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At least 109 records · Page 6Linked to original sources
PubMed2002

Transfection of liver in vivo by biolistic particle delivery: its use in the investigation of cytochrome P450 gene regulation.

We describe the delivery of reporter gene constructs to rat liver through the use of the Helios Gene Gun system. The effectiveness of this transfection method is illustrated by describing its use for determining in vivo the role of a DNA element that regulates cytochrome P450 2B1 (CYP2B1) gene expression in response to xenobiotics. DNA was delivered to the liver of an anesthetized animal via DNA-coated gold microcarriers. The highest level of reporter gene expression was obtained about six hours posttransfection; however, at this time endogenous CYP2B1 mRNA is transiently induced by the anesthetic treatment. The optimal time for investigating expression of a reporter gene under the control of CYP2B1 regulatory elements was 24 h after transfection, by which time the inductive effect of the anesthetic had ceased. Reporter gene expression subsequently declined rapidly to a low level by 48 h. In the transfected liver the heterologous SV40 promoter was about eight-fold stronger than the minimal CYP2B1 promoter. However, when attached to the phenobarbital response element both promoters give the same fold-induction of reporter activity in response to phenobarbital.

Animals↗
PubMed1996

Biolistic-mediated gene expression in guinea pigs and cattle tissue in vivo.

Foreign genes were introduced and expressed in vivo in guinea pigs and cattle utilizing a new hand-held device based on high-pressure helium gas to accelerate DNA-coated microparticles. Guinea pigs were used to evaluate the physical parameters to introduce and express the exogenous DNA. The best conditions were applied to conduct bombardments in cattle. The results showed a high frequency of gene expression in all the bombarded cattle. This procedure could be used to study the immune responses in cattle and in a wide variety of animals through genetic immunization.

Animals↗
PubMed1995

Biolistic transformation of the obligate plant pathogenic fungus, Erysiphe graminis f.sp. hordei.

Particle gun acceleration appears to be a possible way to transform mycelium cells of obligate plant parasites growing on host surfaces. GUS expression was obtained in E. graminis f.sp. hordei cells after bombardment with the GUS gene under the control of the E. graminis f.sp. hordei β-tubulin promoter. Three heterologous promoters, onefrom Aspergillus nidulans and two from Cochliobolus heterostrophus, gave very low or no expression of GUS.

Ascomycota↗
PubMed1990

Transient foreign gene expression in chloroplasts of cultured tobacco cells after biolistic delivery of chloroplast vectors.

Expression of chloramphenicol acetyltransferase (cat) by suitable vectors in chloroplasts of cultured tobacco cells, delivered by high-velocity microprojectiles, is reported here. Several chloroplast expression vectors containing bacterial cat genes, placed under the control of either psbA promoter region from pea (pHD series) or rbcL promoter region from maize (pAC series) have been used in this study. In addition, chloroplast expression vectors containing replicon fragments from pea, tobacco, or maize chloroplast DNA have also been tested for efficiency and duration of cat expression in chloroplasts of tobacco cells. Cultured NT1 tobacco cells collected on filter papers were bombarded with tungsten particles coated with pUC118 (negative control), 35S-CAT (nuclear expression vector), pHD312 (repliconless chloroplast expression vector), and pHD407, pACp18, and pACp19 (chloroplast expression vectors with replicon). Sonic extracts of cells bombarded with pUC118 showed no detectable cat activity in the autoradiograms. Nuclear expression of cat reached two-thirds of the maximal 48 hr after bombardment and the maximal at 72 hr. Cells bombarded with chloroplast expression vectors showed a low level of expression until 48 hr of incubation. A dramatic increase in the expression of cat was observed 24 hr after the addition of fresh medium to cultured cells in samples bombarded with pHD407; the repliconless vector pHD312 showed about 50% of this maximal activity. The expression of nuclear cat and the repliconless chloroplast vector decreased after 72 hr, but a high level of chloroplast cat expression was maintained in cells bombarded with pHD407. Organelle-specific expression of cat in appropriate compartments was checked by introducing various plasmid constructions into tobacco protoplasts by electroporation. Although the nuclear expression vector 35S-CAT showed expression of cat, no activity was observed with any chloroplast vectors.

Cells, Cultured↗
PubMed2003

Accelerated dendritic development of rat cortical pyramidal cells and interneurons after biolistic transfection with BDNF and NT4/5.

Neurotrophins are candidate molecules for regulating dendritogenesis. We report here on dendritic growth of rat visual cortex pyramidal and interneurons overexpressing 'brain-derived neurotrophic factor' BDNF and 'neurotrophin 4/5' NT4/5. Neurons in organotypic cultures were transfected with plasmids encoding either 'enhanced green fluorescent protein' EGFP, BDNF/EGFP or NT4/5/EGFP either at the day of birth with analysis at 5 days in vitro, or at 5 days in vitro with analysis at 10 days in vitro. In pyramidal neurons, both TrkB ligands increased dendritic length and number of segments without affecting maximum branch order and number of primary dendrites. In the early time window, only infragranular neurons were responsive. Neurons in layers II/III became responsive to NT4/5, but not BDNF, during the later time window. BDNF and NT4/5 transfectants at 10 days in vitro had still significantly shorter dendrites than adult pyramidal neurons, suggesting a massive growth spurt after 10 days in vitro. However, segment numbers were already in the range of adult neurons. Although this suggested a role for BDNF, long-term activity-deprived, and thus BDNF-deprived, pyramidal cells developed a dendritic complexity not different from neurons in active cultures except for higher spine densities on neurons of layers II/III and VI. Neutralization of endogenous NT4/5 causes shorter and less branched dendrites at 10 days in vitro suggesting an essential role for NT4/5. Neutralization of BDNF had no effect. Transfected multipolar interneurons became identifiable during the second time window. Both TrkB ligands significantly increased number of segments and branch order towards the adult state with little effects on dendritic length. The results suggested that early in development BDNF and NT4/5 probably accelerate dendritogenesis in an autocrine fashion. In particular, branch formation was advanced towards the adult pattern in pyramidal cells and interneurons.

