本文選題：膿毒癥 + 腦　； 參考：《南方醫(yī)科大學(xué)》2017年碩士論文

【摘要】：研究背景及目的解偶聯(lián)蛋白2(UCP2)是位于線粒體內(nèi)膜的陰離子轉(zhuǎn)運(yùn)蛋白,它可調(diào)節(jié)能量代謝并減少ROS的生成。UCP2的神經(jīng)保護(hù)作用在帕金森病、腦缺血、癲癇等多種神經(jīng)系統(tǒng)疾病中得到廣泛證實。研究表明,膿毒癥時,胰島素不僅有降低血糖、調(diào)節(jié)代謝的作用,還具有減少ROS產(chǎn)生、對抗氧化應(yīng)激、抗凋亡、保護(hù)線粒體等作用,但其具體機(jī)制并不十分明確。既往有體外研究表明胰島素可能通過上調(diào)UCP2的表達(dá),減少活性氧(ROS)的生成,進(jìn)而發(fā)揮抗氧化的保護(hù)作用。本研究擬探討胰島素對膿毒癥腦組織損傷的保護(hù)作用,探討UCP2在膿毒癥大鼠腦組織中的表達(dá),及胰島素對其表達(dá)的影響,探討UCP2在胰島素保護(hù)作用中的可能作用,進(jìn)一步為膿毒癥時胰島素的臨床應(yīng)用提供實驗及理論依據(jù)。方法行胰島素濃度梯度預(yù)實驗,根據(jù)大鼠血糖情況及生存率確定胰島素濃度后,將50只SPF級雄性SD大鼠,按隨機(jī)數(shù)字表法隨機(jī)分為正常對照組(n=10)、膿毒癥組(n=20)和胰島素組(n=20)。通過腹腔注射革蘭陰性菌脂多糖(LPS)15 mg/kg建立膿毒癥大鼠模型。胰島素組在造模前30分鐘予皮下注射長效胰島素1U/kg(預(yù)實驗確定),膿毒癥組皮下注射等量生理鹽水。LPS造模后24h,每組各取8只處死,留取大腦皮質(zhì)。提取大腦皮質(zhì)線粒體測定線粒體活性氧(ROS)、丙二醛(MDA)水平及超氧化物歧化酶(SOD)活力;Western blot檢測 Bcl-2、Bax、活化的 Caspase-9(Cleaved Caspase-9)的蛋白表達(dá);TUNEL染色檢測大腦皮質(zhì)神經(jīng)細(xì)胞凋亡;蘇木精-伊紅(HE)染色觀察大腦皮質(zhì)病理改變;RT-PCR檢測大腦皮質(zhì)UCP2的mRNA表達(dá);Western blot和免疫組化檢測皮質(zhì)組織UCP2蛋白的表達(dá)。結(jié)果1.與正常對照組相比,膿毒癥組及胰島素組大鼠皮質(zhì)組織線粒體ROS和MDA水平明顯升高,而SOD活力明顯下降,均P0.05;而與膿毒癥組相比,胰島素組大鼠皮質(zhì)組織線粒體ROS和MDA水平下降,且SOD活力回升,均P0.05;2.與正常對照組相比,膿毒癥組及胰島素組大鼠大腦皮質(zhì)組織神經(jīng)細(xì)胞凋亡增加,膿毒癥組抗凋亡蛋白Bcl-2表達(dá)下降,而促凋亡蛋白Bax和CleavedCaspase-9表達(dá)升高,均P0.05;而與膿毒癥組相比,胰島素組大鼠皮質(zhì)組織神經(jīng)細(xì)胞凋亡減少,抗凋亡蛋白Bcl-2表達(dá)升高,而促凋亡蛋白Bax和Cleaved Caspase-9 表達(dá)下降,均 P0.05;3.與正常對照組相比,膿毒癥組和胰島素組大鼠大腦皮質(zhì)組織HE染色可見明顯異常病理改變,而與膿毒癥組相比,胰島素組大鼠大腦皮質(zhì)組織病理改變較膿毒癥組減輕;4.與正常對照組相比,膿毒癥組大鼠皮質(zhì)組織UCP2 mRNA和蛋白表達(dá)均升高;而與膿毒癥組相比,胰島素組大鼠皮質(zhì)組織UCP2 mRNA和蛋白表達(dá)較均膿毒癥組升高。結(jié)論1.U/kg長效胰島素皮下注射可有效改善膿毒癥大鼠生存2.UCP2在膿毒癥大鼠腦組織中表達(dá)上調(diào);3.胰島素可上調(diào)膿毒癥大鼠腦組織中UCP2的表達(dá);4.胰島素通過減少ROS生成,減輕線粒體氧化應(yīng)激,抑制線粒體凋亡通路的激活并減少神經(jīng)細(xì)胞凋亡,在膿毒癥大鼠腦組織損傷中發(fā)揮保護(hù)作用,其機(jī)制之一可能是通過上調(diào)UCP2的表達(dá),進(jìn)而減少ROS產(chǎn)生、減輕氧化應(yīng)激并減少細(xì)胞凋亡,從而在膿毒癥腦組織損傷中發(fā)揮保護(hù)作用。
[Abstract]:Background and objective uncoupling protein 2 (UCP2) is an anion transporter in the mitochondrial membrane. It regulates energy metabolism and reduces the neuroprotective effect of ROS producing.UCP2 in many kinds of nervous system diseases, such as Parkinson's disease, cerebral ischemia, and epilepsy. Sugar, which regulates metabolism, also reduces the effect of ROS production, anti oxidative stress, anti apoptosis, and mitochondria protection, but its specific mechanism is not very clear. In the past, in vitro studies have shown that insulin may reduce the production of active oxygen (ROS) by up regulation of UCP2 expression and then play a protective role in antioxidant. This study is to explore the pancreas. The protective effect of isisin on the brain tissue injury of sepsis, to explore the expression of UCP2 in the brain tissue of sepsis rats, and the effect of insulin on its expression, explore the possible role of UCP2 in the protective effect of insulin, and further provide experimental and theoretical basis for the clinical application of insulin in sepsis. After determining the insulin concentration in rats, 50 SPF grade male SD rats were randomly divided into normal control group (n=10), sepsis group (n=20) and insulin group (n=20). The rat model of sepsis was established by intraperitoneal injection of gram-negative lipopolysaccharide (LPS) 15 mg/kg by intraperitoneal injection. The insulin group was 