本文選題：術后認知功能障礙 + NMDA受體　； 參考：《中國人民解放軍醫(yī)學院》2016年博士論文

【摘要】：第一部分 手術創(chuàng)傷對老年小鼠POCD行為學影響目的：通過左肝外葉切除術構建C57BL/6老年小鼠術后認知功能障礙(postoperative cognitive dysfunction or decline,POCD)模型,探索老年小鼠POCD模型的構建,以及手術創(chuàng)傷對老年小鼠術后認知功能的影響。方法：15月齡健康C57BL/6老年小鼠共80只,按隨機數(shù)組表法分為空白對照組、麻醉組、假手術組和手術組。小鼠在術前5天接受訓練,每天3次,尋找固定隱藏平臺。第6天接受左肝外葉切除術。術后第1,3,7天用“空間探索實驗”評價其空間記憶能力,記錄穿臺次數(shù)和目標象限停留時間百分比。結果：在空間獲得性實驗中,各組C57BL/6老年小鼠學習記憶能力隨著時間的延長顯著提高(P0.05)并在第5天趨于穩(wěn)定,且組間沒有差異。Morris水迷宮測試老年小鼠穿臺次數(shù)和目標象限停留百分比結果顯示：與空白對照組比較,麻醉組無差異,假手術組(除術后7d)和手術組明顯下降(P0.05)：與假手術組相比,手術組在各時間點明顯下降(P0.05)。組內(nèi)比較,與術后1d比較,空白組和麻醉組術后3d、7d無差異;假手術組術后3d、7d逐漸恢復,至術后7d時間點基本恢復正常(P0.05);手術組術后3d、7d雖有所恢復,但仍較正常值明顯下降(P0.05)。結論：左肝外葉切除術能成功建立C57BL/6老年小鼠POCD模型,且空間記憶能力的損傷在肝切除術老年小鼠中較假手術老年小鼠重。第二部分POCD與NMDA受體-NF-Kappa B-nNOS信號通路的關系實驗一目的：對C57BL/6老年小鼠進行手術并抑制圍術期應激因素(鎮(zhèn)痛,鎮(zhèn)靜),研究中樞神經(jīng)POCD過程中NMDA受體-NFKB-nNOS信號通路的激活。方法：15月齡健康C57BL/6老年小鼠共80只,按隨機數(shù)字表法分為空白對照組、麻醉組、假手術組和手術組。觀察內(nèi)容如下：分別于術后1,3,7d三個時間點,各組分別有8只動物斷頭取腦,ELISA測定左側腦組織NMDA受體NR2B,nNOS,NF-κB,NOS以及NO含量測定;右側腦組織RT-PCR測定突觸素mNRA, PSD-95蛋白mNRA.結果：檢測結果顯示,與空白對照組比較,各檢測指標(NMDA受體NR2B，NF-κB, nNOS, NOS)蛋白水平以及NO含量,麻醉組在術后各時間點無差異;假手術組(除外術后7d時間點)和手術組在術后各時間點明顯下降(P0.05)：與假手術組比較,各時間點手術組明顯下降(P0.05)。組內(nèi)比較：與術后1d比較,假手術組至7d時基本恢復正常(P0.05);手術組則逐漸下降,至7d時降至最低(P0.05)。RT-PCR測定老年小鼠腦內(nèi)突觸素mRNA和PSD-95mRNA表達水平發(fā)現(xiàn),與空白對照組比較,麻醉組在術后各時間點無差異;假手術組(除外術后7d時間點)和手術組在術后各時間點明顯下降(P0.05);與假手術組比較，各時間點手術組明顯下降(P0.05)。組內(nèi)比較：與術后1d比較,假手術組老年小鼠至7d時基本恢復正常(P0.05);手術組則逐漸下降,至7d時降至最低(P0.05)。結論：C57BL/6老年小鼠在構建POCD模型后,腦內(nèi)NMDA受體、NF-κB、 nNOS蛋白水平的變化趨勢均呈漸進性下降,且7d為最低值。這與腦內(nèi)與突觸相關的突觸素mRNA和PSD-95蛋白mRNA表達趨勢相同,因此提示腦內(nèi)NMDA受體、NF-κB、nNOS蛋白水平的變化是引起老年小鼠POCD的可能機制。實驗二目的：對C57BL/6老年小鼠進行手術,抑制圍術期應激因素(鎮(zhèn)痛,鎮(zhèn)靜),通過應用NMDA受體激動劑(15mg/kg NMDA)驗證老年小鼠POCD過程中存在NMDA受體-NFκB-nNOS通路的激活。方法：15月齡健康C57BL/6老年小鼠共56只，按隨機數(shù)字表法分為空白對照組、假手術組和手術組。腹腔內(nèi)注射NMDA受體激動劑(15mg/kgNMDA) 1小時后行手術。觀察內(nèi)容如下：分別于術后1,3,7d三個時間點,各組分別有8只動物斷頭取腦,ELISA測定左側腦組織NMDA受體NR2B,nNOS,NF-KB,NOS以及NO含量測定;右側腦組織RT-PCR測定突觸素mRNA,PSD-95蛋白mRNA.結果：檢測結果顯示,與空白對照組比較,各檢測指標(NMDA受體NR2B、 NF-κB、nNOS、NOS)蛋白水平以及NO含量，，假手術組在術后各時間點無差異：手術組在術后各時間點明顯下降(P0.05)。手術組內(nèi)：與術后1d比較,其余時間點明顯下降(P0.05),但術后7d與3d比較則無差異(P0.05)。RT-PCR測定老年小鼠腦內(nèi)突觸素mRNA和PSD-95mRNA表達水平發(fā)現(xiàn)，與空白對照組比較，假手術組在術后各時間點(除術后1d)無差異;手術組在術后各時間點明顯下降(P0.05)。手術組內(nèi)：與術后1d比較,其余時間點明顯下降(P0.05),但術后7d與3d比較則無差異(P0.05)。結論：腹腔內(nèi)注射NMDA受體激動劑后，發(fā)現(xiàn)老年POCD小鼠腦內(nèi)海馬組織NMDA受體NR2B、NF-κB、nNOS蛋白水平以及NO含量的降低趨勢減緩,且7d與3d比較無明顯差異，這與神經(jīng)元突觸素mRNA以及PSD-95mRNA表達趨勢相同。提示腦內(nèi)NMDA受體,NF-κB,nNOS蛋白水平的變化是引起老年小鼠POCD的可能機制，并為我們治療POCD提供了新的思路。
[Abstract]:Part one the effect of surgical trauma on the behavior of POCD in aged mice objective by left hepatic lobectomy to construct C57BL/6 aged mice of postoperative cognitive dysfunction (postoperative cognitive dysfunction or decline, POCD) model was constructed to explore the old mice POCD model, as well as the influence of surgical trauma on postoperative cognitive function in elderly mice. Methods: 15 month old a total of 80 healthy aged C57BL/6 mice, according to the random array table were divided into blank control group, anesthesia group, sham operation group and operation group. The mice in the 5 days before training, 3 times a day, looking for hidden fixed platform. Sixth days underwent left hepatic lobectomy. 