LDHB 유전자 삭제에 의한 허혈성 신경세포 사멸 감소
LDHB deficiency ameliorates ischemic neuronal cell death
- 주제(키워드) lactate dehydrogenase , ischemic stress , neuronal cell death , VEGFA
- 발행기관 아주대학교
- 지도교수 박찬배
- 발행년도 2017
- 학위수여년월 2017. 2
- 학위명 석사
- 학과 및 전공 일반대학원 의생명과학과
- 실제URI http://www.dcollection.net/handler/ajou/000000024174
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Although brain occupies only 2% of body weight, it consumes approximately 25% of the body's total energy and neuronal activity comprise 80% of brain energy requirement. Thus, neurons are particularly vulnerable to situations where energy source is lacking, such as ischemia. It is well known that lactate constitutes important energy source of brain energy metabolism. How lactate supports the survival of neuronal cells is well explained by astrocyte-to-neuron lactate-shuttle hypothesis. This study focuses the role of lactate dehydrogenase B (LDHB) in neuronal cell homeostasis, which is the major enzyme in lactate-shuttle theory. For this purpose, LDHB knockout mice were generated. Both in vivo and in vitro assays show that ischemic neuronal cell death is attenuated by LDHB loss. In vivo ischemic condition, PcomA (posterior communicating artery) size of LDHB KO mice is bigger than that of WT mice. These results suggest that increased PcomA size in LDHB KO mice may be contribute to neuronal cell survival. The increase of PcomA size in LDHB KO brain is further supported by the fact that mRNA level of VEGFA (vascular endothelial growth factor A) is highly increased in LDHB KO neuronal primary cells after treatment with cobalt chloride (CoCl2), which is hypoxia mimetic agent. Taken together, loss of LDHB might cause metabolic changes and that lead to decrease the ischemic neuronal cell death by increasing PcomA size and upregulating VEGFA level.
more목차
I. INTRODUCTION ----------------------------------------------------------------------- 1
II. MATERIALS AND METHODS ---------------------------------------------------- 7
A. Generation of LDHB knock-out mouse ---------------------------------------- 7
B. Induction of global forebrain ischemia ---------------------------------------- 7
C. Evaluation of the PcomA plasticity and tissue preparation & cresyl violet staining ------------------------------------------------------------------------------ 8
D. Analysis of neuronal damage ----------------------------------------------- 8
E. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay -------------------------------------------------------------------- 9
F. Neuron cell culture ---------------------------------------------------------------- 9
G. Isoenzyme polymorphisms ----------------------------------------------------- 10
H. MTT assay ------------------------------------------------------------------------ 10
I. Western blotting assay --------------------------------------------------------- 11
J. Quantitative (q)RT-PCR--------------------------------------------------------- 11
K. Data analysis ---------------------------------------------------------------------- 12
III. RESULTS ----------------------------------------------------------------------------- 13
1. The generation of LDHB KO mouse ------------------------------------------ 13
2. Ischemic cell death was attenuated in LDHB KO mice via increased PcomA size in vivo -------------------------------------------------------------- 15
3. Hypoxia response was more upregulated in LDHB KO neuron cells after treatment with CoCl2 than LDHB expressing neuron cells ---------------- 21
IV. DISCUSSION ------------------------------------------------------------------------- 27
V. CONCLUSION ------------------------------------------------------------------------ 30
VI. REFERENCES ----------------------------------------------------------------------- 31
국문요약 ----------------------------------------------------------------------------------- 35
