Koji Iida

Tohoku University, Sendai-shi, Miyagi-ken, Japan

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Publications (31)31.21 Total impact

  • Nano-Biomedical Engineering 2012 - The Tohoku University Global Centre of Excellence Programme; 01/2012
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    ABSTRACT: In the present study, purified prestin was observed by atomic force microscopy (AFM). First, the lipid bilayer was formed on a freshly cleaved mica disk by the deposition of small unilamellar lipid vesicles. Such bilayer was destabilized by incubation with a detergent. Afterward, purified prestin was added to the destabilized lipid bilayer. Finally, a high resolution image of the prestin-reconstituted lipid bilayer was acquired by AFM. As a result, densely embedded prestin with a diameter of 11.0±1.3 nm was visualized. From the obtained image, cytoplasmic side of prestin was found to form a ring-like structure with four peaks and one valley at its center.
    11/2011;
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    ABSTRACT: The SLC26A4 gene encodes the transmembrane protein pendrin, which is involved in the homeostasis of the ion concentration of the endolymph of the inner ear, most likely by acting as a chloride/bicarbonate transporter. Mutations in the SLC26A4 gene cause sensorineuronal hearing loss. However, the mechanisms responsible for such loss have remained unknown. Therefore, in this study, we focused on the function of ten missense pendrin mutations (p.P123S (Pendred syndrome), p.M147V (NSEVA), p.K369E (NSEVA), p.A372V (Pendred syndrome/NSEVA), p.N392Y (Pendred syndrome), p.C565Y (NSEVA), p.S657N (NSEVA), p.S666F (NSEVA), p.T721M (NSEVA) and p.H723R (Pendred syndrome/NSEVA)) reported in Japanese patients, and analyzed their cellular localization and anion exchanger activity using HEK293 cells transfected with each mutant gene. Immunofluorescent staining of the cellular localization of the pendrin mutants revealed that p.K369E and p.C565Y, as well as wild-type pendrin, were transported to the plasma membrane, while 8 other mutants were retained in the cytoplasm. Furthermore, we analyzed whether salicylate, as a pharmacological chaperone, restores normal plasma membrane localization of 8 pendrin mutants retained in the cytoplasm to the plasma membrane. Incubation with 10 mM of salicylate of the cells transfected with the mutants induced the transport of 4 pendrin mutants (p.P123S, p.M147V, p.S657Y and p.H723R) from the cytoplasm to the plasma membrane and restored the anion exchanger activity. These findings suggest that salicylate might contribute to development of a new method of medical treatment for sensorineuronal hearing loss caused by the mutation of the deafness-related proteins, including pendrin.
    Hearing research 12/2010; 270(1-2):110-8. · 2.18 Impact Factor
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    ABSTRACT: Prestin is the motor protein of cochlear outer hair cells and is essential for mammalian hearing. The present study aimed to clarify the structure of prestin by atomic force microscopy (AFM). Prestin was purified from Chinese hamster ovary cells which had been modified to stably express prestin, and then reconstituted into an artificial lipid bilayer. Immunofluorescence staining with anti-prestin antibody showed that the cytoplasmic side of prestin was possibly face up in the reconstituted lipid bilayer. AFM observation indicated that the cytoplasmic surface of prestin was ring-like with a diameter of about 11 nm.
    FEBS letters 07/2010; 584(13):2872-6. · 3.54 Impact Factor
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    ABSTRACT: Prestin is a key molecule for mammalian hearing. The present study investigated changes in characteristics of prestin by culturing prestin-transfected cells with salicylate, an antagonist of prestin. As a result, the plasma membrane localization of prestin bearing a mutation in the GTSRH sequence, which normally accumulates in the cytoplasm, was recovered. Moreover, the nonlinear capacitance of the majority of the mutants, which is a signature of prestin activity, was also recovered. Thus, the present study discovered a new effect of salicylate on prestin, namely, the promotion of the plasma membrane expression of prestin mutants in an active state.
    FEBS letters 04/2010; 584(11):2327-32. · 3.54 Impact Factor
