系統識別號 U0026-2805202011021300
論文名稱(中文) 探討氯化鈣與窄縮共同處理所誘發的大鼠腹主動脈瘤中血管對於擾流壓力的應對
論文名稱(英文) Investigating vascular responses against hemodynamic stress in the calcium chloride and coarctation co-treatment-induced rat abdominal aortic aneurysm model
校院名稱 成功大學
系所名稱(中) 細胞生物與解剖學研究所
系所名稱(英) Institute of Cell Biology and Anatomy
學年度 108
學期 2
出版年 109
研究生(中文) 林冠呈
研究生(英文) Guan-Cheng Lin
學號 T96994024
學位類別 碩士
語文別 英文
論文頁數 33頁
口試委員 指導教授-江美治
中文關鍵字 腹主動脈瘤  血管窄化  血管平滑肌細胞收縮力  粘著斑 
英文關鍵字 Abdominal aortic aneurysm  Aortic coarctation  Vascular smooth muscle cell contractility  Focal adhesion 
中文摘要 腹主動脈瘤是一種主動脈末端的退化性疾病。經過數十年的研究,目前尚未發展出有效的藥物治療。因此,學界需要嶄新的觀點來研發更好的診斷指引與療法。目前已知血管平滑肌細胞的失能是胸主動脈瘤的主要致病機轉,但是血管平滑肌細胞的收縮與放鬆在腹主動脈瘤的病理變化中的角色仍然不清楚。本研究團隊先前建立了以血管窄化誘發腹主動脈瘤的蘭嶼迷你豬動物模式,此模式與人類腹主動脈瘤有類似的組織病變,但後續發展卻經常受限,主因是缺乏合適的試劑與儀器以進行分子生物學與生物力學分析。因此,我們嘗試以血管窄化加上氯化鈣共同處理(簡稱共處理)在Sprague-Dawley大鼠上誘發腹主動脈瘤。在評估氯化鈣單獨處理與共處理的劑量反應實驗中,我們發現了共處理組的主動脈管腔周長隨著劑量增加而逐漸增加。在0.5M共處理組之中,60%的血管管壁呈現明顯的彈性纖維崩壞與厚度不均,而其餘40%的血管結構有輕微損壞與顯著的管壁增厚。為了探討血管膨大和血管硬度之間的關聯,我們定期測量Circumferential strain,它是血管硬度的一個反向指標。在術後兩週,所有實驗組別都被偵測到血管硬化,但是術後十二與十六週所測到的Circumferential strain與血管膨大的程度並無關聯。接著,我們檢視血管內血壓是否能作為血管結構變化的指標。雖然血壓在血管窄化當下顯著降低,實驗結束時的血壓在膨大的血管與對照組之間並無差別。為了評估血管平滑肌細胞的功能,我們測量動脈瘤血管的離體等長收縮力。在共處理組,我們發現以甲型腎上腺素受體致活劑所刺激的最大收縮力有顯著下降,且某些共處理組表現出內皮細胞功能缺損的現象。此外,我們偵測到動脈瘤血管的最大管壁張力有明顯下降。近期的報告指出由Src所調控的Focal adhesion訊息傳遞會影響血管平滑肌的收縮力與血管硬度。因此,我們檢視了Src抑制劑PP2對於甲型腎上腺素受體致活劑所刺激的等長收縮力的影響。相較於對照組,PP2的前處理對動脈瘤血管的收縮力有更大的抑制效果,此現象暗示了Focal adhesion訊息傳遞鏈與收縮力激發機制之間的脫鉤。總結來說,這些結果提供了第一個在大鼠腹主動脈瘤模式中有血管平滑肌收縮功能異常的離體證據,也意味著Focal adhesion相關蛋白質具有成為治療標靶的潛力。
英文摘要 Abdominal aortic aneurysm (AAA) is a degenerative disease of the terminal aorta. After decades of research, effective drug therapy is still not available, pointing to the need for novel concepts to develop better diagnostic metrics and therapeutics. Loss of functional vascular smooth muscle cells (VSMC) is the primary underlying mechanism of thoracic aortic aneurysm. In contrast, the roles of VSMC contraction and relaxation in AAA pathogenesis remain elusive. Previously, we established a coarctation-induced AAA model in Lanyu minipigs, which shared histopathology with human AAA but was often limited by lack of analytic tools for molecular and biomechanical investigations. Therefore, we tried to induce AAA in Sprague-Dawley rat by combining aortic coarctation and calcium chloride treatment (co-treated). The dose response (0.15M, 0.25M, and 0.5M) of calcium chloride alone and co-treatment was evaluated. Dose-dependent increase of lumen perimeter was found in co-treated groups. In the 0.5M CaCl2 co-treated group, 60% exhibited prominent elastin disruption with heterogeneous thickness of media, whereas the other 40% showed moderate damage of elastic lamellae with markedly thickened media. To examine the relationship between aortic dilation and aortic stiffness, we monitored temporal changes of circumferential strain, a negative index of aortic stiffness. Aortic stiffening was found in all experimental groups at 2 weeks post-surgery, but no correlation was found between lumen dilation and circumferential strain at 12 or 16 weeks post-surgery. Next, we measured intravascular blood pressure (BP) as a potential marker of aortic structural change. In spite of significant decline of BP