biomechanics at micro- and nanoscale levels, v.iv, 2007, p.181

[...]... with that of WT prestin by Western blotting The results of Western blotting are shown in Fig 7 A strong 100 kDa band was detected in the lane of WT prestin Two distinct bands of 100 kDa and 80 kDa were observed in the lane of G127A In the lanes of T128A, S129A and R130A, weak 100 kDa bands and 80 kDa and 70 kDa bands were recognized In the lanes of H131A and S129T, however, 80 kDa and 70 kDa bands were... detected, but no 100 kDa bands were observed Based on our previous results [10], bands of 100 kDa and 80 kDa were thought to show prestin glycosylated with complex-type oligosaccharides and with high-mannose-type oligosaccharides, respectively, while 70 kDa bands indicated that no glycosylation occurred In the present results, the 100 kDa bands indicating prestin mutants glycosylated with complex-type... cells that was modified from that reported previously [4] The tester consists of an inverted microscope (IX-70, Olympus), a thermostatic test chamber, two micromanipulators, a linear actuator (UCM410-5C, Oriental Motor), a CCD camera and a DVD recorder A micropipette was attached to one of the micromanipulators (MHW-103, Narishige) that can be moved by the linear actuator A glass microplate was attached... substrates affect the locomotion of fibroblasts and epithelial cells [5], the proliferation and apoptosis of fibroblasts [6], and the cytoskeletal reorganization, cytoskeleton–ECM coupling, and focal adhesion of ECs [7] These results suggest that the mechanical properties of substrates influence angiogenesis through a complex process that affects many cellular responses, including migration, proliferation,... rigid as the pH of the collagen polymerization solution increased, with Young’s modulus values increasing linearly from pH 5 to 8 and then plateauing at pH 8 The Young’s modulus at pH 9 was 4.6 times greater than that at pH 5 In subsequent experiments, collagen gels that were polymerized at pH 5 (E = 4500 ± 1500 Pa) were used as “flexible” gels, and those polymerized at pH 9 (E = 20800 ± 3100 Pa) were used... the stress relaxation test Time t (s) Time t (s) Figure 5 Typical examples of the time-course changes of tension (left) and relative length (right) of cultured aortic smooth muscle cells during the stress relaxation test L*, preset length of each cell; L(t), cell length at time t 3.2 Materials and methods Rat aortic smooth muscle cells (SMCs) isolated using enzymatic digestion method [5] and passaged... centrifugation at 250 × g for 5 min and the supernatant was removed The cells were then agitated with an external solution (140 mM KCl, 3.5 mM MgCl2, 5 mM EGTA, 5 mM HEPES and 0.1 mM CaCl2; pH 7.3) and deposited on plastic dishes After ten minutes, these cells were sonicated in a hypotonic buffer (10 mM PIPES, 10 mM MgCl2, 0.5 mM EGTA; pH 7.2) The membranes attached to the substrate were then incubated... expression and localization of 3×FLAG-tagged prestin in the generated cell lines, immunofluorescence experiments were performed The untransfected CHO cells and those transfected with 3×FLAG-tagged prestin were fixed with 4% formaldehyde in phosphate buffer for 5 min at room temperature and washed with PBS The samples were then incubated with skimmed milk and fetal bovine serum for 30 min at 37°C After... specimens, untreated SMCs and SMCs whose actin filaments were disrupted with cytochalasin D (SMCs-CD) SMCs cultured on dishes in DMEM supplemented with 10% fetal bovine serum and 1% Penicillin-Streptomycin at 37ºC in 5% CO2 and 95% air were harvested with trypsinization, and were diluted by at least 1/100 with Ca2+-Mg2+-free Hank’s balanced salt solution (HBSS(-)) and placed in a dish at 37ºC to remove... under relatively large deformation using a micropipette aspiration technique, and reported that the viscosity of the cells decreased up to 80% with actin disruption The cells in this study were also exposed to relatively large deformation (ε = 70–85%) These results may indicate that actin filaments have significant effects not only on elastic but also viscous properties of cells under large deformation . Street, Covent Garden, London WC2H 9HE Printed in Singapore. BIOMECHANICS AT MICRO- AND NANOSCALE LEVELS Volume IV v PREFACE A project on Biomechanics at Micro- and Nanoscale Levels, . Wada (Tohoku University, Sendai, Japan) Published Vol. I: Biomechanics at Micro- and Nanoscale Levels Edited by Hiroshi Wada ISBN 981-256-098-X Vol. II: Biomechanics at Micro- and Nanoscale Levels Edited. w0 h0" alt="" BIOMECHANICS AT MICRO- AND NANOSCALE LEVELS Volume IV 6593 tp.indd 16593 tp.indd 1 8/2/07 11:57:03 AM8/2/07 11:57:03 AM Biomechanics at Micro- and Nanoscale Levels Editor-in-Charge: