Distinctive collagen maturation process in fibroblasts derived from rabbit anterior cruciate ligament, medial collateral ligament, and patellar tendon in vitro.
OBJECTIVE
Differences in the tissue-specific collagen maturation process between tendon and ligament are still unknown. Collagen cross-link formation is crucial for the collagen maturation process. The aim of this study is to examine collagen maturation processes of anterior cruciate ligament (ACL), medial collateral ligament (MCL), and patellar tendon (PT) in vitro, in order to determine the optimal cell source for tissue engineering of ligament.


METHODS
Cells derived from the ACL, MCL, and PT of New Zealand white rabbits were isolated. Each cell type was cultured for up to 4 weeks after reaching confluence. Cell-matrix layers were evaluated and compared for their morphology, collagen cross-links, and gene expression levels of lysine hydroxylase 1 and 2, lysyl oxidase (LOX), tenomodulin, collagen1A1 (Col1A1), and collagen3A1 (Col3A1).


RESULTS
Transmission electron microscopy photomicrographs verified that collagen fibrils were secreted from all three types of fibroblasts. A higher ratio of dihydroxylysinonorleucine/hydroxylysinonorleucine was evident in the ligament compared to the tendon, which was consistent with lysine hydroxylase 2/lysine hydroxylase 1 gene expression. The gene expression of LOX, which regulates the total amount of enzymatic cross-linking, and the gene expression levels of Col1A1 and Col3A1 were higher in the ACL matrix than in the MCL and PT matrices.


CONCLUSIONS
ACL, MCL, and PT cells have distinct collagen maturation processes at the cellular level. In addition, the collagen maturation of ACL cells is not necessarily inferior to that of MCL and PT cells in that all three cell types have a good ability to synthesize collagen and induce collagen maturation. This bioactivity of ACL cells in terms of ligament-specific mature collagen induction can be applied to tissue-engineered ACL reconstruction or remnant preserving procedure with ACL reconstruction.
Department of Orthopaedic Surgery, The Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo, 105-8461, Japan. soki@jikei.ac.jp.
Soki
S
Kato
Department of Orthopaedic Surgery, The Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo, 105-8461, Japan.
Mitsuru
M
Saito
Department of Orthopaedic Surgery, The Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo, 105-8461, Japan.
Hiroki
H
Funasaki
Department of Orthopaedic Surgery, The Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo, 105-8461, Japan.
Keishi
K
Marumo
D000818:Animals; D016118:Anterior Cruciate Ligament / Q000166:cytology* / Q000378:metabolism; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics*; D017885:Collateral Ligaments; D004247:DNA / Q000235:genetics; D005109:Extracellular Matrix / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism* / Q000648:ultrastructure; D005786:Gene Expression Regulation; D017888:Medial Collateral Ligament, Knee / Q000166:cytology / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D011817:Rabbits; D060888:Real-Time Polymerase Chain Reaction; D013710:Tendons / Q000166:cytology / Q000378:metabolism*; D023822:Tissue Engineering / Q000379:methods
D003094:Collagen; D004247:DNA