本文選題：阿爾茨海默病　切入點(diǎn)：硫氧還蛋白　出處：《蘇州大學(xué)》2010年碩士論文　論文類型：學(xué)位論文

【摘要】： 目的:根據(jù)Aβ1-42及D-半乳糖致衰老理論,在亞急性衰老模型基礎(chǔ)上側(cè)腦室注射Aβ1-42建立一種復(fù)合式阿爾茨海默病(Alzheimer’s disease,AD)動(dòng)物模型。在此基礎(chǔ)上觀察神經(jīng)細(xì)胞凋亡的線粒體通路與硫氧還蛋白(Thioredoxin,Trx)的關(guān)系,旨在為神經(jīng)退行性疾病的發(fā)病機(jī)制、臨床防治等提供科學(xué)理論依據(jù)。 方法: (1)側(cè)腦室注射Aβ1-42建立單一因素AD模型(B組)、采用D-半乳糖腹腔注射SD大鼠建立亞急性衰老模型(C組)、腹腔注射D-半乳糖聯(lián)合側(cè)腦室注射Aβ1-42建立復(fù)合式AD模型(D組)、選用正常大鼠作為對(duì)照組(A組); (2)行為學(xué)采用Morris水迷宮檢測(cè)大鼠學(xué)習(xí)和記憶能力; (3)甲苯胺藍(lán)染色觀察神經(jīng)元形態(tài)及數(shù)量的變化; (4)生化指標(biāo)檢測(cè)血清及大腦組織勻漿中CAT、T-AOC及GSH/GSSG; (5)免疫組織化學(xué)檢測(cè)衰老相關(guān)蛋白AGEs及AGER的表達(dá)變化,腦組織各區(qū)Trx,Cyto-C及Caspase-9表達(dá)采用免疫組織化學(xué)及免疫印跡觀察。 結(jié)果: (1)Morris水迷宮檢測(cè)顯示:與A組相比,B組、C組及D組大鼠學(xué)習(xí)和記憶能力明顯下降,有顯著性差異(P0.05或P0.01);與B組、C組相比,D組大鼠學(xué)習(xí)記憶能力也明顯下降,差異有顯著性(P0.01); (2)尼氏染色結(jié)果顯示:與A組相比,B組、C組及D組染色神經(jīng)元的數(shù)量有不同程度的減少,差異具有統(tǒng)計(jì)學(xué)意義(P0.05或P0.01)。鏡下可見(jiàn)核固縮、胞體變小等形態(tài)學(xué)改變,以D組最為明顯,其次是B組,再者是C組; (3)生化指標(biāo)檢測(cè)顯示:與A組相比,B組、C組及D組血清和腦組織勻漿中CAT、T-AOC和血清中GSH/GSSG顯著下降(P0.05或P0.01),與B組、C組相比,D組血清和腦組織勻漿中CAT、T-AOC及勻漿中GSH/GSSG都明顯下降(P0.01); (4)免疫組化結(jié)果顯示:與A組相比,B組、C組及D組AGEs、AGER在皮質(zhì)、海馬、小腦的表達(dá)明顯增多,有顯著性差異(P0.05或P0.01);與B組、C組相比,D組AGEs、AGER在皮質(zhì)、海馬、小腦的表達(dá)顯著增多(P0.01); (5)免疫組織化學(xué)法和免疫印跡檢測(cè)發(fā)現(xiàn)B組、C組及D組與A相比,在皮質(zhì)、海馬、小腦中Trx的表達(dá)明顯下降(P0.05或P0.01),Cyto-C和Caspase-9的表達(dá)明顯上升(P0.05或P0.01);D組與B組、C組相比,在皮質(zhì)、海馬、小腦中Trx的表達(dá)有所下降(P0.01),Cyto-C和Caspase-9的表達(dá)上升(P0.05或P0.01)。 結(jié)論: (1)較好的在衰老的基礎(chǔ)上建立了一種AD復(fù)合動(dòng)物模型,初步探討了衰老與AD之間存在的潛在關(guān)系。 (2)在D-半乳糖亞急性衰老模型基礎(chǔ)上可建立Aβ1-42復(fù)合式AD動(dòng)物模型。復(fù)合式動(dòng)物模型在行為學(xué),生化指標(biāo)及組織學(xué)上優(yōu)于單一因素AD動(dòng)物模型。此種復(fù)合式動(dòng)物模型不僅可以用于老年癡呆的基礎(chǔ)研究;而且也可用于探討老年癡呆的防治途徑。 (3)D-半乳糖聯(lián)合Aβ1-42建立AD大鼠模型中AGEs和AGER的表達(dá)明顯增多,AGEs和AGER的增加反過(guò)來(lái)會(huì)加速AD的發(fā)病進(jìn)程。AGEs和AGER二者的抑制劑可能是AD的臨床研究藥物靶點(diǎn)。 (4)在不同模型組Trx都有不同程度的下降,線粒體凋亡途徑中重要蛋白Cyto-C、Caspase-9都有所上降。Trx的表達(dá)與Cyto c、caspase-9的表達(dá)負(fù)相關(guān),Cyto c的表達(dá)與caspase-9的表達(dá)正相關(guān)。提示AD中Trx的下降與細(xì)胞凋亡的發(fā)生之間有重要的聯(lián)系,并且有可能是通過(guò)Cyto-C的釋放和Caspase-9的激活而引起凋亡。
[Abstract]:Objective: according to the A beta 1-42 and D- galactose induced aging theory, in the subacute aging model based on intraventricular injection of A beta 1-42 build a composite type of Alzheimer's disease (Alzheimer 's disease, AD) animal model. Based on the observation of nerve cell apoptosis pathway and mitochondrial thioredoxin (Thioredoxin, Trx) the relation to pathogenesis of neurodegenerative diseases, provide scientific theoretical basis for clinical prevention and treatment.
Method:
(1) intracerebroventricular injection of A beta 1-42 build single factor AD model (group B), established by the subacute aging model induced by D- galactose intraperitoneal injection of SD rats (C group), intraperitoneal injection of D- galactose combined with intracerebroventricular injection of A beta 1-42 build complex AD model (D group), normal rats as for the control group (A group);
(2) the learning and memory ability of rats was detected by Morris water maze.
(3) to observe the changes of morphology and number of neurons toluidine blue staining;
(4) biochemical indexes were used to detect CAT, T-AOC and GSH/GSSG in serum and brain tissue homogenate.
