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

Traditionally, skeletal muscle and liver are the preferred target organs for gene transfer to supply a transgene product into the systemic circulation. In this respect, adipose tissue presents a number of attractive features. However, adipose tissue transduction in vivo has not been feasible by conventional methods. To solve this issue, we tested the utility of excipients in adeno-associated virus (AAV) vector-mediated gene transfer and found that Pluronics are suitable for this purpose. In a histological analysis of adipose tissue in db/db mice, Pluronic F88 showed the greatest augmentative effect on beta-galactosidase expression in combination with the AAV1 vector. When the vector encoding mouse erythropoietin (Epo) was used in the same manner, increased plasma Epo concentrations were observed (230 +/- 80 versus 58 +/- 14 mU/ml). Moreover, the plasma Epo concentration returned to the normal level after the surgical removal of transduced adipose tissue. No damage was observed in the transduced tissue. Our results indicate that the proposed method is safe and efficient for gene transfer into adipose tissues, thus providing an alternative for supplemental gene therapy.
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PMID:Adipose tissue as a novel target for in vivo gene transfer by adeno-associated viral vectors. 1697 60

phi C31 integrase has emerged as a potent tool for achieving long-term gene expression in different tissues. The present study aimed at optimizing elements of phi C31 integrase system for alveolar type II cells. Luciferase and beta-galactosidase activities were measured at different time points post transfection. 5-Aza-2'deoxycytidine (AZA) and trichostatin A (TSA) were used to inhibit DNA methyltransferase and histone deacetylase complex (HDAC) respectively. In A549 cells, expression of the integrase using a CMV promoter resulted in highest integrase activity, whereas in MLE12 cells, both CAG and CMV promoter were equally effective. Effect of polyA site was observed only in A549 cells, where replacement of SV40 polyA by bovine growth hormone (BGH) polyA site resulted in an enhancement of integrase activity. Addition of a C-terminal SV40 nuclear localization signal (NLS) did not result in any significant increase in integrase activity. Long-term expression studies with AZA and TSA, provided evidence for post-integrative gene silencing. In MLE12 cells, both DNA methylases and HDACs played a significant role in silencing, whereas in A549 cells, it could be attributed majorly to HDAC activity. Donor plasmids comprising cellular promoters ubiquitin B (UBB), ubiquitin C (UCC) and elongation factor 1 alpha (EF1 alpha) in an improved backbone prevented post-integrative gene silencing. In contrast to A549 and MLE12 cells, no silencing could be observed in human bronchial epithelial cells, BEAS-2B. Donor plasmid coding for murine erythropoietin under the EF1 alpha promoter when combined with phi C31 integrase resulted in higher long-term erythropoietin expression and subsequently higher hematocrit levels in mice after intravenous delivery to the lungs. These results provide evidence for cell specific post integrative gene silencing with C31 integrase and demonstrate the pivotal role of donor plasmid in long-term expression attained with this system.
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PMID:Optimization of Streptomyces bacteriophage phi C31 integrase system to prevent post integrative gene silencing in pulmonary type II cells. 1974 1


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