Changes in Gene Expression of Matrix Constituents with Respect to Passage of Ligament and Tendon Fibroblasts

• Published in: Annals of biomedical engineering 2008-12, Vol.36 (12), p.1927-1933
• Main Authors: Almarza, Alejandro J., Augustine, Serena M., Woo, Savio L-Y.
• Format: Article
• Language: English
• Subjects: Animals; Biochemistry; Biological and Medical Physics; Biomedical and Life Sciences; Biomedical Engineering and Bioengineering; Biomedicine; Biophysics; Cells, Cultured; Classical Mechanics; Collagen Type I - genetics; Collagen Type I - metabolism; Collagen Type III - genetics; Collagen Type III - metabolism; Extracellular Matrix - genetics; Extracellular Matrix - metabolism; Extracellular Matrix Proteins - genetics; Extracellular Matrix Proteins - metabolism; Female; Fibroblasts - metabolism; Fibronectins - genetics; Fibronectins - metabolism; Gene Expression; Ligaments - metabolism; Matrix Metalloproteinase 13 - genetics; Matrix Metalloproteinase 13 - metabolism; Rats; Rats, Long-Evans; Relative abundance; RNA, Messenger - metabolism; Tendons - metabolism; Tissue Inhibitor of Metalloproteinase-1 - genetics; Tissue Inhibitor of Metalloproteinase-1 - metabolism
• ISSN: 0090-6964; 1573-9686
• DOI: 10.1007/s10439-008-9565-1

Trauma to the knee joint often results in injury to one or more supporting soft tissue structures, such as the medial collateral (MCL) and anterior cruciate (ACL) ligaments. Also, a portion of the patellar tendon (PT) is frequently used as a replacement graft for the ACL, resulting in a PT defect. The healing responses of these tissues are dramatically different and range from spontaneous healing to little or no healing. Studies have suggested that native cell behavior could be responsible for differences in healing potential. However, it is difficult to make comparisons as the reported results are based on different cellular passages which could have a dramatic effect on their potential to form healing tissues. Therefore, the objective of this study was to quantify the gene expression of collagen and other matrix constituents of fibroblasts from the MCL, ACL, and PT to document how they change with cell passage. We hypothesized that MCL fibroblasts would possess higher potential for matrix production through passages than ACL and PT cells because the MCL mounts a robust healing response unlike the ACL and PT. These differences in matrix expression would be dependent on passage because at earlier passages all cells would mostly be proliferating while at later passages they would tend to become senescent. Cells were isolated from the MCL, ACL, and PT of three rats and passaged a total of five times (Passage 1 to Passage 5). Using real time RT-PCR, expression of all genes of interest (Collagen Type I (ligament/tendon’s main matrix constituent), Collagen Type III, Fibronectin, Metalloprotease-13 [MMP-13], and Tissue Inhibitor of Metallopreotease-1 [TIMP-1]) were quantitatively assessed. It was found that cell number for all three fibroblast types remained high from Passage 1 to Passage 5. There was a statistically significant increase in Collagen Type I of rat MCL fibroblasts throughout passage ( p