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系統識別號 U0026-2908201820344400
論文名稱(中文) 探討內生性大麻素分解酵素單酰甘油脂肪酶抑制劑併用環氧化酶-2抑制劑或鴉片類藥物嗎啡在小鼠急性和慢性疼痛模型之協同止痛作用
論文名稱(英文) The synergistic antinociceptive effects of the monoacylglycerol lipase inhibitor JZL-184 and the COX-2 inhibitor parecoxib or the opioid morphine in the murine models of pain
校院名稱 成功大學
系所名稱(中) 心理學系
系所名稱(英) Department of Psychology
學年度 106
學期 2
出版年 107
研究生(中文) 陳弈凝
研究生(英文) Yi-Ning Chen
學號 U76051019
學位類別 碩士
語文別 中文
論文頁數 55頁
口試委員 召集委員-劉彥青
口試委員-王祈斐
口試委員-蕭富仁
指導教授-胡書榕
中文關鍵字 單酰甘油脂肪酶抑制劑JZL184  環氧化酶抑制劑帕瑞昔布  嗎啡  內生性大麻素  2-花生四烯酸甘油酯  痛覺  耐藥性 
英文關鍵字 JZL184  parecoxib  morphine  pain  tolerance 
學科別分類
中文摘要 植物性大麻素以及我們體內所產生的內生性大麻素皆可透過活化大麻素受體來抑制疼痛,但因內生性大麻素會被其分解酵素快速分解,因而作用時間短暫。值得一提的是,內生性大麻素2-花生四烯酸甘油酯(2-arachidonoyglyceral,2-AG)可分別被單酰甘油脂肪酶(monoacylglycerol lipase,MAGL)和環氧化酶(cyclooxygenase-2,COX-2)分解產生花生四烯酸、甘油及前列腺素衍生物(prostaglandin glycerol esters,PG-Gs)。2-AG經由COX-2氧化而生成之前列腺素衍生物PGE2-G卻反而有促痛效果,與2-AG的止痛作用恰好相反。因此,本研究嘗試給予閾下劑量之MAGL抑制劑JZL184,以延長2-AG的作用時間,並合併使用環氧化酶抑制劑帕瑞席布(parecoxib)或者鴉片類藥物嗎啡(morphine),預期在急性和慢性疼痛小鼠模型中達到藥物之協同止痛作用。研究結果發現,合併給予JZL184和parecoxib在伯倫南式腳掌切口手術(Brennan’s paw incision,BPI)引發之急性切口疼痛小鼠模型中沒有止痛效果,但在佛朗式完全佐劑(complete Freund’s adjuvant,CFA)引發之慢性發炎疼痛小鼠模型中則有協同止痛的作用;再者,合併給予JZL184和嗎啡或是單獨給予嗎啡,不僅可有效抑制BPI所引發之急性疼痛,對於CFA所引發之長期發炎疼痛亦有協同止痛作用;最重要的是,當以100 mg/kg嗎啡引發小鼠急性耐受性之後,合併給予JZL184和嗎啡在CFA之慢性發炎疼痛產生卓越的協同止痛效果。最後,我們以西方墨點法來探討,此一協同止痛作用是否導致痛覺傳遞路徑上之受體蛋白質量的變化,結果發現,在急性嗎啡耐受性產生後,合併給予JZL184和嗎啡,有效降低發炎側脊髓背角和背根神經節之MAGL蛋白質表現量,並減少發炎側之背根神經節鴉片類μ受體蛋白質。綜上結果顯示,合併使用JZL184和嗎啡之協同止痛作用相較合併使用JZL184和parecoxib為佳,尤其於產生嗎啡急性耐受性之小鼠身上,二者透過內生性大麻素系統和鴉片系統的交互作用,產生極優的協同止痛功效,此結果暗示JZL184在產生嗎啡耐受性病患的臨床治療上有抑制長期疼痛之潛力。
英文摘要 SUMMARY

