本文選題：VCI + 丁苯酞注射液��； 參考：《湖南師范大學(xué)》2013年碩士論文

【摘要】：目的： 本研究在Wistar大鼠認(rèn)知功能障礙(VCI)模型的基礎(chǔ)上,結(jié)合病理檢測、認(rèn)知功能檢測和免疫組化蛋白定量、熒光定量PCR-mRNA檢測、流式細(xì)胞儀Ca2+定量,以及用丁苯酞注射液干預(yù)后的Calbindin-D28k (CaBP)表達(dá)的對比。初步探討CaBP在凋亡神經(jīng)細(xì)胞中的作用及丁苯酞注射液的保護(hù)作用。 方法： 所有動物造模前先用Morris水迷宮定位航行實(shí)驗(yàn)進(jìn)行篩選,淘汰學(xué)習(xí)記憶能力差的大鼠,獲得250g-300g健康雄性Wistar大鼠36只。將36只合格大鼠稱重,排序,采用隨機(jī)數(shù)字表法分為3組：正常組(normal group)(n=4)、VCI模型組(model group)(n=16)、藥物組(drug inerwention group)(n=16)。利用雙側(cè)頸總動脈永久結(jié)扎術(shù)(2-VO)建立大鼠VCI模型。按照造模后處死時(shí)間將3組隨機(jī)分為一周組(1W)、2周組(2W)、4周組(4W)、8周組(8W)四個(gè)亞組,正常組每組1只,其余分組每個(gè)亞組4只動物。用Morris水迷宮定位航行及空間探索實(shí)驗(yàn)對8W大鼠的記憶及學(xué)習(xí)能力進(jìn)行測定；采用HE染色觀察病理學(xué)變化；應(yīng)用免疫組織化學(xué)檢測CaBP在神經(jīng)細(xì)胞內(nèi)表達(dá)水平的動態(tài)變化；采用熒光定量PCR檢測細(xì)胞內(nèi)CaBP-mRNA表達(dá)的變化。采用流式細(xì)胞儀測定海馬細(xì)胞內(nèi)Ca2+濃度。 結(jié)果： 1、免疫組化：正常組各時(shí)間點(diǎn)可見CaBP免疫陽性細(xì)胞正常表達(dá),模型組1W時(shí)表達(dá)高于正常組,1W以后免疫陽性細(xì)胞表達(dá)逐漸下降,且均低于正常組。藥物組較正常組高表達(dá),1W-8W逐漸下降,至8W時(shí)降至正�；蛏缘陀谡＝M(P0.05)。其中1W、2W、4W組與正常組差異有統(tǒng)計(jì)學(xué)意義(P0.05)。同時(shí)間點(diǎn)藥物組與模型組比較,藥物組CaBP免疫陽性細(xì)胞數(shù)目較多,表達(dá)差異均有顯著意義(P0.05)。 2、熒光定量PCR:CaBP-mRNA的熒光定量PCR檢測結(jié)果示1-8W總體表達(dá)逐漸下降,藥物組1W、2W高表達(dá),4W組稍高表達(dá),8W組降至正常或稍低于正常組(P0.05),與免疫組化結(jié)果吻合。模型組總體表達(dá)亦呈下降趨勢,1W時(shí)高表達(dá),1W后明顯低于正常組。藥物組1、2、4W組與正常組表達(dá)有差異(P0.05),8W組與正常組同一水平(P0.05)。模型組1W、4W、8W組較正常組表達(dá)有差異(P0.05),2W組與正常組差異無統(tǒng)計(jì)學(xué)意義。模型組與藥物組各時(shí)間點(diǎn)表達(dá)均有差異,表達(dá)差異均有統(tǒng)計(jì)學(xué)差異意義(P0.05)。 3、細(xì)胞內(nèi)Ca2+：模型組各時(shí)間點(diǎn)均高表達(dá),較正常組、與藥物組有明顯的差異(P0.05)。藥物組1W、2W組稍高表達(dá),較正常組無明顯差異(P0.05),4W、8W組高表達(dá),較正常組表達(dá)差異有統(tǒng)計(jì)學(xué)意義(P0.05)。 結(jié)論： 1、大鼠慢性腦缺血損傷后CaBP參與了大鼠腦缺血神經(jīng)元凋亡的病理生理過程,可降低細(xì)胞內(nèi)Ca2+濃度,避免鈣超載,起到神經(jīng)保護(hù)作用； 2、丁苯酞注射液可促進(jìn)海馬細(xì)胞內(nèi)CaBP的表達(dá),改善血管性認(rèn)知功能障礙大鼠的學(xué)習(xí)和記憶能力。
[Abstract]:Objective: In this study, based on the Wistar rat model of cognitive dysfunction, combined with pathological examination, cognitive function test and immunohistochemical protein quantification, fluorescence quantitative PCR-mRNA detection, flow cytometry Ca2 quantitative analysis. The expression of Calbindin-D28k was compared with that of butyphthalide injection. To explore the role of CaBP in apoptosis of nerve cells and the protective effect of butyphthalide injection. Methods: The Morris water maze navigation experiment was used to screen all the animals before modeling, and 36 healthy male Wistar rats with 250g-300g were obtained by eliminating the rats with poor learning and memory ability. Thirty-six eligible rats were weighed and sequenced. They were divided into 3 groups by random number table method: normal group (normal group), normal group (n = 4), model group (n = 16) and drug group (inerwention) group (n = 16). Rat VCI model was established by permanent ligation of bilateral common carotid artery (2-VOO). According to the time of death, the three groups were randomly divided into four subgroups, one in each group and four animals in each subgroup. The memory and learning ability of 8W rats were measured by Morris water maze navigation and