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    ABSTRACT: The motor protein prestin in cochlear outer hair cells is a member of the solute carrier 26 family, but among the proteins of that family, only prestin can confer the cells with nonlinear capacitance (NLC) and motility. In the present study, to clarify contributions of unique amino acids of prestin, namely, Met-122, Met-225 and Thr-428, to the characteristics of prestin, mutations were introduced into those amino acids. As a result, NLC remained unchanged by both replacement of Met-122 by isoleucine and that of Thr-428 by leucine, suggesting that those amino acids were not important for the generation of NLC. Surprisingly, the replacement of Met-225 by glutamine statistically increased NLC as well as the motility of prestin-expressing cells without an increase in the amount of prestin expression in the plasma membrane. This indicates that Met-225 in prestin somehow adjusts NLC and the motility of prestin-expressing cells.
    Biochemical and Biophysical Research Communications 10/2009; 389(4):569-74. · 2.28 Impact Factor
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    ABSTRACT: Prestin is the motor protein of cochlear outer hair cells, which exhibit elongation and contraction in response to acoustic stimuli. In this study, the effects of mutations in unique amino acids, which among members of the SLC26 family were only present in prestin, on the characteristics of prestin were investigated. As a result, a mutation in Met-225 of prestin was found to increase nonlinear capacitance, which is a signature of prestin activity.
    02/2009;
  • 02/2009;
  • 02/2009;
  • Conference Paper: INNER EAR BIOMECHANICS
    Nano-Biomedical Engineering 2009 - The Tohoku University Global Centre of Excellence Programme; 01/2009
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    ABSTRACT: Prestin, a membrane protein of the outer hair cells (OHCs), is known to be the motor which drives OHC somatic electromotility. Electron microscopic studies showed the lateral membrane of the OHCs to be densely covered with 10-nm particles, they being believed to be a motor protein. Imaging by atomic force microscopy (AFM) of prestin-transfected Chinese hamster ovary (CHO) cells revealed 8- to 12-nm particle-like structures to possibly be prestin. However, since there are many kinds of intrinsic membrane proteins other than prestin in the plasma membranes of OHCs and CHO cells, it was impossible to clarify which structures observed in such membranes were prestin. In the present study, an experimental approach combining AFM with quantum dots (Qdots), used as topographic surface markers, was carried out to detect individual prestin molecules. The inside-out plasma membranes were isolated from the prestin-transfected and untransfected CHO cells. Such membranes were then incubated with antiprestin primary antibodies and Qdot-conjugated secondary antibodies. Fluorescence labeling of the prestin-transfected CHO cells but not of the untransfected CHO cells was confirmed. The membranes were subsequently scanned by AFM, and Qdots were clearly seen in the prestin-transfected CHO cells. Ring-like structures, each with four peaks and one valley at its center, were observed in the vicinity of the Qdots, suggesting that these structures are prestin expressed in the plasma membranes of the prestin-transfected CHO cells.
    Pflügers Archiv - European Journal of Physiology 09/2008; 457(4):885-98. · 4.87 Impact Factor
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    ABSTRACT: Acetylation and deacetylation of proteins occur in cells in response to various stimuli, and are reversibly catalyzed by histone acetyltransferase and histone deacetylase (HDAC), respectively. EoL-1 cells have an FIP1L1-PDGFRA fusion gene that causes transformation of eosinophilic precursor cells into leukemia cells. The HDAC inhibitors apicidin and n-butyrate suppress the proliferation of EoL-1 cells and induce differentiation into eosinophils by a decrease in the protein level of FIP1L1-PDGFRalpha without affecting the mRNA level for FIP1L1-PDGFRA. In this study, we analyzed the mechanism by which the protein level of FIP1L1-PDGFRalpha is decreased by apicidin and n-butyrate. EoL-1 cells were incubated in the presence of the HDAC inhibitors apicidin, trichostatin A or n-butyrate. The protein levels of FIP1L1-PDGFRalpha and phosphorylated eIF-2alpha were determined by Western