Novel Cell-Free Strategy for Therapeutic Angiogenesis: In Vitro Generated Conditioned Medium Can Replace Progenitor Cell Transplantation

• Published in: PloS one Vol. 4; no. 5; p. e5643
• Main Authors: Di Santo, Stefano, Yang, Zijiang, Wyler von Ballmoos, Moritz, Voelzmann, Jan, Diehm, Nicolas, Baumgartner, Iris, Kalka, Christoph
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 21.05.2009; Public Library of Science (PLoS)
• Subjects: Adult; Angiogenesis; Animals; Aorta; Apoptosis; Blood flow; Blood vessels; Bone marrow; Bone Marrow Cells - cytology; Bone marrow transplantation; Cardiology; Cardiovascular diseases; Cardiovascular Disorders/Peripheral Vascular Disease; Cardiovascular Disorders/Vascular Biology; CD34 antigen; CD45 antigen; Cell culture; Cell Hypoxia; Cell Line; Cell Survival; Cell-Free System; Cells (biology); Comparative analysis; Conditioning; Culture Media, Conditioned; Developmental Biology/Stem Cells; Endothelial cells; Endothelial Cells - cytology; Endothelial Cells - metabolism; Endothelium; Health aspects; Heart attacks; Heart diseases; Hematopoietic Stem Cell Mobilization; Hindlimb - blood supply; Hindlimb - pathology; Hindlimb - physiopathology; Hospitals; Human subjects; Humans; Hypoxia; Injection; Intercellular Signaling Peptides and Proteins - metabolism; Ischemia; Ischemia - chemically induced; Ischemia - pathology; Ischemia - physiopathology; Leukocytes (mononuclear); Maturation; Medicine; Muscles; Muscles - physiology; Neovascularization, Physiologic; Paracrine signalling; Perfusion; Permeability; Progenitor cells; Rats; Rats, Nude; Recovery of function; Rodents; Secretion; Stem Cell Transplantation; Stem cells; Stem Cells - cytology; Therapy; Transplantation; Vascular endothelial growth factor; Viability
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0005643

Current evidence suggests that endothelial progenitor cells (EPC) contribute to ischemic tissue repair by both secretion of paracrine factors and incorporation into developing vessels. We tested the hypothesis that cell-free administration of paracrine factors secreted by cultured EPC may achieve an angiogenic effect equivalent to cell therapy. EPC-derived conditioned medium (EPC-CM) was obtained from culture expanded EPC subjected to 72 hours of hypoxia. In vitro, EPC-CM significantly inhibited apoptosis of mature endothelial cells and promoted angiogenesis in a rat aortic ring assay. The therapeutic potential of EPC-CM as compared to EPC transplantation was evaluated in a rat model of chronic hindlimb ischemia. Serial intramuscular injections of EPC-CM and EPC both significantly increased hindlimb blood flow assessed by laser Doppler (81.2+/-2.9% and 83.7+/-3.0% vs. 53.5+/-2.4% of normal, P<0.01) and improved muscle performance. A significantly increased capillary density (1.62+/-0.03 and 1.68+/-0.05/muscle fiber, P<0.05), enhanced vascular maturation (8.6+/-0.3 and 8.1+/-0.4/HPF, P<0.05) and muscle viability corroborated the findings of improved hindlimb perfusion and muscle function. Furthermore, EPC-CM transplantation stimulated the mobilization of bone marrow (BM)-derived EPC compared to control (678.7+/-44.1 vs. 340.0+/-29.1 CD34(+)/CD45(-) cells/1x10(5) mononuclear cells, P<0.05) and their recruitment to the ischemic muscles (5.9+/-0.7 vs. 2.6+/-0.4 CD34(+) cells/HPF, P<0.001) 3 days after the last injection. Intramuscular injection of EPC-CM is as effective as cell transplantation for promoting tissue revascularization and functional recovery. Owing to the technical and practical limitations of cell therapy, cell free conditioned media may represent a potent alternative for therapeutic angiogenesis in ischemic cardiovascular diseases.