HSP22在肺缺血再灌注損傷中作用研究
發(fā)布時(shí)間：2019-03-18 14:04
【摘要】：[背景與目的]：在肺移植術(shù)、肺栓塞溶栓治療等臨床情況下,肺缺血再灌注損傷可引起急性肺功能不全,具有很高的發(fā)病率和死亡率。HSP22過(guò)表達(dá)在心肌缺血再灌注損傷中有保護(hù)作用,但其在肺缺血再灌注損傷中是否有保護(hù)作用未知。主要研究HSP22過(guò)表達(dá)在小鼠肺缺血再灌注損傷中的作用。 [方法]：采用WesternBlot方法檢測(cè)HSP22轉(zhuǎn)基因C57BL小鼠和野生型C57BL小鼠肺組織中HSP22的表達(dá)情況。取HSP22轉(zhuǎn)基因C57BL小鼠12只,隨機(jī)分為2組：假手術(shù)組、缺血再灌注組；野生型C57BL小鼠12只按相同方法分組：假手術(shù)組、缺血再灌注組。建立小鼠左肺原位缺血再灌注損傷模型。于缺血45分鐘并再灌注120分鐘后取動(dòng)脈血行氧分壓測(cè)定,留取左肺組織標(biāo)本分別測(cè)定各組肺組織濕干重比值,丙二醛(MDA)含量,作光鏡觀察組織形態(tài)變化,免疫組織化學(xué)方法觀察各組HSP22蛋白的表達(dá)情況,TUNEL法測(cè)定肺組織細(xì)胞凋亡的變化。 [結(jié)果]：WesternBlot法檢測(cè)轉(zhuǎn)基因小鼠肺組織中HSP22蛋白含量明顯高于野生型小鼠組(P0.05),且表達(dá)量為野生型小鼠組的2.5倍。四組小鼠的動(dòng)脈血氧分壓無(wú)統(tǒng)計(jì)學(xué)差異(P0.05)。WT-I/R組小鼠的濕干重比值、肺泡損傷指數(shù)較WT-SO組顯著升高(P0.01、P0.01), TG-I/R組小鼠的濕干重比值、肺泡損傷指數(shù)較TG-SO組明顯升高(P0.05、P0.01),均提示轉(zhuǎn)基因小鼠和野生型小鼠的肺缺血再灌注損傷模型成功建立。TG-I/R組小鼠的肺組織濕干重比值、MDA含量、肺泡損傷指數(shù)較WT-I/R組明顯降低(均P0.05),提示過(guò)表達(dá)HSP22可減輕小鼠肺缺血再灌注損傷。免疫組化法示TG-I/R組小鼠HSP22積分光密度值較TG-SO組明顯升高(P0.01),WT-I/R組小鼠HSP22積分光密度值較WT-SO組亦升高(P0.01),說(shuō)明缺血再灌注損傷時(shí)HSP22表達(dá)上調(diào)。TUNEL法結(jié)果示：WT-I/R組較WT-SO組小鼠肺組織細(xì)胞熒光強(qiáng)度增強(qiáng),TG-I/R組也較TG-SO組的熒光強(qiáng)度強(qiáng),然而TG-I/R組較WT-I/R組熒光強(qiáng)度減弱,提示缺血再灌注損傷時(shí)細(xì)胞凋亡增加,但過(guò)表達(dá)HSP22可抑制損傷組織的細(xì)胞凋亡。 [結(jié)論]：1.HSP22轉(zhuǎn)基因小鼠在F4代穩(wěn)定傳代,明確HSP22在轉(zhuǎn)基因小鼠的肺組織內(nèi)高表達(dá)。 2.在國(guó)內(nèi)首次建立HSP22轉(zhuǎn)基因小鼠肺缺血再灌注損傷模型。 3.過(guò)表達(dá)HSP22在肺缺血再灌注損傷中具有保護(hù)作用,可能的機(jī)制是HSP22抑制脂質(zhì)過(guò)氧化和細(xì)胞凋亡。
[Abstract]:[background & objective]: in the clinical conditions of lung transplantation and thrombolytic therapy, pulmonary ischemia-reperfusion injury can cause acute pulmonary dysfunction. The overexpression of HSP22 has protective effect in myocardial ischemia-reperfusion injury, but it is unknown whether HSP22 has protective effect in lung ischemia-reperfusion injury. The aim of this study was to investigate the role of HSP22 overexpression in lung ischemia-reperfusion injury in mice. [methods]: WesternBlot method was used to detect the expression of HSP22 in lung tissue of HSP22 transgenic C57BL mice and wild type C57BL mice. Twelve HSP22 transgenic C57BL mice were randomly divided into two groups: sham-operation group and ischemia-reperfusion group, and wild-type C57BL mice were divided into two groups according to the same method: sham-operation group and ischemia-reperfusion group. The model of in situ ischemia reperfusion injury of left lung in mice was established. After 45 minutes of ischemia and 120 minutes of reperfusion, arterial blood was taken to measure the partial pressure of oxygen. The ratio of wet to dry weight of lung tissue and the content of malondialdehyde (MDA) in the lung tissue of each group were measured respectively, and the histological changes were observed under light microscope. Immunohistochemical method was used to observe the expression of HSP22 protein and TUNEL method was used to detect the changes of apoptosis in lung tissue. [results]: the content of HSP22 protein in lung tissue of transgenic mice was significantly higher than that of wild-type mice by WesternBlot (P0.05), and the expression level was 2.5 times higher than that of wild-type mice. There was no significant difference in arterial oxygen pressure among the four groups (P0.05) .Wet-dry weight ratio and alveolar injury index in WT-I / R group were significantly higher than those in WT-SO group (P 0.01, P0.01). The wet dry weight ratio and alveolar damage index in TG-I/R group were significantly higher than those in TG-SO group (P0.05, P0.01). The results showed that the model of lung ischemia-reperfusion injury in transgenic mice and wild-type mice was successfully established. The wet-dry weight ratio of lung tissue, the content of MDA and the index of alveolar injury in TG-I R group were significantly lower than those in WT-I/R group (P 0.05). It is suggested that over-expression of HSP22 can alleviate lung ischemia-reperfusion injury in mice. Immunohistochemical method showed that the integral optical density of HSP22 in TG-I/R group was significantly higher than that in TG-SO group (P0.01), and the integral optical density of HSP22 in WT-I/R group was also higher than that in WT-SO group (P0.01). The results of Tunel showed that the fluorescence intensity of lung cells in WT-I/R group was higher than that in WT-SO group, and the fluorescence intensity in TG-I/R group was higher than that in TG-SO group, and the fluorescence intensity of HSP22 in TG-I/R group was higher than that in TG-SO group. However, the fluorescence intensity of TG-I/R group was lower than that of WT-I/R group, suggesting that apoptosis increased during ischemia-reperfusion injury, but over-expression of HSP22 could inhibit the apoptosis of injured tissue. [conclusion]: 1.HSP22 transgenic mice were stably subcultured in F4 generation, and the high expression of HSP22 in lung tissue of transgenic mice was confirmed. 2. The model of lung ischemia-reperfusion injury in HSP22 transgenic mice was established for the first time in China. 3. Overexpression of HSP22 plays a protective role in lung ischemia-reperfusion injury. The possible mechanism is that HSP22 inhibits lipid peroxidation and cell apoptosis.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R363