1,3,7 days after operation with "space to explore the spatial memory ability evaluation experiment, record the times across the platform and the percentage of target quadrant time. Results: experiments in space, each C57BL/6 in aged mice learning and memory ability with The time significantly improve (P0.05) and tend to be stable in fifth days, and there were no group differences in the.Morris water maze test in aged mice Taiwan number and target quadrant percentage results show: compared with the control group, no difference in anesthesia group, sham operation group (except 7D) and the operation group decreased significantly (P0.05): compared with sham operation group, operation group was significantly decreased at each time point (P0.05). Compared with the comparison group, postoperative 1D, blank group and anesthesia group after 3D, 7d no difference; sham operation group after 3D, 7d gradually recovered to the recovery time of postoperative normal base 7d (P0.05); surgery group 3D 7d, despite the recovery, but still lower than the normal value decreased significantly (P0.05). Conclusion: left hepatic lobectomy can be successfully established C57BL/6 POCD in the senile mice model, and the spatial memory ability of injury during liver resection in aged mice compared with sham operated mice. Second elderly part POCD The relationship between NMDA and -NF-Kappa receptor B-nNOS signaling pathway Objective: a surgical C57BL/6 in old mice and inhibit the perioperative stress factors (analgesia, sedation), activation of central nerves in the process of POCD NMDA receptor -NFKB-nNOS signaling pathway. Methods: 15 month old healthy C57BL/6 mice aged 80, anesthesia group randomly divided into blank control group, sham operation group and operation group. Observe the contents are as follows: respectively after 1,3,7d three time points, each group has 8 animal brains were removed for determination of NMDA, ELISA receptor NR2B, left brain tissue nNOS, NF- kappa B, NOS and NO determination of content; the right side of the brain tissue was determined by RT-PCR the expression of mNRA, PSD-95 protein mNRA. results: test results show that, compared with the control group, all indexes (NMDA receptor NR2B, NF- kappa B, nNOS, NOS) protein level and the content of NO, anesthesia group at each time point after surgery no difference Different; sham operation group (except after 7d time point) and the operation group at each time point after operation was significantly decreased (P0.05) compared with the sham operation group, operation group at each time point was significantly decreased (P0.05). Group comparison: compared with postoperative 1D, sham operation group and 7d returned to normal (P0.05); operation group decreased to the lowest at 7d (P0.05).RT-PCR expression level was found the expression of mRNA and PSD-95mRNA in elderly mice, compared with the blank control group, anesthesia group at each time point after operation there is no difference; sham operation group (except after 7d time point) and the operation group in at each time point after operation was significantly decreased (P0.05); compared with the sham operation group, operation