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研究生:黃元麒
研究生(外文):Yuan-Chi Huang
論文名稱:由肝免疫生物學觀點探討肝疾病藥物的開發平台
論文名稱(外文):Study on the platform development for therapeutics of liver disease involving liver immunobiology
指導教授:吳榮燦
指導教授(外文):Rong-Tsun Wu
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:生物藥學研究所
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
畢業學年度:97
語文別:中文
論文頁數:90
中文關鍵詞:肝臟脂多醣發炎細胞自噬鹼性纖維母細胞生長因子
外文關鍵詞:liverLPSinflammationautophagybFGF
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肝臟長期暴露在無害抗原環境下,形成特殊的免疫耐受(tolerance)狀態。經由耐受和免疫的動態平衡,反應出生理和病理的情況。在眾多肝臟相關疾病當中,有許多與脂多醣(lipopolysaccharides 〔 LPS〕)有關。 肝臟被認為是主要清除並阻止LPS進入體內循環的最後屏障。此外,肝細胞(hepatocytes)內細胞自噬(autophagy)的機制參與在維護肝臟的正常功能並幫助清除氧化壓力。
本研究主要建立肝臟發炎以及促進autophagy的藥物篩選平台。利用動物體內分離的hepatocytes或細胞株和肝臟切片(liver slices),以LPS處理誘發發炎反應,建立藥物篩選的平台。在眾多中草藥粗萃物中,觀察到蒲公英及椰子水萃取物,能夠有效降低一氧化氮(NO)產生及腫瘤壞死因子(TNF-α)基因的表現,抑制hepatocytes的發炎反應。而透過注射LPS誘發發炎反應的動物模式中,也確定篩選出藥物的療效。另一方面,利用hepatocytes以各種不同中草藥粗萃物處理,運用溶酶體染色的方式,初步篩選可能會促進autophagy的藥物。其中觀察到牛蒡有促進細胞溶酶體活化的功效。而且以autophagy專一性的生物標記(LC3)也證實牛蒡會促進autophagy。此外,在LPS與二甲基亞硝酸胺(DMN)誘發的發炎動物模式中,觀察到鹼性纖維母細胞生長因子(bFGF)大量增加的現象。由於先前實驗室中,以低濃度的微脂粒包裹沙利竇邁(thalidomide)為工具,發現thalidomide抑制癌症的功效是透過對bFGF的作用。為驗證是否thalidomide的免疫調節功效也是經由相同的途徑,在細胞實驗中,利用LPS引發發炎反應,再外給予bFGF後,發炎情形更趨嚴重。而利用抗體阻斷bFGF的作用,也發現發炎的情況有趨緩的現象。證明bFGF可能參與在發炎反應中。未來可利用基因剔除等技術,進一步確認bFGF在發炎反應中扮演的角色。
由本研究可知蒲公英和椰子水萃取物可以有效降低肝臟的發炎相關反應,而牛蒡則可以增進hepatocytes的autophagy,可能可以避免肝藏受損。此外本研究也發現bFGF可能參與在發炎反應的發生中。
The liver maintains in a special condition, called “tolerance”. The pathology and physiology of the liver are dependent on balance between tolerance and immunity. There are many liver diseases involving in lipopolysaccharides (LPS). The liver is the major site of LPS clearance. In addition, the autophagy is involving in clearance and protection liver form oxidative stress.
In this study, a drug-screening platform for anti-inflammation and autophagy inducer in liver is developed. First, the primary hepatocytes and cell lines are treated with LPS. Then the liver slices which maintain the correct architectural relationship of all cells present are applyed. In the present study, the extract of Taraxacum mongolicum and Cocos nucifera can reduce the LPS-induced inflammation significantly. The effects of Cocos nucifera on the acute liver inflammation in mice induced by LPS/D-galactosamine (D-GalN) are then investigated. The results showed that Cocos nucifera protect against LPS/D-GalN-induced liver injury, including decrease of ALT/AST release and the expression of TNF-α. In addition, MDC staining of hepatocytes are used for screening drugs that increase autophagy. Arctium lappa increased tendency for MDC staining, suggesting increased lysosomal and/or autophagy activity. The conversion of LC3-I to LC3-II is significantly increased, confirming the effect on inducing autophagy. In addition, the expression of bFGF is high in LPS/D-GalN-induced inflammation and in LPS-treated hepatocytes. Thalidomide has the property for reducing this inflammation. In the previous study, thalidomide (at 0.1 μg/ml) can reduce cellular bFGF levels and affect tumor anchorage-independent growth. Whether the immunomodulatory effect of thalidomide is also through decreasing bFGF pathway? In the study, the bFGF can increase the LPS-induce inflammation. When neutralizing antibodies are used to block bFGF pathway, the expression of TNF-α and production of NO are reduced.
