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Biomedical subjects

K W Leong

Publications and source records attributed to K W Leong.

At least 19 recordsLinked to original sources
PubMed2004

CNS gene transfer mediated by a novel controlled release system based on DNA complexes of degradable polycation PPE-EA: a comparison with polyethylenimine/DNA complexes.

Nonviral gene delivery systems based upon polycation/plasmid DNA complexes are quickly gaining recognition as an alternative to viral gene vectors for their potential in avoiding immunogenicity and toxicity problems inherent in viral systems. We investigated in this study the feasibility of using a controlled release system based on DNA complexed with a recently developed polymeric gene carrier, polyaminoethyl propylene phosphate (PPE-EA), to achieve gene transfer in the brain. A unique feature of this gene delivery system is the biodegradability of PPE-EA, which can provide a sustained release of DNA at different rates depending on the charge ratio of the polymer to DNA. PPE-EA/DNA complexes, naked DNA, and DNA complexed with polyethylenimine (PEI), a nondegradable cationic polymer known to be an effective gene carrier, were injected intracisternally into the mouse cerebrospinal fluid. Transgene expression mediated by naked DNA was mainly detected in the brain stem, a region close to the injection site. With either PPE-EA or PEI as a carrier, higher levels of gene expression could be detected in the cerebral cortex, basal ganglia, and diencephalons. Transgene expression in the brain mediated by PPE-EA/DNA complexes at an N/P ratio of 2 persisted for at least 4 weeks, with a significant higher level than that produced by either naked plasmid DNA or PEI/DNA at the 4-week time point. Furthermore, PPE-EA displayed much lower toxicity in cultured neural cells as compared to PEI and did not cause detectable pathological changes in the central nervous system (CNS). The results established the potential of PPE-EA as a new and biocompatible gene carrier to achieve sustained gene expression in the CNS.

Animals↗
PubMed2004

Polyphosphoramidate gene carriers: effect of charge group on gene transfer efficiency.

Cationic polymeric carriers have been widely used for gene delivery. However, the structure-function relationship, especially the effect of charge groups of cationic polymeric carriers on the transfection activity, is poorly understood. To examine this important parameter, a series of cationic polymers, polyphosphoramidates (PPAs) with an identical backbone, same side chain spacer, similar molecular weights but different charge groups containing primary to quaternary amino groups (PPA-EA, PPA-MEA, PPA-DMA and PPA-TMA, Figure 1) were synthesized. The DNA-binding affinity of these four PPAs increased in the order of PPA-EA PPA-MEA>PPA-DMA>PPA-TMA. Particle size and zeta potential of four different types of PPA/DNA nanoparticles did not show significant correlation with PPA structure. These PPAs did not show significant buffering capacity within pH 5-7, even though transfection mediated by PPA-EA was the only one that seemed to be limited by endolysomal escape. Endocytosis of DNA mediated by PPAs was also similar (17-22%) for all four PPAs. However, the transfection efficiency of these PPAs varied significantly. In vitro transfection efficiency of PPAs decreased in the order of PPA-EA>PPA-MEA>PPA-DMA approximately PPA-TMA. Nanoparticles with PPA-EA containing primary amino groups gave the highest transfection efficiency in cell lines at the charge ratios from 6/1 to 20/1 (+/-). Matching the trend of transfection efficiency observed in vitro, PPA-EA mediated the highest transgene expression, comparable to that of polyethylenimine, in the spinal cord following intrathecal injection of the nanoparticles. These results establish that PPA gene carriers with primary amino group side chains are more potent than those with secondary, tertiary or quaternary amino groups in vitro and in the intrathecal gene delivery model.

Animals↗
PubMed2003

Repeated intrathecal administration of plasmid DNA complexed with polyethylene glycol-grafted polyethylenimine led to prolonged transgene expression in the spinal cord.

