Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The present work utilizes electrospinning to fabricate synthetic polymer/DNA composite scaffolds for therapeutic application in gene delivery for tissue engineering. The scaffolds are non-woven, nano-fibered, membranous structures composed predominantly of poly(lactide-co-glycolide) (PLGA) random copolymer and a poly(D,L-lactide)-poly(ethylene glycol) (PLA-PEG) block copolymer. Release of plasmid DNA from the scaffolds was sustained over a 20-day study period, with maximum release occurring at approximately 2 h. Cumulative release profiles indicated amounts released were approximately 68-80% of the initially loaded DNA. Variations in the PLGA to PLA-PEG block copolymer ratio vastly affected the overall structural morphology, as well as both the rate and efficiency of DNA release. Results indicated that DNA released directly from these electrospun scaffolds was indeed intact, capable of cellular transfection, and successfully encoded the protein beta-galactosidase. When tested under tensile loads, the electrospun polymer/DNA composite scaffolds exhibited tensile moduli of approximately 35 MPa, with approximately 45% strain initially. These values approximate those of skin and cartilage. Taken together, this work represents the first successful demonstration of plasmid DNA incorporation into a polymer scaffold using electrospinning.
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PMID:Development of a nanostructured DNA delivery scaffold via electrospinning of PLGA and PLA-PEG block copolymers. 1271 56

The use of stem cells is promising for future cell-based therapy such as tissue regeneration and engineering. Although Embryonic Stem Cells (ESCs) are theoretically highly beneficial, there are some potential limitations of cell regulations and ethical consideration. Mesenchymal Stem Cells (MSCs) isolated from bone marrow stroma have been shown to possess adipogenic, osteogenic, chondrogenic, myogenic and neurogenic potential in vitro. However, bone marrow procurement is severely painful for donors and often requires general anesthesia. Moreover, only small numbers of cells can be harvested. We previously hypothesized that human adipose tissue obtained from liposuction procedures also contains the same cell population as MSCs, because adipose tissue is mesenchymal in origin, like bone marrow stroma. Subsequent studies revealed that: (1) cell population (which we termed Processed Lipoaspirate [PLA] cells), observed by indirect immunofluorescence study of adipose tissue, consist of cells of mesenchymal origin that have little contamination with endothelial cells, smooth muscle cells and pericytes; (2) these PLA cells exhibit low levels of cell senescence even after multiple passage, as demonstrated by beta-galactosidase staining assay; and (3) PLA cells can differentiate into adipogenic, osteogenic, chondrogenic and myogenic cells in vitro in lineage-specific culture media. These findings suggest that human PLA might have a mesodermal stem cell population. Since human adipose tissue is plentiful, easily harvested in large quantity under local anesthesia with little patient discomfort, it may be an alternative stem cell source for mesenchymal tissue regeneration and engineering. This review highlights our previous research work on PLA cells and future clinical perspectives, particularly in the field of plastic and reconstructive surgery.
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PMID:Mesengenic potential and future clinical perspective of human processed lipoaspirate cells. 1292 9

The entrapment of beta-galactosidase (Escherichia coli) in PLA and PLGA microspheres using a double emulsion technique resulted to significant reduction of protein antigenicity. The extent of antigenicity loss depended on the conditions of microsphere preparation. Most of antigenicity loss occurred on the first emulsification step. Only the effects of microsphere preparation factors having an important influence on protein antigenicity, such as the type of organic phase (polymer solvent) and homogenization, could be predicted (on a qualitative basis) by antigenicity data obtained after the first emulsification step. The type of polymer and polymer solvent used to prepare the microspheres affected beta-galactosidase immunogenicity. The PLA microspheres prepared using ethyl acetate was the most immunogenic microsphere formulation, eliciting similar total antibody responses as the alum formulation of beta-gal. This formulation was the only microsphere formulation that induced an IgG1/IgG2a ratio lower than 1, indicating an immune response biased towards a Th1 type. The results obtained indicate that large protein molecules with complex tertiary structure such as beta-galactosidase can be entrapped in PLA and PLGA microspheres with retention of protein immunogenic potential, providing that appropriate conditions of microsphere preparation are applied, and that the formulation of microspheres might influence the Th1/Th2 type of immune response against the encapsulated antigen.
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PMID:PLA and PLGA microspheres of beta-galactosidase: Effect of formulation factors on protein antigenicity and immunogenicity. 1517 18