本文編號(hào)：2442934
[Abstract]:[background & objective]: in the clinical conditions of lung transplantation and thrombolytic therapy, pulmonary ischemia-reperfusion injury can cause acute pulmonary dysfunction. The overexpression of HSP22 has protective effect in myocardial ischemia-reperfusion injury, but it is unknown whether HSP22 has protective effect in lung ischemia-reperfusion injury. The aim of this study was to investigate the role of HSP22 overexpression in lung ischemia-reperfusion injury in mice. [methods]: WesternBlot method was used to detect the expression of HSP22 in lung tissue of HSP22 transgenic C57BL mice and wild type C57BL mice. Twelve HSP22 transgenic C57BL mice were randomly divided into two groups: sham-operation group and ischemia-reperfusion group, and wild-type C57BL mice were divided into two groups according to the same method: sham-operation group and ischemia-reperfusion group. The model of in situ ischemia reperfusion injury of left lung in mice was established. After 45 minutes of ischemia and 120 minutes of reperfusion, arterial blood was taken to measure the partial pressure of oxygen. The ratio of wet to dry weight of lung tissue and the content of malondialdehyde (MDA) in the lung tissue of each group were measured respectively, and the histological changes were observed under light microscope. Immunohistochemical method was used to observe the expression of HSP22 protein and TUNEL method was used to detect the changes of apoptosis in lung tissue. [results]: the content of HSP22 protein in lung tissue of transgenic mice was significantly higher than that of wild-type mice by WesternBlot (P0.05), and the expression level was 2.5 times higher than that of wild-type mice. There was no significant difference in arterial oxygen pressure among the four groups (P0.05) .Wet-dry weight ratio and alveolar injury index in WT-I / R group were significantly higher than those in WT-SO group (P 0.01, P0.01). The wet dry weight ratio and alveolar damage index in TG-I/R group were significantly higher than those in TG-SO group (P0.05, P0.01). The results showed that the model of lung ischemia-reperfusion injury in transgenic mice and wild-type mice was successfully established. The wet-dry weight ratio of lung tissue, the content of MDA and the index of alveolar injury in TG-I R group were significantly lower than those in WT-I/R group (P 0.05). It is suggested that over-expression of HSP22 can alleviate lung ischemia-reperfusion injury in mice. Immunohistochemical method showed that the integral optical density of HSP22 in TG-I/R group was significantly higher than that in TG-SO group (P0.01), and the integral optical density of HSP22 in WT-I/R group was also higher than that in WT-SO group (P0.01). The results of Tunel showed that the fluorescence intensity of lung cells in WT-I/R group was higher than that in WT-SO group, and the fluorescence intensity in TG-I/R group was higher than that in TG-SO group, and the fluorescence intensity of HSP22 in TG-I/R group was higher than that in TG-SO group. However, the fluorescence intensity of TG-I/R group was lower than that of WT-I/R group, suggesting that apoptosis increased during ischemia-reperfusion injury, but over-expression of HSP22 could inhibit the apoptosis of injured tissue. [conclusion]: 1.HSP22 transgenic mice were stably subcultured in F4 generation, and the high expression of HSP22 in lung tissue of transgenic mice was confirmed. 2. The model of lung ischemia-reperfusion injury in HSP22 transgenic mice was established for the first time in China. 3. Overexpression of HSP22 plays a protective role in lung ischemia-reperfusion injury. The possible mechanism is that HSP22 inhibits lipid peroxidation and cell apoptosis.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R363
【參考文獻(xiàn)】
相關(guān)博士學(xué)位論文 前1條
1 張付峰;HSP22轉(zhuǎn)基因小鼠模型的建立[D];中南大學(xué);2008年
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