Abstract 2713: Molecular Imaging of Cardiac Stem Cell Mediated Angiogenic Gene Therapy
Introduction: Cardiac stem cell therapy remains hampered by acute donor cell death post transplant and the lack of reliable methods for tracking cell survival in vivo. We hypothesize that cells capable of inducible vascular endothelial growth factor 165 (VEGF165) expression may better tolerate ischemia resulting in improved cell survival.
Methods: Mouse embryonic stem cell (mESC) line carrying a double fusion (DF) reporter gene with firefly luciferase and green fluorescence protein (Fluc-GFP) was transfected with an inducible tetracycline (Tet)-on bi-directional promoter driving VEGF165 and renilla luciferase (Rluc). Doxycycline dosage for gene induction was optimized by ELISA and enzyme assays. After transplantation into SCID mice (n=8), cell survival was monitored by bioluminescence imaging using D-Luciferin reporter probe (125 mg/kg).
Results: Expression of DF and Tet-on systems did not affect mESC viability (>95%). At day 14, the differentiation efficiency of embryoid bodies into beating cells was consistently ~5±2%. After Percoll separation, mESC derived cardiac cells stained positive for troponin, MEF, and MHC markers. Addition of doxycycline (500 ng/ml) induced Tet-on expression of VEGF165 and Rluc by >5-fold compared to baseline (P<0.05). Expression of the two genes were strongly correlated (r2=0.85). Longitudinal survival within living mice was imaged over 4 weeks and confirmed by postmortem histology.
Conclusions: This is the first study to demonstrate imaging of stem cell mediated gene therapy targeting cardiovascular disease. With further validation, this platform may have broad applications for current basic research and future clinical studies.