeutic to a specific cell type. These research endeavors have concentrated mainly on the development of new degradable polymers and molecule-labeled drug delivery vehicles. Recent interest in biomaterials that respond to their environment have opened new methods to trigger the release of drugs and localize the therapeutic within a particular site. These novel biomaterials, usually termed "smart" or "intelligent", are able to deliver a therapeutic agent based on either environmental cues or a remote stimulus. Stimuli-responsive materials could potentially elicit a therapeutically effective dose without adverse side effects. Polymers responding to different stimuli, such as pH, light, temperature, ultrasound, magnetism, or biomolecules have been investigated as potential drug delivery vehicles. This review describes the most recent advances in "smart" drug delivery systems that respond to one or multiple stimuli.

12.
Biomaterials ; 31(26): 6859-66, 2010 Sep.
Article in English | MEDLINE | ID: mdl-20493524

ABSTRACT

We synthesized a series of pH-sensitive vehicles, composed of dimethylaminoethyl methacrylate (DMAEMA) and 2-hydroxyethyl methacrylate (HEMA), to optimize the triggered release of DNA for gene transfection. The purpose of this study was to assess the role of swelling and cationic character independently on transfection; both of which may affect DNA release. Gene transfection was performed by delivering plasmid DNA (pDNA) encoding for luciferase. DNA release was controlled via volumetric swelling by regulating the endosomal pH as a result of inhibiting V ATPases using bafilomycin A1. Increasing the cationic character from 10 to 30 mol% DMAEMA did not increase transfection when swelling was inhibited. Transfection was significantly affected by the rate of pDNA release. pH-sensitive nanocarriers were also compared to vehicles comprised of polyethyleneimine (PEI), dioleoyl triammonium propane (DOTAP), and poly(lactic-co-glycolic acid) (PLGA, 50:50). pDNA encapsulating DMAEMA/HEMA nanoparticles and PEI/pDNA complexes had reduced transfection when V ATPases were inhibited, whereas pDNA encapsulating PLGA nanoparticles showed no endosomal pH dependence. DMAEMA/HEMA nanoparticles cross-linked with 3 mol% tetraethylene glycol dimethacrylate (TEGDMA) reported equivalent or greater gene transfection relative to the nanocarriers tested at 24 and 48 h.


Subject(s)
Drug Carriers/chemistry , Luciferases/genetics , Methacrylates/chemistry , Nanoparticles/chemistry , Transfection/methods , Cations , Cell Survival/drug effects , DNA/genetics , Electrophoresis, Agar Gel , Endosomes/drug effects , Endosomes/metabolism , HeLa Cells , Humans , Hydrogen-Ion Concentration/drug effects , Luciferases/metabolism , Methacrylates/pharmacology , Plasmids/genetics , Vacuolar Proton-Translocating ATPases/antagonists & inhibitors
13.
Langmuir ; 26(7): 4607-12, 2010 Apr 06.
Article in English | MEDLINE | ID: mdl-20199077

ABSTRACT

Coupling molecular sensing with electrical conductivity may provide an important and valuable resource in assessing disease pathology. Here, we introduce pH-responsive hydrogels with homogeneously synthesized gold (Au) nanoparticles that reversibly alter conductivity through pH-induced volumetric swelling. These intelligent hybrid materials respond to physiological pH shifts (pH 7.4 to 5.5) that can (1) alter the conductivity of the gel or (2) create conductive conduits via micropatterned arrays.


Subject(s)
Electric Conductivity , Hydrogels/chemistry , Gold/chemistry , Gold Colloid/chemistry , Hydrogen-Ion Concentration , Models, Theoretical , Nanoparticles/chemistry , Nanotechnology
14.
Nano Lett ; 9(12): 4467-73, 2009 Dec.
Article in English | MEDLINE | ID: mdl-19842673

ABSTRACT

Treatment of diseases on the molecular level by genetic material is limited by effective delivery mechanisms. We focused on the synthesis of a pH-sensitive gene delivery vehicle based on dimethylaminoethyl methacrylate (DMAEMA) with tunable swelling, cross-linking density, and DNA release kinetics within the endosomal pH range. Our strategy, which utilized a single step for DNA encapsulation, enhanced gene transfection efficiency and reduced cytotoxicity relative to polyethyleneimine (PEI) and poly-L-lysine (PLL).


