Cyclic Mechanical Stretching of Human Tendon Fibroblasts Increases the Production of Prostaglandin E 2 and Levels of Cyclooxygenase Expression: A Novel In Vitro Model Study

Abstract

Prostaglandin E(2) (PGE(2)) is a known inflammatory mediator of tendinitis, for which mechanical loading on tendons is believed to be one of the most prominent causation factors. Previous in vitro studies have shown that cyclic mechanical stretching of cells can cause changes in cell morphology and alteration of both DNA and protein syntheses. In our study, a novel system was used whereby tendon fibroblasts are cultured on microgrooved silicone surfaces and are subjected to cyclic uniaxial stretching along their long axes to mimic in vivo conditions. Using this unique model system, the cell shape and alignment can be controlled. Further, this study was designed to test the hypotheses that PGE(2) production increases in a stretching magnitude-dependent manner and that cyclooxygenase (COX) is responsible for the increased PGE(2) production in tendon fibroblasts. Human patellar tendon fibroblasts were cultured on the microgrooved silicone membranes and cyclically stretched at 4%, 8%, or 12% of nominal dish length for 24 hr. PGE(2) production was found to be increased 1.7-fold at 8% cyclic stretching and 2.2-fold at 12% cyclic stretching compared with nonstretched controls. In addition, human tendon fibroblasts had increased expression of both COX-1 and COX-2 for all three applied stretching magnitudes, with the exception of COX-1 at 4% cyclic stretching. Also, cellular PGE(2) production, after 8% cyclic stretching, was significantly decreased with the addition of indomethacin (25 microM), a COX competitive inhibitor, compared with stretched cells without indomethacin treatment. These findings suggest that the increase in PGE(2) production by the human tendon fibroblasts is stretching magnitude-dependent, and that the increase in COX expression contributes to the increased production of PGE(2) after cyclic stretching. As PGE(2) is a known inflammatory mediator of tendinitis, the contribution of COX-1 and COX-2 to PGE(2) production and their roles in tendon inflammation are clearly indicated.

Mechanoregulation of gene expression in fibroblasts

“…Besides IL-1β, PGE 2 is another major catabolic factor that is considered to be involved in the development of tendinopathy and joint diseases (Almekinders et al, 1993;Laufer, 2003;Wang et al, 2003Wang et al, , 2004. Previously, it was shown that tendon fibroblasts under repetitive uniaxial mechanical stretching at 8% and 12% strains increased COX-2 expression and release of PGE 2 (Wang et al, 2003).…”

“…Previously, it was shown that tendon fibroblasts under repetitive uniaxial mechanical stretching at 8% and 12% strains increased COX-2 expression and release of PGE 2 (Wang et al, 2003). Microsomal PGE synthase type 1 (mPGES-1), which is involved in the conversion of PGH 2 to PGE 2 , is an inducible enzyme functionally linked to COX-2.…”

Leukotriene B4 at low dosage negates the catabolic effect of prostaglandin E2 in human patellar tendon fibroblasts

“…The protocol for obtaining tendon samples was approved by the University of Pittsburgh Institutional Review Board. HPTFs were isolated and maintained in Dulbecco's Modified Eagle Medium (DMEM) containing 10% FBS, penicillin (50 U/ml), and streptomycin (50 U/ml) as previously described (Wang et al, 2003). The fibroblasts were subcultured for 4-6 passages and were used for the experiments.…”

“…Tendon inflammation is marked by the presence of inflammatory mediators, including prostaglandin E 2 (PGE 2 ) and leukotriene B 4 (LTB 4 ) (Almekinders et al, 1993). Although the exact amounts of both PGE 2 and LTB 4 in inflamed tendons are not known, their levels are markedly increased when tendons or tendon fibroblasts are subjected to repetitive mechanical loading conditions (Langberg et al, 1999;Wang et al, 2003;Li et al, 2004). These data suggest that PGE 2 and LTB 4 may play a role in tendon degeneration, the late stages of tendinopathy (Wang, 2005).…”

Coordinate Regulation of IL-1β and MMP-13 in Rat Tendons Following Subrupture Fatigue Damage

“…Clinical histopathologic findings demonstrate altered expression of several collagenases in tendinopathy [16,27,32,33], while animal exercise and overuse have produced both inflammatory and degenerative changes in tendons [5,9,29,34,38]. Similarly, mechanical overloading of tendon explants [13,15] and tendon cells [2,4,8,26,36,41,45,46] in vitro led to increases in both collagenase and inflammatory mediators, in conjunction with a loss of mechanical integrity. MMP-13 (collagenase-3), which has been implicated in osteoarthritis, rheumatoid One or more of the authors (HBS, MBS, ELF) has received funding from the National Institutes of Health.…”