blotting. Actinomycin D and cycloheximide were used to block RNA synthesis and protein synthesis, respectively, in the chasing experiment of the amount of FIP1L1-PDGFRalpha protein. When apicidin- and n-butyrate-treated EoL-1 cells were incubated in the presence of actinomycin D, the decrease in the protein level of FIP1L1-PDGFRalpha was significantly enhanced when compared with controls. In contrast, the protein levels were not changed by cycloheximide among these groups. Apicidin and n-butyrate induced the continuous phosphorylation of eIF-2alpha for up to 8 days. The decrease in the level of FIP1L1-PDGFRalpha protein by continuous inhibition of HDAC may be due to the decrease in the translation rate of FIP1L1-PDGFRA.
    International Archives of Allergy and Immunology 02/2008; 146 Suppl 1:7-10. · 2.25 Impact Factor
  • Journal of Biomechanical Science and Engineering 01/2008; 3(2):287-298.
  • Journal of Biomechanical Science and Engineering 01/2008; 3(2):221-234.
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    ABSTRACT: When the ear is exposed to traumatic loud noise, outer hair cells (OHCs) are damaged and thus permanent hearing loss occurs. Recently, prior conditioning with heat stress has been reported to protect OHCs from traumatic noise exposure by increasing the stiffness of the OHC soma and has also been reported to enhance distortion product otoacoustic emissions [DPOAEs; Murakoshi, M., Yoshida, N., Kitsunai, Y., Iida, K., Kumano, S., Suzuki, T., Kobayashi, T., Wada, H., 2006. Effects of heat stress on Young's modulus of outer hair cells in mice. Brain Res. 1107, 121-130]. In the present study, to further investigate the heat stress-induced protective mechanism of hearing and such stress-induced DPOAE enhancement mechanism, the amount of filamentous actin (F-actin), which is concerned with cell stiffness, and the amount of prestin, which is concerned with the generation of DPOAEs, were examined in OHCs, with and without heat stress. Heat stress was found to increase the amount of F-actin 6-24 h after heat stress. The greatest increase in the amount of F-actin was observed at the cuticular plate where F-actin anchors the roots of the stereocilia to the cell body. Based on this result, the part of the stereocilia most reinforced and protected by heat stress was concluded to be the roots of the stereocilia. In contrast with F-actin, heat stress did not affect the amount of prestin.
    Brain Research 11/2007; 1177:47-58. · 2.88 Impact Factor
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    01/2007;
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    ABSTRACT: The high sensitivity of mammalian hearing is achieved by amplification of the motion of the cochlear partition. This cochlear amplification is thought to be generated by the elongation and contraction of outer hair cells (OHCs) in response to acoustical stimulation. This motility is made possible by a membrane protein embedded in the lateral membrane of OHCs. Although a fructose transporter, GLUT-5, was initially proposed to be this protein, a later study identified the gene of the motor protein distributed throughout the OHC plasma membrane. This protein has been named "prestin." However, although previous morphological studies by electron microscopy and atomic force microscopy (AFM) found the lateral wall of OHCs to be covered with 10-nm particles, believed to be motor proteins, it is unknown whether such particles consist only of prestin or are a complex of GLUT-5 and prestin molecules. To determine if the 10-nm particles are indeed constituted only of prestin, plasma membranes of prestin-transfected and untransfected Chinese hamster ovary (CHO) cells, which do not express GLUT-5, were observed by AFM. First, the cells attached to a substrate were sonicated so that only the plasma membrane remained on the substrate. The cytoplasmic face of the cell was observed by the tapping mode of the AFM in liquid. As a result, particle-like structures were recognized on the plasma membranes of both the prestin-transfected and untransfected CHO cells. Comparison of the difference in the frequency distribution of these structures between those two cells showed approximately 75% of the particle-like structures with a diameter of 8-12 nm in the prestin-transfected CHO cells to be possibly constituted only by prestin molecules. Our data suggest that the densely packed 10-nm particles observed on the OHC lateral wall are likely to be constituted only of prestin molecules.