We used the MAGL inhibitor JZL184 to prolong the action of 2-AG and combined with the COX-2 inhibitor parecoxib or with the opioid morphine to achieve the synergistic antinociceptive effects on acute and chronic pain in mice. We took mechanical stimuli to evaluate analgesic effect by using electric von Frey instrument. Our results showed that co-administration the subthreshold dosage of JZL184 (4 mg/kg, i.p.) and parecoxib (5 mg/kg, i.p.) did not reduce acute pain in the Brennan’s paw incision (BPI) model, but worked synergistically to relieve the chronic inflammatory pain induced by complete Freund’s adjuvant (CFA). On the other hand, Co-administration of JZL184 (4 mg/kg, i.p.) and morphine (2.5 mg/kg, i.p.) synergistically alleviate the BPI-induced acute pain and CFA-induced chronic pain. Most importantly, comparing with morphine alone, co-administration of JZL184 and morphine displayed excellent pain relief in acute morphine tolerance mice which received 100 mg/kg morphine. Finally, by using Western blotting, we observed that the combined administration of JZL184 and morphine decreased the MAGL and μ-opioid receptor protein levels in the dorsal horn of spinal cord and dorsal root ganglia (DRG). Taken together, our results indicate that the synergistic antinociceptive effect of JZL184 and morphine is better than that of JZL184 and parecoxib, especially in the acute morphine tolerance mice. JZL184 and morphine may work synergistically via the interaction between the endocannabinoid system and opioid system to alleviate inflammatory pain in these mice.

INTRODUCTION

Both cannabis and endocannabinoids alleviate pain via activating cannabinoid receptors, however, the action of endocannabinoids is transient due to their degradation enzymes. The endocannabinoid 2-arachidonoyglyceral (2-AG) can be catalyzed by monoacylglycerol lipase (MAGL) and cyclooxygenase-2 (COX-2), respectively, into arachidonic acid, glycerol, and prostaglandin glycerol esters (PG-Gs). PGE2-G, one of the COX-2 metabolite of 2-AG, exerts a pro-nociceptive action in contrast to the antinociceptive property of 2-AG (Hu et al., 2008; Naidu et al., 2009; Alhouayek and Muccioli, 2014). Therefore, the MAGL inhibitor JZL184 can extend the actions of 2-AG including analgesia. Synergistic effect of analgesics produces greater effects than the sum of individual effects so that we combined low dosages of two analgesic to diminish side effects of high dosage drugs, such as tolerance. In the other hand, endocannabinoids and morphine also work synergistically to produce analgesic effect (Pacher et al., 2006; Scavone et al., 2013). In this study, we combine JZL184 and COX-2 inhibitor parecoxib or the opioid morphine to achieve the synergistic antinociceptive effects on acute and chronic pain in mice, and examining which combination exhibits more powerful potential.

MATERIALS AND METHODS

At least 9-weeks-old male ICR mice from NCKU laboratory animal center (Taiwan) and BioLASCO Taiwan co., ltd (Taiwan) served as subjects. We use electric von Frey instrument to assess mechanical pain threshold of mice’ left hind paw before and after injections of analgesics. There are acute Brennan’s paw incision (BPI)-induced incision and chronic complete Freund’s adjuvant (CFA)-induced inflammatory pain models in this study. Tests take place immediately after BPI surgery and 3 days after CFA injection. The grouping of JZL184 and parecoxib experiment sets are vehicle, JZL184 alone, parecoxib alone, and JZL184 + parecoxib groups; the grouping of JZL184 and morphine experiment sets are vehicle, JZL184 alone, morphine alone, and JZL184 + morphine groups. In acute morphine tolerance model, mice were tested 24 hours after 100 mg/kg morphine injection (i.p.) at day 3 of CFA injection. Finally, we analysis CB1 receptor, MAGL, and opioid μ receptor protein level of PAG, dorsal horn, dorsal root ganglia, and paw by Western blotting.

RESULTS AND DISCUSSION

Our results first showed that co-administration the subthreshold dosage of JZL184 (4 mg/kg, i.p.) and parecoxib (5 mg/kg, i.p.) did not reduce acute pain in the Brennan’s paw incision (BPI) model. However, JZL184 and parecoxib synergistically relieved the chronic inflammatory pain induced by complete Freund’s adjuvant (CFA). On the other hand, Co-administration of JZL184 (4 mg/kg, i.p.) and morphine (2.5 mg/kg, i.p.) synergistically alleviate the BPI-induced acute pain and CFA-induced chronic pain. Most importantly, comparing with morphine alone, co-administration of JZL184 and morphine displayed excellent pain relief in acute morphine tolerance mice which received 100 mg/kg morphine. Finally, by using Western blotting, we observed that the combined administration of JZL184 and morphine decreased the MAGL and μ-opioid receptor protein levels in the dorsal horn of spinal cord and dorsal root ganglia (DRG) of those acute morphine tolerance mice. The decrease of μ-opioid receptor protein levels might represent the exist of interactions of the endocannabinoid system and the endogenous opioid system.

