七氟醚后處理對(duì)糖尿病大鼠心肌保護(hù)失效的機(jī)制與Drp1活性的關(guān)系
發(fā)布時(shí)間：2018-08-30 17:49
【摘要】：目的:干預(yù)動(dòng)力相關(guān)蛋白-1(dynamin related protein-1,Drp1)的活性,探究糖尿病大鼠心肌缺血再灌注損傷(myocardial ischemia reperfusion injury,MIRI)模型不可逆損傷程度之差別;探求七氟醚后處理對(duì)糖尿病大鼠心肌保護(hù)失效的機(jī)制是否與Drp1的活性有關(guān)。方法:健康成年雄性SD大鼠,凈重為225~275 g,以高碳水化合物-高脂肪飼料飼喂并行鏈脲佐菌素30 mg/kg腹腔內(nèi)給藥制備II型糖尿病(T2DM)模型。對(duì)大鼠每個(gè)星期測(cè)定-次空腹血糖,連續(xù)四星期不低于300 mgl/dL者認(rèn)為模型合格。取T2DM模型大鼠40只,依據(jù)計(jì)算機(jī)隨機(jī)數(shù)劃分為4組(n=10):(1)單純穿線組(Sham組)、(2)缺血/再灌注組(I/R組)、(3)七氟醚后處理組(SP組)、(4)Drp1活性抑制劑Mdivi-1+七氟醚后處理組(M-SP組)。采取在體阻斷左前降支(left anterior descending,LAD)血流0.5 h,再灌注2 h法建立MIRI模型,Sham組單純穿線而不阻斷LAD血流。缺血前15 min M-SP組靜脈注射Mdivi-1 1.2 mg/kg,再灌注初期5 min內(nèi)SP組、M-SP組吸入2.5%七氟醚進(jìn)行后處理。再灌注2 h時(shí)采集大鼠右頸內(nèi)靜脈血樣,高速離心后取血清,采用ELISA法檢測(cè)并計(jì)算血清cTnI濃度;隨后處死各組大鼠并取其心肌組織,采用TTC法測(cè)定心肌梗死區(qū)面積(infarct size,IS)和缺血危險(xiǎn)區(qū)(area at risk,AAR)面積,采用TUNEL法檢測(cè)凋亡并計(jì)算細(xì)胞凋亡指數(shù)(apoptotic index,AI);采用western blot法測(cè)定心肌組織中活化型caspase-3表達(dá)水平。結(jié)果:1.AAR和IS測(cè)定結(jié)果:與Sham組比較,其余3組心肌AAR面積增大很顯著(P0.01),但3組之間兩兩比較AAR面積不構(gòu)成統(tǒng)計(jì)學(xué)差異(P0.05)。與Sham組比較,其余3組IS增大(P0.01);與I/R組比較,M-SP組IS減小(P0.05),而SP組IS差異無統(tǒng)計(jì)學(xué)意義(P0.05);與SP組比較,M-SP組IS減小(P0.05)。2.血清cTnI濃度檢測(cè)結(jié)果:與Sham組比較,其余3組血清cTnI濃度升高(P0.05);與I/R組比較,M-SP組血清cTnI濃度降低(P0.05),而SP組血清cTnI濃度差異無統(tǒng)計(jì)學(xué)意義(P0.05);與SP組比較,M-SP組血清cTnI濃度降低(P0.05)。3.TUNEL法檢測(cè)結(jié)果:與Sham組比較,其余3組心肌組織AI值升高(P0.05);與I/R組比較,M-SP組心肌組織AI值降低(P0.05),而SP組心肌組織AI值差異無統(tǒng)計(jì)學(xué)意義(P0.05);與SP組比較,M-SP組心肌組織AI值降低(P0.05)。4.western blot法檢測(cè)結(jié)果:與Sham組比較,其余3組活化型caspase-3表達(dá)上調(diào)(P0.05);與I/R組比較,M-SP組活化型caspase-3表達(dá)下調(diào)(P0.05),SP組活化型caspase-3表達(dá)差異無統(tǒng)計(jì)學(xué)意義(P0.05);與SP組比較,M-SP組活化型caspase-3表達(dá)下調(diào)(P0.05)。結(jié)論:在糖尿病條件下,七氟醚后處理對(duì)大鼠心肌保護(hù)作用失效,抑制Drp1活性后T2DM大鼠MIRI模型的心肌不可逆損傷程度有所減輕;提示七氟醚后處理對(duì)糖尿病大鼠心肌保護(hù)失效的機(jī)制可能與Drp1活性有關(guān)。
[Abstract]:Objective: to investigate the irreversible degree of myocardial ischemia-reperfusion injury (myocardial ischemia reperfusion injury,MIRI) model in diabetic rats by interfering with the activity of dynamic associated protein 1 (dynamin related protein-1,Drp1). To explore whether the mechanism of myocardial protection failure after sevoflurane treatment in diabetic rats is related to the activity of Drp1. Methods: healthy adult male SD rats with a net weight of 225? 275g were fed with high carbohydrate-high fat diet and intraperitoneal administration of streptozotocin for 30 mg/kg to establish II diabetes mellitus (T2DM) model. Rats who measured fasting blood glucose for four weeks or more per week were considered to be eligible for the model. Forty T2DM model rats were randomly divided into 4 groups (n = 10): (1, Sham group (), (2), I / R group (), (3), SP group), (4, Drp1 activity inhibitor Mdivi-1 sevoflurane post-treatment group, M-SP group). The left anterior descending branch (left anterior descending,LAD) was occluded in vivo for 0.5 h, and the MIRI model was established by reperfusion for 2 h. In the sham group, the blood flow of LAD was not blocked. 15 min M-SP before ischemia, 2.5% sevoflurane was inhaled into the SP group within 5 min after Mdivi-1 1.2 mg/kg, reperfusion. The blood samples of right internal jugular vein were collected at 2 h after reperfusion, the serum samples were collected after high speed centrifugation, the serum cTnI concentration was detected and calculated by ELISA method, then the rats in each group were killed and their myocardial tissues were taken. The area of myocardial infarction area (infarct size,IS) and ischemic risk area (area at risk,AAR) were measured by TTC method, apoptosis index (apoptotic index,AI) was calculated by TUNEL method, and the expression of activated caspase-3 was measured by western blot method. Results 1. The results of AAR and IS: compared with Sham group, the area of myocardial AAR in the other three groups increased significantly (P0.01), but there was no statistical difference in AAR area between the three groups (P0.05). Compared with Sham group, IS in the other three groups increased (P0.01), IS in M-SP group decreased (P0.05) compared with I / R group, but IS in SP group was not significantly different (P0.05), IS in M-SP group decreased (P0.05) compared with SP group (P0.05). Results of serum cTnI concentration: compared with Sham group, The serum cTnI concentration in the other three groups was increased (P0.05), the serum cTnI concentration in the M-SP group was lower than that in the I / R group (P0.05), but the serum cTnI concentration in the SP group was not significantly different (P0.05), and the serum cTnI concentration in the M-SP group was lower than that in the SP group (P0.05). 