Gene delivery into the spinal cord provides a potential approach to the treatment of spinal cord traumatic injury, amyotrophic lateral sclerosis, and spinal muscular atrophy. These disorders progress over long periods of time, necessitating a stable expression of functional genes at therapeutic levels for months or years. We investigated in this study the feasibility of achieving prolonged transgene expression in the rat spinal cord through repeated intrathecal administration of plasmid DNA complexed with 25 kDa polyethylenimine (PEI) into the lumbar subarachnoid space. With a single injection, DNA/PEI complexes could provide transgene expression in the spinal cord 40-fold higher than naked plasmid DNA. The transgene expression at the initial level persisted for about 5 days, with a low-level expression being detectable for at least 8 weeks. When repeated dosing was tested, a 70% attenuation of gene expression was observed following reinjection at a 2-week interval. This attenuation was associated with apoptotic cell death and detected even using complexes containing a noncoding DNA that did not mediate any gene expression. When each component of the complexes, PEI polymer or naked DNA alone, were tested in the first dosing, no reduction was found. Using polyethylene glycol (PEG)-grafted PEI for DNA complexes, no attenuation of gene expression was detected after repeated intrathecal injections, even in those rats receiving three doses, administered 2 weeks apart. Lumbar puncture is a routine and relatively nontraumatic clinical procedure. Repeated administration of DNA complexed with PEG-grafted PEI through this less invasive route may prolong the time span of transgene expression when needed, providing a viable strategy for the gene therapy of spinal cord disorders.

Animals↗
PubMed2002

Multi-layered microcapsules for cell encapsulation.

Mechanical stability, complete encapsulation, selective permeability, and suitable extra-cellular microenvironment, are the major considerations in designing microcapsules for cell encapsulation. We have developed four types of multi-layered microcapsules that allow selective optimization of these parameters. Primary hepatocytes were used as model cells to test these different microcapsule configurations. Type-1 microcapsules with an average diameter of 400 microm were formed by complexing modified collagen with a ter-polymer shell of 2-hydroxyethyl methylacrylate (HEMA), methacrylic acid (MAA) and methyl methacrylate (MMA), resulting in a capsule thickness of 2-5 microm. Cells in these microcapsules exhibited improved cellular functions over those cultured on collagen monolayers. Type-II microcapsules were formed by encapsulating the Type-I microcapsules in another 2-5 microm ter-polymer shell and a approximately 5 microm collagen layer between the two ter-polymer shells to ensure complete cell encapsulation. Type-II microcapsules comprised of a macro-porous exoskeleton with materials such as alumina sol-gel coated on the Type-I microcapsules. Nano-indendation assay indicated an improved mechanical stability over the Type-I microcapsules. Type-IV microcapsules were created by encapsulating Type-III microcapsules in another 2-5 microm ter-polymer shell, with the aim of imparting a negatively charged smooth surface to minimize plasma protein absorption and ensure complete cell encapsulation. The permeability for nutrient exchange, cellular functions in terms of urea production and mechanical stability of the microcapsules were characterized. The advantages and limitations of these microcapsules for tissue engineering are discussed.

Animals↗
PubMed2002

Enhanced gene expression in mouse muscle by sustained release of plasmid DNA using PPE-EA as a carrier.

Delivery of plasmid DNA by nanoparticles improves the DNA bioavailability, for instance in intramuscular administration, by localizing the DNA in the muscle tissue. Extracellular sustained release of the DNA may lead to more prolonged transgene expression. The present study describes a novel controlled gene delivery system based on a water soluble and biodegradable polyphosphoester, poly(2-aminoethyl propylene phosphate) (PPE-EA). The polymer degraded in PBS at 37 degrees C through the cleavage of the backbone phosphate bonds, and it was synthesized with a relative high molecular weight to ensure a suitable hydrolytic stability as a gene carrier. The tissue response and cytotoxicity study demonstrated a better tissue compatibility of PPE-EA in mouse muscle compared with commonly used polyethylenimine and poly-L-lysine. PPE-EA condensed DNA efficiently and protected DNA from nuclease and serum degradation. Sustained release of plasmid was achieved from PPE-EA/DNA complexes as a result of PPE-EA degradation. The DNA release profiles appear to be predominantly controlled by carrier degradation and the release rate of plasmid could be adjusted by varying the charge ratio of PPE-EA to DNA. At an N/P (amino to phosphate groups) ratio of 1, a 46% burst was observed for the first day, followed by about 4% release per day (24 microg DNA/day/mg of complex) for 12 days. Higher charge ratios reduced both the DNA release rate and the burst effect. The released DNA retained its structural and functional integrity. Intramuscular injection of PPE-EA-p43-LacZ complexes at N/P ratios of 0.5 and 1 resulted in enhanced beta-galactosidase expression in anterior tibialis muscle in Balb/c mice, as compared with naked DNA injections. Similarly, PPE-EA/IFN(alpha)2b DNA complexes generated an increased systemic level of interferon-alpha2b in mouse serum following intramuscular injection, as compared with naked DNA injection.