CONCLUSION

This study attempts to examine the potency of JLZ184 as analgesic, and we found that the synergistic antinociceptive effect of JZL184 and morphine is better than that of JZL184 and parecoxib, especially in the acute morphine tolerance mice. JZL184 and morphine may work synergistically via the interaction between the endocannabinoid system and opioid system to alleviate inflammatory pain in these mice. Our findings hence suggest that JZL184 has therapeutic potential to treat chronic pain in morphine tolerance patients in the clinical setting.
論文目次 目錄

摘要 i
致謝 vi
目錄 vii
表目錄 ix
圖目錄 x
第一章、緒論 1
第一節、疼痛的分類 1
第二節、內生性大麻素的止痛療效 3
第三節、內生性大麻素抑制劑和COX-2抑制劑的協同止痛機制 5
第四節、內生性大麻素抑制劑和嗎啡的協同止痛現象 8
第五節、總結 10
第二章、材料及方法 11
第一節、實驗動物 11
第二節、藥品之製備及劑量 11
第三節、機械性疼痛行為測試 12
第四節、Brennan’s Paw Incision(BPI)引發之急性切口疼痛模型 12
第五節、Complete Freuns’s adjuvant(CFA)引發之慢性發炎疼痛模型 13
第六節、實驗組別 13
第七節、CFA引發慢性發炎疼痛合併急性嗎啡耐受性小鼠模型 14
第八節、西方墨點法 15
第九節、統計分析 16
第三章、實驗結果 17
實驗一:合併使用JZL184和parecoxib對於BPI引發小鼠急性切口疼痛之止痛效果 17
實驗二:合併使用JZL184和Parecoxib對於CFA引發小鼠慢性發炎疼痛之止痛效果 17
實驗三:合併使用JZL184和嗎啡對於BPI引發小鼠急性切口疼痛之止痛效果 19
實驗四:合併使用JZL184和嗎啡對於CFA引發小鼠慢性發炎疼痛之止痛效果 20
實驗五:合併使用JZL184和嗎啡對於調節急性嗎啡耐受性之CFA慢性發炎小鼠疼痛之影響 22
實驗六:不同劑量之JZL184在BPI引發小鼠急性切口疼痛模型之作用 25
實驗七:合併使用JZL184和嗎啡對於調節急性嗎啡耐受性之CFA慢性發炎疼痛小鼠導水管周圍灰質、脊髓背角、背根神經節及腳掌MAGL、大麻素受體第一型及鴉片類μ受體蛋白質表現量之影響 26
第四章 討論 28
參考文獻 52

表目錄

表一、BPI引發之急性術後疼痛模型及CFA引發之長期性發炎疼痛模型分組及實驗藥劑說明表 50
表二、CFA長期性發炎疼痛合併急性嗎啡耐受性小鼠實驗分組及實驗藥物說明表 51