immediately following aortic coarctation, BP was restored to similar levels in both dilated and non-dilated aortas. To assess VSMC function, we measured ex vivo isometric force in aneurysmal aorta. Pronounced decrease of maximal force stimulated by α1-adrenoreceptor agonist phenylephrine was found in the co-treated group, and part of the co-treated group displayed endothelial dysfunction. Moreover, marked decreases of maximum wall tension were detected in the aneurysmal segment. Recently, focal adhesion (FA) signaling mediated by tyrosine kinase Src was shown to regulate VSMC force production and aortic stiffness. Therefore, we examined the effect of PP2, a Src inhibitor, on phenylephrine stimulated isometric force. PP2 pretreatment led to greater inhibition of contractility in aneurysmal aortas than their counterpart of sham group, suggesting uncoupling of FA signaling to force development. To sum up, these results provide the first ex vivo evidence for VSMC contractile dysfunction in a rat AAA model and suggest that FA-related proteins may be potential AAA therapeutic target.
論文目次 Abstract------------------ii
Chinese abstract----------iv
Materials and Methods-----4
Table and Figures---------24
參考文獻 1. Bogunovic N, Meekel JP, Micha D, Blankensteijn JD, Hordijk PL, and Yeung KK. Impaired smooth muscle cell contractility as a novel concept of abdominal aortic aneurysm pathophysiology. Scientific reports 9: 6837, 2019.
2. Chen hsuan Liu YhH, Wen fa Chang, Chih ping Chu, Hsiu chen Wang. An Atlas and Manual of Histopathological Staining Methods: Animal Technology Laboratories, Agricultural Technology Research Institute, 1996.
3. Daugherty A, Rateri DL, Charo IF, Owens AP, Howatt DA, and Cassis LA. Angiotensin II infusion promotes ascending aortic aneurysms: attenuation by CCR2 deficiency in apoE-/- mice. Clinical science (London, England : 1979) 118: 681-689, 2010.
4. Eliason JL, Hannawa KK, Ailawadi G, Sinha I, Ford JW, Deogracias MP, Roelofs KJ, Woodrum DT, Ennis TL, Henke PK, Stanley JC, Thompson RW, and Upchurch GR, Jr. Neutrophil depletion inhibits experimental abdominal aortic aneurysm formation. Circulation 112: 232-240, 2005.
5. Emeto TI, Moxon JV, Au M, and Golledge J. Oxidative stress and abdominal aortic aneurysm: potential treatment targets. Clinical science (London, England : 1979) 130: 301-315, 2016.
6. Galle C, Schandene L, Stordeur P, Peignois Y, Ferreira J, Wautrecht JC, Dereume JP, and Goldman M. Predominance of type 1 CD4+ T cells in human abdominal aortic aneurysm. Clinical and experimental immunology 142: 519-527, 2005.
7. Gao L, Siu KL, Chalupsky K, Nguyen A, Chen P, Weintraub NL, Galis Z, and Cai H. Role of uncoupled endothelial nitric oxide synthase in abdominal aortic aneurysm formation: treatment with folic acid. Hypertension 59: 158-166, 2012.
8. Gao YZ, Saphirstein RJ, Yamin R, Suki B, and Morgan KG. Aging impairs smooth muscle-mediated regulation of aortic stiffness: a defect in shock absorption function? American journal of physiology Heart and circulatory physiology 307: H1252-1261, 2014.
9. Golledge J, Norman PE, Murphy MP, and Dalman RL. Challenges and opportunities in limiting abdominal aortic aneurysm growth. Journal of vascular surgery 65: 225-233, 2017.
