Cyclic mechanical strain stimulates the expression of heat shock protein 72 (HSP 72) in cultured human flbroblasts

Purpose of the study: To study the effect of cyclic stretching of human fibroblasts on expression of HSP 72 by Western-blot. Material and methods: Patellar tendon specimens were obtained from 5 patients (mean age=35 years). Third passage fibroblasts were plated at a density of 5×10⁵ on flexible silicone dishes. After the cells had reached subconfluency, the PCS concentration was reduced 24 hours before stimulation to 1% to synchronize the cells and to diminish the serum-induced stimulation of cell proliferation. The silicone dishes were stretched cyclically in the long axis by an electromechanical device. Strain 5%, frequency 1 Hz, strain duration 15, 60 minutes. Final measurements were made after 2, 4 and 8 hours. Cellular protein was extracted, applied onto the SDSPAGE, transferred to nitrocellulose membrane, and stained after reaction with a monoclonal antibody to HSP 72. Parallel series of cells not subjected to cyclic stretching served as controls. Results: A strain period of 15 minutes resulted in a decrease of HSP 72 synthesis after 2 hours compared to control level (0.86±0.23). There was an increase in HSP 72 synthesis after 4 hours (1.17±0.22) and 8 hours (1.11 ±0.22). A biphasic response was observed for the 60 minutes strain period. HSP 72 synthesis was increased after 2 hours ( 1.42±0.73). After 4 hours it was lower than in the controls (0.91 ±0.2), but increased after 8 hours (1.45±0.79). Conclusion: Application of mechanical stress results in an alteration of HSP 72 synthesis depending on the stress time. HSP 72 is an essential element in cell homeostasis and plays an important role in the synthesis, folding and transport of macromolecules. It exerts a major role in the maintenance of cellular metabolism by protecting cellular organelles from alteration of environment. Increased expression of HSP 72 after cyclic mechanical strain may have a protective effect on stressed human fibroblasts by stabilization basic cellular processes. HSP 72 may represent an important marker for the mechanobiology on engineered tendon and ligament tissues.