去鐵胺預(yù)處理對(duì)大鼠肺缺血再灌注損傷的保護(hù)作用研究
發(fā)布時(shí)間：2018-08-22 07:06
【摘要】：目的：肺缺血再灌注損傷是指中斷血供肺臟發(fā)生的缺血、缺氧性損傷隨血流灌注恢復(fù)進(jìn)一步加重的病理現(xiàn)象，廣泛存在于心胸外科領(lǐng)域。研究表明，缺血再灌注時(shí)期游離鐵增多，后者可以催化氧自由基生成，介導(dǎo)炎癥反應(yīng)。提自于多絨鏈霉菌中的去鐵敏是一種高選擇性的鐵離子螯合劑，最初用于治療血液疾病。其特定的羥胺基團(tuán)具有同機(jī)體內(nèi)游離鐵離子相結(jié)合的能力，減少由其參與產(chǎn)生的活性氧基團(tuán)對(duì)生物膜的攻擊；同時(shí)誘導(dǎo)低氧預(yù)適應(yīng)，調(diào)節(jié)機(jī)體經(jīng)受缺血、缺氧、炎癥攻擊的能力，減少組織凋亡。目前去鐵胺(Deferoxamine,DFO)逐漸成為組織保護(hù)領(lǐng)域焦點(diǎn)，國(guó)內(nèi)外實(shí)驗(yàn)均證明在心肌保護(hù)，缺血缺氧性腦損傷治療、肝臟保存液改良等的有較可靠的效果，但尚缺乏對(duì)肺缺血再灌注損傷保護(hù)作用研究的報(bào)道。本實(shí)驗(yàn)旨在觀察DFO預(yù)處理對(duì)大鼠原位肺缺血再灌注損傷的作用，并對(duì)其可能機(jī)制做初步探討。方法：取健康清潔SD大鼠90只,體重300g士25g,采用隨機(jī)數(shù)字表法分為3組(n=30)：假手術(shù)組(NC組)，生理鹽水預(yù)處理組(NS組),去鐵胺預(yù)處理組(DFO組)。NS組開(kāi)胸前連續(xù)3d經(jīng)腹腔注射生理鹽水，阻斷左肺門(mén)45min再灌注120min；DFO組術(shù)前連續(xù)3d經(jīng)腹腔注射DFO預(yù)處理，再建立鼠原位肺缺血再灌注模型；NC組左前外側(cè)胸壁第3-5肋間切口開(kāi)胸顯露肺門(mén)，不進(jìn)行肺缺血再灌注處理。各組均于再灌注30min、60min、120min共3個(gè)時(shí)間點(diǎn)分別留取左心尖血、肺組織標(biāo)本后處死動(dòng)物10只，各時(shí)間點(diǎn)血液標(biāo)本檢測(cè)動(dòng)脈血氧分壓(Pa02)、TNF-α濃度，評(píng)估肺氧合功能、炎癥水平。各時(shí)間點(diǎn)肺組織標(biāo)本計(jì)算濕／干重比值(W／D)評(píng)價(jià)肺水腫程度；制作肺組織勻漿并檢測(cè)其中丙二醛含量觀察脂質(zhì)氧化反應(yīng)；末端轉(zhuǎn)移酶標(biāo)記法測(cè)定肺臟組織內(nèi)細(xì)胞凋亡水平；光鏡觀察組織和細(xì)胞病理學(xué)改變。結(jié)果：1.肺氧合功能：所測(cè)得的動(dòng)脈血氧分壓（Pa02）中，NS組和DFO組Pa02值出現(xiàn)下降，與NC組比較，在再灌注R60min、R120min時(shí)間點(diǎn)均存在統(tǒng)計(jì)學(xué)差異（P0.05）；與NS組相比，DFO組Pa02值在R60min、R120min時(shí)間點(diǎn)增高(P0.05)。2.肺干濕重比值：所測(cè)得的肺干濕重比（W/D）中，NS組和DFO組表現(xiàn)為W/D值增高，與NC組比較，在再灌注R30min、R60min、 R120min時(shí)間點(diǎn)均存在統(tǒng)計(jì)學(xué)差異（P0.05）；與NS組相比，W/D值在R30min、R60min、R120min時(shí)間點(diǎn)降低(P0.05)。3.脂質(zhì)氧化反應(yīng)： DFO組和NS組肺組織MDA含量隨時(shí)間逐步上升，與NC組比較在再灌注R30min、R60min、 R120min時(shí)間點(diǎn)均存在統(tǒng)計(jì)學(xué)差異（P0.05）；與NS組相比，DFO組中R60min、R120min時(shí)間點(diǎn)肺組織MDA含量降低(P0.05)。4.炎癥反應(yīng)強(qiáng)度： DFO組和NS組肺組織血清TNF-α濃度隨時(shí)間逐步上升，與NC組比較在再灌注R30min、R60min、 R120min時(shí)間點(diǎn)均存在統(tǒng)計(jì)學(xué)差異（P0.05）；與NS組相比，DFO組中R30min、R60min、R120min時(shí)間點(diǎn)血清TNF-α濃度濃度降低(P0.05)。5.病理評(píng)價(jià)：各時(shí)間點(diǎn)NC組大鼠肺組織結(jié)構(gòu)清晰肺泡完整，肺泡無(wú)充血、水腫、滲出等改變。NS組各時(shí)間點(diǎn)大鼠肺組織均見(jiàn)肺泡萎陷或不張，大量炎性細(xì)胞浸潤(rùn)表現(xiàn)，泡腔內(nèi)紅細(xì)胞滲出，隨再灌注時(shí)間延長(zhǎng)加重，肺泡損傷指數(shù)較NC組增高(P0.05)。DFO組與NS組相比各時(shí)間點(diǎn)大鼠肺組織亦有炎性浸潤(rùn)、肺泡結(jié)構(gòu)破壞伴少量出血，DFO組各時(shí)間點(diǎn)肺損傷指數(shù)低于NS組(P0.05)。6.TUNEL染色：各時(shí)間點(diǎn)上NS組大鼠肺組織均有大量棕褐色顆粒，呈明顯凋亡表現(xiàn)，隨再灌注時(shí)間延長(zhǎng)加重，凋亡指數(shù)較NC組顯著增高（P0.05)。DFO組與NS組相比各時(shí)間點(diǎn)大鼠肺組織亦有上述改變，但R30min、R120min凋亡指數(shù)低于NS組(P0.05)，，NC組存在凋亡現(xiàn)象不明顯。結(jié)論：1.DFO對(duì)大鼠原位肺缺血再灌注損傷具有改善通氣功能、減輕水腫，具備一定的保護(hù)作用。2.在大鼠LIRI模型中，DFO干預(yù)后可有效抑制脂質(zhì)氧化反應(yīng)，減輕炎癥反應(yīng)的作用。3.在大鼠LIRI模型中, DFO預(yù)處理有抑制肺組織因LIRI導(dǎo)致的凋亡。