In conclusion, the extract of Taraxacum mongolicum and Cocos nucifera can reduce LPS-induced liver inflammation. Furthermore, bFGF may play a role in inflammation.
中文摘要 i
英文摘要 ii
總目錄 iii
圖表目錄 iv
縮寫檢索表 vi
緒論 1
材料方法 15
結果 30
討論 41
參考文獻 47
圖表 54
1. Crispe, I.N., Hepatic T cells and liver tolerance. Nat Rev Immunol, 2003. 3(1): p. 51-62.
2. Crispe, I.N., et al., Cellular and molecular mechanisms of liver tolerance. Immunol Rev, 2006. 213: p. 101-18.
3. Lumsden, A.B., J.M. Henderson, and M.H. Kutner, Endotoxin levels measured by a chromogenic assay in portal, hepatic and peripheral venous blood in patients with cirrhosis. Hepatology, 1988. 8(2): p. 232-6.
4. You, Q., et al., Mechanism of T cell tolerance induction by murine hepatic Kupffer cells. Hepatology, 2008. 48(3): p. 978-90.
5. Karrar, A., et al., Human liver sinusoidal endothelial cells induce apoptosis in activated T cells: a role in tolerance induction. Gut, 2007. 56(2): p. 243-52.
6. Mathison, J.C. and R.J. Ulevitch, The clearance, tissue distribution, and cellular localization of intravenously injected lipopolysaccharide in rabbits. J Immunol, 1979. 123(5): p. 2133-43.
7. Su, G.L., Lipopolysaccharides in liver injury: molecular mechanisms of Kupffer cell activation. Am J Physiol Gastrointest Liver Physiol, 2002. 283(2): p. G256-65.
8. Scott, M.J., et al., Endotoxin uptake in mouse liver is blocked by endotoxin pretreatment through a suppressor of cytokine signaling-1-dependent mechanism. Hepatology, 2009.
9. Vodovotz, Y., et al., Inflammatory modulation of hepatocyte apoptosis by nitric oxide: in vivo, in vitro, and in silico studies. Curr Mol Med, 2004. 4(7): p. 753-62.
10. Hibbs JB Jr, T.R., Vavrin Z, Granger DL, Drapier JC, Amber IJ, Lancaster LR, Synthesis of nitric oxide from a terminal guanidino nitrogen atom of L-arginine: a molecular mechanism regulating cellular proliferation that target intracellular iron. The Netherlands: Elsevier, 1990: p. 189-223.
11. Ding, A.H., C.F. Nathan, and D.J. Stuehr, Release of reactive nitrogen intermediates and reactive oxygen intermediates from mouse peritoneal macrophages. Comparison of activating cytokines and evidence for independent production. J Immunol, 1988. 141(7): p. 2407-12.
12. Laskin, J.D., et al., Prooxidant and antioxidant functions of nitric oxide in liver toxicity. Antioxid Redox Signal, 2001. 3(2): p. 261-71.
13. Joshi, M.S., J.L. Ponthier, and J.R. Lancaster, Jr., Cellular antioxidant and pro-oxidant actions of nitric oxide. Free Radic Biol Med, 1999. 27(11-12): p. 1357-66.
14. Kim, P.K. and T.R. Billiar, Give me iNOS or give me death. Hepatology, 2001. 34(2): p. 436-7.
15. Wajant, H., K. Pfizenmaier, and P. Scheurich, Tumor necrosis factor signaling. Cell Death Differ, 2003. 10(1): p. 45-65.