group at each time point was significantly decreased (P0.05). Group comparison: compared with postoperative 1D, sham operation group aged mice to 7d normal (P0.05); operation group decreased to the lowest at 7d (P0.05). Conclusion: the old C57BL/6 In mice in the POCD model, NMDA receptors in the brain, NF- kappa B, the change trend of nNOS protein level showed a gradual decline, and 7d is the lowest. The brain and synaptic associated synaptophysin mRNA and PSD-95 protein expression of mRNA in the same trend, thus suggesting that NMDA receptors in the brain, the expression of NF- B. Changes in the level of nNOS protein is the possible mechanism of aged mice caused by POCD. Experiment two objective: surgical C57BL/6 in old mice, inhibition of perioperative stress factors (analgesia, sedation), through the application of NMDA receptor agonist (15mg/kg NMDA) of NMDA receptor activation -NF B-nNOS pathway of the validation process of aged mice. Methods POCD 15 month old: C57BL/6 total of 56 healthy aged mice, were randomly divided into control group, sham operation group and operation group. Intraperitoneal injection of NMDA receptor agonist (15mg/kgNMDA) 1 hours after surgery. Observe the contents are as follows: after operation. 1,3,7d three time points, each group has 8 animal brains were removed for determination of NMDA, ELISA receptor NR2B, left brain tissue nNOS, NF-KB, NOS and NO determination of content; the right side of the brain tissue RT-PCR determination of synapsin mRNA, PSD-95 protein mRNA. results: test results show that, compared with the control group, all indexes (NMDA receptor NR2B, NF-, nNOS, NOS) kappa B protein level and the content of NO, the sham operation group at each time point after operation, no difference in operation group at each time point after operation was significantly decreased (P0.05). The operation group: compared with 1D after operation, the rest of the time points were significantly decreased (P0.05), but after the operation 7d and 3D were no difference (P0.05) determination of.RT-PCR expression levels of synaptophysin mRNA and PSD-95mRNA found in the brain of aged mice, compared with the blank control group, sham operation group at each time point after operation (except 1D) no difference; surgical group at each time point after operation was significantly decreased (P0.05) operation group. In: Compared with the 1D after the operation, the rest of the time points were significantly decreased (P0.05), but the postoperative 7d compared with 3D had no difference (P0.05). Conclusion: intraperitoneal injection of NMDA receptor agonist, was found in the hippocampus of aged POCD mice NMDA receptor NR2B, NF- kappa B, slow decrease the protein level of nNOS and NO the content of 7D and 3D had no significant difference, this is the same as the neurons of synapsin mRNA and the expression of PSD-95mRNA receptors in the brain. The trend suggests that NMDA, NF- kappa B, nNOS protein level changes is the possible mechanism of aged mice caused by POCD, and provides a new way for us to treat POCD.

【學位授予單位】：中國人民解放軍醫(yī)學院
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R619

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