Subject(s)
DNA/chemistry , DNA/pharmacokinetics , Drug Carriers/chemistry , Methacrylates/chemistry , Nanostructures/chemistry , Nanostructures/ultrastructure , Transfection/methods , Cross-Linking Reagents/chemistry , DNA/ultrastructure , Hydrogen-Ion Concentration , Materials Testing
15.
Biomaterials ; 29(12): 1950-7, 2008 Apr.
Article in English | MEDLINE | ID: mdl-18255142

ABSTRACT

pH-Sensitive poly(N,N-dimethylaminoethyl methacrylate (DMAEMA)/2-hydroxyethyl methacrylate (HEMA)) nanoparticles were prepared for the triggered release of paclitaxel within a tumor microenvironment. Tumors exhibit a lower extracellular pH than normal tissues. We show that paclitaxel release from DMAEMA/HEMA particles can be actively triggered by small, physiological changes in pH (within 0.2-0.6 pH units). Monodispersed nanoparticles were synthesized by forming an O/W emulsion followed by photopolymerization. Particles were characterized by transmission electron microscopy, dynamic light scattering, electrophoresis, and cytotoxicity. High release rates and swelling ratios are achieved at low pH, low crosslinking density, and high content of DMAEMA. Paclitaxel release is limited to 9% of the payload at pH 7.4 after a 2-h incubation at 37 degrees C. After adjusting to pH 6.8, 25% of the payload is released within 2h. Cell viability studies indicate that pH-sensitive DMAEMA/HEMA nanoparticles are not cytotoxic and may be used as an efficient, feedback-regulated drug delivery carrier.


Subject(s)
Cell Survival/drug effects , Delayed-Action Preparations/administration & dosage , Methacrylates/chemistry , Nanoparticles/chemistry , Nanoparticles/therapeutic use , Paclitaxel/administration & dosage , Antineoplastic Agents, Phytogenic/administration & dosage , Antineoplastic Agents, Phytogenic/chemistry , Delayed-Action Preparations/chemistry , Diffusion , Drug Carriers/chemistry , HeLa Cells , Humans , Materials Testing , Nanoparticles/ultrastructure , Paclitaxel/chemistry , Particle Size
16.
Int J Nanomedicine ; 1(1): 59-64, 2006.
Article in English | MEDLINE | ID: mdl-17722262

ABSTRACT

Developing methods to label viruses with fluorescent moieties has its merits in elucidating viral infection mechanisms and exploring novel antiviral therapeutics. Fluorescent quantum dots (QDs), an emerging probe for biological imaging and medical diagnostics, were employed in this study to tag retrovirus encoding enhanced green fluorescent protein (EGFP) genes. Electrostatic repulsion forces generated from both negatively charged retrovirus and QDs were neutralized by cationic Polybrene, forming colloidal complexes of QDs-virus. By examining the level of EGFP expression in 3T3 fibroblast cells treated with QDs-tagged retroviruses for 24 hours, the infectivity of retrovirus incorporated with QDs was shown to be only slightly decreased. Moreover, the imaging of QDs can be detected in the cellular milieu. In summary, the mild method developed here makes QDs-tagged virus a potential imaging probe for direct tracking the infection process and monitoring distribution of viral particles in infected cells.


Subject(s)
Microscopy, Fluorescence/methods , Quantum Dots , Retroviridae/ultrastructure , Staining and Labeling/methods , Animals , Cations , Green Fluorescent Proteins , Mice , NIH 3T3 Cells , Retroviridae/chemistry , Retroviridae/isolation & purification
17.
Int J Nanomedicine ; 1(2): 173-80, 2006.
Article in English | MEDLINE | ID: mdl-17722533

ABSTRACT

Reverse microemulsion was used as a template to fabricate chitosan-alginate core-shell nanoparticles encapsulated with enhanced green fluorescent protein (EGFP)-encoded plasmids. The average size of DNA-entrapped nanoparticles measured by dynamic light scattering was increased proportionally, with the N/P ratios ranging from 5 to 20. These alginate-coated chitosan nanoparticles endocytosed by NIH 3T3 cells trigged swelling of transport vesicles which render gene escape before entering digestive endolysosomal compartment and concomitantly promote gene transfection rate. Results showed that DNA-encapsulated chitosan-alginate nanoparticles with average size of 64nm (N/P ratio of 5) could achieve the level of gene expression comparable with the one obtained by using polyethyleneimine-DNA complexes.


Subject(s)
Alginates/chemistry , Chitosan/chemistry , DNA/administration & dosage , DNA/pharmacokinetics , Drug Carriers/chemistry , Nanoparticles/chemistry , Transfection/methods , Animals , DNA/genetics , Diffusion , Emulsions/chemistry , Glucuronic Acid/chemistry , Hexuronic Acids/chemistry , Materials Testing , Mice , NIH 3T3 Cells , Nanoparticles/ultrastructure , Particle Size
18.
Biotechnol Prog ; 21(2): 372-6, 2005.
Article in English | MEDLINE | ID: mdl-15801773

