Open AccessIn Vivo, Cardiac-Specific Knockdown of a Target Protein, Malic Enzyme- 1, in Rat via Adenoviral Delivery of DNA for Non-Native miRNA
Author(s) -
J. Michael O’Donnell,
Asha Kalichira,
Jian Bi,
E. Douglas Lewandowski
Publication year - 2012
Publication title -
current gene therapy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.704
H-Index - 71
eISSN - 1875-5631
pISSN - 1566-5232
DOI - 10.2174/156652312803519760
Subject(s) - in vivo , gene knockdown , microbiology and biotechnology , gene silencing , messenger rna , biology , chemistry , biochemistry , gene
This study examines the feasibility of using the adenoviral delivery of DNA for a non-native microRNA to suppress expression of a target protein (cytosolic NADP(+)-dependent malic-enzyme 1, ME1) in whole heart in vivo, via an isolated-heart coronary perfusion approach. Complementary DNA constructs for ME1 microRNA were inserted into adenoviral vectors. Viral gene transfer to neonatal rat cardiomyocytes yielded 65% suppression of ME1 protein. This viral package was delivered to rat hearts in vivo (Adv.miR_ME1, 10(13) vp/ml PBS) via coronary perfusion, using a cardiac-specific isolation technique. ME1 mRNA was reduced by 73% at 2-6 days post-surgery in heart receiving the Adv.miR_ME1. Importantly, ME1 protein was reduced by 66% (p < 0.0002) at 5-6 days relative to sham-operated control hearts. Non-target protein expression for GAPDH, calsequestrin, and mitochondrial malic enzyme, ME3, were all unchanged. The non-target isoform, ME2, was unchanged at 2-5 days and reduced at day 6. This new approach demonstrates for the first time significant and acute silencing of target RNA translation and protein content in whole heart, in vivo, via non-native microRNA expression.