番茄紅素對血管性癡呆大鼠的抗凋亡作用
發(fā)布時間：2018-09-03 08:56
【摘要】：目的:血管性癡呆(Vascular dementia,VD)是指由于各種腦血管因素(如缺血、出血、急慢性腦供血障礙等)引起腦組織損傷后導(dǎo)致認知功能障礙和記憶力下降的一種臨床綜合征。缺血性腦血管病在血管性癡呆的發(fā)病原因中居首位。近年,隨著人口老齡化的進程,腦卒中和心血管疾病的發(fā)病率持續(xù)上升,血管性癡呆的患病率也隨之增高,但其確切的發(fā)病機制仍未闡明。目前認為氧化應(yīng)激、興奮性氨基酸毒性、炎癥反應(yīng)、細胞凋亡等共同參與血管性癡呆的發(fā)生發(fā)展。細胞凋亡(Apoptosis)為細胞的程序性死亡,是細胞受到傷害性刺激因素導(dǎo)致細胞死亡的重要機制。細胞凋亡分為外源性途徑和內(nèi)源性途徑,外源性凋亡途徑,即死亡受體途徑,經(jīng)Fas蛋白與細胞膜上的受體結(jié)合并激活,招募位于細胞內(nèi)的半胱氨酸天冬氨酸蛋白酶(Caspase),啟動細胞凋亡;內(nèi)源性凋亡途徑,即線粒體途徑,內(nèi)外刺激使線粒體膜的通透性增強,引起細胞色素C的釋放,其與凋亡激活因子1(Apaf-1)形成復(fù)合體,啟動凋亡程序。Caspase9和caspase3是凋亡途徑級聯(lián)反應(yīng)鏈上的關(guān)鍵性生物分子。Caspase3通過切割多聚ADP核糖聚合酶(PARP)等引起DNA損傷,最終導(dǎo)致細胞死亡。番茄紅素(Lycopene,LP)是一種高度不飽和的脂溶性色素,廣泛存在于番茄、西瓜等蔬菜水果中。番茄紅素具有強大的抗氧化能力,能夠清除過多的氧自由基,抵抗衰老、增強人體免疫力、減少心腦血管疾病發(fā)生、抑制腫瘤生長等。近年來,多項有關(guān)番茄紅素在腦血管病方面作用的研究表明,番茄紅素可通過其強大的抗氧化作用對腦血管起到保護作用。但番茄紅素是否可通過抗細胞凋亡發(fā)揮腦保護作用鮮有報道。本實驗通過結(jié)扎大鼠雙側(cè)頸總動脈法(2-VO)制備血管性癡呆動物模型,應(yīng)用番茄紅素進行干預(yù),觀察番茄紅素對血管性癡呆大鼠行為認知功能及海馬CA1區(qū)凋亡指標caspase9、caspase3表達水平的影響。方法:對2月齡健康雄性Sprague-Dawley(SD)大鼠進行雙側(cè)頸總動脈結(jié)扎制備血管性癡呆動物模型。假手術(shù)組10只,只分離頸部肌肉,暴露血管,不結(jié)扎動脈。將手術(shù)后存活動物按照癡呆動物標準進行篩選,符合標準的動物隨機分為模型組、番茄紅素組,每組約10只。番茄紅素組大鼠每日給予番茄紅素10mg/kg,模型組和假手術(shù)組大鼠給予等容量植物油,持續(xù)給藥6周。干預(yù)結(jié)束后,行水迷宮實驗,測定各組大鼠逃避潛伏期及穿越平臺的次數(shù)。后留取大鼠海馬組織,應(yīng)用免疫組織化學方法(Immunohistochemistry)和免疫印跡法(Western blot)測定海馬CA1區(qū)caspase3、caspase9的表達水平。結(jié)果:1番茄紅素對血管性癡呆大鼠的行為學影響水迷宮定位航行實驗結(jié)果表明,三組大鼠的逃避潛伏期從第2天開始均較前1天縮短,表明三組大鼠都有一定的學習能力,但同時三組大鼠之間的學習能力又有一定的差別,三組大鼠之間的逃避潛伏期有顯著差異(P0.05)。模型組與假手術(shù)組相比,前4天的逃避潛伏期明顯延長(P0.05);番茄紅素組與模型組相比,前4天的逃避潛伏期明顯縮短(P0.05);番茄紅素組與假手術(shù)組相比,前4天的逃避潛伏期延長(P0.05)。第5天進行穿越平臺次數(shù)實驗,穿越平臺次數(shù)的多少與大鼠的記憶功能有關(guān),穿越平臺次數(shù)越多說明大鼠的記憶能力越好。模型組與假手術(shù)組相比,穿越平臺次數(shù)明顯減少(P0.05);番茄紅素組與模型組相比,穿越平臺次數(shù)明顯增多(P0.05);番茄紅素組與假手術(shù)組相比,穿越平臺次數(shù)減少,但無顯著統(tǒng)計學差異(P0.05)。2海馬CA1區(qū)caspase3、caspase9表達水平western blot結(jié)果大鼠海馬CA1區(qū)caspase3、caspase9表達情況:模型組與假手術(shù)組相比,大鼠海馬CA1區(qū)caspase3、caspase9蛋白表達水平明顯增加(P0.05);番茄紅素組與模型組相比,海馬CA1區(qū)caspase3、caspase9蛋白表達水平明顯減少(P0.05);番茄紅素組與假手術(shù)組相比,凋亡蛋白表達水平增加,但無明顯統(tǒng)計學差異(P0.05)。3海馬CA1區(qū)caspase3免疫組織化學結(jié)果大鼠海馬CA1區(qū)caspase3蛋白的免疫組化結(jié)果:模型組與假手術(shù)組相比,海馬神經(jīng)元萎縮,caspase3蛋白表達陽性細胞數(shù)明顯增加(P0.05);番茄紅素組與模型組相比,caspase3蛋白表達陽性細胞數(shù)明顯減少(P0.05);番茄紅素組與假手術(shù)組相比,caspase3蛋白表達陽性細胞數(shù)明顯增加(P0.05)。結(jié)論:1番茄紅素能夠明顯縮短血管性癡呆大鼠的逃避潛伏期,使其穿越平臺次數(shù)明顯增加,表明番茄紅素可提高血管性癡呆大鼠的學習記憶能力。2番茄紅素可以下調(diào)血管性癡呆大鼠海馬組織CA1區(qū)caspase3、caspase9的表達水平,提示番茄紅素可能通過抗細胞凋亡對海馬神經(jīng)元起到保護作用。
