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研究生:郭佩雯
研究生(外文):Pei-Wen Kuo
論文名稱:RAB18過度表現在精神分裂症致病機轉上的角色
論文名稱(外文):The Role of RAB18 over Expression in the Pathogenesis of Schizophrenia
指導教授:洪成志洪成志引用關係
指導教授(外文):Chen-Jee Hong
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:腦科學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:42
中文關鍵詞:精神分裂症Rab18社交行為抗精神分裂症藥物負向症狀
外文關鍵詞:Rab18Schizophreniasociabilityantipsychotic drugsnegative symptom
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背景:精神分裂症是個思考,情緒,知覺障礙的疾病,Ayalew et al. (2012)用綜合基因體分析的方法發現RAB18為精神分裂症相關的前十名候選基因之一。我們發現精神分裂症患者的血中單核球的RAB18 mRNA表現量高於一般人,也在患者發現RAB18 -1700 A→T的變異,在282個正常人身上則沒有發現此變異。
假說: Rab18表現量增加在精神分裂症的病理機轉上,扮演重要的角色。
方法與材料: 利用冷光報導載體分析檢測RAB18 -1700A/T與-968G/A的啟動子活性。再使用C57BL/6品系的Rab18基因轉殖鼠做曠野實驗、前脈衝抑制測試、社交互動測試、高腳十字迷宮測試、多重T型迷宮測試以及強迫游泳測試。並施打抗精神症藥物做西方墨點法檢測Rab18的大腦表現量。
結果: RAB18 -1700T相較於普遍的-1700A構築體有較高啟動子活化。在社交互動行為上,WT老鼠對裝陌生老鼠容器的探索時間顯著高於空容器的探索時間,但Rab18過度表現的小鼠則無此差異。在曠野實驗、前脈衝抑制測試、高腳十字迷宮測試、多重T型迷宮測試以及強迫游泳測試中,WT與Rab18過度表現的小鼠無顯著差異。正常老鼠施打抗精神症藥物clozapine(10 mg/kg)會使前額葉Rab18表現量降低。而兩種抗精神症藥物clozapine (2mg/kg)和haloperidol(0.2mg/kg)都可使Rab18過度表現的小鼠提高對陌生老鼠的社交互動行為。
結論: 精神分裂症患者才有的RAB18啟動子-1700T變異會有較高啟動子活化。Rab18過度表現的基因轉殖鼠會有符合精神分裂症負向症狀的社交互動降低的特徵。抗精神症藥物clozapine不只可以恢復Rab18過度表現的基因轉殖鼠的社交缺乏行為,也可改變正常老鼠的Rab18表現量。我們的發現顯示RAB18在精神分裂症的致病機轉上扮演著重要的角色。

Background: Schizophrenia is a mental disorder with thinking, emotional, sensory and cognitive disturbances. Ayalew et al. (2012) used a convergent functional genomics approach to prioritize genes associated with schizophrenia and found that RAB18 is among the top 10 candidates. We previously found that schizophrenia patient’s RAB18 mRNA expression in the mononuclear cells was higher than that from normal controls. We also identified a specific polymorphism (-1700A→T) in the RAB18 promoter region of a schizophrenia patient, which was not found in 282 normal people .
Hypothesis: Rab18 overexpression plays a role in the pathogenesis of schizophrenia
Materials and Methods: We used dual-luciferase reporter assay to compare the effect of RAB18 -1700A/T on promoter activity. Mouse behaviors were evaluated with the open field test, prepulse inhibition, the sociability test, elevated plus maze test, multiple T maze test, and force swim test. The effects of antipsychotic drugs on Rab18 protein expression in the brain was assayed by Western blotting.
Results: RAB18 -1700T construct had a higher promoter activity than the construct with the common allele -1700A. In the sociability test, WT mice spent more time approaching the cage containing a stranger mouse than did Rab18-tg mice. Rab18-tg mice’s performances were not significantly different from WT mice in the open-field test, prepulse inhibition test, elevated plus maze, multiple T maze test, and force swim test. Clozapine (10mg/kg) reduced Rab18 protein expression in the prefrontal cortex of the WT mice. Both Clozapine (2mg/kg) and haloperidol (0.2mg/kg) improved Rab18-tg mice’s social interactions with stranger mice.