The immune response induced in mice by beta-galactosidase (beta-gal) adsorbed or encapsulated on poly(lactic acid) (PLA) and poly(lactic-co-glycolic acid) (PLGA) microspheres was investigated. The encapsulated protein elicited higher antibody response than the protein adsorbed on the microspheres in the case of the PLA microspheres. However, the encapsulated protein elicited weaker antibody response than the adsorbed protein in the case of the PLGA (50:50) microspheres, probably because, in this case, the encapsulation process adversely affected protein immunogenicity. In the case of adsorbed beta-gal, higher antibody response was obtained with the PLA microspheres than with the PLGA (50:50) microspheres. This may be related to the lower rate of beta-gal desorption from the PLA microspheres. Based on the immunoglobulin G1/immunoglobulin G2a ratios and the stimulation indices for interferon-gamma and interleukin-4, beta-gal encapsulated or adsorbed on PLA microspheres induced a Th(1)-biased immune response whereas beta-gal encapsulated or adsorbed on PLGA (50:50) microspheres induced a Th(2)-biased immune response. The results obtained indicate that more potent immune responses are obtained when the protein is encapsulated than adsorbed on the microspheres, providing that the encapsulation process does not adversely affect protein immunogenicity. Also, the type of polymer used to prepare the microspheres, but not the method of protein association with the microspheres, may affect the type of immune response.
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PMID:Immune responses in mice of beta-galactosidase adsorbed or encapsulated in poly(lactic acid) and poly(lactic-co-glycolic acid) microspheres. 1579 20

The objective of this study was to investigate the effect of formulation parameters (i.e. polymer molecular weight and homogenization speed) on various physicochemical and biological properties of cationic nanoparticles. Cationic nanoparticles were prepared using different molecular weights of poly(DL-lactide-co-glycolide) (PLGA) and poly(DL-lactic acid) (PLA) by double emulsion solvent evaporation at two different homogenization speeds, and were characterized in terms of size, surface charge, morphology, loading efficiency, plasmid release, plasmid integrity, cytotoxicity, and transfection efficiency. Cationic surfactant, cetyltrimethylammonium bromide (CTAB), was used to provide positive charge on the surface of nanoparticles. Reporter plasmid gWIZ Beta-gal was loaded on the surface of nanoparticles by incubation. Use of higher homogenization speed and lower molecular weight polymer led to a decrease in mean particle size, increase in zeta potential, increase in plasmid loading efficiency, and a decrease in burst release. The nanoparticles displayed good morphology as evident from scanning electron micrographs. In vitro cytotoxicity study by MTT assay showed a low toxicity. Structural integrity of the pDNA released from nanoparticles was maintained. Transfecting human embryonic kidney (HEK293) cells with nanoparticles prepared from low molecular weight PLGA and PLA resulted in an increased expression of beta-galactosidase as compared to those prepared from high molecular weight polymer. Our results demonstrate that the PLGA and PLA cationic nanoparticles can be used to achieve prolonged release of pDNA, and the plasmid release rate and transfection efficiency are dependent on the formulation variables.
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PMID:Preparation, characterization, cytotoxicity and transfection efficiency of poly(DL-lactide-co-glycolide) and poly(DL-lactic acid) cationic nanoparticles for controlled delivery of plasmid DNA. 1761 Oct 54

