Expression of calcitonin gene-related peptide following adenovirus-mediated gene transfer to cos-1 cells


Calcitonin gene-related peptide (CGRP) is a potent dilator in the cerebral circulation. Vasospasm following subarachnoid hemorrhage is associated with depletion of CGRP in cerebral perivascular neurons. Gene transfer of CGRP using an adenoviral vector may potentially restore CGRP levels in vessels, and thereby attenuate vasospasm. We have prepared a recombinant adenoviral vector containing a transgene encoding prepro-CGRP, driven by an RSV promoter (AdRSVCGRP). We determined whether AdRSVCGRP would release CGRP into media of cultured cells after transfection. COS-1 cells were incubated with vehicle, AdRSVCGRP, or AdRSVLacZ (control virus) for 1 hr. Following incubation with AdRSVCGRP, the cells were incubated for 48 hrs to allow transgene expression. Transgene expression was evaluated by radioimmunoassay. CGRP in medium was 0.05±0.03 ng/ dish (mean±SE) after treatment with vehicle and 0.0310.03 after transfection with AdRSVLacZ (100 pfu/cell). In contrast, when COS-1 cells were transfected with AdRSVCGRP, CGRP in medium was 8.1±5.5 (10 pfu/ cell), 94±35 (100 pfu/ cell), and 121±54 (500 pfu/cell) (p<0.05 vs. vehicle). These data provide evidence that CGRP can be expressed and released from COS-1 cells in vitro following adenovirus-mediated gene transfer of prepro-CGRP.

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FASEB Journal