Conclusions: RAB18 -1700T polymorphism uniquely found in a schizophrenia patient had a higher promoter activity than the common -1700A allele. Transgenic mice with over-expressed Rab18 displayed less social interaction that is a common negative symptom of schizophrenia. Clozapine not only improve WT mice’s Rab18 expression but also improve Rab18-tg mice’s social withdrawal. Antipsychotic drug could alter WT mice’s Rab18 expression. Our findings, implicate a significant role of Rab18 in the pathogenesis of schizophrenia.

目錄 i
圖目錄 iv
致謝 v
Abstract vi
中文摘要 viii
導論(Introduction) 1
Rab18在細胞內的功能 1
Rab18跟精神分裂症的關係 2
前人對於精神分裂症的研究 2
精神分裂症的動物模式與假說 2
初步的發現 4
假設與研究目的 6
實驗材料與方法 7
1. 冷光報導載體分析之製備(Plasmid DNA prepartion) 7
1-1. RAB18 promoter的質體構築(RAB18 promoter construct) 8
1-2. 質體DNA萃取(Plasmid extration) 8
1-3. 細胞培養(Cell culture) 9
1-4. 質體轉染細胞(Cells transfected with vectors) 9
1-5. 冷光報導基因分析(Dual luciferase assay) 10
2. 實驗動物基因型鑑定及蛋白質萃取 10
2-1. Rab18基因轉殖鼠(Rab18 transgenic mouse) 10
2-2. 小鼠尾巴DNA萃取 (DNA extraction) 11
2-3. 基因型鑑定 (Genotyping) 11
2-4. 組織蛋白萃取(Protein extraction) 11
2-5. 西方墨點法(Western blot) 13
3. 小鼠行為實驗(Behavioral tests) 14
3-1. 曠野實驗(Open field test) 14
3-2. 前脈衝抑制測驗 (Prepulse inhibition test) 15
3-3. 社交行為測驗(Sociability test) 16
3-4. 多重T型迷宮測試(Multiple T-Maze test) 17
3-5. 高腳迷宮測驗(Elevated plus maze test;EPM) 18
3-6. 強迫游泳實驗(Forced swim test ; FST) 19
4. 藥物施打 19
5. 統計分析 20
實驗結果 21
1. RAB18啟動子-1700 SNP之變異會影響啟動子活性 21
2-1. Rab18基因轉殖鼠鑑定與蛋白表現量分析 22
2-2. Rab18 基因轉殖鼠的活動力及體重 24
2-3. Rab18 基因轉殖鼠的前脈衝抑制反應 25
2-4. Rab18基因轉殖鼠的社交行為 26
2-5. Rab18基因轉殖鼠的認知行為 27
2-6. Rab18基因轉殖鼠的情緒行為 29
3-1. 抗精神症藥物clozapine對前額葉的Rab18表現量之影響 30
3-2. 抗精神症藥物對Rab18 -tg之影響 31
討論 33
RAB18啟動子-1700T有較高活化 33
RAB18表現量的變化與精神分裂症的關聯 33
Rab18-tg小鼠顯性出類似精神分裂症的負向症狀 34
Rab18可能透過調控血清素來影響小鼠的負向症狀 34
Rab18過度表現不會造成認知功能異常 35
抗精神症藥物減少前額葉Rab18表現量 36
結論 36
未來展望 37
參考文獻 38

圖目錄

圖 1. RAB18與精神分裂症的關聯 5
圖 2. Dual-luciferase assay示意圖 7
圖 3. 前額葉取法 12
圖 4. 行為實驗流程圖 14
圖 5. 前脈衝抑制實驗的四種刺激形式 (trial) 16
圖 6. 社交互動測驗示意圖 17
圖 7. T-maze示意圖 18
圖 8. RAB18啟動子上-1700 SNP會影響luciferase的表現 22
圖 9. 基因型鑑定與Rab18蛋白表現量分析 23
圖 10. Rab18 基因轉殖鼠的活動力與體重的測量 24
圖 11. Rab18 基因轉殖鼠的前脈衝抑制測量 25
圖 12. Rab18基因轉殖鼠社交行為之測量 26
圖 13. 利用雙重T型迷宮測試Rab18小鼠之空間記憶 27
圖 14. 測試Rab18-tg小鼠的情感行為測驗 29
圖 15. Clozapine(10mg/kg)對腦部Rab18蛋白表現的影響 30
圖 16. 施打clozapine 或haloperidol會影響 Rab18-tg的社交互動行為 31