Absorbable Implants↗
PubMed2001

Chitosan-DNA nanoparticles as gene carriers: synthesis, characterization and transfection efficiency.

Chitosan-DNA nanoparticles were prepared using a complex coacervation process. The important parameters for the nanoparticle synthesis were investigated, including the concentrations of DNA, chitosan and sodium sulfate, temperature of the solutions, pH of the buffer, and molecular weights of chitosan and DNA. At an amino group to phosphate group ratio (N/P ratio) between 3 and 8 and a chitosan concentration of 100 microg/ml, the size of particles was optimized to approximately 100--250 nm with a narrow distribution, with a composition of 35.6 and 64.4% by weight for DNA and chitosan, respectively. The surface charge of these particles was slightly positive with a zeta potential of +12 to +18 mV at pH lower than 6.0, and became nearly neutral at pH 7.2. The chitosan-DNA nanoparticles could partially protect the encapsulated plasmid DNA from nuclease degradation as shown by electrophoretic mobility analysis. The transfection efficiency of chitosan-DNA nanoparticles was cell-type dependent. Typically, it was three to four orders of magnitude, in relative light units, higher than background level in HEK293 cells, and two to ten times lower than that achieved by LipofectAMINE-DNA complexes. The presence of 10% fetal bovine serum did not interfere with their transfection ability. Chloroquine could be co-encapsulated in the nanoparticles at 5.2%, but with negligible enhancement effect despite the fact that chitosan only showed limited buffering capacity compared with PEI. The present study also developed three different schemes to conjugate transferrin or KNOB protein to the nanoparticle surface. The transferrin conjugation only yielded a maximum of four-fold increase in their transfection efficiency in HEK293 cells and HeLa cells, whereas KNOB conjugated nanoparticles could improve gene expression level in HeLa cells by 130-fold. Conjugation of PEG on the nanoparticles allowed lyophilization without aggregation, and without loss of bioactivity for at least 1 month in storage. The clearance of the PEGylated nanoparticles in mice following intravenous administration was slower than unmodified nanoparticles at 15 min, and with higher depositions in kidney and liver. However, no difference was observed at the 1-h time point.

Animals↗
PubMed2001

Transgene expression in the brain stem effected by intramuscular injection of polyethylenimine/DNA complexes.

Gene delivery into the CNS without tissue destruction is challenging. As neurons are capable of taking up exogenous particulates from the muscles that they innervate, we investigated the feasibility of achieving gene transfer in CNS neurons by peripheral intramuscular injection of plasmid DNA complexed with the cationic polymer polyethylenimine (PEI) in the rat hypoglossal system. Using the luciferase reporter gene driven by a Rous sarcoma virus promoter, transgene expression of up to 4 x 10(6) RLU per brain stem at 20 microg of plasmid DNA was achieved after tongue injection. Using lacZ as a reporter gene, transgene expression in the brain stem was detected in hypoglossal motor neurons, a group of neurons that innervate tongue muscles. The plasmid DNA was detected by PCR analysis in the brain-stem samples, demonstrating that the PEI/DNA complexes had migrated by retrograde axonal transport to neuronal cell bodies in the brain stem after being internalized by nerve terminals in the tongue muscle. Using a therapeutic bcl-2 gene driven by a cytomegalovirus promoter and Western blotting, transgene expression was detectable in the brain stem as early as 18 h after tongue injection and lasted for at least 2 weeks. Two lipid transfection agents, GenePORTER and TransFast, mediated a weak gene expression in the hypoglossal system, but not two polymers, poly-l-lysine and chitosan. The nonviral neuronal gene delivery method established in this study bypasses the blood-brain barrier and suggests a possible therapeutic strategy for noninvasive CNS gene transfer.

Animals↗
PubMed2001

Fabrication of poly(phosphoester) nerve guides by immersion precipitation and the control of porosity.