[Abstract]:AIM: Pulmonary ischemia-reperfusion injury (PIRI) refers to the pathological phenomenon of ischemia and hypoxic injury aggravated with the recovery of blood perfusion. It is widely found in cardiothoracic surgery. Deferrin in mold is a highly selective iron chelating agent originally used in the treatment of blood diseases. Its specific hydroxylamine groups have the ability to combine with free iron ions in the body, reducing the aggression of reactive oxygen species produced by them to biofilms, and inducing hypoxic preconditioning to regulate the body's ability to withstand ischemia and hypoxia. Deferoxamine (DFO) has gradually become the focus in the field of tissue protection. Domestic and foreign experiments have proved that it has a reliable effect on myocardial protection, treatment of ischemic and hypoxic brain injury, and improvement of liver preservation fluid. However, there is no report on the protective effect of DFO on lung ischemia-reperfusion injury. Methods: Ninety healthy and clean SD rats weighing 300 g and 25g were randomly divided into three groups (n=30), sham operation group (NC group), normal saline group (NS group) and desferriamine pretreatment group (DFO group). NS group received intraperitoneal injection of normal saline for 3 days before thoracotomy to block the left hilum for 45 minutes and reperfusion for 120 minutes; DFO group received intraperitoneal injection of DFO preconditioning for 3 days before operation to establish in situ pulmonary ischemia-reperfusion model in rats; NC group received thoracotomy to expose the hilum through the third-fifth intercostal incision on the left anterolateral chest wall, without ischemia-reperfusion treatment. Blood samples of left ventricular apex were collected at 3 time points of in, 60 min and 120 min, and then 10 animals were sacrificed. The arterial oxygen partial pressure (Pa02), TNF-a concentration, pulmonary oxygenation function and inflammation level were measured at each time point. Results: 1. Pulmonary oxygenation function: Pa02 values in NS and DFO groups decreased, compared with NC group, in reperfusion R60. Compared with NS group, the Pa02 value in DFO group increased at R60 min and R120 min (P 0.05). 