10. Guo DC, Pannu H, Tran-Fadulu V, Papke CL, Yu RK, Avidan N, Bourgeois S, Estrera AL, Safi HJ, Sparks E, Amor D, Ades L, McConnell V, Willoughby CE, Abuelo D, Willing M, Lewis RA, Kim DH, Scherer S, Tung PP, Ahn C, Buja LM, Raman CS, Shete SS, and Milewicz DM. Mutations in smooth muscle alpha-actin (ACTA2) lead to thoracic aortic aneurysms and dissections. Nature genetics 39: 1488-1493, 2007.
11. Ho YC, Wu ML, Gung PY, Chen CH, Kuo CC, and Yet SF. Heme oxygenase-1 deficiency exacerbates angiotensin II-induced aortic aneurysm in mice. Oncotarget, 2016.
12. Huang J, Davis EC, Chapman SL, Budatha M, Marmorstein LY, Word RA, and Yanagisawa H. Fibulin-4 deficiency results in ascending aortic aneurysms: a potential link between abnormal smooth muscle cell phenotype and aneurysm progression. Circulation research 106: 583-592, 2010.
13. Huang J, Yamashiro Y, Papke CL, Ikeda Y, Lin Y, Patel M, Inagami T, Le VP, Wagenseil JE, and Yanagisawa H. Angiotensin-converting enzyme-induced activation of local angiotensin signaling is required for ascending aortic aneurysms in fibulin-4-deficient mice. Science translational medicine 5: 183ra158, 181-111, 2013.
14. Jespersen B, Knupp L, and Northcott CA. Femoral Arterial and Venous Catheterization for Blood Sampling, Drug Administration and Conscious Blood Pressure and Heart Rate Measurements. e3496, 2012.
15. Jespersen B, Tykocki NR, Watts SW, and Cobbett PJ. Measurement of Smooth Muscle Function in the Isolated Tissue Bath-applications to Pharmacology Research. e52324, 2015.
16. Kent KC, Zwolak RM, Egorova NN, Riles TS, Manganaro A, Moskowitz AJ, Gelijns AC, and Greco G. Analysis of risk factors for abdominal aortic aneurysm in a cohort of more than 3 million individuals. Journal of vascular surgery 52: 539-548, 2010.
17. Kuzmik GA, Sang AX, and Elefteriades JA. Natural history of thoracic aortic aneurysms. Journal of vascular surgery 56: 565-571, 2012.
18. Lederle FA, Johnson GR, Wilson SE, Chute EP, Hye RJ, Makaroun MS, Barone GW, Bandyk D, Moneta GL, and Makhoul RG. The aneurysm detection and management study screening program: validation cohort and final results. Aneurysm Detection and Management Veterans Affairs Cooperative Study Investigators. Archives of internal medicine 160: 1425-1430, 2000.
19. Li H, Bai S, Ao Q, Wang X, Tian X, Li X, Tong H, Hou W, and Fan J. Modulation of Immune-Inflammatory Responses in Abdominal Aortic Aneurysm: Emerging Molecular Targets. Journal of immunology research 2018: 7213760, 2018.
20. Lin PY, Wu YT, Lin GC, Shih YH, Sampilvanjil A, Chen LR, Yang YJ, Wu HL, and Jiang MJ. Coarctation-induced degenerative abdominal aortic aneurysm in a porcine model. Journal of vascular surgery 57: 806-815.e801, 2013.
21. Lindeman JHN, Ashcroft BA, Beenakker J-WM, van Es M, Koekkoek NBR, Prins FA, Tielemans JF, Abdul-Hussien H, Bank RA, and Oosterkamp TH. Distinct defects in collagen microarchitecture underlie vessel-wall failure in advanced abdominal aneurysms and aneurysms in Marfan syndrome. Proceedings of the National Academy of Sciences 107: 862-865, 2010.
22. Liu B, Granville DJ, Golledge J, and Kassiri Z. Pathogenic mechanisms and the potential of drug therapies for aortic aneurysm. American Journal of Physiology-Heart and Circulatory Physiology 318: H652-H670, 2020.
23. Longo GM, Xiong W, Greiner TC, Zhao Y, Fiotti N, and Baxter BT. Matrix metalloproteinases 2 and 9 work in concert to produce aortic aneurysms. J Clin Invest 110: 625-632, 2002.
24. Maiellaro-Rafferty K, Weiss D, Joseph G, Wan W, Gleason RL, and Taylor WR. Catalase overexpression in aortic smooth muscle prevents pathological mechanical changes underlying abdominal aortic aneurysm formation. American Journal of Physiology - Heart and Circulatory Physiology 301: H355-H362, 2011.