Immersion precipitation was employed as a method for the fabrication of polymeric conduits from P(BHET-EOP/TC), a poly(phosphoester) with an ethylene terephthalate backbone, to be applied as guidance channels for nerve regeneration. Coatings of various porosities could be obtained by immersing mandrels coated with a solution of the polymer in chloroform into non-solvent immersion baths, followed by freeze or vacuum-drying. The porosity of the coatings decreased with an increase in polymer molecular weight, drying time before precipitation and concentration of polymer solution. The effects of these parameters can be rationalized by employing ternary phase diagrams, where porosity is directly related to the degree of phase separation available to the system before gelation occurs. To afford improved porosity control, a new system was developed which employed the contrasting phase-separation behavior of P(BHET-EOP/TC)/chloroform solution in methanol and water. As water is essentially a non-solvent for the polymer, the demixing boundary of the P(BHET-EOP/TC)-CHCl3-H2O system is located close to the polymer-solvent edge of the phase diagram, while that of the P(BHET-EOP/TC)-CHCl3-MeOH system is located further away. A mixture of methanol and water allows the demixing boundary to be shifted to intermediate coordinates. By immersing P(BHET-EOP/TC) coatings in immersion baths containing different ratios of water and methanol, then gradually titrating the bath with methanol to a concentration of 70% (v/v) methanol, surface porosities ranging from 2 to 58% could be achieved.

Biocompatible Materials↗
PubMed2001

A new nerve guide conduit material composed of a biodegradable poly(phosphoester).

There is a resurgence of interest in the development of degradable and biocompatible polymers for fabrication of nerve guide conduits (NGCs) in recent years. Poly(phosphoester) (PPE) polymers are among the attractive candidates in this context, in view of their high biocompatibility, adjustable biodegradability, flexibility in coupling fragile biomolecules under physiological conditions and a wide variety of physicochemical properties. The feasibility of using a biodegradable PPE, P(BHET-EOP/TC), as a novel NGC material was investigated. Two types of conduits were fabricated by using two batches of P(BHET-EOP/TC) with different weight-average molecular weights (Mw) and polydispersity indexes (PI). The polymers as well as conduits were non-toxic to all six types of cells tested, including primary neurones and neuronally differentiated PC12 cells. After in situ implantation in the sciatic nerve of the rat, two types of conduits triggered a similar tissue response, inducing the formation of a thin tissue capsule composed of approximately eight layers of fibroblasts surrounding the conduits at 3 months. Biological performances of the conduits were examined in the rat sciatic nerve model with a 10 mm gap. Although tube fragmentation, even tube breakage, was observed within less than 5 days post-implantation, successful regeneration through the gap occurred in both types of conduits, with four out of 10 in the Type I conduits (Mw 14,900 and PI 2.57) and 11 out of 12 in the Type II conduits (Mw 18,900 and PI 1.72). The degradation of conduits was further evidenced by increased roughness on the tube surface in vivo under scanning electron microscope and a mass decrease in a time-dependent manner in vitro. The Mw of the polymers dropped 33 and 24% in the Type I and II conduits, respectively, in vitro within 3 months. Among their advantages over other biodegradable NGCs, the PPE conduits showed negligible swelling and no crystallisation after implantation. Thus, these PPE conduits can be effective aids for nerve regeneration with potential to be further developed into more sophisticated NGCs that have better control of the conduit micro-environment for improved nerve regeneration.

Animals↗
PubMed2001

Interactions of phospholipid bilayer with chitosan: effect of molecular weight and pH.

Chitosan has demonstrated its potentials as a gene carrier and a membrane perturbant for subsequent drug delivery to cells. However, there is currently a lack of experimental correlation between the physiochemical properties of chitosan and the resulting degree of lipid bilayer destabilization. In this study, the effect of pH and chitosan molecular weight on the interaction between chitosan and dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayer was examined with cross-polarization microscopy, differential scanning calorimetry (DSC), and Fourier transform- (FT-) Raman spectroscopy. Cross-polarized images showed that the direct hydration of the DPPC/chitosan mixture led to the formation of larger DPPC multilamellar vesicles (MLV), and pure chitosan also induced fusions of individual MLV. Under the influence of chitosan, the calorimetric enthalpy of DPPC was reduced in a concentration-dependent manner, and a new phase appeared at 28 degrees C during sample cooling. Even the lowest chitosan mole fraction of 0.04% reduced the cooperative unit of the DPPC bilayer by more than 70%. In addition, the electrostatic effect between chitosan and DPPC tuned the degree of membrane bilayer perturbation. Reduction of pH increased the number of protonated amines on the chitosan backbone and caused further disruption on the membrane organization. Mixing DPPC with chitosan in an organic medium before hydration enhanced the hydrophobic interactions between the two molecules and greatly reduced the cooperative unit among individual lipids during the main phase transition. The increase of chitosan molecular weight also affected the cooperativity in the thermotropic transition of DPPC bilayer. FT-Raman spectroscopy provided additional evidence that chitosan directly perturbed the organizations of the hydrophobic inner core of the DPPC bilayer.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2001

Controlled local delivery of interleukin-2 by biodegradable polymers protects animals from experimental brain tumors and liver tumors.