16. Grell, M., et al., Induction of cell death by tumour necrosis factor (TNF) receptor 2, CD40 and CD30: a role for TNF-R1 activation by endogenous membrane-anchored TNF. EMBO J, 1999. 18(11): p. 3034-43.
17. Schwabe, R.F. and D.A. Brenner, Mechanisms of Liver Injury. I. TNF-alpha-induced liver injury: role of IKK, JNK, and ROS pathways. Am J Physiol Gastrointest Liver Physiol, 2006. 290(4): p. G583-9.
18. Brun, P., et al., Increased intestinal permeability in obese mice: new evidence in the pathogenesis of nonalcoholic steatohepatitis. Am J Physiol Gastrointest Liver Physiol, 2007. 292(2): p. G518-25.
19. Everson, C.A., Clinical assessment of blood leukocytes, serum cytokines, and serum immunoglobulins as responses to sleep deprivation in laboratory rats. Am J Physiol Regul Integr Comp Physiol, 2005. 289(4): p. R1054-63.
20. Everson, C.A. and L.A. Toth, Systemic bacterial invasion induced by sleep deprivation. Am J Physiol Regul Integr Comp Physiol, 2000. 278(4): p. R905-16.
21. Wang, D., et al., Cardiovascular hazard and non-steroidal anti-inflammatory drugs. Curr Opin Pharmacol, 2005. 5(2): p. 204-10.
22. O'Neill, L.A., Targeting signal transduction as a strategy to treat inflammatory diseases. Nat Rev Drug Discov, 2006. 5(7): p. 549-63.
23. Haslett, P.A., et al., Thalidomide costimulates primary human T lymphocytes, preferentially inducing proliferation, cytokine production, and cytotoxic responses in the CD8+ subset. J Exp Med, 1998. 187(11): p. 1885-92.
24. Moreira, A.L., et al., Thalidomide exerts its inhibitory action on tumor necrosis factor alpha by enhancing mRNA degradation. J Exp Med, 1993. 177(6): p. 1675-80.
25. Tseng, S., et al., Rediscovering thalidomide: a review of its mechanism of action, side effects, and potential uses. J Am Acad Dermatol, 1996. 35(6): p. 969-79.
26. Delrieu, I., The high molecular weight isoforms of basic fibroblast growth factor (FGF-2): an insight into an intracrine mechanism. FEBS Lett, 2000. 468(1): p. 6-10.
27. Cotton, L.M., M.K. O'Bryan, and B.T. Hinton, Cellular signaling by fibroblast growth factors (FGFs) and their receptors (FGFRs) in male reproduction. Endocrine Reviews, 2008. 29(2): p. 193-216.
28. Wang, F. and W.L. McKeehan, The fibroblast growth factor (FGF) signaling complex. Handbook of Cell Signaling, 2003.
29. Mei, S.C. and R.T. Wu, The G-rich promoter and G-rich coding sequence of basic fibroblast growth factor are the targets of thalidomide in glioma. Molecular Cancer Therapeutics, 2008. 7(8): p. 2405-14.
30. Muddasani, P., et al., Basic fibroblast growth factor activates the MAPK and NFkappaB pathways that converge on Elk-1 to control production of matrix metalloproteinase-13 by human adult articular chondrocytes. J Biol Chem, 2007. 282(43): p. 31409-21.
31. Tang, C., et al., Basic fibroblast growth factor stimulates fibronectin expression through phospholipase C gamma, protein kinase C alpha, c-Src, NF-kappaB, and p300 pathway in osteoblasts. J. Cell. Physiol., 2007. 211(1): p. 45-55.
32. Herseth, J., et al., IL-1beta differently involved in IL-8 and FGF-2 release in crystalline silica-treated lung cell co-cultures. Part Fibre Toxicol, 2008. 5: p. 16.
33. Kanazawa, S., et al., VEGF, basic-FGF, and TGF-beta in Crohn's disease and ulcerative colitis: a novel mechanism of chronic intestinal inflammation. Am J Gastroenterol, 2001. 96(3): p. 822-8.
34. Nagashima, M., et al., Effects of combinations of anti-rheumatic drugs on the production of vascular endothelial growth factor and basic fibroblast growth factor in cultured synoviocytes and patients with rheumatoid arthritis. Rheumatology (Oxford), 2000. 39(11): p. 1255-62.