[Abstract]:Objective: Vascular dementia (VD) is a kind of clinical syndrome that causes cognitive impairment and memory loss after brain tissue injury caused by various cerebrovascular factors (such as ischemia, hemorrhage, acute or chronic cerebral blood supply disorders, etc.). Ischemic cerebrovascular disease is the most common cause of vascular dementia. In recent years, with the development of human beings, the incidence of VD is increasing. As the aging of the mouth progresses, the incidence of stroke and cardiovascular disease continues to rise, and the incidence of vascular dementia increases, but the exact pathogenesis remains unclear. S) Procedural cell death is an important mechanism of cell death induced by noxious stimuli. Apoptosis is divided into exogenous and endogenous pathways, and exogenous pathways of apoptosis, i.e. death receptor pathway, are activated by Fas protein binding to receptors on cell membrane to recruit cysteine aspartate methionine in cells. Caspase initiates apoptosis; endogenous apoptosis pathway, i.e. mitochondrial pathway, enhances the permeability of mitochondrial membrane and induces the release of cytochrome C. Caspase 9 and caspase 3 are key biological molecules in the cascade reaction chain of apoptosis pathway. Lycopene (LP) is a highly unsaturated lipid-soluble pigment, widely found in tomatoes, watermelons and other vegetables and fruits. Lycopene has a strong antioxidant capacity, can scavenge excessive oxygen free radicals, resist aging, increase. In recent years, a number of studies on the role of lycopene in cerebrovascular diseases have shown that lycopene can protect cerebrovascular through its strong antioxidant effect. But whether lycopene can play a fresh role in brain protection by anti-apoptosis In this study, vascular dementia animal model was established by ligating bilateral common carotid arteries (2-VO) in rats. Lycopene was used to interfere with the behavior and cognitive function of vascular dementia rats and the expression of apoptotic markers caspase 9 and caspase 3 in hippocampal CA1 region. Methods: Sprague-Dawley (2-month-old male) was used to observe the effect of lycopene on the behavior and cognitive function of vascular dementia rats. SD) Rats were ligated bilateral common carotid arteries to establish vascular dementia animal model. In sham operation group, 10 rats were isolated neck muscles, exposed blood vessels, and not ligated arteries. The rats in the model group and sham operation group were given the same volume of vegetable oil for 6 weeks. After the intervention, the escape latency and the number of crossing platforms were measured by water maze test. The hippocampus tissues of the rats were harvested by immunohistochemistry and Western blotting. The expression levels of caspase 3 and caspase 