本文選題：卡托普利 + 氯沙坦�。� 參考：《瀘州醫(yī)學(xué)院》2012年碩士論文

【摘要】：目的：分別觀察腎素一血管緊張素系統(tǒng)抑制藥卡托普利和氯沙坦對(duì)血管性癡呆(VD)模型大鼠的學(xué)習(xí)記憶的改善及治療效果,并初步探討其可能作用機(jī)制。方法：Morris水迷宮篩選學(xué)習(xí)記憶功能正常的雄性SD大鼠60只；隨機(jī)分手術(shù)組(48只)和假手術(shù)組(12只)；手術(shù)組采用不同時(shí)點(diǎn)分別永久性結(jié)扎大鼠左、右側(cè)頸總動(dòng)脈法建立VD模型。術(shù)后4周,篩選制模成功大鼠36只,隨機(jī)分為2.5mg/Kg卡托普利組、2.0mg/Kg氯沙坦組和模型組,每組12只。用藥組每天1次灌胃給藥,模型組與假手術(shù)組給予等容量生理鹽水,連續(xù)4周。藥后4周,Morris水迷宮測(cè)試各組大鼠學(xué)習(xí)記憶能力。部分大鼠4%多聚甲醛灌注固定,石蠟包埋腦組織,連續(xù)冠狀切片,HE染色和免疫組織化學(xué)染色(SABC法),TUNEL法檢測(cè)海馬神經(jīng)元凋亡,光鏡觀察海馬神經(jīng)元的損傷、Bcl-2與Bax蛋白表達(dá)及海馬神經(jīng)元凋亡情況。GD-10.0多媒體彩色病理圖文分析系統(tǒng)分析Bcl-2及Bax陽性表達(dá)的平均灰度值,灰度值越高表達(dá)產(chǎn)物越多。部分大鼠斷頭取腦,分離皮質(zhì)和海馬,冰上勻漿,3000r/min轉(zhuǎn)速離心10min制備組織勻漿上清液。放射免疫法測(cè)定大腦皮層及海馬組織中TNF-α、IL-1β含量；采用羥胺法測(cè)定大腦皮層及海馬組織中超氧化物歧化酶(SOD)活性；硫代巴比妥酸(TBA)法測(cè)定大腦皮層及海馬組織中丙二醛(MDA)含量；硝酸還原酶法測(cè)定大腦皮層及海馬組織中一氧化氮(NO)含量；化學(xué)比色法測(cè)定大腦皮層及海馬組織中一氧化氮合酶(NOS)和海馬組織中膽堿脂酶(AChE)活性。腹主動(dòng)脈取血,全自動(dòng)血流變快測(cè)儀測(cè)定檢測(cè)血液流變學(xué)。結(jié)果：(1)手術(shù)后4周：①定位航行實(shí)驗(yàn)：隨著訓(xùn)練次數(shù)的增加,各組大鼠平均逃避潛伏期均縮短。與假手術(shù)組比較,手術(shù)組大鼠逃避潛伏期均明顯延長(P0.05)。②空間搜索實(shí)驗(yàn)：與假手術(shù)組比較,模型組在120s內(nèi)穿越平臺(tái)的次數(shù)明顯減少(P0.05)。(2)藥后4周：①定位航行實(shí)驗(yàn)：經(jīng)過5天水迷宮訓(xùn)練,大鼠逃避潛伏期,卡托普利和氯沙坦治療組與模型組比較明顯縮短(P0.05)；②空間搜索實(shí)驗(yàn)：120s內(nèi)穿越平臺(tái)的次數(shù)卡托普利和氯沙坦治療組與模型組比較均有明顯增多(P0.05)。(3)病理組織學(xué)觀察(HE染色)：假手術(shù)組海馬組織神經(jīng)元排列整齊,形態(tài)完整；細(xì)胞數(shù)量較多,細(xì)胞形態(tài)正常,體積較大,核居中,大而圓,染為淡藍(lán)色,核仁核膜清晰,核仁染為紫色,胞質(zhì)著色淺而均勻。模型組大鼠海馬組織神經(jīng)元排列紊亂,數(shù)目減少,胞核固縮濃染。各用藥組均能改善上述神經(jīng)元損傷。(4)TUNEL法檢測(cè)：假手術(shù)組大鼠海馬可見少數(shù)細(xì)胞胞核呈棕黃色,模型組大鼠海馬許多細(xì)胞胞核著成棕黃色,不均勻,并呈環(huán)指樣結(jié)構(gòu),與假手術(shù)組比較,其凋亡細(xì)胞陽性率顯著增強(qiáng)(P0.05)。卡托普利和氯沙坦治療組組海馬胞核著成棕黃色的細(xì)胞數(shù)明顯減少,凋亡細(xì)胞陽性率亦明顯降低(P0.05)。(5)免疫組織化學(xué)檢測(cè)：各組大鼠海馬Bal-2、Bax陽性神經(jīng)元染色均以胞漿有棕黃色物質(zhì)沉積為主,也有部分胞膜及核膜著色,并可顯示神經(jīng)元輪廓。假手術(shù)組大鼠海馬Bal-2、Bax均呈少量表達(dá)。模型組大鼠Bcl-2及Bax蛋白表達(dá)均明顯增加,與假手術(shù)組比較有顯著意義(P0.05)。卡托普利組和氯沙坦組大鼠Bcl-2及Bax蛋白表達(dá)有明顯改善(P0.05)。(6)皮層及海馬組織勻漿：卡托普利組和氯沙坦治療組大腦皮質(zhì)和海馬SOD活性較模型組明顯升高(P0.05),兩治療組大腦皮層及海馬MDA含量較模型組明顯降低(P0.05)�？ㄍ衅绽M和氯沙坦治療組大腦皮質(zhì)及海馬NOS活力及NO含量較模型組明顯降低(P0.05)�？ㄍ衅绽吐壬程怪委熃M對(duì)大腦皮質(zhì)及海馬TNF-α含量和IL-1p含量較模型組明顯下降(P0.05)。與模型組比較,卡托普利和氯沙坦治療組大鼠可降低大腦海馬AChE活性(P0.05)。(7)血液流變學(xué)：與模型空白組比較,卡托普利與氯沙坦組大鼠全血黏度的低切、中切和血漿黏度均有顯著性降低(P0.05),對(duì)血液流變學(xué)具有一定改善作用。結(jié)論：(1)不同時(shí)點(diǎn)分別永久性結(jié)扎大鼠左右側(cè)頸總動(dòng)脈成功制備了VD模型。(2)卡托普利及氯沙坦可改善VD模型大鼠學(xué)習(xí)記憶能力,減輕海馬神經(jīng)元損傷；其機(jī)制與以下因素有關(guān)：①可能通過影響B(tài)cl-2及Bax蛋白表達(dá),減少海馬神經(jīng)元凋亡；②抑制腦組織AChE活性,減少ACh的降解；③增加SOD活性增強(qiáng)氧自由基的清除,降低NOS活性,減少NO的含量,減輕氧自由基和NO對(duì)腦組織的損害；④減少腦組織TNF-α和IL-1β含量,減輕免疫炎癥反應(yīng)對(duì)神經(jīng)元的損傷；⑤改善血液流變學(xué),增加腦組織的供血供氧。
[Abstract]:Objective: To observe the improvement and therapeutic effect of renin and angiotensin system inhibitor Kato Pury and losartan on the learning and memory of vascular dementia (VD) model rats, and to explore its possible mechanism. Methods: 60 male rats with normal learning and memory function were screened by Morris water maze, and 48 rats were randomly divided into operation group (48 rats). And sham operation group (12); VD model was established in the operation group with permanent ligation of rats and right common carotid artery method. 