PURPOSE: The purpose of our study was to develop an injectable polymeric system for the long-term localized delivery of bioactive interleukin-2 for antitumor immunotherapy. METHODS: IL-2 was encapsulated into gelatin and chondroitin-6-sulfate using an aqueous-based complex coacervation. CTLL-2 cells were used to measure the bioactivity of released IL-2 and radiolabeled IL-2 was used for release studies in the rat brain and mouse liver. Antitumor efficacy studies were carried out in primary (9L gliosarcoma) and metastatic (B16-F10 melanoma) brain tumor models in rats and mice, respectively, as well as a murine liver tumor model (CT26 carcinoma). Survivors of the metastatic brain tumor challenge were rechallenged with tumor in the opposite lobe of the brain to confirm that antitumor immunologic memory had developed. RESULTS: Bioactive IL-2 was released for over 2 weeks in vitro and in vivo IL-2 release showed significant IL-2 levels for up to 21 days. Polymeric IL-2 microspheres injected intratumorally were statistically more effective in protecting animals challenged with fatal tumor doses in the brain and the liver than placebo or autologous tumor cells genetically engineered to secrete IL-2. Immunologic memory was induced following IL-2 microsphere therapy in the B16-F10 brain tumor model that was capable of protecting 42% of animals from a subsequent intracranial tumor challenge, suggesting that tumor destruction was mediated by the immune system. CONCLUSIONS: Local IL-2 therapy using novel polymeric carriers. aimed at stimulating long-lasting antitumor immunity, may provide an improved method of treating a variety of cancers.

Animals↗
PubMed2000

Antigen-specific induction of peripheral T cell tolerance in vivo by codelivery of DNA vectors encoding antigen and Fas ligand.

Fas ligand (FasL, CD95L) induces apoptosis in activated T cells with upregulated Fas (CD95) expression through the process termed activation-induced cell death (AICD). We postulated that coexpression of antigen and FasL within individual antigen-presenting cells would lead to antigen-specific activation of T cells and to their consequent deletion by FasL-mediated AICD. A DNA-gelatin coacervate containing transferrin cell ligand, calcium, and the lysosomatropic agent chloroquine, a formulation previously shown to achieve high-level transfection of immune and muscle cells in vivo, was used to codeliver plasmids encoding FasL and antigen. Mice developed a strong cytolytic T cell response to beta-Gal when injected with DNA encoding beta-galactosidase (LacZ) model antigen, either as naked DNA or DNA nanoparticles, but failed to respond when there was concomitant injection of nanoparticles containing both the LacZ and murine FasL DNA vectors. This loss of T cell response was systemic, specific for beta-Gal, complete when nanoparticles were administered before antigen challenge, and decreased the T cell response from prior immunization with LacZ DNA. In effect, this "tolerization" injection induced antigen-specific peripheral tolerance in study mice, and represents a possible approach to the treatment of autoimmune diseases and transplantation rejection.

Animals↗
PubMed2000

Hepatocyte encapsulation for enhanced cellular functions.

An efficient bioartificial liver-assisted device can sustain the lives of patients with acute liver failure. Among different configurations of the bioreactor design, hepatocyte encapsulation has important features that satisfy most requirements of the device. We have encapsulated rat hepatocytes in a two-layer polymeric membrane by complex coacervation using a simple setup and demonstrated enhanced cellular functions up to three times higher than those of the monolayer control. These microcapsules of the functioning hepatocytes have a 2- to 3-microm outer layer of synthetic polymer with 25% 2-hydroxyethyl methacrylate, 25% methacrylic acid, and 50% methyl methacrylate and an inner layer of positively charged modified collagen as a suitable substrate for the enhanced cellular functions. Permeable only to small molecules up to albumin, the microcapsules should allow unimpeded exchange of nutrients, oxygen, growth factors, and metabolites but prevent attack by immunoglobulins of the immune system, and no "skin effect" of the collagen has been observed. Mechanical properties of the microcapsules measured with a nano-indentation method suggest that the microcapsules should be suitable for use in a bioartificial liver-assisted device.

Animals↗
PubMed1999

Gene transfer by DNA-gelatin nanospheres.

