本文選題：甲基苯丙胺 + 凋亡�。� 參考：《第一軍醫(yī)大學(xué)》2007年碩士論文

【摘要】： 甲基苯丙胺(Methamphetamine，METH)屬于苯丙胺類興奮劑(Amphetaminetype stimulants，ATS)，因其原料外觀為純白結(jié)晶體，晶瑩剔透，故被吸毒、販毒者稱為“冰”(Ice)。由于它的毒性劇烈，人們便稱之為“冰毒”。它具有藥物依賴性(主要是精神依賴性)、中樞神經(jīng)興奮、致幻、食欲抑制和擬交感效應(yīng)等藥理、毒理學(xué)特性，是聯(lián)合國(guó)精神藥品公約管制的精神活性物質(zhì)。由于該毒品可一次成癮，其商品名稱為SPEED(快速丸)。甲基苯丙胺可造成精神偏執(zhí)，行為舉止咄咄逼人，并引發(fā)反社會(huì)及性暴力傾向，還可引起吸服者失眠、幻覺、情緒低落，，同時(shí)嚴(yán)重?fù)p害內(nèi)臟器官和腦組織，嚴(yán)重時(shí)導(dǎo)致腎機(jī)能衰竭及精神失常，甚至造成死亡。近年來，甲基苯丙胺在世界上不少國(guó)家和地區(qū)已成為主流毒品，制販甲基苯丙胺的犯罪活動(dòng)正日益猖獗。在國(guó)內(nèi)，其濫用現(xiàn)象也在不斷蔓延擴(kuò)大。 METH對(duì)中樞神經(jīng)系統(tǒng)的損害在哺乳動(dòng)物體內(nèi)的實(shí)驗(yàn)已經(jīng)進(jìn)行了廣泛的研究，但是其確切的作用機(jī)制還沒有完全的清楚。本實(shí)驗(yàn)通過體外海馬細(xì)胞培養(yǎng)模型來研究METH的作用機(jī)制，去除動(dòng)物體內(nèi)的干擾因素，更加利于實(shí)驗(yàn)的控制，可以更加系統(tǒng)和直接的研究其對(duì)海馬神經(jīng)細(xì)胞的毒性作用，為METH的研究提供一條新的思路，從而更好的了解METH的作用機(jī)制。方法：用終濃度分別為0、0.8、1.6、3.2、6.4、12.8mmol／L的METH處理海馬神經(jīng)元48h，用MTT比色法分析細(xì)胞存活率，倒置顯微鏡和電鏡觀察凋亡的形態(tài)學(xué)改變，用流式細(xì)胞儀(FCM)檢測(cè)細(xì)胞凋亡百分率。結(jié)果：METH可劑量依賴性的降低神經(jīng)元的存活率，當(dāng)METH濃度為6.4mmol／L時(shí)，其存活率為45.9％；倒置顯微鏡觀察可見神經(jīng)元胞體固縮，突起斷裂，網(wǎng)絡(luò)消失；電鏡可見其細(xì)胞核明顯固縮和凝聚；用流式細(xì)胞儀檢查可見在0.8、1.6、3.2、6.4、12.8mmol／L METH處理海馬神經(jīng)元，細(xì)胞的凋亡率分別為(8.250±1.702)％、(19.525±2.495)％、(32.500±2.028)％、(49.975±1.603)％、(75.150±2.689)％，與對(duì)照組相比(3.150±0.603)％均有均顯著性差異(P=0.000＜0.01)，且隨劑量增加其凋亡百分率逐漸增大。(P=0.000＜0.01)。結(jié)論：一定濃度的甲基苯丙胺對(duì)體外培養(yǎng)的海馬神經(jīng)細(xì)胞具有細(xì)胞毒性，并且誘導(dǎo)神經(jīng)細(xì)胞死亡具有劑量相關(guān)效應(yīng)。 煙酰胺，又名：維生素B3，是一種必須的營(yíng)養(yǎng)物質(zhì)，存在于谷物和家畜的肉中，常被用來作為一種營(yíng)養(yǎng)補(bǔ)充劑。煙酰胺是煙酰胺腺嘌呤二核苷酸(輔酶I-NAD)的前體，并在線粒體電子傳遞鏈傳遞電子至氧，并釋放能量，氧化磷酸化作用利用這些能量制造三磷酸腺苷(ATP)。參與體內(nèi)葡萄糖酵解、脂肪代謝、丙酮酸代謝等過程，為脂質(zhì)代謝，組織呼吸的氧化作用和糖元分解所需之成分。 最近，對(duì)煙酰胺的研究表明，煙酰胺不僅是一種機(jī)體必須的營(yíng)養(yǎng)物質(zhì)，而且對(duì)細(xì)胞有著保護(hù)作用。對(duì)于煙酰胺的細(xì)胞保護(hù)作用，在越來越多的實(shí)驗(yàn)中得到了證實(shí)。Suman K. Mukherjee在小鼠的帕金森病模型中發(fā)現(xiàn)通過給C57BL／6的腹腔注射煙酰胺(500 mg／kg)，能夠抑制腦的神經(jīng)細(xì)胞的凋亡；并且注射煙酰胺8個(gè)小時(shí)后，發(fā)現(xiàn)腦的中腦區(qū)域的NAD水平從0.28±0.08 umol／g上升到0.38±0.04umol／g，在腦的紋狀體區(qū)域，NAD的水平從0.35±0.04umol／g上升到0.45±0.04 umol／g，并且NAD的水平在注射煙酰胺13h后仍然保持著高水平。