Targeting the respiratory muscles of fetal sheep for prenatal gene therapy for Duchenne muscular dystrophy.
OBJECTIVE
Duchenne muscular dystrophy (DMD) is a lethal degenerative muscular disease. Fetal gene therapy may correct the primary genetic defect. Our aim was to achieve expression of a reporter gene in the respiratory muscles of early gestation fetal sheep.


METHODS
An adenovirus vector containing the beta-galactosidase reporter gene (AdRSVbetagal) was injected into the thoracic musculature (n = 3) and pleural cavity (n = 6) of fetal sheep (61-67 days' gestation) under ultrasound guidance. Tissues were harvested after 48 hours and site and intensity of beta-galactosidase expression were assessed.


RESULTS
Limited transgene expression observed after a single injection was improved by multiple injections, but remained localized. Ultrasound-guided creation of a hydrothorax led to an increase in the intensity of beta-galactosidase expression (ELISA). X-gal staining and immunohistochemistry showed that vector spread was confined to the innermost intercostal musculature.


CONCLUSIONS
Ultrasound-guided injection can deliver gene therapy vectors to the fetal pleural cavity and achieve transduction of the respiratory muscles.
Weisz
Boaz
B
Department of Obstetrics and Gynaecology, University College London, United Kingdom. b.weisz@ucl.ac.uk
David
Anna L
AL
Gregory
Lisa G
LG
Perocheau
Dany
D
Ruthe
Ali
A
Waddington
Simon N
SN
Themis
Mike
M
Cook
Terry
T
Coutelle
Charles
C
Rodeck
Charles H
CH
Peebles
Donald M
DM
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D005260:Female; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000008:administration & dosage*; D006876:Hydrothorax; D007150:Immunohistochemistry; D007273:Injections, Intramuscular; D020388:Muscular Dystrophy, Duchenne / Q000196:embryology / Q000628:therapy*; D011247:Pregnancy; D012132:Respiratory Muscles / Q000378:metabolism*; D012756:Sheep; D019076:Transgenes / Q000502:physiology; D016216:Ultrasonography, Prenatal; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001616:beta-Galactosidase