Mechanical loading regulates protease production by fibroblasts in three‐dimensional collagen substrates

Abstract

Mechanical loading is important in tissue formation and remodelling, notably in wound repair. The aim of this study was to measure the effects of controlled loading on the release of extracellular matrix protease activities by fibroblasts. Fibroblast populated collagen lattices were subjected to external cyclical loads through a computer controlled unit incorporated into a culture system, a tensioning-Culture Force Monitor. Cyclical loading was compared to untensioned and statically loaded gels (tethered endogenous contraction). Overall changes in a range of protease activities were monitored (chiefly by zymography) as measures of the cyto-mechanical response to these loads. Under static load, 2.5- and 13-fold more matrix metalloproteinase-2 was produced than matrix metalloproteinase-9, at 24 and 48 hours. Total matrix metalloproteinase-9 increased 37 fold on cyclical loading. Total matrix metalloproteinase-3 and urokinase plasminogen activator activities were dramatically reduced on cyclical loading while tissue type plasminogen activator activity was increased. Comparison with cell responses on stiffer substrates (collagen sponges) identified similar matrix metalloproteinase responses to load, but at much reduced levels (4-6 fold matrix metalloproteinase-9 stimulation on loading), showing the importance of matrix compliance to this mechano-response. In conclusion, physiological mechanical loading of fibroblasts in three dimensional collagen lattices elicited complex and substantial changes in matrix modifying proteases. These changes suggest that cells switch between expression of comparable protease activities mainly influencing cell-matrix interactions associated with migration or more generalized extracellular matrix remodelling.

Collagen network strengthening following cyclic tensile loading

“…In vivo, the changes in tissue structure and mechanics over time can partly be attributed to cellular metabolic activity in response to dynamic mechanical stimulation, such as synthesis of extracellular matrix (ECM) protein molecules (e.g. collagen [9,10], proteoglycans [11]) and proteinases, such as the MMP family [12] which modify, replace or remove components of the ECM. However, even without the active involvement of cells, the intrinsic mechanical properties of the tissue itself can change with time and history of the applied loads [13].…”

In vitro alterations in cytoskeletal tensional homeostasis control gene expression in tendon cells

“…This was characterized by the presence of organized stress fibers within the cytoskeleton and an up-regulation of the anabolic gene (al(1) collagen). Loss of cytoskeletal organization through chemical disruption or a detachment of the gel resulted in an upregulation in the expression of the catabolic gene (interstitial collagenase) and an inhibition in the expression of the anabolic gene (al (1) [33,40,41,43,56].…”

Biomechanical Determinants of Abdominal Aortic Aneurysm Rupture

“…Similarly, expression of several other MMPs, tissue plasminogen activator, and urokinase plasminogen activator is enhanced by mechanical stimulation of SMCs and/or fibroblasts. 76,79 We are especially intrigued by macrophages present in the AAA wall 19,80 because they anchor themselves to the surrounding ECM and would hence "feel" any stress that is transmitted locally. It appears that these cells, when subjected to local stress concentrations, may produce greater levels of proteolytic enzymes than when under lower stress.…”