    Journal of the Association for Research in Otolaryngology 10/2006; 7(3):267-78. · 2.95 Impact Factor
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    ABSTRACT: Intense sound exposure causes permanent hearing loss due to hair cell and cochlear damage. Prior conditioning with sublethal stressors, such as nontraumatic sound, heat stress and restraint protects the ear from acoustic injury. However, the mechanisms underlying conditioning-related cochlear protection remain unknown. In this paper, Young's modulus and the amount of filamentous actin (F-actin) of outer hair cells (OHCs) with/without heat stress were investigated by atomic force microscopy and confocal laser scanning microscopy, respectively. Conditioning with heat stress resulted in a statistically significant increase in Young's modulus of OHCs at 3-6 h after application, and such modulus then began to decrease by 12 h and returned to pre-conditioning level at 48 h after heat stress. The amount of F-actin began to increase by 3 h after heat stress and peaked at 12 h. It then began to decrease by 24 h and returned to the pre-conditioning level by 48-96 h after heat stress. These time courses are consistent with a previous report in which heat stress was shown to suppress permanent threshold shift (PTS). In addition, distortion product otoacoustic emissions (DPOAEs) were confirmed to be enhanced by heat stress. These results suggest that conditioning with heat stress structurally modifies OHCs so that they become stiffer due to an increase in the amount of F-actin. As a consequence, OHCs possibly experience less strain when they are exposed to loud noise, resulting in protection of mammalian hearing from traumatic noise exposure.
    Brain Research 09/2006; 1107(1):121-30. · 2.88 Impact Factor
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    ABSTRACT: Outer hair cells (OHCs) are capable of altering their cell length in response to changes in membrane potential. Due to this electromotility, OHCs probably subject the basilar membrane to force, resulting in cochlear amplification. To understand the mechanism of such amplification, knowledge of the mechanical properties of OHCs is required since the force produced by OHC electromotility is thought to depend on such properties. Various studies have been conducted to investigate the mechanical properties of guinea pig OHCs. With regard to mice, however, although various kinds of transgenic and knockout mice possess great potential as research models, the mechanical properties of mouse OHCs have not as yet been reported since the cells and/or tissues in the mouse hearing organ are relatively small and vulnerable to external stimuli, rendering sample preparation difficult. In this study, therefore, to establish indicators of the mechanical properties of OHCs in mice, such properties were measured by atomic force microscopy (AFM). CBA/JNCrj strain male mice aged 10-12 weeks (25-30 g) were used. Cochleae were dissected out from the animal and both the basilar membrane and the organ of Corti were simultaneously unwrapped from the modiolus with forceps. Dissected coiled tissue was then incubated with an enzymatic digestion medium for 15 min. After digestion, OHCs were isolated by gently triturating the coiled tissue. Local mechanical properties of the OHCs were then measured by an indentation test using an AFM. Young's modulus and stiffness of the OHC in the apical turn of the mouse cochlea were 2.1+/-0.5 kPa and 4.4+/-1.2 mN/m, respectively. Young's modulus of the OHC in the apical turn of the cochlea in mice was roughly the same as that in the apical turn of the cochlea in guinea pigs; however, the stiffness of the former was about two times greater than that of the latter because the cell length of the former was shorter than that of the latter.
    Auris Nasus Larynx 07/2006; 33(2):149-57. · 0.95 Impact Factor
  • Future Medical Engineering Based on Bionanotechnology - The Final Symposium of the Tohoku University 21st Century Center of Excellence Program; 01/2006

Publication Stats

64 Citations
31.21 Total Impact Points

Institutions

  • 2005–2010
    • Tohoku University
      • Department of Bioengineering and Robotics
      Sendai-shi, Miyagi-ken, Japan
  • 2007
    • The University of Tokyo
      • Department of Medical Genome Sciences
      Edo, Tōkyō, Japan