Abstract
765. The Efficacy of Adenovirus Vector-Mediated Gene Transfer into Human Osteoblasts In Vitro and Osseous Tissue In Vivo
Molecular therapy, Vol.9(S1), pp.S290-S291
05/01/2004
DOI: 10.1016/j.ymthe.2004.06.680
Abstract
Adenovirus has been extensively used in experimental and clinical models as a vector for gene therapy and has been used successfully to infect a variety of bone cells. However, gene transfer to osteoblasts in unfractured bone has not been adequately investigated. The purpose of this study is to demonstrate the feasibility and efficacy of adenovirus-mediated gene transfer into bone cells in vitro and in vivo.HeLa, human embryonic palatal mesenchymal (HEPM), and primary human osteoblast cells from cancellous bone were transduced with adenovirus expressing green fluorescent protein (GFP) or murine erythropoietin (mEPO). The efficiency of adenoviral transduction was determined by calculating the percentage of GFP positive cells using FACS analysis. To determine the duration of expression, media from the cells expressing mEPO were collected over thirty days and gene expression was quantitatively determined by ELISA. For in vivo studies, mouse tibia was directly injected intra-bone marrow with adenovirus expressing either mEPO or β-galactosidase (lacZ). The hematocrit of the mice was measured once a week for three weeks.Based on the data from FACS analysis, our studies show that HeLa, HEPM and human osteoblasts can be transduced with adenovirus. The transduction efficiency of HeLa cells by adenovirus is known to be high. In comparison to HeLa cells, the transduction efficiency of both HEPM and human osteoblasts is low. HEPM had less than half the number of positive GFP cells than HeLa (40.1%), and human osteoblasts had less than one seventh (13.7%). The results of the ELISA assay for mEPO show that gene expression in human osteoblasts is more persistent than in HEPM cells. The concentration of mEPO expression in HEPM peaked by day 10 (517.7mIU/ML) and returned to near baseline by day 31. In contrast, the concentration of mEPO secreted by human osteoblasts stayed near its peak level (350mIU/mL) throughout the study period. The injection of mice tibia bone with adenovirus expressing mEPO resulted in a gradual increase in hematocrit from 48.6% to 83.9% at 3 weeks in all mice, while control Ad-lacZ injected mice remained at baseline level.Our results show that adenovirus-mediated gene transfer to osteoblasts can be a useful method for gene therapy in bone diseases.
Details
- Title: Subtitle
- 765. The Efficacy of Adenovirus Vector-Mediated Gene Transfer into Human Osteoblasts In Vitro and Osseous Tissue In Vivo
- Creators
- Keun LeeCharles SaltzmanPeter TaftMichael KlotzJoseph Zabner
- Resource Type
- Abstract
- Publication Details
- Molecular therapy, Vol.9(S1), pp.S290-S291
- Publisher
- Elsevier Limited
- DOI
- 10.1016/j.ymthe.2004.06.680
- ISSN
- 1525-0016
- eISSN
- 1525-0024
- Language
- English
- Date published
- 05/01/2004
- Academic Unit
- Pulmonary, Critical Care, and Occupational Medicine; Internal Medicine
- Record Identifier
- 9984363457302771
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