Transient extremity ischemia augments CD34+ progenitor cell availability.

Peripheral blood is an easily accessed source for stem cell production; however, the number of cells produced is relatively low. We hypothesized that ischemic preconditioning may serve as a safe method to increase the number of CD34+ cells that can be harvested and cultured in a short period. This study was conducted to test this hypothesis by examining the safety and efficacy of brief, transient ischemia of the lower limbs to augment the number of cells that can be produced from blood of healthy volunteers. Following induction of ischemia, blood samples were withdrawn at baseline, 30 min, 12 h and 24 h. The number of progenitor cells was determined by flow cytometry after the harvested cells were cultured for 5 days. We also analyzed the blood samples to determine IL-8 and VEGF concentrations. No serious adverse events were observed. The total number of cells increased from 0.46 ± 0.1 × 10(6) cells/ml in the pretreatment blood samples to 0.7 ± 0.1 × 10(6) cells/ml in blood taken 12 h after the conclusion of transient ischemia, p = 0.0029. The number of CD34+ cells increased from 4.23 ± 0.8 × 10(4) cells/ml in the pretreatment samples to 7.17 ± 1.34 × 10(4) cells/ml in blood taken 12 h after ischemia, p = 0.0001. The harvested stem cells maintained their ability to construct tubular structures. The augmentation in the number of CD34+ cells was positively correlated with the increase of IL-8, but not with VEGF concentrations. Ischemic preconditioning is a safe and effective technique to increase the availability of stem cells for therapeutic purposes.

Human angiogenic cell precursors restore function in the infarcted rat heart: a comparison of cell delivery routes.

BACKGROUND: We recently isolated angiogenic cell precursors (ACPs) from human blood, which can induce angiogenesis in vitro. AIMS: In the present study, we used a nude rat model of ischaemic cardiomyopathy to compare the efficacy of intramyocardial and intracoronary ACP implantation, and to evaluate effects on cardiac function, scar size and angiogenesis. METHODS AND RESULTS: Adult nude rats underwent coronary artery ligation. Six days later, ACPs (characterized in vitro prior to implantation) or culture media were injected directly into the ischaemic myocardial region or into the coronary artery via the aorta. Cardiac function was measured by echocardiography prior to and at 2 and 4 weeks after implantation. Scar morphology, cell engraftment, and myocardial angiogenesis were evaluated at 4 weeks. Two and four weeks after implantation, cardiac function declined in both of the control groups but improved in both the intramyocardial and intracoronary ACP groups. Significant reductions in myocardial scar area were only observed in the intramyocardial ACP group, while increases in blood vessel density, which were observed in all ACP recipients, were greatest in the intracoronary ACP group. CONCLUSIONS: Human ACPs, delivered via intramyocardial or intracoronary injection, engrafted into damaged cardiac tissue and improved cardiac function within 4 weeks through effects on scar morphology and blood vessel formation.

Isolation of an adult blood-derived progenitor cell population capable of differentiation into angiogenic, myocardial and neural lineages.

Blood-derived adult stem cells were previously considered impractical for therapeutic use because of their small numbers. This report describes the isolation of a novel human cell population derived from the peripheral blood, termed synergetic cell population (SCP), and defined by the expression of CD31Bright, CD34+, CD45-/Dim and CD34Bright, but not lineage-specific features. The SCP was capable of differentiating into a variety of cell lineages upon exposure to defined culture conditions. The resulting cells exhibited morphological, immunocytochemical and functional characteristics of angiogenic, neural or myocardial lineages. Angiogenic cell precursors (ACPs) expressed CD34, CD133, KDR, Tie-2, CD144, von Willebrand factor, CD31Bright, concomitant binding of Ulex-Lectin and uptake of acetylated low density lipoprotein (Ac-LDL), secreted interleukin-8, vascular endothelial growth factor and angiogenin and formed tube-like structures in vitro. The majority of CD31Bright ACP cells demonstrated Ac-LDL uptake. Neural cell precursors (NCPs) expressed the neuronal markers Nestin, betaIII-Tubulin, and Neu-N, the glial markers GFAP and O4, and responded to neurotransmitter stimulation. Myocardial cell precursors (MCPs) expressed Desmin, cardiac Troponin and Connexin 43. In conclusion, the simple and rapid method of SCP generation and the resulting considerable quantities of lineage-specific precursor cells makes it a potential source of autologous treatment for a variety of diseases.

Clinical experience using incubated autologous macrophages as a treatment for complete spinal cord injury: phase I study results.

OBJECT: A Phase I, open-label nonrandomized study was conducted to assess the safety and tolerability of incubated autologous macrophages administered to patients with acute complete spinal cord injury (SCI). METHODS: This therapy was first tested in rat models of spinal cord transection and contusion, in which it was shown to promote motor recovery. The procedure developed for clinical use consists of isolating monocytes from patient blood and incubating them ex vivo with autologous dermis. The resulting incubated autologous macrophages were injected into the patient's spinal cord immediately caudal to the lesion within 14 days of injury. Patients underwent preoperative and follow-up neurological assessment (American Spinal Injury Association [ASIA] standards), electrophysiological monitoring (motor evoked and/or somatosensory evoked potentials), magnetic resonance imaging, and safety monitoring. Before macrophage administration, complete neurological functional loss (ASIA Grade A) was confirmed in all patients. Of the eight patients in the study, three recovered clinically significant neurological motor and sensory function (ASIA Grade C status). During the study period, some adverse events were encountered, the most serious of which involved two cases of pulmonary embolism and one case of osteomyelitis that were treated and resolved without further complication. These and other adverse events appear to be similar to those encountered in other spinal cord-injured patients and are not considered a consequence of the experimental therapy. CONCLUSIONS: It is concluded that incubated autologous macrophage cell therapy is well tolerated in patients with acute SCI. Further clinical evaluation is warranted.

Features of skin-coincubated macrophages that promote recovery from spinal cord injury.

Uncontrolled inflammation is considered to exacerbate the neuronal loss that follows spinal cord trauma. However, controlled inflammation response appears to be beneficial. Skin-coincubated macrophages injected into contused spinal cord of rats resulted in improved motor recovery and reduced spinal cyst formation. The macrophages express elevated levels of cell-surface molecules CD80, CD86, CD54 and MHC-II, markers characteristic of antigen presenting cells (APCs). Additionally, skin-coincubation elevates secretion of interleukin-1 beta (IL-1 beta) and Brain-Derived Neurotrophic Factor (BDNF), and reduces secretion of tumor necrosis factor alpha (TNF-alpha). We propose that macrophages activated by skin-coincubation bolster neuroprotective immune activity in the spinal cord, making the environment less cytotoxic and less hostile to axonal regeneration.