SIRT3通過調(diào)控CypD乙�；閷�(dǎo)膿毒癥相關(guān)性腦病的研究
發(fā)布時(shí)間：2018-07-22 13:03
【摘要】：目的:探討CypD及其乙�；谀摱景Y相關(guān)性腦病小鼠海馬組織中的表達(dá)水平及其影響。方法3-4月齡雄性C57BL/6J小鼠隨機(jī)分為假手術(shù)組(Sham組,n=21)和盲腸結(jié)扎術(shù)組(CLP組,n=21)。CLP組實(shí)施盲腸結(jié)扎術(shù),建造膿毒癥相關(guān)性腦病模型。造模后第4d至第6d每天行Morris水迷宮測(cè)試空間學(xué)習(xí)記憶能力,術(shù)后第7d行空間探索實(shí)驗(yàn),記錄小鼠的游泳速度、逃避潛伏期和探索時(shí)間。在術(shù)后第1d、4d和7d分別使用HE染色切片觀察兩組小鼠海馬組織的損傷;ELISA法檢測(cè)兩組小鼠海馬組織中炎癥因子IL-6和TNF-α的表達(dá)水平,Western Blot法分別測(cè)定兩組小鼠海馬組織中CypD和CypD乙�；谋磉_(dá)水平。結(jié)果:1.與Sham組比較,CLP組小鼠的逃避潛伏期延長(zhǎng)(P0.05),探索時(shí)間縮短(P0.05),游泳速度比較差異無統(tǒng)計(jì)學(xué)意義(P0.05)。2.與Sham組比較,CLP組小鼠在術(shù)后第1d、4d和7d海馬組織損傷明顯加重,其中在術(shù)后第4d時(shí)損傷最顯著。3.與Sham組比較,CLP組小鼠在術(shù)后第1d、4d和7d海馬組織中的IL-6和TNF-α的表達(dá)水平均增加(P0.05),其中在術(shù)后第4d時(shí)炎癥因子增加最顯著。4.與Sham組比較,CLP組小鼠在術(shù)后第1d、4d和7d海馬組織中CypD和CypD乙�；谋硭骄黾�(P0.05),其中術(shù)后第4d時(shí)兩者增加最顯著。結(jié)論:膿毒癥相關(guān)性腦病小鼠的學(xué)習(xí)記憶能力下降,其海馬組織的損傷明顯加重,IL-6、TNF-α以及CypD和CypD乙酰化的表達(dá)水平增加,且其表達(dá)水平的增加均在術(shù)后第4d時(shí)最顯著。目的:探討SIRT3是否通過調(diào)控CypD乙酰化水平而介導(dǎo)小鼠膿毒癥相關(guān)性腦病的發(fā)生發(fā)展。方法:雄性C57BL/6J小鼠,3-4月齡,隨機(jī)分為六組,每組16只,分別為假手術(shù)組(Sham組)、盲腸結(jié)扎術(shù)組(CLP組)、CypDsiRNA轉(zhuǎn)染組(CypD-si組)、CypD siRNA 對(duì)照組(CypD-c 組)、SIRT3 質(zhì)粒轉(zhuǎn)染組(SIRT3-p 組)和 SIRT3質(zhì)�？蛰d組(SIRT3-v組)。除Sham組外,其余五組的小鼠均實(shí)施盲腸結(jié)扎穿孔術(shù),建立膿毒癥相關(guān)性腦病模型。CypD-si組和CypD-c組分別用體內(nèi)轉(zhuǎn)染的方法尾靜脈注射siRNA和control siRNA。SIRT3-p組和SIRT3-v組分別尾靜脈注射pcDNA3.1和vector pcDNA3.1。造模后第4d每天行Morris水迷宮測(cè)試空間學(xué)習(xí)記憶能力,術(shù)后第7d行空間探索實(shí)驗(yàn),記錄小鼠的游泳速度、逃避潛伏期和探索時(shí)間。在術(shù)后第4d用TUNEL法測(cè)定六組小鼠海馬組織中細(xì)胞凋亡水平,測(cè)定六組小鼠海馬線粒體膜電位的水平以及mPTP的開放程度。用Western Blot法分別測(cè)定兩組小鼠海馬組織中IL-6、TNF-α、caspase-3、CypD、CypD乙酰化和SIRT3的表達(dá)水平。結(jié)果:1.與Sham組比較,其余五組小鼠的逃避潛伏期明顯延長(zhǎng)(P0.05),探索時(shí)間縮短(P0.05)。與CLP組比較,CypD-si組、SIRT3-P組小鼠的潛伏期縮短(P0.05),探索時(shí)間延長(zhǎng)(P0.05)。游泳速度的比較差異無統(tǒng)計(jì)學(xué)意義(P0.05)。2.與Sham組比較,其余五組小鼠海馬組織中的細(xì)胞凋亡均增加(P0.05)。與CLP組比較,CypD-si組、SIRT3-p組小鼠的細(xì)胞凋亡水平減輕(P0.05)。3.與Sham組比較,其余五組小鼠海馬組織中mPTP的開放程度增加(P0.05)。與CLP組比較,CypD-si組、SIRT3-p組小鼠海馬組織中mPTP的開放程度減少(P0.05)。4.與Sham組比較,其余五組小鼠海馬組織中IL-6、TNF-α、CypD和CypD乙�；谋磉_(dá)水平增加,SIRT3的表達(dá)水平降低(P0.05)。與CLP組比較,CypD-si 組、SIRT3-P 組小鼠海馬組織中的 IL-6、TNF-α、caspase-3、CypD和CypD乙酰化的表達(dá)水平降低,SIRT3的表達(dá)水平增加(P0.05)。結(jié)論:在膿毒癥相關(guān)性腦病小鼠的海馬組織中,神經(jīng)細(xì)胞壞死和凋亡增加,mPTP的開放增加,IL-6、TNF-α、caspase-3、CypD和CypD乙酰化的表達(dá)水平增加,SIRT3的表達(dá)水平降低。