4 weeks after operation, 36 rats were selected and divided into 2.5mg/Kg captopril group, 2.0mg/Kg losartan group and model group, 12 rats in each group. The drug group was administered 1 times a day, model group and sham operation group. The learning and memory ability of rats in each group was tested by Morris water maze for 4 weeks after 4 weeks. Some rats were perfused with 4% polyformaldehyde, paraffin embedded brain tissue, continuous coronal section, HE staining and immunohistochemical staining (SABC). TUNEL method was used to detect the apoptosis of hippocampal neurons, and the damage of hippocampal neurons was observed by light microscope. The expression of Bcl-2 and Bax protein and the apoptosis of hippocampal neurons were analyzed by.GD-10.0 multimedia color pathological graphic analysis system. The average gray value of Bcl-2 and Bax positive expression was analyzed. The higher the gray value, the more products were expressed. Some rats broke head to take brain, separated cortex and hippocampus, on ice homogenate, and 3000r/min rotational speed centrifugation 10min was used to prepare tissue homogenate supernatant. Radioimmunoassay was used to determine the content of TNF- alpha and IL-1 beta in the cerebral cortex and hippocampus; the activity of superoxide dismutase (SOD) in the cerebral cortex and hippocampus was measured by hydroxylamine method; the content of malondialdehyde (MDA) in the cerebral cortex and hippocampus was measured by thiobarbituric acid (TBA); the nitrate reductase method was used to determine the cerebral cortex and hippocampus. Nitric oxide (NO) content; chemical colorimetric assay for the activity of nitric oxide synthase (NOS) and hippocampal cholinesterase (AChE) in the cerebral cortex and hippocampus. Blood rheology was measured by abdominal aorta and automatic hemorrheology rapid measuring instrument. Results: (1) 4 weeks after operation: (1) positioning navigation experiment: with the increase of training times, each The average escape latency of the rats in the group was shortened. Compared with the sham operation group, the escape latency of the operation group was significantly prolonged (P0.05). (2) the space search experiment: compared with the sham group, the number of the model group passed the platform in 120s significantly decreased (P0.05). (2) 4 weeks after the medicine: (1) the positioning navigation experiment: after 5 days water maze training, rats escape The incubation period, Kato Pury and losartan treatment group was significantly shorter than the model group (P0.05); (2) the spatial search experiment: the number of Kato Pury and losartan treatment groups in 120s and losartan group were significantly increased (P0.05). (3) histopathological observation (HE staining): the hippocampal neurons in the sham operation group were arranged neatly and shape. Complete state, more cell number, normal cell morphology, large volume, large and round nucleus, large and round, dyed light blue, clear nucleolus membrane, nucleolus dye purple, light and homogeneous cytoplasm. The hippocampus neurons in the model group were arranged in disorder, the number was reduced, and the nuclei were condensed and concentrated. (4) TUNEL In the sham operation group, the hippocampus of the rats in the sham operation group was found to