A DNA and gelatin nanoparticle coacervate containing chloroquine and calcium, and with the cell ligand transferrin covalently bound to the gelatin, has been developed as a gene delivery vehicle. In this study, the coacervation conditions which resulted in the formation of distinct nanoparticles are defined. Nanospheres formed within a narrow range of DNA concentrations and achieved incorporation of more than 98% of the DNA in the reaction. Crosslinking of gelatin to stabilize the particles does not effect the electrophoretic mobility of the DNA. DNA in the nanosphere is partially resistant to digestion with concentrations of DNase I that result in extensive degradation of free DNA but is completely degraded by high concentrations of DNase. Optimum cell transfection by nanosphere DNA required the presence of calcium and nanospheres containing transferrin. The biological integrity of the nanosphere DNA was demonstrated with a model system utilizing DNA encoding the cystic fibrosis transport regulator (CFTR). Transfection of cultured human tracheal epithelial cells (9HTEo) with nanospheres containing this plasmid resulted in CFTR expression in over 50% of the cells. Moreover, human bronchial epithelial cells (IB-3-1) defective in CFTR-mediated chloride transport were complemented with effective transport activity when transfected with nanospheres containing the CFTR transgene.

Calcium↗
PubMed1999

Oral gene delivery with chitosan--DNA nanoparticles generates immunologic protection in a murine model of peanut allergy.

Food allergy is a common and often fatal disease with no effective treatment. We describe here a new immunoprophylactic strategy using oral allergen-gene immunization to modulate peanut antigen-induced murine anaphylactic responses. Oral administration of DNA nanoparticles synthesized by complexing plasmid DNA with chitosan, a natural biocompatible polysaccharide, resulted in transduced gene expression in the intestinal epithelium. Mice receiving nanoparticles containing a dominant peanut allergen gene (pCMVArah2) produced secretory IgA and serum IgG2a. Compared with non-immunized mice or mice treated with 'naked' DNA, mice immunized with nanoparticles showed a substantial reduction in allergen-induced anaphylaxis associated with reduced levels of IgE, plasma histamine and vascular leakage. These results demonstrate that oral allergen-gene immunization with chitosan-DNA nanoparticles is effective in modulating murine anaphylactic responses, and indicate its prophylactic utility in treating food allergy.

2S Albumins, Plant↗
PubMed1998

Controlled gene delivery by DNA-gelatin nanospheres.

A novel system for gene delivery, based on the use of DNA-gelatin nanoparticles (nanospheres) formed by salt-induced complex coacervation of gelatin and plasmid DNA, has been developed. These particles were spherical, with a size range of 200-700 nm, contained 25-30% (w/w) DNA, and were stabilized by cross-linking of gelatin. As a consequence of being controlled by the cross-linking density of the gelatin matrix, the average release rate of DNA from nanospheres synthesized under standard conditions was 2.2%/day in serum. Nanosphere DNA incubated in bovine serum was more resistant to nuclease digestion than was naked DNA. Various bioactive agents could be encapsulated in the nanospheres by ionic interaction with the matrix components, physical entrapment, or covalent conjugation. Transfection of cultured cells with a luciferase plasmid was enhanced by conjugating human transferrin onto the nanosphere and coencapsulating the endolysolytic agent chloroquine. Under our experimental conditions, gene expression in mice subsequent to intramuscular injection of nanospheres containing 1 microg of a beta-galactosidase plasmid was greater and more prolonged than was observed after injection of an equal amount of naked DNA or DNA complexed with Lipofectamine.

Animals↗
PubMed1998

DNA-polycation nanospheres as non-viral gene delivery vehicles.

Nanospheres synthesized by salt-induced complex coacervation of cDNA and polycations such as gelatin and chitosan were evaluated as gene delivery vehicles. DNA-nanospheres in the size range of 200-750 nm could transfect a variety of cell lines. Although the transfection efficiency of the nanospheres was typically lower than that of lipofectamine and calcium phosphate controls in cell culture, the beta-gal expression in muscle of BALB/c mice was higher and more sustained than that achieved by naked DNA and lipofectamine complexes. This gene delivery system has several attractive features: (1) ligands can be conjugated to the nanosphere for targeting or stimulating receptor-mediated endocytosis; (2) lysosomolytic agents can be incorporated to reduce degradation of the DNA in the endosomal and lysosomal compartments; (3) other bioactive agents or multiple plasmids can be co-encapsulated; (4) bioavailability of the DNA can be improved because of protection from serum nuclease degradation by the polymeric matrix; (5) the nanosphere can be lyophilized for storage without loss of bioactivity.

Animals↗