35. Zhang, H. and A.C. Issekutz, Growth factor regulation of neutrophil-endothelial cell interactions. J Leukoc Biol, 2001. 70(2): p. 225-32.
36. Zittermann, S.I. and A.C. Issekutz, Basic fibroblast growth factor (bFGF, FGF-2) potentiates leukocyte recruitment to inflammation by enhancing endothelial adhesion molecule expression. Am J Pathol, 2006. 168(3): p. 835-46.
37. Thiele, A., et al., Cytokine modulation and suppression of liver injury by a novel analogue of thalidomide. Eur J Pharmacol, 2002. 453(2-3): p. 325-34.
38. Fernández-Martínez, E., et al., Immunomodulatory effects of thalidomide analogs on LPS-induced plasma and hepatic cytokines in the rat. Biochemical Pharmacology, 2004. 68(7): p. 1321-9.
39. Chong, L., et al., Anti-fibrotic effects of thalidomide on hepatic stellate cells and dimethylnitrosamine-intoxicated rats. J Biomed Sci, 2006. 13(3): p. 403-18.
40. Antifibrotic effects of a herbal combination regimen on hepatic fibrotic rats. 2007: p. 8.
41. Schmid, D. and C. Munz, Innate and Adaptive Immunity through Autophagy. Immunity, 2007. 27(1): p. 11-21.
42. Yin, X., W. Ding, and W. Gao, Autophagy in the liver. Hepatology, 2008. 47(5): p. 1773-85.
43. Perlmutter, D.H., The role of autophagy in alpha-1-antitrypsin deficiency: a specific cellular response in genetic diseases associated with aggregation-prone proteins. Autophagy, 2006. 2(4): p. 258-63.
44. Ding, W. and X. Yin, Sorting, recognition and activation of the misfolded protein degradation pathways through macroautophagy and the proteasome. Autophagy, 2008. 4(2): p. 141-50.
45. Richardson, S.D., et al., Occurrence, genotoxicity, and carcinogenicity of regulated and emerging disinfection by-products in drinking water: a review and roadmap for research. Mutat Res, 2007. 636(1-3): p. 178-242.
46. Schümann, J. and G. Tiegs, Pathophysiological mechanisms of TNF during intoxication with natural or man-made toxins. Toxicology, 1999. 138(2): p. 103-26.
47. Dybing, E., et al., Risk assessment of dietary exposures to compounds that are genotoxic and carcinogenic--an overview. Toxicology Letters, 2008. 180(2): p. 110-7.
48. Schook, L.B., et al., Dimethylnitrosamine (DMN)-induced IL-1 beta, TNF-alpha, and IL-6 inflammatory cytokine expression. Toxicol Appl Pharmacol, 1992. 116(1): p. 110-6.
49. Corcoran, G.B. and S.D. Ray, The role of the nucleus and other compartments in toxic cell death produced by alkylating hepatotoxicants. Toxicol Appl Pharmacol, 1992. 113(2): p. 167-83.
50. Josephy, P.D., H.L. Lord, and V.A. Snieckus, Dimethylnitrosamine genotoxicity: does N-acetyltransferase activity play a role? Carcinogenesis, 1994. 15(3): p. 479-82.
51. Virgin, H.W. and B. Levine, Autophagy genes in immunity. Nat Immunol, 2009. 10(5): p. 461-70.
52. Xu, Y., et al., Toll-like receptor 4 is a sensor for autophagy associated with innate immunity. Immunity, 2007. 27(1): p. 135-44.
53. Saitoh, T., et al., Loss of the autophagy protein Atg16L1 enhances endotoxin-induced IL-1beta production. Nature, 2008. 456(7219): p. 264-8.
54. Nedjic, J., et al., Autophagy in thymic epithelium shapes the T-cell repertoire and is essential for tolerance. Nature, 2008. 455(7211): p. 396-400.
55. Cadwell, K., et al., A key role for autophagy and the autophagy gene Atg16l1 in mouse and human intestinal Paneth cells. Nature, 2008. 456(7219): p. 259-63.
