Effect of cytoskeleton reorganization inhibition on the activation of extracellular signal-regulated kinase in osteoblasts by fluid shear stress

Abstract

To investigate the effect of cytoskeleton reorganization inhibition with RNA interference on the activation of extracellular signal-regulated kinase (ERK1/2) in primary osteoblasts induced by fluid shear stress (FSS). BALB/c mouse primary cultured osteoblasts were isolated by enzyme digestion technique. Osteoblasts were treated with LIM domain kinase 2 (LIM-2) specific siRNA or negative control siRNA, and then were loaded or unloaded by FSS of 1.2 Pa for 0, 5, 15, 30 and 60 min, respectively. The Western blotting was performed to detect the protein expression levels of P-ERK1/2 and ERK1/2, respectively. Two-way ANOVA and one-way ANOVA were used in data analysis. FSS loading for different time (0, 5, 15, 30, 60 min) treated with negative RNA inteference had significant effect on the levels of P-ERK/ERK ratio (0.047 ± 0.031, 0.253 ± 0.137, 0.390 ± 0.155, 0.613 ± 0.123, 0.680 ± 0.108, respectively, P < 0.01). Statistical analysis showed that there was significant interaction between FSS and cytoskeleton reorganization inhibition treated with RNA inteference on the levels of P-ERK/ERK ratio (P < 0.01). The levels of P-ERK/ERK ratio increased when osteoblasts were loaded for 5 - 15 min (0.623 ± 0.129 and 0.623 ± 0.064, respectively, P < 0.05) and returned to baseline at 30 min (0.333 ± 0.086), and then reached the peak at 60 min (0.667 ± 0.064, P < 0.01). FSS could activate ERK1/2 rapidly in primary cultured osteoblasts. Cytoskeleton reorganization inhibition treated with RNA interference speeded-up the activation of ERK1/2 by FSS, which could increase the sensitivity of ERK1/2 to FSS.