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Distinctive collagen maturation process in fibroblasts derived from rabbit anterior cruciate ligament, medial collateral ligament, and patellar tendon in vitro.

Kato S, Saito M, Funasaki H, Marumo K - Knee Surg Sports Traumatol Arthrosc (2013)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: 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.

ABSTRACT

Purpose: 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.

Conclusion: 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.

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Transmission electron microscopy (TEM) photomicrographs showed collagen fibrils secreted from rabbit anterior cruciate ligament (ACL), medial collateral ligament (MCL), and patellar tendon (PT) cells at 0, 2, and 4 weeks after reaching confluence in culture. The TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence in culture. Two weeks after reaching confluence, all of the cells secreted large amounts of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment
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Fig4: Transmission electron microscopy (TEM) photomicrographs showed collagen fibrils secreted from rabbit anterior cruciate ligament (ACL), medial collateral ligament (MCL), and patellar tendon (PT) cells at 0, 2, and 4 weeks after reaching confluence in culture. The TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence in culture. Two weeks after reaching confluence, all of the cells secreted large amounts of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment

Mentions: TEM photomicrographs showed collagen fibrils secreted from rabbit ACL, MCL, and PT cells at 0, 2, and 4 weeks after reaching confluence in culture. TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence. Rough-surfaced endoplasmic reticula synthesize proteins. After 2 weeks in culture, all three types of fibroblasts secreted a large amount of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured, releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment (Fig. 4).Fig. 4


Distinctive collagen maturation process in fibroblasts derived from rabbit anterior cruciate ligament, medial collateral ligament, and patellar tendon in vitro.

Kato S, Saito M, Funasaki H, Marumo K - Knee Surg Sports Traumatol Arthrosc (2013)

Transmission electron microscopy (TEM) photomicrographs showed collagen fibrils secreted from rabbit anterior cruciate ligament (ACL), medial collateral ligament (MCL), and patellar tendon (PT) cells at 0, 2, and 4 weeks after reaching confluence in culture. The TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence in culture. Two weeks after reaching confluence, all of the cells secreted large amounts of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4555208&req=5

Fig4: Transmission electron microscopy (TEM) photomicrographs showed collagen fibrils secreted from rabbit anterior cruciate ligament (ACL), medial collateral ligament (MCL), and patellar tendon (PT) cells at 0, 2, and 4 weeks after reaching confluence in culture. The TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence in culture. Two weeks after reaching confluence, all of the cells secreted large amounts of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment
Mentions: TEM photomicrographs showed collagen fibrils secreted from rabbit ACL, MCL, and PT cells at 0, 2, and 4 weeks after reaching confluence in culture. TEM photomicrographs showed a random orientation of the deposited fibrous matrix. The cytoplasm of the cells contained large quantities of rough-surfaced endoplasmic reticulum when the cells reached confluence. Rough-surfaced endoplasmic reticula synthesize proteins. After 2 weeks in culture, all three types of fibroblasts secreted a large amount of collagen. However, the cytoplasm of ACL cells contained many lacunae. After 4 weeks in culture, the plasma membrane of the MCL and PT cells had ruptured, releasing the contents of the cytoplasm from the cells because of deterioration in the conditions of the culture environment (Fig. 4).Fig. 4

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: 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.

ABSTRACT

Purpose: 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.

Conclusion: 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.

Show MeSH
Related in: MedlinePlus