2. The ratio of dry to wet lung weight: The W/D value in NS group and DFO group increased significantly at R30 min, R60 min and R120 min of reperfusion compared with NC group. Compared with NS group, W/D value decreased at R30 min, R60 min, R120 min time point (P 0.05). 3. Lipid oxidation reaction: MDA content in lung tissue of DFO group and NS group increased gradually with time, and there were statistical differences between DFO group and NC group at R30 min, R60 min, R120 min time point of reperfusion (P 0.05); compared with NS group, there were statistical differences between DFO group at R60 min, R120 min time point of lung. Tissue MDA content decreased (P 0.05). 4. Inflammation intensity: The serum TNF-alpha concentration in DFO group and NS group increased gradually with time, and there were significant differences between the two groups at the time points of reperfusion R30 min, R60 min, R120 min (P 0.05); compared with NS group, the serum TNF-alpha concentration in DFO group decreased at the time points of R30 min, R60 min, R120 min (P 0.05). 5. Pathological evaluation: The pulmonary tissue of NC group was clear, alveolar intact, alveolar no congestion, edema, exudation and other changes. In NS group, alveolar atrophy or atelectasis, a large number of inflammatory cells infiltration, alveolar erythrocyte exudation, with the extension of reperfusion time, alveolar injury index was higher than NC group (P 0.05). Compared with NS group, the lung tissue of DFO group also had inflammatory infiltration and alveolar structure damage with a little bleeding. The lung injury index of DFO group was lower than NS group (P 0.05). 6. TUNEL staining showed that there were a lot of brown granules in lung tissue of NS group at each time point, showing obvious apoptosis. With the prolongation of reperfusion time, the apoptosis index increased. Compared with NC group, DFO group also had the above changes in lung tissue at each time point, but the apoptosis index at R30 min and R120 min was lower than NS group (P 0.05). There was no obvious apoptosis in NC group. Conclusion: 1. DFO can improve ventilation function and reduce edema in rats with lung ischemia-reperfusion injury in situ. 2. In rat LIRI model, DFO intervention can effectively inhibit lipid oxidation and alleviate inflammation. 3. In rat LIRI model, DFO preconditioning can inhibit lung tissue apoptosis induced by LIRI.