ABSTRACT

Ultrasound-induced cavitation has been extensively used to enhance the efficiency of nonviral-based gene delivery. Although such unique mechanical force could possibly augment the efficacy of retrovirus-mediated gene transfer, we harnessed an alternative approach, a resonant acoustic field, to facilitate the retroviral transduction rate. NIH 3T3 fibroblast cells suspended in a culture well and mixed with ecotropic retroviruses were co-treated with megahertz resonant acoustic fields (RAF). Suspended NIH 3T3 cells under RAF treatment agglomerated at acoustic nodal planes by primary radiation force within a short exposure time. These first arrived and agglomerated cells formed bands as nucleating sites for nanometer-sized ecotropic retroviruses circulated between nodal planes to attach on and thereby increased cell-virus encounters. According to the neomycin-resistant colony assay, 2-fold increment of retroviral transduction rate was obtained by exposing cells and retroviruses in the RAF for 6 min in the presence of 8 microg/mL Polybrene.


Subject(s)
Acoustics , Cell Adhesion , Retroviridae/genetics , Transduction, Genetic , 3T3 Cells , Animals , Mice
19.
J Gene Med ; 7(4): 398-406, 2005 Apr.
Article in English | MEDLINE | ID: mdl-15619287

ABSTRACT

BACKGROUND: Efficiency of retrovirus-mediated gene transfer has been hampered by short retroviral half-life due to shedding of retroviral envelope proteins which is of utmost importance to the interaction between retrovirus and cell. METHODS: Calcium-alginate microspheres with average size of 780 nm were prepared by reverse emulsification and characterized by scanning electron microscopy and microelectrophoresis. To obtain deactivated retroviruses produced from 293 packaging cells, retrovirus-containing media were pre-incubated at 37 degrees C for 6, 12, 18, and 24 h, respectively. Murine J774A.1 macrophages were co-treated with Ca-alginate microspheres and deactivated retroviruses encoding the enhanced green fluorescent protein (eGFP) gene. Through phagocytosis, deactivated retroviruses and Ca-alginate microspheres were co-internalized into macrophages. After retroviral transduction for 24 and 48 h, the percentages of macrophages with eGFP expression were determined by bright-field and fluorescence microscopy. RESULTS: After 48-h incubation with Ca-alginate microspheres and deactivated retroviruses, phagosomes turned into large vacuoles occupied almost half of the cytoplasmic space. This was probably attributed to the erosion of Ca-alginate microspheres by destructive agents within vacuolar compartments and concomitant osmotic swelling. It was reasoned that deactivated retroviruses escaped such enlarged vesicles easily and underwent reverse transcription in the cytosol. The expression of eGFP in macrophages infected by retroviruses pre-incubated for 24 h in the presence of Ca-alginate microspheres was thereby augmented up to tenfold in comparison with the cells treated with 24-h deactivated retroviruses only. CONCLUSIONS: Ca-alginate microspheres performed as auxiliary agents for the enhancement of retrovirus-mediated gene transfer efficiency even though retroviruses had been deactivated due to the loss of envelope proteins.


Subject(s)
Alginates , Macrophages/metabolism , Phagocytosis , Retroviridae/genetics , Transduction, Genetic , Animals , Cell Line , Glucuronic Acid , Hexuronic Acids , Mice , Microspheres
20.
Article in Korean | WPRIM (Western Pacific) | ID: wpr-213821

ABSTRACT

OBJECTIVE: To determine whether unexplained elevation of maternal serum alpha-fetoprotein(MSAFP) is associated with adverse outcomes in pregnancies complicated by placenta previa. METHODS: One hundred and five pregnant women with placenta previa between January 1995 and March 1999 were included in this study. They underwent the triple test at 14 to 22 gestational weeks. These 105 pregnant women were stratified into two groups by MSAFP at 2.0MOM. The outcomes of pregnancy in high MSAFP(>or=2.0MOM) group were compared with those in normal MSAFP(or=2.0 MOM). Eleven out of 632 pregnant women with high MSAFP were having placenta previa. 3. The incidence of preterm birth was significantly high in high MSAFP group with placenta previa.(p<0.001) 4. Women with unexplained elevated MSAFP and having placenta previa showed the increased risk for preterm delivery (p<0.05), preeclamsa (p<0.001), and bleeding in third trimester (p<0.001), but not placenta accreta, placenta abruption, cesarean hysterctomy and intrauterine fetal death. CONCLUSION: We concluded that unexplained elevated levels of midtrimester MSAFP in complicated placenta previa were associated with the elevated risk of preterm delivery, preeclamsia and bleeding in the early third trimester, but not placenta accreta, placenta abruption, cearean hysterectomy and intrauterine fetal death.


Subject(s)
Female , Humans , Pregnancy , alpha-Fetoproteins , Fetal Death , Hemorrhage , Hysterectomy , Incidence , Placenta Accreta , Placenta Previa , Placenta , Pregnancy Trimester, Second , Pregnancy Trimester, Third , Pregnant Women , Premature Birth
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