[Abstract]:Objective: To investigate the expression level and effect of CypD and its acetylation in the hippocampus of mice with sepsis related encephalopathy. Methods male C57BL/6J mice of 3-4 month old were randomly divided into sham operation group (group Sham, n=21) and cecum ligation group (group CLP, n=21).CLP group to carry out cecal ligation and construct sepsis related encephalopathy model. 6D was used to test space learning and memory ability by Morris water maze every day. After 7d, the swimming speed of mice was recorded and the latency and time of exploration were recorded. The damage of hippocampus tissue in two groups of mice was observed in 1D, 4D and 7d after operation, and ELISA method was used to detect the inflammatory factor IL-6 in the hippocampus of the two groups. The expression level of TNF- alpha and Western Blot method was used to determine the expression level of CypD and CypD acetylation in the hippocampus of two mice. Results: 1. compared with the Sham group, the escape latency of the CLP group was prolonged (P0.05), the exploration time was shortened (P0.05), and the difference of swimming speed was not statistically significant (P0.05).2. compared with the Sham group. After 1D, 4D and 7d, the damage of hippocampal tissue was obviously aggravated, and the most significant injury of.3. was compared with that of the Sham group at the post operation 4D. The expression level of IL-6 and TNF- alpha in the CLP group was increased in 1D, 4D and 7d hippocampus in the CLP group. The level of surface water of acetylation of CypD and CypD in 4D and 7d hippocampus increased (P0.05), and both of them increased most significantly at the time of postoperative 4D. Conclusion: the learning and memory ability of mice associated with sepsis related encephalopathy was decreased, the damage of hippocampus tissue was obviously aggravated, the expression level of IL-6, TNF- a, CypD and CypD increased, and the expression level of the hippocampus was increased. The increase was most significant after operation 4D. Objective: To investigate whether SIRT3 can mediate the development of sepsis related encephalopathy in mice by regulating the level of CypD acetylation. Methods: male C57BL/6J mice were divided into six groups at 3-4 months of age, 16 rats in each group, respectively, the sham operation group (group Sham), the cecum ligation group (group CLP), and the CypDsiRNA transfection group (CypD-si). Group (group CypD-c), CypD siRNA control group (group CypD-c), SIRT3 plasmid transfection group (SIRT3-p group) and SIRT3 plasmid unloaded group (group SIRT3-v). Except for Sham group, the other five groups of mice were performed cecum ligation and perforation, and a sepsis related encephalopathy model was set up in the.CypD-si group and CypD-c components in vivo. Group A.SIRT3-p and group SIRT3-v were injected with pcDNA3.1 and vector pcDNA3.1., respectively, to test space learning and memory ability by Morris water maze every day after 4D and Morris water maze was tested every day. The swimming speed of mice was recorded, the incubation period and the time of exploration were avoided. The cells in the hippocampus of six groups of mice were determined by TUNEL method at the end of the operation. The level of apoptosis, the level of mitochondrial membrane potential and the opening degree of mPTP in the hippocampus of six groups of mice were measured. The expression level of IL-6, TNF- alpha, Caspase-3, CypD, CypD acetylation and SIRT3 in the hippocampus of two mice was measured by Western Blot method. Results: 1. and Sham group compared with the Sham group, the escape latency of the remaining five groups of mice was obviously prolonged (P0.05). Time shortened (P0.05). Compared with group CLP, the latency of mice in group CypD-si and SIRT3-P was shortened (P0.05), and the time of exploration was prolonged (P0.05). The difference of swimming speed was not statistically significant (P0.05) compared with group Sham (P0.05), the apoptosis in the hippocampus of the other five groups increased (P0.05). The decrease of apoptosis level (P0.05).3. was compared with that in group Sham, and the degree of opening of mPTP in the hippocampus of the other five groups increased (P0.05). Compared with the CLP group, the opening degree of mPTP in the hippocampal tissues of the CypD-si group and the SIRT3-p group was reduced (P0.05).4. compared with the Sham group, and the other five groups of mice hippocampus The level of expression of SIRT3 decreased (P0.05). Compared with group CLP, IL-6, IL-6, TNF- a, Caspase-3, CypD and CypD acetylation in group CypD-si and SIRT3-P group decreased, and SIRT3 expression level increased (P0.05). Conclusion: in the hippocampus of mice with sepsis related encephalopathy, the necrosis and apoptosis of nerve cells in the hippocampus of mice with sepsis related encephalopathy. The expression of mPTP, IL-6, TNF-, Caspase-3, CypD and CypD increased and the expression level of SIRT3 decreased.