圖目錄

圖一、機械性疼痛行為測試裝置示意圖 38
圖二、BRENNAN’S PAW INCISION(BPI)手術說明圖 38
圖三、BPI引發小鼠急性切口疼痛實驗流程圖 39
圖四、CFA引發小鼠長期性發炎疼痛實驗流程圖 39
圖五、CFA慢性發炎疼痛合併急性嗎啡耐受性小鼠實驗流程圖 40
圖六、合併使用JZL184及PARECOXIB或者單獨給予各別藥物對於BPI引發之小鼠急性切口疼痛之影響 40
圖七、合併使用JZL184和PARECOXIB或者單獨給予各別藥物對於CFA引發之慢性發炎疼痛小鼠之影響 41
圖八、合併使用JZL184及嗎啡或者單獨給予各別藥物對於BPI引發之小鼠急性切口疼痛之影響 42
圖九、合併使用JZL184和嗎啡或者單獨給予各別藥物對於CFA引發之慢性發炎疼痛小鼠之影響 43
圖十、合併使用JZL184和嗎啡或者單獨給予嗎啡對於產生急性嗎啡耐受性之CFA慢性發炎疼痛小鼠之影響 44
圖十一、4、16及40 MG/KG之JZL184在BPI引發之急性切口疼痛小鼠的止痛效果 45
圖十二、合併給予JZL184和嗎啡及單獨給予嗎啡對嗎啡耐受性小鼠導水管周圍灰質(PAG)大麻素受體第一型、MAGL及鴉片類Μ受體蛋白質表現量的影響 46
圖十三、合併給予JZL184和嗎啡及單獨給予嗎啡對嗎啡耐受性小鼠脊髓背角大麻素受體第一型、MAGL及鴉片類Μ受體蛋白質表現量的影響 47
圖十四、合併給予JZL184和嗎啡及單獨給予嗎啡對嗎啡耐受性小鼠背根神經節(DRG)大麻素受體第一型、MAGL及鴉片類Μ受體蛋白質表現量的影響 48
圖十五、合併給予JZL184和嗎啡及單獨給予嗎啡對嗎啡耐受性小鼠發炎側腳掌皮膚大麻素受體第一型、MAGL及鴉片類Μ受體蛋白質表現量的影響 49

