Early onset of age related hearing loss in LDHB knock out mouse

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dc.contributor.advisorChan Bae Park-
dc.description학위논문(석사)--아주대학교 일반대학원 :의생명학과,2014. 8-
dc.description.tableofcontentsABSTRACT ---------------------------------------------------------------------------------------------- i TABLE OF CONTENTS ---------------------------------------------------------------------------- iii LIST OF FIGURES ------------------------------------------------------------------------------------- v I.INTRODUCTION -------------------------------------------------------------------------------------1 II.MATERIALS AND METHOD A. Creation of LDHB knockout mice---------------------------------------------------- 4 B. Auditory brainstem response (ABR)-------------------------------------------------- 4 C. Cell Culture -------------------------------------------------------------------------------- 5 D. siRNA Transfection ----------------------------------------------------------------------- 5 E. Cell viability assay ------------------------------------------------------------------------ 5 F. ATP level measurement ------------------------------------------------------------------ 6 G. Mitochondrial membrane potential------------------------------------------------------ 6 H. Reactive oxygen species (ROS) --------------------------------------------------------- 7 I. NAD+/NADH ratio measurement -------------------------------------------------------- 7 J. LDH activity---------------------------------------------------------------------------------- 7 K. Lactate production assay ---------------------------------------------------------------- 8 L. LDH isozyme pattern assay ----------------------------------------------------------- 8 M. Western blot assay ----------------------------------------------------------------------- 9 III.RESULTS A. Generation of whole body LDHB knock out mouse ------------------------------11 B. LDHB knock out mouse grows normally---------------------------------------------13 C. LDHB knock out mouse show early onset of age related hearing loss------15 D. Differentiated hair cell express LDHB and show increased mitochondrial function ------------------------------------------------------------------------------------- 17 E. LDHB knockdown decreased the mitochondrial function in differentiated UB-OC1-----------------------------------------------------------------------------------21 F. LDHB knockdown differentiated UB-OC1 cells stabilized the Hif1a expression -23 G. LDHB knockdown differentiated UB-OC1cells are sensitive to mitochondrial toxins ----------------------------------------------------------------------------------------24 IV.DISCUSSION --------------------------------------------------------------------------------------27 V. CONCLUSION -------------------------------------------------------------------------------------30 VI. RRFERNCES --------------------------------------------------------------------------------------31-
dc.publisherThe Graduate School, Ajou University-
dc.rights아주대학교 논문은 저작권에 의해 보호받습니다.-
dc.titleEarly onset of age related hearing loss in LDHB knock out mouse-
dc.contributor.affiliation아주대학교 일반대학원-
dc.contributor.department일반대학원 의생명과학과-
dc.date.awarded2014. 8-
dc.subject.keywordHearing loss-
dc.description.alternativeAbstractAge related hearing loss (AHL) is a main feature of mammalian aging and is the most common sensory dysfunction in the elderly. AHL is associated with an age depended loss of sensory hair cell, spiral ganglion neurons, and stria vascularis cell in the inner ear(Gates and Mills,2005 and Yamasoba et al., 2007). The progressive loss of these cells finally results in hearing loss because hair cells and cochlear neuron cells do not regenerate. Mitochondria are key of energy supply, cellular redox balance, signaling and regulation of intrinsic apoptosis in inner ear, especially in the high energy demanding cells, such as hair cells and neuron cells (Hengchao, Chen et al., 2014). Lactate dehydrogenates B is important enzyme in maintaining high level of pyruvate which in the presence of oxygen will be further metabolized in the TCA cycle to produce NADH and FADH2 for oxidative phosphorylation in the mitochondria. This study aimed to observe the metabolism study of Age-related hearing loss in LDHB knockout mice. In response to deletion of LDHB gene in mouse caused early onset of age related hearing loss, as increase of high frequency threshold, hair cell death, which are the typical features of Age-related hearing loss. We investigate the molecular mechanism of hearing loss caused by LDHB deficiency by using in vitro hair cell UB-OC1. We found that differentiated UB-OC1 showed LDHB and increase of the mitochondrial function: mitochondrial respiratory subunits and MMP enhanced, increase of ATP and NAD+/NADH ratio were accompanied. LDHB knock down decreased mitochondrial function in differentiated UB-OC1 cell, the decrease of NAD+ and increase of HIF1a which probably cause mitochondrial dysfunction. LDHB knock down sensitized hair cell to antibiotics which support important clinical information.-
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Graduate School of Ajou University > Department of Biomedical Sciences > 3. Theses(Master)
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