Role And Its Mechanisms Of Cyclic Strain Frequency In Regulating The Alignment Of Vascular Smooth Muscle Cells In Vitro

The mechanical stimuli from the pulsatile blood flow are known to regulate vascular remodeling. Each cyclic mechanical strain loading includes at least three elements, magnitude, frequency, and exposure time. The circumferential orientation and structural network of the muscle layer are very important for maintaining the mechanical strength and function of the arterial wall. Mechanical stress can affect the alignment of vascular smooth muscle cells (VSMCs) in vitro, but the role and its mechanisms of cyclic strain frequency in regulating on the alignment of VSMCs remain unclear.In this study, VSMCs were stretched under the same elongation but different frequencies. As a control, the same elongation stretch was constantly applied throughout the experiment with no variation in frequency. Cells were photographed and their orientation angles were measured by software. Their alignment was evaluated by changes of the VSMC angle information entropy (AIE), and the time courses of these changes were modeled mathematically. The changes of integrin-β1, p38 and actin filaments in VSMCs under different frequencies of cyclic strain were observed by western blotting, flow cytometry and real time RT-PCR. The functions of them in signal transduction were also observed with the use of anti-integrin-β1 blocking antibody, specific inhibitor PD98059 and Cytochalasin D respectively.The results indicated that different frequency of cyclic strain can affect VSMC alignment obviously. AIEs were decreased by cyclic strain within 12 hours in all groups treated with different frequencies, and then reached their respective asymptotic values after 24 hours, with a lower frequency causing a lower AIE. The trend of AIE change was described by logistic equations. Values of AIE at a given frequency was predicted and verified with experiments. The changes of the activation state ofβ1 integrin and the F/G actin ratio of filaments system were consistent with VSMC alignment;β1-integrin antibody, PD98059 and Cytochalasin D can inhibit VSMC alignment, and the effect of the frequency of cyclic strain could be inhibited by Cytochalasin D.It is concluded that the strain frequency is a significant regulating factor in cell alignment; within a certain time frame, an optimum frequency exists to induce the most effective cell alignment; after sufficient exposure time, more regular alignment of VSMCs can be obtained under a lower strain frequency within a certain frequency range;β1 integrin, p38 and actin filament were involved in the cascade of signal transduction induced by cyclic strain, and the effect of frequency is dependent on an intact filaments system in VSMCs.