其它的研究顯示對(duì)于神經(jīng)細(xì)胞，煙酰胺在氧化應(yīng)激、暫時(shí)性腦缺血、永久性局灶性腦缺血等動(dòng)物模型中能夠防止神經(jīng)細(xì)胞的變性，并且還能防止內(nèi)皮細(xì)胞的損傷，影響血管的生理學(xué)。 本實(shí)驗(yàn)通過建立METH給藥模型，通過注射煙酰胺，探討煙酰胺對(duì)METH給藥模型的保護(hù)作用及其可能的保護(hù)機(jī)制，對(duì)于METH中毒機(jī)制的研究具有一定的啟發(fā)作用，為進(jìn)一步探索METH治療和干預(yù)提供一定的基礎(chǔ)。方法：用高效液相色譜檢測(cè)海馬區(qū)和紋狀體區(qū)DA含量；用NO、SOD、MDA試劑盒檢測(cè)海馬區(qū)和紋狀體區(qū)中NO、SOD、MDA的含量變化；用原位缺口末端標(biāo)記法(TUNEL)觀察細(xì)胞凋亡情況及煙酰胺干預(yù)的影響。結(jié)果：METH給藥模型中多巴胺和SOD的含量有顯著性下降，預(yù)先應(yīng)用煙酰胺能使多巴胺和SOD的下降有顯著性減輕，煙酰胺+METH組與METH組間比較差異有顯著性意義(P=0.000＜0.01)；METH給藥模型中NO和MDA均有顯著性上升，預(yù)先應(yīng)用煙酰胺能使NO和MDA的下降有顯著性減輕，煙酰胺+METH組與METH組間比較差異有顯著性意義(P=0.000＜0.01)；METH給藥模型中海馬區(qū)和紋狀體區(qū)可見大量的凋亡細(xì)胞，預(yù)先應(yīng)用煙酰胺能使凋亡細(xì)胞顯著性減少，煙酰胺+METH組與METH組間比較差異有顯著性意義(P=0.000＜0.01)。結(jié)論：煙酰胺可以減輕METH對(duì)腦中DA系統(tǒng)和SOD的損傷，清除NO、MDA等自由基，對(duì)METH導(dǎo)致的大鼠腦神經(jīng)細(xì)胞凋亡有明顯的抑制作用，具有神經(jīng)保護(hù)效應(yīng)。
[Abstract]:Methamphetamine (METH) belongs to Amphetaminetype stimulants (ATS). Because its raw material is pure white crystalline and crystal clear, it is drug addicts and drug dealers call "Ice". Because of its toxicity, people call it "ice poison". It has drug dependence (mainly mental dependence). The central nervous excitement, phantom, appetite suppression, and quasi sympathetic effects, such as pharmacological, toxicological properties, are the psychoactive substances controlled by the United Nations Convention on psychotropic drugs. As the drug can be addicted to one time, the name of the drug is SPEED (rapid pill). Methamphetamine can cause mental obsession, aggressive behavior, and cause antisocial and sexual violence. Propensity can also cause insomnia, hallucination, low mood, and serious damage to viscera and brain tissue, which seriously cause renal failure and mental disorders, and even cause death. In recent years, methamphetamine has become the mainstream drug in many countries and regions in the world, and the crime of making methamphetamine is increasingly rampant. At home, its abuse is spreading.