space exploration experiment, the pathological changes were observed by HE staining, the dynamic changes of CaBP expression in nerve cells were detected by immunohistochemistry. Fluorescence quantitative PCR was used to detect the expression of CaBP-mRNA. The concentration of Ca2 in hippocampal cells was measured by flow cytometry. Results: 1Immunohistochemistry: normal expression of CaBP immunoreactive cells was observed at each time point in the normal group, and the expression of CaBP immunoreactive cells in the model group was lower than that in the normal group after 1 week, and was lower than that in the normal group. The high expression of 1W-8W in the drug group was gradually decreased than that in the normal group, and decreased to normal or slightly lower than that of the normal group at 8W. Among them, there was a significant difference between the 1W 2 W 4 W group and the normal group (P 0.05). At the same time, the number of CaBP immunoreactive cells in the drug group was more than that in the model group, and the difference was significant (P 0.05). 2. The results of fluorescence quantitative PCR:CaBP-mRNA showed that the total expression of 1-8W decreased gradually, and that of the 4W group decreased to normal or slightly lower than that of the normal group, which coincided with the immunohistochemical results. The overall expression of the model group also showed a downward trend after 1 W high expression was significantly lower than that of the normal group. There was significant difference in the expression of P0. 05 and P0. 05 between the two groups (P 0. 05) and the normal group (P 0. 05) at the same level as that in the normal control group (P 0. 05 and P 0. 05). There was no significant difference between the model group and the normal group. There were significant differences in expression between model group and drug group at each time point, and there was significant difference in expression between model group and drug group (P 0.05). (3) intracellular Ca2: the expression of Ca2 in the model group was higher than that in the normal group, and there was a significant difference between the model group and the drug group (P 0.05). There was no significant difference between the drug group and the control group in the expression of 1W ~ 2W group, but there was no significant difference between the two groups (P 0.05 ~ 4W ~ 8W group), and there was a significant difference between the two groups in the expression of P _ (0.05) P _ (0.05) P _ (0.05). Conclusion: 1. CaBP was involved in the pathophysiological process of apoptosis of rat cerebral ischemic neurons after chronic cerebral ischemia injury, which could reduce the concentration of intracellular Ca2, avoid calcium overload and play a neuroprotective role. 2, butyphthalide injection can promote the expression of CaBP in hippocampal cells and improve the learning and memory ability of rats with vascular cognitive impairment.
【學(xué)位授予單位】：湖南師范大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R749.13

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