30 before the model. Subcutaneous injection of long acting insulin 1U/kg (pre test), the sepsis group was subcutaneously injected with the same amount of normal saline.LPS to make 24h, each group took 8 dead and left the cerebral cortex. The mitochondria of the cerebral cortex were extracted and the mitochondrial reactive oxygen species (ROS), the malondialdehyde (MDA) water and the superoxide dismutase (SOD) activity were measured, and Western blot detected Bcl-2, B. Ax, the protein expression of activated Caspase-9 (Cleaved Caspase-9); TUNEL staining to detect the apoptosis of cerebral cortex nerve cells; hematoxylin eosin (HE) staining to observe the pathological changes in cerebral cortex; RT-PCR detection of mRNA expression in cerebral cortex UCP2; Western blot and immunohistochemical detection of the expression of cortical tissue UCP2 protein. Results 1. and normal control group phase In the sepsis group and the insulin group, the level of mitochondrial ROS and MDA increased significantly, while the activity of SOD decreased significantly, all P0.05, while the level of ROS and MDA in the cortical mitochondria of the insulin group decreased and the SOD activity increased, all P0.05, compared with the sepsis group, 2. compared with the normal control group, the sepsis group and the insulin group were larger than the normal control group. The apoptosis of neural cells in cerebral cortex was increased, the expression of anti apoptotic protein Bcl-2 in sepsis group decreased, while the expression of apoptotic protein Bax and CleavedCaspase-9 increased, all of which were P0.05. Compared with the sepsis group, the apoptosis of cortical neurons in the insulin group decreased and the expression of anti apoptotic protein Bcl-2 increased, while apoptotic protein Bax and Cleaved Caspase were promoted. The expression of -9 decreased in P0.05. 3. compared with the normal control group, the HE staining in the cerebral cortex of the sepsis group and the insulin group showed obvious abnormal pathological changes. Compared with the sepsis group, the pathological changes in the cerebral cortex of the insulin group were less than that of the sepsis group, and 4. compared with the normal control group, the cortical tissue of the sepsis group was UCP2. The expression of mRNA and protein increased, but compared with the sepsis group, the expression of UCP2 mRNA and protein in the cortical tissue of the insulin group was higher than that of the sepsis group. Conclusion the subcutaneous injection of 1.U/kg long acting insulin can effectively improve the expression of 2.UCP2 in the brain tissue of sepsis rats, and 3. insulin can up regulate the brain of sepsis rats. The expression of UCP2 in the weave; 4. insulin can reduce oxidative stress by reducing ROS, inhibit the activation of mitochondrial apoptosis pathway and reduce the apoptosis of neural cells. It may play a protective role in the brain tissue damage of sepsis rats. One of the mechanisms may be to reduce the expression of UCP2 and then reduce the production of ROS and reduce oxidative stress and decrease Apoptosis can play a protective role in sepsis brain tissue injury.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R459.7

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本文編號：2065908