25. McCormick ML, Gavrila D, and Weintraub NL. Role of oxidative stress in the pathogenesis of abdominal aortic aneurysms. Arterioscler Thromb Vasc Biol 27: 461-469, 2007.
26. Min J, Reznichenko M, Poythress RH, Gallant CM, Vetterkind S, Li Y, and Morgan KG. Src modulates contractile vascular smooth muscle function via regulation of focal adhesions. Journal of cellular physiology 227: 3585-3592, 2012.
27. Moehle CW, Bhamidipati CM, Alexander MR, Mehta GS, Irvine JN, Salmon M, Upchurch GR, Jr., Kron IL, Owens GK, and Ailawadi G. Bone marrow-derived MCP1 required for experimental aortic aneurysm formation and smooth muscle phenotypic modulation. The Journal of thoracic and cardiovascular surgery 142: 1567-1574, 2011.
28. Morgan S, Yamanouchi D, Harberg C, Wang Q, Keller M, Si Y, Burlingham W, Seedial S, Lengfeld J, and Liu B. Elevated protein kinase C-delta contributes to aneurysm pathogenesis through stimulation of apoptosis and inflammatory signaling. Arterioscler Thromb Vasc Biol 32: 2493-2502, 2012.
29. Nicholson CJ, Singh K, Saphirstein RJ, Gao YZ, Li Q, Chiu JG, Leavis P, Verwoert GC, Mitchell GF, Porter T, and Morgan KG. Reversal of Aging-Induced Increases in Aortic Stiffness by Targeting Cytoskeletal Protein-Protein Interfaces. J Am Heart Assoc 7, 2018.
30. Parastatidis I, Weiss D, Joseph G, and Taylor WR. Overexpression of catalase in vascular smooth muscle cells prevents the formation of abdominal aortic aneurysms. Arterioscler Thromb Vasc Biol 33: 2389-2396, 2013.
31. Pyo R, Lee JK, Shipley JM, Curci JA, Mao D, Ziporin SJ, Ennis TL, Shapiro SD, Senior RM, and Thompson RW. Targeted gene disruption of matrix metalloproteinase-9 (gelatinase B) suppresses development of experimental abdominal aortic aneurysms. J Clin Invest 105: 1641-1649, 2000.
32. Qin Y, Cao X, Guo J, Zhang Y, Pan L, Zhang H, Li H, Tang C, Du J, and Shi GP. Deficiency of cathepsin S attenuates angiotensin II-induced abdominal aortic aneurysm formation in apolipoprotein E-deficient mice. Cardiovascular research 96: 401-410, 2012.
33. Raaz U, Zollner AM, Schellinger IN, Toh R, Nakagami F, Brandt M, Emrich FC, Kayama Y, Eken S, Adam M, Maegdefessel L, Hertel T, Deng A, Jagger A, Buerke M, Dalman RL, Spin JM, Kuhl E, and Tsao PS. Segmental Aortic Stiffening Contributes to Experimental Abdominal Aortic Aneurysm Development. Circulation, 2015.
34. Renard M, Holm T, Veith R, Callewaert BL, Ades LC, Baspinar O, Pickart A, Dasouki M, Hoyer J, Rauch A, Trapane P, Earing MG, Coucke PJ, Sakai LY, Dietz HC, De Paepe AM, and Loeys BL. Altered TGFbeta signaling and cardiovascular manifestations in patients with autosomal recessive cutis laxa type I caused by fibulin-4 deficiency. European journal of human genetics : EJHG 18: 895-901, 2010.
35. Romary DJ, Berman AG, and Goergen CJ. High-frequency murine ultrasound provides enhanced metrics of BAPN-induced AAA growth. American journal of physiology Heart and circulatory physiology 317: H981-h990, 2019.
36. Sakalihasan N, Michel J-B, Katsargyris A, Kuivaniemi H, Defraigne J-O, Nchimi A, Powell JT, Yoshimura K, and Hultgren R. Abdominal aortic aneurysms. Nature Reviews Disease Primers 4: 34, 2018.
37. Salmon M, Johnston WF, Woo A, Pope NH, Su G, Upchurch GR, Jr., Owens GK, and Ailawadi G. KLF4 regulates abdominal aortic aneurysm morphology and deletion attenuates aneurysm formation. Circulation 128: S163-174, 2013.
38. Saphirstein RJ, Gao YZ, Jensen MH, Gallant CM, Vetterkind S, Moore JR, and Morgan KG. The focal adhesion: a regulated component of aortic stiffness. PLoS One 8: e62461, 2013.
