Heart, Lung and Circulation

Assessment of Optimal Cell Therapy for the Angiogenesis Response in a Murine Hindlimb Ischaemia Model using CD34+ cells and Endothelial Progenitor Cells

      Purpose: Therapeutic angiogenesis using stem/progenitor cells has been the focus of recent research. CD34+ cells and endothelial progenitor cells (EPCs) have been found to promote angiogenesis. This study aimed to assess the angiogenic potential of CD34+ cells and EPCs using different modes of delivery in vivo.
      Methods: Human EPCs were isolated from cell cultures and CD34+ cells were purified from buffy coat using microbeads. Unilateral hindlimb ischaemia was introduced on BalbC nu/nu mice. At 24 hours post-surgery 2 × 105 CD34+ cells, EPCs, or PBS control were injected intravascularly (IV) to the tail vein, or intramuscularly (IM) into adductor muscle. Laser Doppler perfusion imaging (LDPI) was used to assess flow recovery. Adductor muscle was assessed for capillary density.
      Results: By day 10 post-surgery mice injected IM showed better recovery in LDPI, compared with those mice injected IV (LDPI 0.30 ± 0.06 vs. 0.15 ± 0.05; P < 0.05). This was true for CD34+ cells, EPCs and PBS. Mice receiving CD34+ cells IM recovered better from day 10 to 21 when compared to mice injected IM with EPC or PBS (day 21 LDPI 0.45 ± 0.04, 0.35 ± 0.05 and 0.28 ± 0.05, respectively: P < 0.05). Immunohistochemical staining of adductor muscles revealed capillary density was highest in mice receiving CD34+ cells IM, compared with those mice receiving EPC IM, PBS IM, CD34+ IV, EPC IV, and PBS IV (155 ± 9%, 140 ± 5%, 135 ± 7%, 115 ± 10%, 110 ± 10% vs. 100% for PBS IV, respectively; P < 0.05).
      Conclusions: This study provides evidence that direct IM injection of CD34+ cells into the ischaemic hindlimb delivers the best angiogenesis outcome among various cell therapy techniques investigated in this study.