title: The effect of blocking angiogenesis on anterior cruciate ligament healing following stem cell transplantation.
abstract: Ruptured human anterior cruciate ligaments (ACL) contain vascular stem cells capable of enhancing the healing of tendon grafts. In the current study we explored the role that neo-angiogenesis plays in ACL healing. ACL-derived CD34+ cells were isolated via Fluorescence Activated Cell Sorting (FACS) from the rupture sites of human ACLs. The cells were then virally transduced to express either vascular endothelial growth factor (VEGF) or soluble FLT-1 (sFLT-1), which is an antagonist of VEGF. We established five groups: CD34+VEGF(100%), where 100% of the cells were transduced with VEGF, CD34+VEGF(25%), where only 25% of the cells were transduced with VEGF, CD34+, CD34+sFLT-1, and a No cells group. The CD34+sFLT1 group had a significant reduction in biomechanical strength compared to the CD34+ group at 4 and 8 weeks; whereas the biomechanical strength of the CD34+VEGF(25%) group was significantly greater than the CD34+ group at week 4; however, no difference was observed by week 8. Immunohistochemical staining demonstrated a significantly lower number of isolectin B4 and hCD31 positive cells, markers associated with angiogenesis, in the CD34+sFLT1 group, and a higher number of isolectin B4 and hCD31 positive cells in the CD34+VEGF(100%) and CD34+VEGF(25%) groups compared to the CD34+ group. Graft maturation was significantly delayed in the CD34+sFLT1 group and accelerated in the CD34+VEGF(25%) group compared to the CD34+ group. In conclusion, blocking VEGF reduced angiogenesis, graft maturation and biomechanical strength following ACL reconstruction. Native expression of VEGF by the CD34+ cells improved tendon graft maturation and biomechanical strength; however, over-expression of VEGF impeded improvements in biomechanical strength.
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA; Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors.forename: Koji
authors.initials: K
authors.lastname: Takayama
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA; Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Kawakami
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA; Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Mifune
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA; Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA; Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Matsumoto
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Tang
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors.forename: James H
authors.initials: JH
authors.lastname: Cummins
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA; Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors.forename: Nick
authors.initials: N
authors.lastname: Greco
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Kuroda
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Kurosaka
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors.forename: Bing
authors.initials: B
authors.lastname: Wang
authors.affiliation: Stem Cell Research Center, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors.forename: Freddie H
authors.initials: FH
authors.lastname: Fu
authors.affiliation: Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA. Electronic address: jhuard@pitt.edu.
authors.forename: Johnny
authors.initials: J
authors.lastname: Huard
mesh_terms: D000328:Adult; D053687:Adult Stem Cells / Q000166:cytology / Q000378:metabolism / Q000637:transplantation*; D000818:Animals; D016118:Anterior Cruciate Ligament / Q000098:blood supply* / Q000166:cytology / Q000473:pathology; D000070598:Anterior Cruciate Ligament Injuries; D059549:Anterior Cruciate Ligament Reconstruction / Q000379:methods*; D018952:Antigens, CD34 / Q000032:analysis; D001696:Biomechanical Phenomena; D002478:Cells, Cultured; D015870:Gene Expression; D006801:Humans; D018919:Neovascularization, Physiologic; D051381:Rats; D033581:Stem Cell Transplantation / Q000379:methods*; D023822:Tissue Engineering; D014161:Transduction, Genetic; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics / Q000378:metabolism; D014945:Wound Healing
chemical_list: D018952:Antigens, CD34; D042461:Vascular Endothelial Growth Factor A; C501162:FLT1 protein, human; D040281:Vascular Endothelial Growth Factor Receptor-1
keywords: ACL (anterior cruciate ligament) reconstruction; Angiogenesis; Cell sheet; Stem cells