Non-viral vectors such as liposomes, polycations, and nanoparticles have been used as gene delivery systems. In this study, we prepared and characterized biodegradable poly(L-lactic acid) (PLA)/polyethylenimine (PEI) nanoparticles as gene carriers. pCMV/beta-gal and pEGFP-C1 were utilized as model plasmid DNAs (pDNA). Nanoparticles were prepared using a double emulsion-solvent evaporation technique, and their pDNA binding capacity was assessed by agarose gel electrophoresis. Transfection was studied in HEK 293 and HeLa cell lines, and the transfection efficiencies were determined by beta-galactosidase assay or flow cytometry. Three kinds of PLA/PEI systems were studied by varying the molecular weight of PEI. The PLA/PEI 25K system had a higher transfection efficiency than the PLA/PEI 0.8K or PLA/PEI 750K systems. The transfection efficiency was found to be dependent on the ratio of PLA/PEI nanoparticles to pDNA with an optimum ratio of 60:1 (w/w). The cytotoxicity was dependent on the quantity of PLA/PEI nanoparticles used, but it was comparable to that of commercial Lipofectin. These results demonstrate the potential of PLA/PEI nanoparticles as gene carriers.
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PMID:Poly(L-lactic acid)/polyethylenimine nanoparticles as plasmid DNA carriers. 1827 14

The effect of bcl-2 expression on cell viability and recombinant protein synthesis was investigated in the Spodoptera frugiperda Sf-9 and Trichoplusia ni BTI-Tn-5B1-4 (High Fivetrade mark) insect cell lines. It was found that coinfection with a baculovirus expressing bcl-2 [Autographa californica nuclear polyhedrosis virus (AcNPV)-bcl2] extended the life span of High Fivetrade mark cells but not Sf-9 cells when compared to infection with recombinant baculoviruses expressing either human tissue plasminogen activator (AcNPV-tPA) or Escherichia coli beta-galactosidase (AcNPV-betagal). Similar results were obtained in coinfection experiments; i.e., AcNPV-bcl2 coinfection increased the life span of High Fivetrade mark cells over that of cells infected with either AcNPV-tPA or AcNPV-betagal alone, but they did not affect the life span of coinfected Sf-9 cells. Coinfection of Sf-9 cells with AcNPV-bcl2 and AcNPV-betagal resulted in a decrease in the maximum beta-gal expression levels of over 90% when compared to infection with AcNPV-betagal alone. A similar trend was found in the beta-gal mRNA levels. Coinfection also resulted in a reduced beta-gal expression level in High Fivetrade mark cells, but the reduction was consistent with what would be expected when two recombinant viruses compete for use of the cellular machinery. In contrast to the inhibitory effect of AcNPV-bcl2 coinfection on betagal expression, t-PA expression levels were either not affected (Sf-9 cells) or were increased 50% (High Fivetrade mark cells) over those obtained by infection with AcNPV-tPA alone. These results support the hypotheses that bcl-2 can inhibit transcription of genes under polyhedrin promoter control and that beta-gal expression levels, but not t-PA expression levels, are controlled at the transcriptional level. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 380-390, 1997.
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PMID:bcl-2 expression in Spodoptera Frugiperda Sf-9 and Trichoplusia Ni BTI-Tn-5B1-4 insect cells: effect on recombinant protein expression and cell viability. 1864 41

Cardiopulmonary dirofilariosis (Dirofilaria immitis) is characterized by apparent contradictory events, like the long-term survival of adult worms in the circulatory system of the infected hosts and the development of life-threatening events like thromboembolisms and others. Thus parasite mechanisms, like the activation of fibrinolytic system, are key to the survival of both the worms and the host. The aim of this study was to investigate the interaction between D. immitis adult worms surface-associated antigens (DiSAA) and the fibrinolytic system of the host. We demonstrate that DiSAA extract is able to bind plasminogen and generate plasmin, with the latter occurring in a tissue plasminogen activator (t-PA) dependent manner. Additionally, 11 plasminogen-binding proteins from DiSAA extract were identified by proteomics and mass spectrometry (MS) (actin-5C, actin-1, enolase, fructose-bisphosphate aldolase, GAPDH, MSP domain protein, MSP 2, beta-galactosidase-binding-lectin, galectin, immunoglobulin I-set domain-containing protein and cyclophilin Ovcyp-2). Because in a previous work we have shown the positive interaction between the excretory/secretory antigens of D. immitis (DiES) and the host fibrinolytic system and many of the molecules identified here are shared by both antigens, we hypothesize that DiSAA cooperate in host fibrinolytic system activation promoting the fibrin clot lysis.
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PMID:Surface associated antigens of Dirofilaria immitis adult worms activate the host fibrinolytic system. 2343 49


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