Effects Of Hydrostatic Pressure And IGF-1on The Expression Of F-Actin Of The Goat TMJ Disc Cells

Objective:To investigate F-actin changes of the goat TMJ disc cells in vitro under hydrostatic pressure and insulin-like growth factor-1stimulation.Methods:TMJ discs were dissected from four one-month-old goats under sterile conditions and were digested with collagenase. Immunhistochemical staining for type I and II collagen were performed. The cells at passages2-3were used; the experiment was divided into four groups according to the different intervention. Group A:the cells were cultivated with complete medium; group B:the cells were subjected to a frequency of1Hz and0.2Mpa hydrostatic pressure for3h; group C:the cells cultivated with complete medium which contained insulin-like growth factor-1(10ng/mL); group D:the cells were subjected to combined effects of hydrostatic pressure and insulin-like growth factor-1, like group B and group C. Laser scanning confocal microscope(LSCM) was used to examine the changes of F-actin stained with fluorescein isothiocyanate phalloidne at24h and72h cultivation respectively. The average cell fluorescence intensity was measured with an image analysis system and analyzed.Results:The cultivated cells were identified by morphologic observation and immunohis-tochemical staining to be TMJ disc cells. The F-actin of cytoskeleton was showed to be a green fluorescence material. At24hours, fluorescence intensity of F-actin in group A and group C was strong and clear, which maintained the normal morphology of TMJ disc cells; the arrangement of F-actin in group B was showed to be disordered, and F-actin in group D was thinner compared with group A. At72hours, the F-actin in group A and C is clear and sturdy, however, part of F-actin in group B and group D was thinner and ruptured and pseudopodia was formed. As time going on, fluorescence intensity of F-actin in group A, group B and group D had an increasing trend, and there were significant differences in group A, group B and group D (P<0.05) except group C(P>0.05) between the two time point. At24hours, fluorescence intensity of F-actin was the highest in group C, the lowest in group B, both of group A and C had significant differences compared with group A and group D (P<0.05). At72hours, fluorescence intensity in group B and group D was significantly lower than group A and group C(P<0.05). Meanwhile, there were no significant differences between group B and group D, group A and group C(P>0.05).Conclusions:1. The TMJ disc cells obtained by enzyme digestion were mainly spindle-shaped and partly were polygon-shaped, showed favorable growth and functional character, kept phenotype expression of fibrochondrocytes steadily for passage3. Cells obtained from this way can be used for experiment.2. Hydrostatic pressure may cause the F-actin breakage and rearrangement of goat TMJ disc cells, and IGF-1up-regulated the F-actin expression. Such effects may be correlated with the biological behavior of the TMJ disc cells.