參考文獻 Abrams D, Couey P, Shade S, Kelly M, Benowitz N (2011) Cannabinoid–opioid interaction in chronic pain. Clinical Pharmacology & Therapeutics 90:844-851.
Alhouayek M, Muccioli GG (2014) COX-2-derived endocannabinoid metabolites as novel inflammatory mediators. Trends in pharmacological sciences 35:284-292.
Braune S, Schady W (1993) Changes in sensation after nerve injury or amputation: the role of central factors. Journal of Neurology, Neurosurgery & Psychiatry 56:393-399.
Brennan TJ, Vandermeulen EP, Gebhart G (1996) Characterization of a rat model of incisional pain. Pain 64:493-502.
Caillé S, Alvarez-Jaimes L, Polis I, Stouffer DG, Parsons LH (2007) Specific alterations of extracellular endocannabinoid levels in the nucleus accumbens by ethanol, heroin, and cocaine self-administration. Journal of Neuroscience 27:3695-3702.
Cichewicz D, Welch S (2003) Modulation of oral morphine antinociceptive tolerance and naloxone-precipitated withdrawal signs by oral Delta 9-tetrahydrocannabinol. The Journal of pharmacology and experimental therapeutics 305:812-817.
Cichewicz DL (2004) Synergistic interactions between cannabinoid and opioid analgesics. Life sciences 74:1317-1324.
Cichewicz DL, Martin ZL, Smith FL, Welch SP (1999) Enhancement of μ opioid antinociception by oral Δ9-tetrahydrocannabinol: dose-response analysis and receptor identification. Journal of Pharmacology and Experimental Therapeutics 289:859-867.
Corchero J, Manzanares J, Fuentes JA (2004) Cannabinoid/opioid crosstalk in the central nervous system. Critical Reviews™ in Neurobiology 16.
Cox ML, Haller VL, Welch SP (2007) Synergy between Δ9-tetrahydrocannabinol and morphine in the arthritic rat. European journal of pharmacology 567:125-130.
Crofford LJ (1997) COX-1 and COX-2 tissue expression: implications and predictions. The Journal of Rheumatology Supplement 49:15-19.
Daniels SE, Grossman EH, Kuss ME, Talwalker S, Hubbard RC (2001) A double-blind, randomized comparison of intramuscularly and intravenously administered parecoxib sodium versus ketorolac and placebo in a post—oral surgery pain model. Clinical therapeutics 23:1018-1031.
Davis MP (2014) Cannabinoids in pain management: CB1, CB2 and non-classic receptor ligands. Expert opinion on investigational drugs 23:1123-1140.
Dinh T, Carpenter D, Leslie F, Freund T, Katona I, Sensi S, Kathuria S, Piomelli D (2002a) Brain monoglyceride lipase participating in endocannabinoid inactivation. Proceedings of the national Academy of sciences 99:10819-10824.
Dinh TP, Freund TF, Piomelli D (2002b) A role for monoglyceride lipase in 2-arachidonoylglycerol inactivation. Chemistry and physics of lipids 121:149-158.
Fairbanks CA, Wilcox GL (1999) Spinal antinociceptive synergism between morphine and clonidine persists in mice made acutely or chronically tolerant to morphine. Journal of Pharmacology and Experimental Therapeutics 288:1107-1116.
Ghosh S, Wise LE, Chen Y, Gujjar R, Mahadevan A, Cravatt BF, Lichtman AH (2013) The monoacylglycerol lipase inhibitor JZL184 suppresses inflammatory pain in the mouse carrageenan model. Life sciences 92:498-505.
Glaser ST, Abumrad NA, Fatade F, Kaczocha M, Studholme KM, Deutsch DG (2003) Evidence against the presence of an anandamide transporter. Proceedings of the National Academy of Sciences 100:4269-4274.
Gulyas A, Cravatt B, Bracey M, Dinh T, Piomelli D, Boscia F, Freund T (2004) Segregation of two endocannabinoid‐hydrolyzing enzymes into pre‐and postsynaptic compartments in the rat hippocampus, cerebellum and amygdala. European Journal of Neuroscience 20:441-458.
Hu SSJ, Ho YC, Chiou LC (2014) No more pain upon Gq‐protein‐coupled receptor activation: role of endocannabinoids. European Journal of Neuroscience 39:467-484.
Hu SSJ, Bradshaw H, Chen JC, Tan B, Walker JM (2008) Prostaglandin E2 glycerol ester, an endogenous COX‐2 metabolite of 2‐arachidonoylglycerol, induces hyperalgesia and modulates NFκB activity. British journal of pharmacology 153:1538-1549.
Joshi GP, Viscusi ER, Gan TJ, Minkowitz H, Cippolle M, Schuller R, Cheung RY, Fort JG (2004) Effective treatment of laparoscopic cholecystectomy pain with intravenous followed by oral COX-2 specific inhibitor. Anesthesia & Analgesia 98:336-342.
Kim JA, Bartlett S, He L, Nielsen CK, Chang AM, Kharazia V, Waldhoer M, Ou CJ, Taylor S, Ferwerda M (2008) Morphine-induced receptor endocytosis in a novel knockin mouse reduces tolerance and dependence. Current Biology 18:129-135.
Kinsey SG, Wise LE, Ramesh D, Abdullah R, Selley DE, Cravatt BF, Lichtman AH (2013) Repeated low-dose administration of the monoacylglycerol lipase inhibitor JZL184 retains cannabinoid receptor type 1–mediated antinociceptive and gastroprotective effects. Journal of Pharmacology and Experimental Therapeutics 345:492-501.
Lau BK, Vaughan CW (2014) Descending modulation of pain: the GABA disinhibition hypothesis of analgesia. Current opinion in neurobiology 29:159-164.