【學(xué)位授予單位】：瀘州醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2013
【分類(lèi)號(hào)】：R655.3
本文編號(hào)：2196328
[Abstract]:AIM: Pulmonary ischemia-reperfusion injury (PIRI) refers to the pathological phenomenon of ischemia and hypoxic injury aggravated with the recovery of blood perfusion. It is widely found in cardiothoracic surgery. Deferrin in mold is a highly selective iron chelating agent originally used in the treatment of blood diseases. Its specific hydroxylamine groups have the ability to combine with free iron ions in the body, reducing the aggression of reactive oxygen species produced by them to biofilms, and inducing hypoxic preconditioning to regulate the body's ability to withstand ischemia and hypoxia. Deferoxamine (DFO) has gradually become the focus in the field of tissue protection. Domestic and foreign experiments have proved that it has a reliable effect on myocardial protection, treatment of ischemic and hypoxic brain injury, and improvement of liver preservation fluid. However, there is no report on the protective effect of DFO on lung ischemia-reperfusion injury. Methods: Ninety healthy and clean SD rats weighing 300 g and 25g were randomly divided into three groups (n=30), sham operation group (NC group), normal saline group (NS group) and desferriamine pretreatment group (DFO group). NS group received intraperitoneal injection of normal saline for 3 days before thoracotomy to block the left hilum for 45 minutes and reperfusion for 120 minutes; DFO group received intraperitoneal injection of DFO preconditioning for 3 days before operation to establish in situ pulmonary ischemia-reperfusion model in rats; NC group received thoracotomy to expose the hilum through the third-fifth intercostal incision on the left anterolateral chest wall, without ischemia-reperfusion treatment. Blood samples of left ventricular apex were collected at 3 time points of in, 60 min and 120 min, and then 10 animals were sacrificed. The arterial oxygen partial pressure (Pa02), TNF-a concentration, pulmonary oxygenation function and inflammation level were measured at each time point. Results: 1. Pulmonary oxygenation function: Pa02 values in NS and DFO groups decreased, compared with NC group, in reperfusion R60. Compared with NS group, the Pa02 value in DFO group increased at R60 min and R120 min (P 0.05). 2. The ratio of dry to wet lung weight: The W/D value in NS group and DFO group increased significantly at R30 min, R60 min and R120 min of reperfusion compared with NC group. Compared with NS group, W/D value decreased at R30 min, R60 min, R120 min time point (P 0.05). 3. Lipid oxidation reaction: MDA content in lung tissue of DFO group and NS group increased gradually with time, and there were statistical differences between DFO group and NC group at R30 min, R60 min, R120 min time point of reperfusion (P 0.05); compared with NS group, there were statistical differences between DFO group at R60 min, R120 min time point of lung. Tissue MDA content decreased (P 0.05). 4. Inflammation intensity: The serum TNF-alpha concentration in DFO group and NS group increased gradually with time, and there were significant differences between the two groups at the time points of reperfusion R30 min, R60 min, R120 min (P 0.05); compared with NS group, the serum TNF-alpha concentration in DFO group decreased at the time points of R30 min, R60 min, R120 min (P 0.05). 5. Pathological evaluation: The pulmonary tissue of NC group was clear, alveolar intact, alveolar no congestion, edema, exudation and other changes. In NS group, alveolar atrophy or atelectasis, a large number of inflammatory cells infiltration, alveolar erythrocyte exudation, with the extension of reperfusion time, alveolar injury index was higher than NC group (P 0.05). Compared with NS group, the lung tissue of DFO group also had inflammatory infiltration and alveolar structure damage with a little bleeding. The lung injury index of DFO group was lower than NS group (P 0.05). 6. TUNEL staining showed that there were a lot of brown granules in lung tissue of NS group at each time point, showing obvious apoptosis. With the prolongation of reperfusion time, the apoptosis index increased. Compared with NC group, DFO group also had the above changes in lung tissue at each time point, but the apoptosis index at R30 min and R120 min was lower than NS group (P 0.05). There was no obvious apoptosis in NC group. Conclusion: 1. DFO can improve ventilation function and reduce edema in rats with lung ischemia-reperfusion injury in situ. 2. In rat LIRI model, DFO intervention can effectively inhibit lipid oxidation and alleviate inflammation. 3. In rat LIRI model, DFO preconditioning can inhibit lung tissue apoptosis induced by LIRI.
【學(xué)位授予單位】：瀘州醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2013
【分類(lèi)號(hào)】：R655.3
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