9 in hippocampal CA1 region were determined by tern blot. Results: 1. Lycopene affected the behavior of vascular dementia rats. The results of water maze navigation experiment showed that the escape latency of three groups of rats was shorter from the second day than the first day, indicating that the three groups of rats had certain learning ability, but at the same time, the three groups of rats had certain learning ability. The escape latency of model group was significantly longer than that of sham operation group (P 0.05); the escape latency of lycopene group was significantly shorter than that of model group (P 0.05); the escape latency of lycopene group was significantly shorter than that of sham operation group (P 0.05); and the escape latency of lycopene group was significantly shorter than that of sham operation group (P 0.05). The escape latency was prolonged (P 0.05). The number of times crossing the platform was related to the memory function of rats. The more times crossing the platform, the better the memory ability of rats. The expression of caspase 3 and caspase 9 in hippocampal CA1 region was detected by Western blot. Compared with sham operation group, the expression of caspase 3 and caspase 9 in hippocampal CA1 region of rats in model group was lower than that in sham operation group. The expression of Caspase-3 and caspase-9 protein in hippocampal CA1 region was significantly lower in lycopene group than in model group (P 0.05). The expression of apoptotic protein in lycopene group was significantly higher than that in sham operation group, but there was no significant difference (P 0.05). Immunohistochemical results of caspase-3 protein in CA1 region of horse: Compared with sham-operated group, the number of caspase-3 protein positive cells in hippocampal neurons atrophied and increased significantly in model group (P 0.05); compared with model group, the number of caspase-3 protein positive cells in lycopene group decreased significantly (P 0.05); compared with sham-operated group, the expression of caspase-3 protein in lycopene group decreased significantly (P 0.05); and compared with sham-operated group, the CONCLUSION: Lycopene can significantly shorten the escape latency and increase the number of platforms crossing in vascular dementia rats, indicating that Lycopene can improve the learning and memory ability of vascular dementia rats. 2 Lycopene can down-regulate caspase 3 and caspase 3 in hippocampal CA1 region of vascular dementia rats. The expression level of E9 suggested that lycopene might play a protective role in hippocampal neurons through anti apoptosis.