The effects of METH on the damage of the central nervous system in mammals have been extensively studied, but the exact mechanism of the action has not been fully understood. In this experiment, the mechanism of METH was studied by the model of cultured hippocampal cells in vitro, the interference factors in animals were removed, and the control of the experiment could be more favorable. The systematic and direct study of its toxic effects on hippocampal neurons provides a new way of thinking for the study of METH, so as to better understand the mechanism of METH's action. Methods: the 48h of hippocampal neurons was treated with METH with final concentration of 0,0.8,1.6,3.2,6.4,12.8mmol / L, and the cell survival rate was analyzed by MTT colorimetry, inverted microscope and electron microscopy. Morphological changes of apoptosis were observed and the percentage of apoptosis was detected by flow cytometry (FCM). Results: METH could reduce the survival rate of neurons in a dose-dependent manner. When the concentration of METH was 6.4mmol / L, the survival rate was 45.9%. The nuclear fusion and condensation were obvious. The hippocampal neurons were treated with 0.8,1.6,3.2,6.4,12.8mmol / L METH by flow cytometry. The apoptosis rate of the cells was (8.250 + 1.702)%, (19.525 + 2.495)%, (32.500 + 2.028)%, (49.975 + 1.603)%, (75.150 + 2.689)%, respectively, (P=0) (P=0). .000 < 0.01), and the percentage of apoptosis increased gradually with the increase of dose. (P=0.000 < 0.01). Conclusion: a certain concentration of methamphetamine has cytotoxicity to cultured hippocampal neurons in vitro, and the induction of neural cell death has a dose-dependent effect.
Nicotinamide, also known as: vitamin B3, is a necessary nutrient that exists in the meat of grain and livestock and is often used as a nutritional supplement. Nicotinamide is the precursor of nicotinamide adenine dinucleotide (coenzyme I-NAD), and transmits electricity to oxygen in the mitochondrial electron transfer chain and releases energy, and is used for the acidification of phosphoric acid. Energy produces adenosine triphosphate (ATP). It participates in the process of glycolysis, fat metabolism, pyruvate metabolism and other processes, which are required for lipid metabolism, tissue respiration oxidation and glycogen decomposition.
Recently, studies on nicotinamide have shown that nicotinamide is not only a necessary nutrient for the body, but also a protective effect on cells. For the protective effect of nicotinamide, more and more experiments have shown that.Suman K. Mukherjee was found in the Parkinson's disease model of mice by intraperitoneal injection of C57BL / 6. Amines (500 mg / kg) can inhibit the apoptosis of brain neurons, and after 8 hours of injection of nicotinamide, the level of NAD in the brain region of the brain rises from 0.28 + 0.08 umol / g to 0.38 + 0.04umol / G. In the striatum region of the brain, the level of NAD rises from 0.35 + 0.04umol / g to 0.45 + 0.04 umol / G. Amides remain high after 13h. Other studies have shown that nicotinamide can prevent degeneration of nerve cells in animal models such as oxidative stress, temporary cerebral ischemia, permanent focal cerebral ischemia and other animal models, and also prevent endothelial cell damage and affect the physiology of blood vessels.
In this experiment, by establishing the METH drug delivery model, by injecting nicotinamide, the protective effect of nicotinamide on the METH drug delivery model and its possible protective mechanism are discussed. It has a certain inspiration for the study of the mechanism of METH poisoning and provides a certain basis for further exploration of the treatment and intervention of METH. Method: high performance liquid chromatography is used to detect the hippocampus. DA content in the region and striatum; the changes in the content of NO, SOD and MDA in the hippocampus and striatum were detected with NO, SOD and MDA kits. The apoptosis of cells and the effect of nicotinamide intervention were observed by in situ nick end labeling (TUNEL). Results: the content of dopamine and SOD in the METH administration model decreased significantly, and nicotinamide could be used in advance. The decrease of dopamine and SOD was significantly reduced, the difference of nicotinamide +METH group and METH group was significant (P=0.000 < 0.01); NO and MDA in the METH administration model increased significantly. The decrease of NO and MDA was significantly reduced in pre application of nicotinamide, and there was significant difference between the nicotinamide + METH group and the METH group (P=0.). 000 < 0.01); a large number of apoptotic cells were found in the CNOOC and striatum of the METH drug delivery model. Pre application of nicotinamide could reduce the significant number of apoptotic cells, and there was a significant difference between the nicotinamide +METH group and the METH group (P=0.000 < 0.01). Conclusion: nicotinamide can reduce the damage of METH to the DA system and SOD in the brain, remove NO, MDA and so on. Free radicals can obviously inhibit the apoptosis of rat brain neurons induced by METH, and have neuroprotective effects.

【學(xué)位授予單位】：第一軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2007
【分類號(hào)】：R363

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相關(guān)期刊論文 前3條

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