ABSTRACT: Injection of endothelial progenitor cells (EPCs) into arteries for cell therapy is a promising field in regenerative medicine. However, adhesion of EPCs during capillary passage is restricted, and non-adhering cells are lost into circulation. Here we demonstrate that it is possible to achieve a three- to sevenfold higher rate of EPC adhesion to endothelium and extracellular matrix molecules after short-term activation with phorbol myristate acetate (PMA). In addition, differentiation and toxicity analyses of PMA activated EPCs showed no impact on cell differentiation and negligible impact on cell survival.
Artificial Cells Blood Substitutes and Biotechnology (formerly known as Artificial Cells Blood Substitutes and Immobilization Bi 02/2011; 39(4):214-22. · 0.94 Impact Factor
ABSTRACT: CD133+CD34+ hematopoietic stem cells (HSCs) have been shown to differentiate into cell types of nonhematopoietic lineage. It is unclear whether HSCs target and repair damaged musculoskeletal tissue. We aimed to analyze if HSCs are mobilized after musculoskeletal surgery to circulation, home to surgical wound fluid (SWF)-activated endothelium, and are chemoattracted by SWF under in vitro conditions.
Circulating HSC levels were measured at t = 3, 8, 24, 48 h postoperatively using fluorescence-activated cell sorting (FACS) and compared with preoperative levels (t = 0) and normal volunteers. For adhesion experiments, HSCs were incubated on SWF-activated human umbilical vein endothelial cells (HUVECs) and HSC/HUVEC ratios determined by FACS. Adhesion receptor expression on HSC (L-selectin, lymphocyte function-associated antigen 1 (LFA-1), very late antigen-4) and SWF-activated HUVECs (P-selectin, E-selectin, V-cell adhesion molecules (CAM), I-CAM) was determined and HSC adhesion measured again after blocking upregulated receptors. Using a modified Boyden chamber, HSC chemotaxis was analyzed for an SWF and cytokine-neutralized SWF (vascular endothelial growth factor (VEGF), stromal-derived factor-1, interleukin-8) gradient.
Circulating HSCs were significantly increased 8 h after surgery. Increasing HSC adhesion to HUVECs was shown for SWF isolated at any postoperative time point, and chemoattraction was significantly induced in an SWF gradient with SWF isolated 8 and 24 h postoperatively. Receptor and cytokine blockade experiments with monoclonal antibodies revealed decreased HSC adhesion to SWF-activated endothelium and showed lower chemotaxis after blocking the LFA-1-I-CAM-1 receptor axis (adhesion) and neutralizing VEGF-165 (chemotaxis).
Our data demonstrate that HSCs are mobilized after trauma, target to wound-associated endothelium via the LFA-1-I-CAM-1 axis, and are chemoattracted by VEGF-165 under in vitro conditions.
Langenbeck s Archives of Surgery 03/2010; 396(3):379-87. · 1.81 Impact Factor
ABSTRACT: An initial step in endothelium-derived hyperpolarizing factor-mediated responses is endothelial cell hyperpolarization. Here we address the mechanisms by which cytochrome P450 (CYP)-derived epoxyeicosatrienoic acids (EETs) contribute to this effect in native and cultured endothelial cells.
In native CYP2C-expressing endothelial cells, bradykinin elicited a Ca(2+) influx that was potentiated by the soluble epoxide hydrolase inhibitor, 1-adamantyl-3-cyclohexylurea (ACU), and attenuated by CYP inhibition. Similar effects were observed in cultured endothelial cells overexpressing CYP2C9, but not in CYP2C9-deficient cells, and were prevented by the EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid as well as by the cAMP antagonist, Rp-cAMPS. The effects on Ca(2+) were mirrored by prolongation of the bradykinin-induced hyperpolarization. Ruthenium red and the combination of charybdotoxin and apamin prevented the latter effect, suggesting that Trp channel activation increases Ca(2+) influx and prolongs the activation of Ca(2+)-dependent K(+) (K(Ca)) channels. Indeed, overexpression of CYP2C9 enhanced the agonist-induced translocation of a TrpC6-V5 fusion protein to caveolin-1-rich areas of the endothelial cell membrane, which was prevented by Rp-cAMPS and mimicked by 11,12-EET.
Elevated EET levels regulate Ca(2+) influx into endothelial cells and the subsequent activation of K(Ca) channels, via a cAMP/PKA-dependent mechanism that involves the intracellular translocation of Trp channels.
Arteriosclerosis Thrombosis and Vascular Biology 01/2008; 27(12):2612-8. · 6.37 Impact Factor