【學(xué)位授予單位】：南京醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R459.7;R747.9
本文編號(hào)：2137550
[Abstract]:Objective: To investigate the expression level and effect of CypD and its acetylation in the hippocampus of mice with sepsis related encephalopathy. Methods male C57BL/6J mice of 3-4 month old were randomly divided into sham operation group (group Sham, n=21) and cecum ligation group (group CLP, n=21).CLP group to carry out cecal ligation and construct sepsis related encephalopathy model. 6D was used to test space learning and memory ability by Morris water maze every day. After 7d, the swimming speed of mice was recorded and the latency and time of exploration were recorded. The damage of hippocampus tissue in two groups of mice was observed in 1D, 4D and 7d after operation, and ELISA method was used to detect the inflammatory factor IL-6 in the hippocampus of the two groups. The expression level of TNF- alpha and Western Blot method was used to determine the expression level of CypD and CypD acetylation in the hippocampus of two mice. Results: 1. compared with the Sham group, the escape latency of the CLP group was prolonged (P0.05), the exploration time was shortened (P0.05), and the difference of swimming speed was not statistically significant (P0.05).2. compared with the Sham group. After 1D, 4D and 7d, the damage of hippocampal tissue was obviously aggravated, and the most significant injury of.3. was compared with that of the Sham group at the post operation 4D. The expression level of IL-6 and TNF- alpha in the CLP group was increased in 1D, 4D and 7d hippocampus in the CLP group. The level of surface water of acetylation of CypD and CypD in 4D and 7d hippocampus increased (P0.05), and both of them increased most significantly at the time of postoperative 4D. Conclusion: the learning and memory ability of mice associated with sepsis related encephalopathy was decreased, the damage of hippocampus tissue was obviously aggravated, the expression level of IL-6, TNF- a, CypD and CypD increased, and the expression level of the hippocampus was increased. The increase was most significant after operation 4D. Objective: To investigate whether SIRT3 can mediate the development of sepsis related encephalopathy in mice by regulating the level of CypD acetylation. Methods: male C57BL/6J mice were divided into six groups at 3-4 months of age, 16 rats in each group, respectively, the sham operation group (group Sham), the cecum ligation group (group CLP), and the CypDsiRNA transfection group (CypD-si). Group (group CypD-c), CypD siRNA control group (group CypD-c), SIRT3 plasmid transfection group (SIRT3-p group) and SIRT3 plasmid unloaded group (group SIRT3-v). Except for Sham group, the other five groups of mice were performed cecum ligation and perforation, and a sepsis related encephalopathy model was set up in the.CypD-si group and CypD-c components in vivo. Group A.SIRT3-p and group SIRT3-v were injected with pcDNA3.1 and vector pcDNA3.1., respectively, to test space learning and memory ability by Morris water maze every day after 4D and Morris water maze was tested every day. The swimming speed of mice was recorded, the incubation period and the time of exploration were avoided. The cells in the hippocampus of six groups of mice were determined by TUNEL method at the end of the operation. The level of apoptosis, the level of mitochondrial membrane potential and the opening degree of mPTP in the hippocampus of six groups of mice were measured. The expression level of IL-6, TNF- alpha, Caspase-3, CypD, CypD acetylation and SIRT3 in the hippocampus of two mice was measured by Western Blot method. Results: 1. and Sham group compared with the Sham group, the escape latency of the remaining five groups of mice was obviously prolonged (P0.05). Time shortened (P0.05). Compared with group CLP, the latency of mice in group CypD-si and SIRT3-P was shortened (P0.05), and the time of exploration was prolonged (P0.05). The difference of swimming speed was not statistically significant (P0.05) compared with group Sham (P0.05), the apoptosis in the hippocampus of the other five groups increased (P0.05). The decrease of apoptosis level (P0.05).3. was compared with that in group Sham, and the degree of opening of mPTP in the hippocampus of the other five groups increased (P0.05). Compared with the CLP group, the opening degree of mPTP in the hippocampal tissues of the CypD-si group and the SIRT3-p group was reduced (P0.05).4. compared with the Sham group, and the other five groups of mice hippocampus The level of expression of SIRT3 decreased (P0.05). Compared with group CLP, IL-6, IL-6, TNF- a, Caspase-3, CypD and CypD acetylation in group CypD-si and SIRT3-P group decreased, and SIRT3 expression level increased (P0.05). Conclusion: in the hippocampus of mice with sepsis related encephalopathy, the necrosis and apoptosis of nerve cells in the hippocampus of mice with sepsis related encephalopathy. The expression of mPTP, IL-6, TNF-, Caspase-3, CypD and CypD increased and the expression level of SIRT3 decreased.
【學(xué)位授予單位】：南京醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R459.7;R747.9
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 Qing-Hong Zhang;Zhi-Yong Sheng;Yong-Ming Yao;;Septic encephalopathy: When cytokines interact with acetylcholine in the brain[J];Military Medical Research;2014年02期
，本文編號(hào)：2137550
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