1. Stenmark, H. and V.M. Olkkonen, The Rab GTPase family. Genome Biol, 2001. 2(5): p. REVIEWS3007.
2. Pfeffer, S.R., Rab GTPases: master regulators of membrane trafficking. Curr Opin Cell Biol, 1994. 6(4): p. 522-6.
3. Park, H.H., Structural basis of membrane trafficking by rab family small g protein. Int J Mol Sci, 2013. 14(5): p. 8912-23.
4. Vazquez-Martinez, R., et al., Rab18 inhibits secretory activity in neuroendocrine cells by interacting with secretory granules. Traffic, 2007. 8(7): p. 867-82.
5. Martin, S., et al., Regulated localization of Rab18 to lipid droplets: effects of lipolytic stimulation and inhibition of lipid droplet catabolism. J Biol Chem, 2005. 280(51): p. 42325-35.
6. Ozeki, S., et al., Rab18 localizes to lipid droplets and induces their close apposition to the endoplasmic reticulum-derived membrane. J Cell Sci, 2005. 118(Pt 12): p. 2601-11.
7. Pulido, M.R., et al., Rab18 dynamics in adipocytes in relation to lipogenesis, lipolysis and obesity. PLoS One, 2011. 6(7): p. e22931.
8. Dejgaard, S.Y., et al., Rab18 and Rab43 have key roles in ER-Golgi trafficking. J Cell Sci, 2008. 121(Pt 16): p. 2768-81.
9. Gerondopoulos, A., et al., Rab18 and a Rab18 GEF complex are required for normal ER structure. J Cell Biol, 2014. 205(5): p. 707-20.
10. Lutcke, A., et al., Cloning and subcellular localization of novel rab proteins reveals polarized and cell type-specific expression. J Cell Sci, 1994. 107 ( Pt 12): p. 3437-48.
11. Burre, J., et al., Synaptic vesicle proteins under conditions of rest and activation: analysis by 2-D difference gel electrophoresis. Electrophoresis, 2006. 27(17): p. 3488-96.
12. Bem, D., et al., Loss-of-function mutations in RAB18 cause Warburg micro syndrome. Am J Hum Genet, 2011. 88(4): p. 499-507.
13. Carpanini, S.M., et al., A novel mouse model of Warburg Micro syndrome reveals roles for RAB18 in eye development and organisation of the neuronal cytoskeleton. Dis Model Mech, 2014. 7(6): p. 711-22.
14. Ayalew, M., et al., Convergent functional genomics of schizophrenia: from comprehensive understanding to genetic risk prediction. Mol Psychiatry, 2012. 17(9): p. 887-905.
15. Le-Niculescu, H., et al., Towards understanding the schizophrenia code: an expanded convergent functional genomics approach. Am J Med Genet B Neuropsychiatr Genet, 2007. 144B(2): p. 129-58.
16. Duncan, C.E., A.F. Chetcuti, and P.R. Schofield, Coregulation of genes in the mouse brain following treatment with clozapine, haloperidol, or olanzapine implicates altered potassium channel subunit expression in the mechanism of antipsychotic drug action. Psychiatr Genet, 2008. 18(5): p. 226-39.
17. Mohn, A.R., et al., Mice with reduced NMDA receptor expression display behaviors related to schizophrenia. Cell, 1999. 98(4): p. 427-36.
18. Ban, T.A., W. Guy, and W.H. Wilson, Description and distribution of the subtypes of chronic schizophrenia based on Leonhard's classification. Psychiatr Dev, 1984. 2(3): p. 179-99.
19. Braff, D., et al., Prestimulus effects on human startle reflex in normals and schizophrenics. Psychophysiology, 1978. 15(4): p. 339-43.
20. Borrell, J., et al., Prenatal immune challenge disrupts sensorimotor gating in adult rats. Implications for the etiopathogenesis of schizophrenia. Neuropsychopharmacology, 2002. 26(2): p. 204-15.