(5) immunohistochemistry was used to detect the expression changes of aging related proteins AGEs and AGER. The expression of Trx, Cyto-C and Caspase-9 in different regions of the brain was examined by immunohistochemistry and Western blotting.
Result:
(1) Morris water maze test showed that compared with group A, the learning and memory ability of B group, C group and D group decreased significantly (P0.05 or P0.01). Compared with B group and C group, the learning and memory ability of the D group decreased significantly, with significant difference (P0.01).
(2) Nissl staining showed that: compared with A group, B group, C group and D group the number of neurons was reduced in different degrees, the difference was statistically significant (P0.05 or P0.01). Under the microscope, nuclear pyknosis, change smaller cell body morphology, the D group was most obvious. Followed by the B group, and C group;
(3) biochemical index test showed that compared with group A, GSH/GSSG, CAT, T-AOC and serum GSH/GSSG in serum and brain homogenate of B group, C group and D group decreased significantly (P0.05 or P0.01), compared with those in B group and C group, there was a significant decrease in serum and brain homogenate in group A.
(4) immunohistochemistry showed that compared with group A, the expression of AGEs and AGER in cortex, hippocampus and cerebellum increased significantly in group B, group C and group D (P0.05 or P0.01). Compared with B group and C group, the expression of B and its expression in cortex, hippocampus and cerebellum increased significantly.
(5) immunohistochemistry and Western blot detection showed that B group, C group and D group compared with A in cortex, hippocampus and cerebellum in the expression of Trx decreased significantly (P0.05 or P0.01), the expression of Cyto-C and Caspase-9 increased significantly (P0.05 or P0.01); D group and B group, C group in the cortex, hippocampus, cerebellum, the expression of Trx decreased (P0.01), increased expression of Cyto-C and Caspase-9 (P0.05 or P0.01).
Conclusion:
(1) a AD compound animal model was established on the basis of aging, and the potential relationship between aging and AD was preliminarily discussed.
(2) based on D- D-galactose subacute aging model can be established on the A beta 1-42 composite AD animal model. The compound in animal models of behavior, biochemistry and histology is better than single factor AD animal model. The combined type animal model not only can be used in basic research of senile dementia; but also can be used to investigate ways to prevent and cure Alzheimer's disease.
(3) the expression of AGEs and AGER increased significantly in the establishment of AD rat model with D- galactose and A beta 1-42. The increase of AGEs and AGER in turn would accelerate the pathogenesis of AD. The inhibitors of.AGEs and AGER two might be the clinical drug targets of AD.
(4) have different degrees of decline in different model group Trx, Cyto-C protein in the mitochondrial apoptotic pathway, Caspase-9 has on the expression of Cyto and.Trx reduced C, negative correlation between the expression of caspase-9, expression of caspase-9 and Cyto C positive correlation. There is an important connection between Trx and AD decreased in tips the occurrence of apoptosis, and might induce apoptosis through activation of Cyto-C release and Caspase-9.

【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類號(hào)】：R749.16;R-332

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