56. Mazier, D., et al., Complete development of hepatic stages of Plasmodium falciparum in vitro. Science, 1985. 227(4685): p. 440-2.
57. Liu, S., et al., Role of toll-like receptors in changes in gene expression and NF-kappa B activation in mouse hepatocytes stimulated with lipopolysaccharide. Infect Immun, 2002. 70(7): p. 3433-42.
58. Schmelz, M., V.J. Schmid, and A.R. Parrish, Selective disruption of cadherin/catenin complexes by oxidative stress in precision-cut mouse liver slices. Toxicol Sci, 2001. 61(2): p. 389-94.
59. Moronvalle-Halley, V., et al., Evaluation of cultured, precision-cut rat liver slices as a model to study drug-induced liver apoptosis. Toxicology, 2005. 207(2): p. 203-14.
60. Olinga, P., et al., Comparison of five incubation systems for rat liver slices using functional and viability parameters. Journal of pharmacological and toxicological methods, 1997. 38(2): p. 59-69.
61. Schmieder, P., et al., Optimization of a precision-cut trout liver tissue slice assay as a screen for vitellogenin induction: comparison of slice incubation techniques. Aquat Toxicol, 2000. 49(4): p. 251-268.
62. Wroblewski, F. and J.S. Ladue, Lactic dehydrogenase activity in blood. Proc Soc Exp Biol Med, 1955. 90(1): p. 210-3.
63. Obatomi, D.K., et al., Optimizing preincubation conditions for precision-cut rat kidney and liver tissue slices: effect of culture media and antioxidants. Toxicology in Vitro, 1998. 12(6): p. 725-737.
64. Evdokimova, E., H. Taper, and P.B. Calderon, Effects of bacterial endotoxin (lipopolysaccharides) on survival and metabolism of cultured precision-cut rat liver slices. Toxicology in vitro : an international journal published in association with BIBRA, 2002. 16(1): p. 47-54.
65. Misko, T.P., et al., A fluorometric assay for the measurement of nitrite in biological samples. Anal Biochem, 1993. 214(1): p. 11-6.
66. Munafo, D.B. and M.I. Colombo, A novel assay to study autophagy: regulation of autophagosome vacuole size by amino acid deprivation. J Cell Sci, 2001. 114(Pt 20): p. 3619-29.
67. Yamada, I., et al., Mao (Ephedra sinica Stapf) protects against D-galactosamine and lipopolysaccharide-induced hepatic failure. Cytokine, 2008. 41(3): p. 293-301.
68. Huang, C.H., et al., Chinese herb Radix Polygoni Multiflori as a therapeutic drug for liver cirrhosis in mice. Journal of Ethnopharmacology, 2007. 114(2): p. 199-206.
69. Ewaschuk, J., et al., Probiotic bacteria prevent hepatic damage and maintain colonic barrier function in a mouse model of sepsis. Hepatology, 2007. 46(3): p. 841-50.
70. Annane, D., et al., Corticosteroids in the treatment of severe sepsis and septic shock in adults: a systematic review. JAMA, 2009. 301(22): p. 2362-75.
71. Eriksson, T., et al., Enantiomers of thalidomide: blood distribution and the influence of serum albumin on chiral inversion and hydrolysis. Chirality, 1998. 10(3): p. 223-8.
72. Lu, J., et al., Metabolism of thalidomide in liver microsomes of mice, rabbits, and humans. J Pharmacol Exp Ther, 2004. 310(2): p. 571-7.
73. Arrieta, O., et al., Protective effect of pentoxifylline plus thalidomide against septic shock in mice. International journal of experimental pathology, 1999. 80(1): p. 11-6.
74. Yamashita, A., et al., Fibroblast growth factor-2 determines severity of joint disease in adjuvant-induced arthritis in rats. J Immunol, 2002. 168(1): p. 450-7.
75. Meij, J.T., et al., Exacerbation of myocardial injury in transgenic mice overexpressing FGF-2 is T cell dependent. Am J Physiol Heart Circ Physiol, 2002. 282(2): p. H547-55.
76. Huang, C.H., et al., Chinese herb Radix Polygoni Multiflori as a therapeutic drug for liver cirrhosis in mice. J Ethnopharmacol, 2007. 114(2): p. 199-206.
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