Lin Y-T, Ro L-S, Wang H-L, Chen J-C (2011) Up-regulation of dorsal root ganglia BDNF and trkB receptor in inflammatory pain: an in vivo and in vitro study. Journal of neuroinflammation 8:126.
Long JZ, Li W, Booker L, Burston JJ, Kinsey SG, Schlosburg JE, Pavón FJ, Serrano AM, Selley DE, Parsons LH (2009) Selective blockade of 2-arachidonoylglycerol hydrolysis produces cannabinoid behavioral effects. Nature chemical biology 5:37-44.
Malan TP, Marsh G, Hakki SI, Grossman E, Traylor L, Hubbard RC (2003) Parecoxib sodium, a parenteral cyclooxygenase 2 selective inhibitor, improves morphine analgesia and is opioid-sparing following total hip arthroplasty. Anesthesiology: The Journal of the American Society of Anesthesiologists 98:950-956.
Martini L, Whistler JL (2007) The role of mu opioid receptor desensitization and endocytosis in morphine tolerance and dependence. Current opinion in neurobiology 17:556-564.
McPartland JM, Duncan M, Di Marzo V, Pertwee RG (2015) Are cannabidiol and Δ9‐tetrahydrocannabivarin negative modulators of the endocannabinoid system? A systematic review. British journal of pharmacology 172:737-753.
Naidu PS, Booker L, Cravatt BF, Lichtman AH (2009) Synergy between enzyme inhibitors of fatty acid amide hydrolase and cyclooxygenase in visceral nociception. Journal of Pharmacology and Experimental Therapeutics 329:48-56.
Ng A, Smith G, Davidson A (2003) Analgesic effects of parecoxib following total abdominal hysterectomy. British Journal of Anaesthesia 90:746-749.
Nomura DK, Morrison BE, Blankman JL, Long JZ, Kinsey SG, Marcondes MCG, Ward AM, Hahn YK, Lichtman AH, Conti B (2011) Endocannabinoid hydrolysis generates brain prostaglandins that promote neuroinflammation. Science 334:809-813.
Nussmeier NA, Whelton AA, Brown MT, Langford RM, Hoeft A, Parlow JL, Boyce SW, Verburg KM (2005) Complications of the COX-2 inhibitors parecoxib and valdecoxib after cardiac surgery. New England Journal of Medicine 352:1081-1091.
Pacher P, Bátkai S, Kunos G (2006) The endocannabinoid system as an emerging target of pharmacotherapy. Pharmacological reviews 58:389-462.
Padi SS, Jain NK, Singh S, Kulkarni SK (2004) Pharmacological profile of parecoxib: a novel, potent injectable selective cyclooxygenase-2 inhibitor. European journal of pharmacology 491:69-76.
Pertwee RG (2005) The therapeutic potential of drugs that target cannabinoid receptors or modulate the tissue levels or actions of endocannabinoids. The AAPS journal 7:E625-E654.
Pertwee RG (2006) Cannabinoid pharmacology: the first 66 years. British journal of pharmacology 147.
Ramesh D, Ross GR, Schlosburg JE, Owens RA, Abdullah RA, Kinsey SG, Long JZ, Nomura DK, Sim-Selley LJ, Cravatt BF (2011) Blockade of endocannabinoid hydrolytic enzymes attenuates precipitated opioid withdrawal symptoms in mice. Journal of Pharmacology and Experimental Therapeutics 339:173-185.
Reche I, Fuentes JA, Ruiz-Gayo M (1996) Potentiation of Δ9-tetrahydrocannabinol-induced analgesia by morphine in mice: involvement of μ-and κ-opioid receptors. European journal of pharmacology 318:11-16.
Saario SM, Savinainen JR, Laitinen JT, Järvinen T, Niemi R (2004) Monoglyceride lipase-like enzymatic activity is responsible for hydrolysis of 2-arachidonoylglycerol in rat cerebellar membranes. Biochemical pharmacology 67:1381-1387.
Scavone J, Sterling R, Van Bockstaele E (2013) Cannabinoid and opioid interactions: implications for opiate dependence and withdrawal. Neuroscience 248:637-654.
Sio SW, Ang SF, Lu J, Moochhala S, Bhatia M (2010) Substance p upregulates cyclooxygenase-2 and prostaglandin E metabolite by activating ERK1/2 and NF-κB in a mouse model of burn-induced remote Acute lung injury. The Journal of Immunology 185:6265-6276.
Sugiura T, Kobayashi Y, Oka S, Waku K (2002) Biosynthesis and degradation of anandamide and 2-arachidonoylglycerol and their possible physiological significance. Prostaglandins, Leukotrienes and Essential Fatty Acids (PLEFA) 66:173-192.
Warner TD, Mitchell JA (2004) Cyclooxygenases: new forms, new inhibitors, and lessons from the clinic. The FASEB journal 18:790-804.
Woolf CJ (2011) Central sensitization: implications for the diagnosis and treatment of pain. Pain 152:S2-S15.
Woolf CJ, Doubell TP (1994) The pathophysiology of chronic pain—increased sensitivity to low threshold Aβ-fibre inputs. Current opinion in neurobiology 4:525-534.
Xu J, Brennan TJ (2011) The pathophysiology of acute pain: animal models. Current opinion in Anaesthesiology 24:508.
Yamaguchi T, Hagiwara Y, Tanaka H, Sugiura T, Waku K, Shoyama Y, Watanabe S, Yamamoto T (2001) Endogenous cannabinoid, 2-arachidonoylglycerol, attenuates naloxone-precipitated withdrawal signs in morphine-dependent mice. Brain research 909:121-126.

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