title: In vivo structural and cellular remodeling of engineered bone-ligament-bone constructs used for anterior cruciate ligament reconstruction in sheep.
abstract: Anterior cruciate ligament (ACL) ruptures rank among the most prevalent and costly sports-related injuries. Current tendon grafts used for ACL reconstruction are limited by suboptimal biomechanical properties. We have addressed these issues by engineering multiphasic bone-ligament-bone (BLB) constructs that develop structural and mechanical properties similar to native ACL. The purpose of this study was to examine the acute remodeling process that occurs as the BLB grafts advance toward the adult ligament phenotype in vivo. Thus, we implanted BLB constructs fabricated from male cells into female host sheep and allowed 3, 7, 14, or 28 days (n = 4 at each time point) for recovery. To address whether or not graft-derived cells were even necessary, a subset of BLB constructs (n = 3) were acellularized, implanted, and allowed 28 days for recovery. At each recovery time point, the following histological analyses were performed: picrosirius red staining to assess collagen alignment and immunohistochemistry to assess both graft development and host immune response. Polymerase chain reaction (PCR) analysis, performed on every explanted BLB, was used to detect the presence of graft-derived male cells remaining in the constructs and/or migration into surrounding host tissue. The analysis of the PCR and histology samples revealed a rapid migration of host-derived macrophages and neutrophils into the graft at 3 days, followed by increased collagen density and alignment, vascularization, innervation, and near complete repopulation of the graft with host cells within 28 days. This study provides a greater understanding of the processes of ligament regeneration in our BLB constructs as they remodel toward the adult ligament phenotype.
authors.affiliation: a Department of Molecular & Integrative Physiology , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Shelby E
authors.initials: SE
authors.lastname: Florida
authors.affiliation: a Department of Molecular & Integrative Physiology , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Keith W
authors.initials: KW
authors.lastname: VanDusen
authors.affiliation: a Department of Molecular & Integrative Physiology , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Vasudevan D
authors.initials: VD
authors.lastname: Mahalingam
authors.affiliation: a Department of Molecular & Integrative Physiology , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Aleesa J
authors.initials: AJ
authors.lastname: Schlientz
authors.affiliation: b Medsport Sports Medicine Program, Orthopaedic Surgery , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Edward M
authors.initials: EM
authors.lastname: Wojtys
authors.affiliation: c Department of Cell & Developmental Biology University of Michigan , Ann Arbor , MI , USA.|d Department of Internal Medicine , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Deneen M
authors.initials: DM
authors.lastname: Wellik
authors.affiliation: a Department of Molecular & Integrative Physiology , University of Michigan , Ann Arbor , MI , USA.|e Department of Biomedical Engineering , University of Michigan , Ann Arbor , MI , USA.
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Larkin
mesh_terms: D000818:Animals; D016118:Anterior Cruciate Ligament / Q000601:surgery*; D059549:Anterior Cruciate Ligament Reconstruction / Q000379:methods*; D016723:Bone Remodeling; D001842:Bone and Bones / Q000473:pathology*; D053148:Caspase 3 / Q000378:metabolism; D002452:Cell Count; D016923:Cell Death; D003094:Collagen / Q000378:metabolism; D005260:Female; D007150:Immunohistochemistry; D007382:Intermediate Filaments / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D008297:Male; D009504:Neutrophils / Q000378:metabolism; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012756:Sheep; D013194:Staining and Labeling; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds / Q000737:chemistry*; D014998:Y Chromosome / Q000235:genetics
chemical_list: D019408:Platelet Endothelial Cell Adhesion Molecule-1; D003094:Collagen; D053148:Caspase 3
keywords: ACL reconstruction; remodeling; scaffoldless; tissue engineering