Abstract P027: Preconditioning Cardiosphere-Derived Cells by Hypoxia and Prolyl Hydroxylase Inhibitors to Induce HIF-Related Metabolic Changes and C-Kit Expression
Cardiosphere-derived cells (CDCs) decrease loss of cardiac function following infarction, but the potential of CDC therapy is limited due to the low retention rate and the time required to obtain sufficient cells for transplantation.
AIM: Here, we aimed to precondition CDCs by culturing under hypoxia, or with prolyl hydroxylase inhibitors (PHIs), to prepare the cells for the hypoxic environment within the infarcted heart.
METHODS: Rat heart explants were cultured under hypoxia (2% O2) or normoxia (21% O2) to generate explant-derived cells (EDC). EDCs were isolated, cultured to form cardiosphere and expanded into monolayer CDCs. At 80% confluency, normoxic CDCs were treated with dimethyloxalyl glycine (DMOG), Ethyl 2-(2,3-dihyroxybenzamido) (KKC226) and FG-2216 for 24 h.
RESULTS: Hypoxia, 1 mM DMOG, 0.5 mM KKC226 and 30 μM FG-2216 treated CDCs showed significantly increased HIF-1α protein expression (3.9-fold, 2.6-fold, 2.1-fold and 1.5-fold, respectively) and reduced oxygen consumption (81%, 32%, 47% and 35%, respectively), compared with normoxic CDCs. Hypoxia and PHI treatment increased protein levels of GLUT-1 (6.7-fold, 3.2-fold, 4.0-fold and 2.1-fold, respectively), resulting in significant increases in glucose uptake and lactate accumulation in the culture medium, compared with controls, commensurate with increased glycolytic metabolism after hypoxia or PHI treatment. Further, hypoxia and KKC226 increased c-Kit mRNA expression 5.1-fold and 1.5-fold, respectively. qRT-PCR confirmed increased CXCR-4 mRNA in all hypoxia and PHI-treated CDCs. EDC and CDC proliferation were 1.7-fold faster under hypoxia, compared with normoxia, but did not increase with PHI treatment.
CONCLUSION: Hypoxia and PHIs stabilized and activated HIF, which induced metabolic changes in CDCs, including GLUT-1 upregulation and a switch to anaerobic glycolytic metabolism. All preconditioned cells had reduced oxygen consumption and were better adapted to survive within an hypoxic infarct scar. HIF-induced upregulation of CXCR-4 may increase the homing of these cells to the infarcted myocardium. Finally, increased proliferation and expression of the cardiac stem cell marker, c-Kit, could decrease the time required for cell expansion prior to therapy by a week.
- © 2011 by American Heart Association, Inc.