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Protective effect of netrin-1 on hypoxia-induced mitochondrial apoptosis through HSP27expression in human umbilical cord blood derived mesenchymal stem cells : 저산소 유도 인간 제대혈중간엽 줄기세포의 미토콘드리아성세포자멸사에 대한 netrin-1의 HSP27 발현에 의한 방어 효과
DC Field | Value | Language |
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dc.contributor.advisor | 윤화영 | - |
dc.contributor.author | 손태우 | - |
dc.date.accessioned | 2017-07-19T11:28:43Z | - |
dc.date.available | 2017-07-19T11:28:43Z | - |
dc.date.issued | 2013-02 | - |
dc.identifier.other | 000000009321 | - |
dc.identifier.uri | https://hdl.handle.net/10371/133698 | - |
dc.description | 학위논문 (석사)-- 서울대학교 대학원 : 수의학과(수의내과학전공), 2013. 2. 윤화영. | - |
dc.description.abstract | Netrin (Ntn) has the potential to provide an anti-apoptotic agent with a high affinity for tissue, for therapeutic strategies of human umbilical cord blood derived mesenchymal stem cell (hUCB-MSC), although the mechanism by which Ntn-1protects hypoxic injury has yet to be identified. Therefore, the present study examined the effect of Ntn-1 on hypoxia-induced hUCB-MSC apoptosis, as well as the potential underlying mechanisms.Hypoxia (72 h) induced a decrease in the rate of MTT reduction and [3H]-thymidine incorporation, which were inhibited by Ntn-1 (10 ng/ml). Also, Ntn-1 decreased the increase of hypoxia-induced Bax, cleaved caspase-9, and cleaved caspase-3, but blocked the decrease of hypoxia-reduced Bcl-2.Next, in order to examine the Ntn-1-related signaling cascade in protection of hypoxic injury,six Ntn receptors in hUCB-MSC were analyzed. DCC and integrin(IN) α6β4 except UNC5A-C, and neogenin were existed in hUCB-MSC. Among them, IN α6β4 only was detected in lipid raft fractions. In addition, Ntn-1 induced the dissociation of DCC and APPL-1 complexthereby stimulating the formation of APPL-1 and Akt2 complex. Ntn-1 also reversed the hypoxia-induced decrease ofAkt and GSK-3β phosphorylation, which is involved in HSF-1 expression. Ntn-1-induced phospho-Akt and -GSK-3β was inhibited by DCC and IN α6β4 function-blocking antibody, and Akt inhibitor. Hypoxia and/or Ntn-1 stimulated HSP27 expression, which was blocked by HSF-1-specific siRNA. Furthermore,HSP27-specific siRNA reversedthe Ntn-1-induced increase of phospho-Akt.Additionally,HSP27-specific siRNA attenuated the Ntn-1-reduced loss of mitochondrial membrane injury viainhibition of cytochrome c (cyt c) release and formation of cyt c and HSP27 complex. Moreover, the inhibition of each signaling proteins attenuated Ntn-1-induced blockage of apoptosis. In conclusion, Ntn-1-induced HSP27 protected hypoxic injury-relatedhUCB-MSC apoptosis through DCC- and IN α6β4-dependent Akt, GSK-3β, and HSF-1signaling pathways. | - |
dc.description.tableofcontents | List of Figures vi
List of Tables vi List of Abbreviations vii Introduction 1 Materials and Methods 3 1.Materials 3 2.Isolation and characterization of human UCB-MSCs 4 3.Culture of hUCB-MSCs 5 4.Hypoxic treatment of hUCB-MSCs 5 5.[3H]-thymidine incorporation 6 6.3-(4,5,-dimethyl thiazolyl-2)-2,5-Diphenyl tetrazolium bromide (MTT) 6 7.Gene silencing with small interfering (si)RNA transfection 7 8.Cell Fractionation 7 9.RNA isolation and reverse transcription-polymerase chain reaction (RT-PCR) 8 10.Immunoprecipitation 9 11.Western blot analysis 10 12.Detergent-free purification of caveolin-rich membrane fraction 11 13.Detection of mitochondrial membrane potential 11 14.Isolation of mitochondria 12 15.Statistical analysis 13 Results 14 1.Effect of Ntn-1 on hypoxia-induced hUCB-MSC apoptosis via lipid raft independent DCC and lipid raft dependent IN α6β4 14 2.Effect of Ntn-1 on DCC-related anti-apoptotic protein complex, Akt, GSK-3β, and HSF-1 22 3.Role of heat shock protein 27 (HSP27) and Ntn-1 related signaling proteins in Ntn-1-induced cytoprotective effects 30 Discussion 42 국문초록 55 | - |
dc.format | application/pdf | - |
dc.format.extent | 2298579 bytes | - |
dc.format.medium | application/pdf | - |
dc.language.iso | en | - |
dc.publisher | 서울대학교 대학원 | - |
dc.subject.ddc | 636 | - |
dc.title | Protective effect of netrin-1 on hypoxia-induced mitochondrial apoptosis through HSP27expression in human umbilical cord blood derived mesenchymal stem cells | - |
dc.title.alternative | 저산소 유도 인간 제대혈중간엽 줄기세포의 미토콘드리아성세포자멸사에 대한 netrin-1의 HSP27 발현에 의한 방어 효과 | - |
dc.type | Thesis | - |
dc.contributor.AlternativeAuthor | Son Tae Woo | - |
dc.description.degree | Master | - |
dc.citation.pages | viii, 57 | - |
dc.contributor.affiliation | 수의과대학 수의학과 | - |
dc.date.awarded | 2013-02 | - |
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