Animals↗
PubMed1999

[Studies of improving the frequency of wheat transformation by biolistic bombardment] [In Process Citation]

In the present study immature embryos of spring wheat Zhong-60634 were bombarded with gold particles coated with pAHC25 containing Bar and GUS genes. A total of 17 bialaphos-tolerant plants were obtained from 342 immature embryos which were first cultured for 3-4 days and then bombarded and selected. Integration of the introduced genes into the genome of transgenic wheat plants was shown by PCR and Southern analysis and transformation frequency was 0.88%. Furthermore, the present study showed that calli regeneration ability was improved while the embryo culture medium was supplemented with 2 mg/L dicamba instead of 2 mg/L 2,4-D. Meanwhile, in order to keep the calli differentiation ability calli should be transferred to differentiation medium supplemented with 3 mg/L bialaphos from calli selection medium with 5 mg/L bialaphos within a month.

Journal Article↗
PubMed1993

Chloroplast transformation in plants: polyethylene glycol (PEG) treatment of protoplasts is an alternative to biolistic delivery systems.

Nicotiana plumbaginifolia protoplasts were directly transformed by PEG treatment with a cloned 16S rRNA gene isolated from a double antibiotic-resistant Nicotiana tabacum plastid mutant. Putative plastid transformants were selected in cell culture by their spectinomycin resistance and identified by their unselected streptomycin resistance. Alternatively, cell lines were selected in the presence of both antibiotics. The cell line (and its regenerated plants) selected solely for spectinomycin resistance demonstrated an extensive segregation of streptomycin resistance in subsequent tests, while the double-selected line showed stable resistance for both antibiotics. The resistance markers were inherited maternally. In the putative plastid transformants the origin of the resistance mutations was identified by the absence of an AatII site, missing in the donor N. tabacum plastid gene (spectinomycin resistance site) but present in that of wild-type N. plumbaginifolia, and a sequence analysis of the particular nucleotide changes in both resistance sites. Restriction enzyme analysis of total plastid DNA (ptDNA), and the recloning and full sequencing of the fragment introduced, investigated in one of the plastid transformants, showed no DNA rearrangements accompanied with the integration process. Sequence analysis indicated a targeted, homologous integration of the DNA fragment introduced but an unexpectedly complete homology of the parental ptDNA sequences in this region prevented the location of borders. Although the frequency of plastid transformant colonies (2 x 10(-5)) should still be improved, this method for stable chloroplast DNA transformation is comparable with or more efficient than the particle bombardment techniques.

Base Sequence↗
PubMed2002

Augmentation of wound healing with translation initiation factor eIF4E mRNA.

BACKGROUND: Initiation of translation is the rate-limiting step in protein synthesis; eIF4E increases translational efficiency by facilitating ribosome scanning. eIF4E is present in cells in rate-limiting amounts; chronic overexpression of eIF4E causes cell transformation by upregulating growth-related proteins. Biolistic delivery of epidermal growth factor (EGF) increases wound healing; transiently increasing wound eIF4E levels with biolistic mRNA transmission may further augment wound healing without oncogenesis. PATIENTS AND METHODS: Midline fascial wounds were created in rats and biolistically treated with gold particles carrying mRNA encoding for hEGF with or without eIF4E prior to suture closure; control animals received blank bullets. The animals were sacrificed at 7 or 14 days for determination of peak wound bursting strength on a tensiometer. Results are expressed as means +/- standard deviation; statistics were via analysis of variance. RESULTS: [Table: see text]. CONCLUSIONS: Simultaneous biolistic delivery of EGF mRNA with eIF4E mRNA significantly increases wound breaking strength compared to that in control animals or treatment with EGF mRNA alone without risk of cellular transformation. Further studies of translational activation to augment wound healing are warranted.

Animals↗
PubMed2006

Performance studies of particle acceleration for transdermal drug delivery.

We have proposed a transdermal biolistic method to accelerate a powder formulation of drugs to penetrate human skin for the treatment of a range of diseases. One of the key issues for designing and evaluating transdermal biolistic system is ensuing that the powder drugs are delivered into the skin with a controllable velocity range and spatial distribution. The aerodynamics of supersonic nozzles and performance of the delivery system were initially studied, mainly analytically and experimentally. In this paper, computational fluid dynamics is utilized to characterize two existing prototype devices, in order to further investigate the transient gas and particle dynamics in their supersonic nozzles. To validate the implemented numerical approach, calculated pressure histories, two-dimensional flow structures and particle velocity distributions are made and compared with the reported experimental measurements. The key features of gas dynamics, gas-particle interaction and performance of the prototype transdermal biolistics are discussed and interpreted.

Administration, Cutaneous↗