21. Jaaro-Peled, H., Gene models of schizophrenia: DISC1 mouse models. Prog Brain Res, 2009. 179: p. 75-86.
22. Ho, B.C., et al., Progressive structural brain abnormalities and their relationship to clinical outcome: a longitudinal magnetic resonance imaging study early in schizophrenia. Arch Gen Psychiatry, 2003. 60(6): p. 585-94.
23. Jung, W.H., et al., Structural brain alterations in individuals at ultra-high risk for psychosis: a review of magnetic resonance imaging studies and future directions. J Korean Med Sci, 2010. 25(12): p. 1700-9.
24. Steen, R.G., et al., Brain volume in first-episode schizophrenia: systematic review and meta-analysis of magnetic resonance imaging studies. Br J Psychiatry, 2006. 188: p. 510-8.
25. Lipska, B.K. and D.R. Weinberger, To model a psychiatric disorder in animals: schizophrenia as a reality test. Neuropsychopharmacology, 2000. 23(3): p. 223-39.
26. Robinson, T.E. and J.B. Becker, Enduring changes in brain and behavior produced by chronic amphetamine administration: a review and evaluation of animal models of amphetamine psychosis. Brain Res, 1986. 396(2): p. 157-98.
27. Swerdlow, N.R., et al., Amphetamine disruption of prepulse inhibition of acoustic startle is reversed by depletion of mesolimbic dopamine. Psychopharmacology (Berl), 1990. 100(3): p. 413-6.
28. Thompson, R.F., The Brain. 1993.
29. Stahl, S.M., Stahl. Essential Psychopharmacology: Neuroscientific Basis and Practical Applications. 2008, Cambridge University Press.
30. Lisman, J.E., et al., Circuit-based framework for understanding neurotransmitter and risk gene interactions in schizophrenia. Trends Neurosci, 2008. 31(5): p. 234-42.
31. Kalinichev, M., et al., Comparison between intraperitoneal and subcutaneous phencyclidine administration in Sprague-Dawley rats: a locomotor activity and gene induction study. Prog Neuropsychopharmacol Biol Psychiatry, 2008. 32(2): p. 414-22.
32. Sams-Dodd, F., Distinct effects of d-amphetamine and phencyclidine on the social behaviour of rats. Behav Pharmacol, 1995. 6(1): p. 55-65.
33. Mansbach, R.S. and M.A. Geyer, Effects of phencyclidine and phencyclidine biologs on sensorimotor gating in the rat. Neuropsychopharmacology, 1989. 2(4): p. 299-308.
34. Aghajanian, G.K. and G.J. Marek, Serotonin model of schizophrenia: emerging role of glutamate mechanisms. Brain Res Brain Res Rev, 2000. 31(2-3): p. 302-12.
35. Bleich, A., et al., The role of serotonin in schizophrenia. Schizophr Bull, 1988. 14(2): p. 297-315.
36. Jordan, S., et al., The antipsychotic aripiprazole is a potent, partial agonist at the human 5-HT1A receptor. Eur J Pharmacol, 2002. 441(3): p. 137-40.
37. Meltzer, H.Y., Role of serotonin in the action of atypical antipsychotic drugs. Clin Neurosci, 1995. 3(2): p. 64-75.
38. Meltzer, H.Y., et al., Serotonin receptors: their key role in drugs to treat schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry, 2003. 27(7): p. 1159-72.
39. Kapur, S. and G. Remington, Serotonin-dopamine interaction and its relevance to schizophrenia. Am J Psychiatry, 1996. 153(4): p. 466-76.
40. Coleman, L.G., Jr., et al., Deficits in adult prefrontal cortex neurons and behavior following early post-natal NMDA antagonist treatment. Pharmacol Biochem Behav, 2009. 93(3): p. 322-30.
41. Michel, H.E., et al., Prepulse inhibition (PPI) disrupting effects of Glycyrrhiza glabra extract in mice: A possible role of monoamines. Neurosci Lett, 2013. 544: p. 110-4.
42. Clapcote, S.J., et al., Behavioral phenotypes of Disc1 missense mutations in mice. Neuron, 2007. 54(3): p. 387-402.
43. Olton, D.S., Mazes, maps, and memory. Am Psychol, 1979. 34(7): p. 583-96.
44. Walf, A.A. and C.A. Frye, The use of the elevated plus maze as an assay of anxiety-related behavior in rodents. Nat Protoc, 2007. 2(2): p. 322-8.
