title: Autologous human plasma in stem cell culture and cryopreservation in the creation of a tissue-engineered vascular graft.
abstract: OBJECTIVE
  Previous work demonstrated the effectiveness of autologous adipose-derived stem cells (ASCs) as endothelial cell (EC) substitutes in vascular tissue engineering. We further this work toward clinical translation by evaluating ASC function after (1) replacement of fetal bovine serum (FBS) with autologous human plasma (HP) in culture and (2) cryopreservation.
  
  
  METHODS
  Human ASCs and plasma, isolated from periumbilical fat and peripheral blood, respectively, were collected from the same donors. ASCs were differentiated in endothelial growth medium supplemented with FBS (2%) vs HP (2%). Proliferation was measured by growth curves and MTT assay. Endothelial differentiation was measured by quantitative polymerase chain reaction, assessment of acetylated low-density lipoprotein uptake, and cord formation after plating on Matrigel (BD Biosciences, San Jose, Calif). Similar studies were conducted before and after cryopreservation of ASCs and included assessment of cell retention on the luminal surface of a vascular graft.
  
  
  RESULTS
  ASCs expanded in HP-supplemented medium showed (1) similar proliferation to FBS-cultured ASCs, (2) consistent differentiation toward an EC lineage (increases in CD31, von Willebrand factor, and CD144 message; acetylated low-density lipoprotein uptake; and cord formation on Matrigel), and (3) retention on the luminal surface after seeding and subsequent flow conditioning. Cryopreservation did not significantly alter ASC viability, proliferation, acquisition of endothelial characteristics, or retention after seeding onto a vascular graft.
  
  
  CONCLUSIONS
  This study suggests that (1) replacement of FBS with autologous HP--a step necessary for the translation of this technology into human use--does not significantly impair proliferation or endothelial differentiation of ASCs used as EC substitutes and (2) ASCs are tolerant to cryopreservation in terms of maintaining EC characteristics and retention on a vascular graft.
authors.affiliation: Division of Vascular and Endovascular Surgery, Thomas Jefferson University, Philadelphia, Pa.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhang
authors.affiliation: Division of Vascular and Endovascular Surgery, Thomas Jefferson University, Philadelphia, Pa.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Policha
authors.affiliation: Division of Vascular and Endovascular Surgery, Thomas Jefferson University, Philadelphia, Pa.
authors.forename: Thomas
authors.initials: T
authors.lastname: Tulenko
authors.affiliation: Division of Vascular and Endovascular Surgery, Thomas Jefferson University, Philadelphia, Pa. Electronic address: paul.dimuzio@jefferson.edu.
authors.forename: Paul
authors.initials: P
authors.lastname: DiMuzio
mesh_terms: D000273:Adipose Tissue / Q000166:cytology*; D015415:Biomarkers / Q000378:metabolism; D001705:Bioprosthesis; D001807:Blood Vessel Prosthesis; D051929:CD146 Antigen / Q000235:genetics / Q000378:metabolism; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D019070:Cell Lineage; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D015925:Cryopreservation; D066026:Endothelial Progenitor Cells / Q000378:metabolism*; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism; D018919:Neovascularization, Physiologic; D010641:Phenotype; D010949:Plasma / Q000378:metabolism*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods*; D014841:von Willebrand Factor / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; D051929:CD146 Antigen; D008077:Lipoproteins, LDL; D019408:Platelet Endothelial Cell Adhesion Molecule-1; C032629:acetyl-LDL; D014841:von Willebrand Factor