【學位授予單位】：河北醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R749.13
[Abstract]:Objective: Vascular dementia (VD) is a kind of clinical syndrome that causes cognitive impairment and memory loss after brain tissue injury caused by various cerebrovascular factors (such as ischemia, hemorrhage, acute or chronic cerebral blood supply disorders, etc.). Ischemic cerebrovascular disease is the most common cause of vascular dementia. In recent years, with the development of human beings, the incidence of VD is increasing. As the aging of the mouth progresses, the incidence of stroke and cardiovascular disease continues to rise, and the incidence of vascular dementia increases, but the exact pathogenesis remains unclear. S) Procedural cell death is an important mechanism of cell death induced by noxious stimuli. Apoptosis is divided into exogenous and endogenous pathways, and exogenous pathways of apoptosis, i.e. death receptor pathway, are activated by Fas protein binding to receptors on cell membrane to recruit cysteine aspartate methionine in cells. Caspase initiates apoptosis; endogenous apoptosis pathway, i.e. mitochondrial pathway, enhances the permeability of mitochondrial membrane and induces the release of cytochrome C. Caspase 9 and caspase 3 are key biological molecules in the cascade reaction chain of apoptosis pathway. Lycopene (LP) is a highly unsaturated lipid-soluble pigment, widely found in tomatoes, watermelons and other vegetables and fruits. Lycopene has a strong antioxidant capacity, can scavenge excessive oxygen free radicals, resist aging, increase. In recent years, a number of studies on the role of lycopene in cerebrovascular diseases have shown that lycopene can protect cerebrovascular through its strong antioxidant effect. But whether lycopene can play a fresh role in brain protection by anti-apoptosis In this study, vascular dementia animal model was established by ligating bilateral common carotid arteries (2-VO) in rats. Lycopene was used to interfere with the behavior and cognitive function of vascular dementia rats and the expression of apoptotic markers caspase 9 and caspase 3 in hippocampal CA1 region. Methods: Sprague-Dawley (2-month-old male) was used to observe the effect of lycopene on the behavior and cognitive function of vascular dementia rats. SD) Rats were ligated bilateral common carotid arteries to establish vascular dementia animal model. In sham operation group, 10 rats were isolated neck muscles, exposed blood vessels, and not ligated arteries. The rats in the model group and sham operation group were given the same volume of vegetable oil for 6 weeks. After the intervention, the escape latency and the number of crossing platforms were measured by water maze test. The hippocampus tissues of the rats were harvested by immunohistochemistry and Western blotting. The expression levels of caspase 3 and caspase 9 in hippocampal CA1 region were determined by tern blot. Results: 1. Lycopene affected the behavior of vascular dementia rats. The results of water maze navigation experiment showed that the escape latency of three groups of rats was shorter from the second day than the first day, indicating that the three groups of rats had certain learning ability, but at the same time, the three groups of rats had certain learning ability. The escape latency of model group was significantly longer than that of sham operation group (P 0.05); the escape latency of lycopene group was significantly shorter than that of model group (P 0.05); the escape latency of lycopene group was significantly shorter than that of sham operation group (P 0.05); and the escape latency of lycopene group was significantly shorter than that of sham operation group (P 0.05). The escape latency was prolonged (P 0.05). The number of times crossing the platform was related to the memory function of rats. The more times crossing the platform, the better the memory ability of rats. The expression of caspase 3 and caspase 9 in hippocampal CA1 region was detected by Western blot. Compared with sham operation group, the expression of caspase 3 and caspase 9 in hippocampal CA1 region of rats in model group was lower than that in sham operation group. The expression of Caspase-3 and caspase-9 protein in hippocampal CA1 region was significantly lower in lycopene group than in model group (P 0.05). The expression of apoptotic protein in lycopene group was significantly higher than that in sham operation group, but there was no significant difference (P 0.05). Immunohistochemical results of caspase-3 protein in CA1 region of horse: Compared with sham-operated group, the number of caspase-3 protein positive cells in hippocampal neurons atrophied and increased significantly in model group (P 0.05); compared with model group, the number of caspase-3 protein positive cells in lycopene group decreased significantly (P 0.05); compared with sham-operated group, the expression of caspase-3 protein in lycopene group decreased significantly (P 0.05); and compared with sham-operated group, the CONCLUSION: Lycopene can significantly shorten the escape latency and increase the number of platforms crossing in vascular dementia rats, indicating that Lycopene can improve the learning and memory ability of vascular dementia rats. 2 Lycopene can down-regulate caspase 3 and caspase 3 in hippocampal CA1 region of vascular dementia rats. The expression level of E9 suggested that lycopene might play a protective role in hippocampal neurons through anti apoptosis.
【學位授予單位】：河北醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R749.13
【參考文獻】
相關(guān)期刊論文 前6條
1 包華音;;番茄紅素藥理作用的近五年研究進展[J];食品研究與開發(fā);2014年19期
2 徐平湘;陳芩f，
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