39. Saphirstein RJ, Gao YZ, Lin QQ, and Morgan KG. Cortical actin regulation modulates vascular contractility and compliance in veins. The Journal of physiology 593: 3929-3941, 2015.
40. Sehgel NL, Sun Z, Hong Z, Hunter WC, Hill MA, Vatner DE, Vatner SF, and Meininger GA. Augmented vascular smooth muscle cell stiffness and adhesion when hypertension is superimposed on aging. Hypertension 65: 370-377, 2015.
41. Sehgel NL, Zhu Y, Sun Z, Trzeciakowski JP, Hong Z, Hunter WC, Vatner DE, Meininger GA, and Vatner SF. Increased vascular smooth muscle cell stiffness: a novel mechanism for aortic stiffness in hypertension. American journal of physiology Heart and circulatory physiology 305: H1281-1287, 2013.
42. Shen M, Lee J, Basu R, Sakamuri SS, Wang X, Fan D, and Kassiri Z. Divergent Role of Matrix Metalloproteinase 2 in Pathogenesis of Thoracic Aortic Aneurysm. Arterioscler Thromb Vasc Biol, 2015.
43. Shimizu K, Mitchell RN, and Libby P. Inflammation and cellular immune responses in abdominal aortic aneurysms. Arterioscler Thromb Vasc Biol 26: 987-994, 2006.
44. Siu KL, Miao XN, and Cai H. Recoupling of eNOS with folic acid prevents abdominal aortic aneurysm formation in angiotensin II-infused apolipoprotein E null mice. PLoS One 9: e88899, 2014.
45. Sun J, Sukhova GK, Zhang J, Chen H, Sjoberg S, Libby P, Xia M, Xiong N, Gelb BD, and Shi GP. Cathepsin K deficiency reduces elastase perfusion-induced abdominal aortic aneurysms in mice. Arterioscler Thromb Vasc Biol 32: 15-23, 2012.
46. Sun J, Sukhova GK, Zhang J, Chen H, Sjoberg S, Libby P, Xiang M, Wang J, Peters C, Reinheckel T, and Shi GP. Cathepsin L activity is essential to elastase perfusion-induced abdominal aortic aneurysms in mice. Arterioscler Thromb Vasc Biol 31: 2500-2508, 2011.
47. Trachet B, Fraga-Silva RA, Londono FJ, Swillens A, Stergiopulos N, and Segers P. Performance comparison of ultrasound-based methods to assess aortic diameter and stiffness in normal and aneurysmal mice. PLoS One 10: e0129007, 2015.
48. Wang L, Guo DC, Cao J, Gong L, Kamm KE, Regalado E, Li L, Shete S, He WQ, Zhu MS, Offermanns S, Gilchrist D, Elefteriades J, Stull JT, and Milewicz DM. Mutations in myosin light chain kinase cause familial aortic dissections. American journal of human genetics 87: 701-707, 2010.
49. Yamanouchi D, Morgan S, Kato K, Lengfeld J, Zhang F, and Liu B. Effects of caspase inhibitor on angiotensin II-induced abdominal aortic aneurysm in apolipoprotein E-deficient mice. Arterioscler Thromb Vasc Biol 30: 702-707, 2010.
50. Yamashiro Y, Papke CL, Kim J, Ringuette LJ, Zhang QJ, Liu ZP, Mirzaei H, Wagenseil JE, Davis EC, and Yanagisawa H. Abnormal mechanosensing and cofilin activation promote the progression of ascending aortic aneurysms in mice. Science signaling 8: ra105, 2015.
51. Yu Z, Morimoto K, Yu J, Bao W, Okita Y, and Okada K. Endogenous superoxide dismutase activation by oral administration of riboflavin reduces abdominal aortic aneurysm formation in rats. Journal of vascular surgery 64: 737-745, 2016.
52. Zhang J, Chen H, Liu L, Sun J, Shi MA, Sukhova GK, and Shi GP. Chemokine (C-C motif) receptor 2 mediates mast cell migration to abdominal aortic aneurysm lesions in mice. Cardiovascular research 96: 543-551, 2012.
53. Zhu L, Vranckx R, Khau Van Kien P, Lalande A, Boisset N, Mathieu F, Wegman M, Glancy L, Gasc JM, Brunotte F, Bruneval P, Wolf JE, Michel JB, and Jeunemaitre X. Mutations in myosin heavy chain 11 cause a syndrome associating thoracic aortic aneurysm/aortic dissection and patent ductus arteriosus. Nature genetics 38: 343-349, 2006.
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