Abstract 18401: P53 Silenced, Apoptosis Resistant, Endothelial Progenitor Stem Cells (EPC) Improve Collateral Circulation Post Femoral Artery Occlusion in Diabetic Mice
Literature shows that EPCs contribute to increased collateral vessel formation following vaso-occlusion. However diabetes reduces EPC number and reduces collateral vessel formation. We cultured human EPCs and exposed them to 5.5 mM (equivalent to 99mg%) and 20mM (equivalent to 360mg%, HG) glucose, which resulted in significant EPC death within 48hrs. We noted HG exposure was associated with up-regulation of P53 and its downstream genes such as P21, PUMA and Caspase-3. We hypothesized that EPC reduction in hyperglycemia is secondary to up-regulation of P53. We obtained mouse EPCs from P53 null mice and observed that p53 KO EPCs are more resistant to cell death in HG. P53 null EPCs evolved into mature mouse EC (MEC) and retained endothelial properties. Next, we used Lenti and Adenovirus mediated si-RNA methods to silence P53 (long and short term respectively) in human EPCs and P53 silenced EPc showed better survival in HG. We developed femoral artery occlusion model to mimic peripheral vascular disease in diabetic animals. We used streptozotocin induced type 1 diabetic or Leptin resistant type 2 diabetic mouse model. We delivered saline, P53 WT and P53 null mouse EPCs intra-muscularly around the femoral occlusion in these mouse models (n=7 in each group) and counted CD31+ve number of capillaries in a 20x microscopic field. Highest number was noted in the group that received P53null EPCs. We also quantified increased vessel formation by laser doppler assay of the ischemic and contralateral hind limbs upto 2 weeks and conducted qRT-PCR and western blot of key endothelial genes such as eNOS, p-eNOS, VEGF-A, PECAM-1 and vWF from both quadriceps in all groups of mice.
Summary: Transplantation of P53 null EPCs in hyperglycemic mice with femoral occlusion demonstrate better collateral vessel formation post femoral artery occlusion setting compared to WT-EPC transplanted group. Preliminary results from transplantation of p53 silenced human CD34+ cells also indicate increased vessel formation compared to non p53 silenced hCD34+ cell in NOD-SCID diabetic mice.
Conclusion: This finding indicates towards possible therapeutic benefit in transplanting P53 silenced human EPC or CD34+ cells to prevent peripheral vascular disease in patients with diabetes.
- © 2012 by American Heart Association, Inc.