FIGURE 8.

LIPUS suppresses adipogenesis and promotes osteogenesis through ROCK-Cot/Tpl2-ERK pathway. A, preadipocyte 3T3-L1 cells were induced to differentiate into adipocytes with a combination of dexamethasone, insulin, and IBMX for 12 days with or without the addition of 5 μm Y-27632 and stimulated with daily LIPUS for 20 min. Total RNAs were isolated and reverse transcribed, and the gene expressions of adipogenic markers were analyzed by real time PCR. Relative mRNA expression levels were calculated in comparison with the housekeeping Rpl13a mRNA. Error bars represent the S.D. of triplicate values. Statistical significance was determined by Student's t test (**, p < 0.01). Each experiment was performed at least three times with consistent results. A typical result is shown. B, adipogenic differentiation of ST2 cells was induced with a combination of dexamethasone, insulin, and IBMX for 15 days with or without the addition of 5 μm Y-27632 and/or daily 20-min LIPUS stimulation. Analyses of adipogenic gene marker mRNAs were performed as in A. C, osteogenic differentiation of ST2 cells was induced with a combination of l-ascorbic acid 2-phosphate trisodium and β-glycerophosphate for 23 days with or without 5 μm Y-27632 and/or daily LIPUS stimulation for 20 min. The analysis of osteogenic gene marker mRNAs was performed by real time PCR as in A. *, p <0.05.