45. Can, A., et al., The mouse forced swim test. J Vis Exp, 2012(59): p. e3638.
46. Chu, T.T., Y. Liu, and E. Kemether, Thalamic transcriptome screening in three psychiatric states. J Hum Genet, 2009. 54(11): p. 665-75.
47. Kuzman, M.R., et al., Genome-wide expression analysis of peripheral blood identifies candidate biomarkers for schizophrenia. J Psychiatr Res, 2009. 43(13): p. 1073-7.
48. Ehrlichman, R.S., et al., Neuregulin 1 transgenic mice display reduced mismatch negativity, contextual fear conditioning and social interactions. Brain Res, 2009. 1294: p. 116-27.
49. Jones, C.A., D.J. Watson, and K.C. Fone, Animal models of schizophrenia. Br J Pharmacol, 2011. 164(4): p. 1162-94.
50. Kalueff, A.V., M. Wheaton, and D.L. Murphy, What's wrong with my mouse model? Advances and strategies in animal modeling of anxiety and depression. Behav Brain Res, 2007. 179(1): p. 1-18.
51. Sasson, N., et al., Orienting to social stimuli differentiates social cognitive impairment in autism and schizophrenia. Neuropsychologia, 2007. 45(11): p. 2580-8.
52. DS, R., CNS Receptor Partial Agonists: A New Approach to Drug Discovery. Primary Psychiatry, 2007.
53. French, D.P., Schizophrenic Psychology: New Research. 2006: Nova.
54. Silver, H., Selective serotonin re-uptake inhibitor augmentation in the treatment of negative symptoms of schizophrenia. Expert Opin Pharmacother, 2004. 5(10): p. 2053-8.
55. Schotte, A., et al., Occupancy of central neurotransmitter receptors by risperidone, clozapine and haloperidol, measured ex vivo by quantitative autoradiography. Brain Res, 1993. 631(2): p. 191-202.
56. Meltzer, H.Y., Clinical studies on the mechanism of action of clozapine: the dopamine-serotonin hypothesis of schizophrenia. Psychopharmacology (Berl), 1989. 99 Suppl: p. S18-27.
57. Harrison, P.J., Postmortem studies in schizophrenia. Dialogues Clin Neurosci, 2000. 2(4): p. 349-57.
58. Eggers, A.E., A serotonin hypothesis of schizophrenia. Med Hypotheses, 2013. 80(6): p. 791-4.
59. Arguello, P.A. and J.A. Gogos, Cognition in mouse models of schizophrenia susceptibility genes. Schizophr Bull, 2010. 36(2): p. 289-300.
60. Tandon, R., et al., Definition and description of schizophrenia in the DSM-5. Schizophr Res, 2013. 150(1): p. 3-10.
61. Keefe, R.S., Should cognitive impairment be included in the diagnostic criteria for schizophrenia? World Psychiatry, 2008. 7(1): p. 22-8.
62. MacCabe, J.H., et al., Superior intellectual ability in schizophrenia: neuropsychological characteristics. Neuropsychology, 2012. 26(2): p. 181-90.
63. Xi, D., et al., Group II metabotropic glutamate receptor agonist ameliorates MK801-induced dysfunction of NMDA receptors via the Akt/GSK-3beta pathway in adult rat prefrontal cortex. Neuropsychopharmacology, 2011. 36(6): p. 1260-74.
64. Neff, N.H., et al., Clozapine modulates aromatic L-amino acid decarboxylase activity in mouse striatum. J Pharmacol Exp Ther, 2006. 317(2): p. 480-7.
65. van Os, J., G. Kenis, and B.P. Rutten, The environment and schizophrenia. Nature, 2010. 468(7321): p. 203-12.
66. de Bruin, J.P., H.G. van Oyen, and N. Van de Poll, Behavioural changes following lesions of the orbital prefrontal cortex in male rats. Behav Brain Res, 1983. 10(2-3): p. 209-32.
67. Silva-Gomez, A.B., et al., Decreased dendritic spine density on prefrontal cortical and hippocampal pyramidal neurons in postweaning social isolation rats. Brain Res, 2003. 983(1-2): p. 128-36.

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