be brown and yellow, and the nucleus of many cells in the model group were brown and yellow, inhomogeneous and ring finger like structure. Compared with the sham operation group, the positive rate of apoptotic cells increased significantly (P0.05). The hippocampus nucleus of captopril and losartan group was brownish yellow cells. The number of apoptotic cells decreased significantly (P0.05). (5) immuno histochemical detection: Bal-2 and Bax positive neurons in hippocampus of rats in each group were stained mainly with brown yellow substance in cytoplasm, and some membrane and nuclear membrane were coloured, and the neurons of the neurons were displayed. The hippocampus Bal-2 in the sham operation group was a small amount of expression of Bax. The expression of Bcl-2 and Bax protein in the rat model group increased significantly (P0.05). The expression of Bcl-2 and Bax protein in the captopril group and losartan group was significantly improved (P0.05). (6) the cortex and hippocampus homogenate: the cerebral cortex and hippocampal SOD activity in the captopril group and the losartan treatment group were significantly higher than those in the model group. The content of MDA in cerebral cortex and hippocampus in the two treatment group was significantly lower than that in the model group (P0.05). The NOS activity and NO content in the cerebral cortex and hippocampus of the Kato Pury group and the Losartan group were significantly lower than that of the model group (P0.05). The content of TNF- A and the IL-1p content in the cortex and hippocampus of the cerebral cortex and the hippocampus in Kato Pury and losartan groups were significantly lower than those in the model group. (P0.05). Compared with the model group, the rats of Kato Pury and losartan treatment group could reduce the AChE activity in the hippocampus of the brain (P0.05). (7) Hemorheology: compared with the model blank group, the blood viscosity of the whole blood in the rats of Kato Pury and losartan group was lower than that of the rats in the Losartan group. The viscosity of both the middle and the plasma decreased significantly (P0.05), and the blood rheology was improved. Conclusion: (1) the VD model was successfully prepared by ligating the left and right common carotid artery in rats at different time points. (2) Kato Pury and losartan could improve the learning and memory ability of VD model rats and reduce the damage of hippocampal neurons. The mechanism is related to the following factors: (1) the expression of Bcl-2 and Bax protein may be affected and the hippocampal neurons can be reduced. Apoptosis; (2) inhibiting the activity of AChE in brain tissue and reducing the degradation of ACh; (3) increasing the activity of oxygen free radicals by increasing SOD activity, reducing the activity of NOS, reducing the content of NO, reducing the damage of oxygen free radicals and NO on brain tissue; reducing the content of TNF- A and IL-1 beta in the brain tissue and reducing the damage of the immune inflammatory reaction to the neurons; and 5. Increase the blood supply and oxygen supply in the brain tissue.
【學(xué)位授予單位】：瀘州醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R749.13

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