3. Tunel method: compared with the Sham group, the serum cTnI concentration in the M-SP group was lower than that in the Sham group. Compared with the I / R group, the AI value of myocardial tissue in M-SP group decreased (P0.05), but there was no significant difference in AI value of myocardial tissue in SP group (P0.05). Compared with SP group, the AI value of myocardial tissue in M-SP group decreased (P0.05). 4. Western blot assay results: compared with Sham group, the myocardial tissue AI value of M-SP group decreased (P0.05). The expression of activated caspase-3 in the other three groups was up-regulated (P0.05), the expression of activated caspase-3 in M-SP group was down-regulated (P0.05), the expression of activated caspase-3 in M-SP group was not significantly higher than that in SP group (P0.05), and the expression of activated caspase-3 in M-SP group was lower than that in SP group (P0.05). Conclusion: under the condition of diabetes, the protective effect of sevoflurane on myocardium of rats was ineffective, and the degree of irreversible myocardial damage of MIRI model of T2DM rats after inhibiting the activity of Drp1 was alleviated. The results suggest that the mechanism of myocardial protection failure after sevoflurane treatment in diabetic rats may be related to the activity of Drp1.
【學(xué)位授予單位】：山西醫(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)】：R614
本文編號(hào)：2213818
[Abstract]:Objective: to investigate the irreversible degree of myocardial ischemia-reperfusion injury (myocardial ischemia reperfusion injury,MIRI) model in diabetic rats by interfering with the activity of dynamic associated protein 1 (dynamin related protein-1,Drp1). To explore whether the mechanism of myocardial protection failure after sevoflurane treatment in diabetic rats is related to the activity of Drp1. Methods: healthy adult male SD rats with a net weight of 225? 275g were fed with high carbohydrate-high fat diet and intraperitoneal administration of streptozotocin for 30 mg/kg to establish II diabetes mellitus (T2DM) model. Rats who measured fasting blood glucose for four weeks or more per week were considered to be eligible for the model. Forty T2DM model rats were randomly divided into 4 groups (n = 10): (1, Sham group (), (2), I / R group (), (3), SP group), (4, Drp1 activity inhibitor Mdivi-1 sevoflurane post-treatment group, M-SP group). The left anterior descending branch (left anterior descending,LAD) was occluded in vivo for 0.5 h, and the MIRI model was established by reperfusion for 2 h. In the sham group, the blood flow of LAD was not blocked. 15 min M-SP before ischemia, 2.5% sevoflurane was inhaled into the SP group within 5 min after Mdivi-1 1.2 mg/kg, reperfusion. The blood samples of right internal jugular vein were collected at 2 h after reperfusion, the serum samples were collected after high speed centrifugation, the serum cTnI concentration was detected and calculated by ELISA method, then the rats in each group were killed and their myocardial tissues were taken. The area of myocardial infarction area (infarct size,IS) and ischemic risk area (area at risk,AAR) were measured by TTC method, apoptosis index (apoptotic index,AI) was calculated by TUNEL method, and the expression of activated caspase-3 was measured by western blot method. Results 1. The results of AAR and IS: compared with Sham group, the area of myocardial AAR in the other three groups increased significantly (P0.01), but there was no statistical difference in AAR area between the three groups (P0.05). Compared with Sham group, IS in the other three groups increased (P0.01), IS in M-SP group decreased (P0.05) compared with I / R group, but IS in SP group was not significantly different (P0.05), IS in M-SP group decreased (P0.05) compared with SP group (P0.05). Results of serum cTnI concentration: compared with Sham group, The serum cTnI concentration in the other three groups was increased (P0.05), the serum cTnI concentration in the M-SP group was lower than that in the I / R group (P0.05), but the serum cTnI concentration in the SP group was not significantly different (P0.05), and the serum cTnI concentration in the M-SP group was lower than that in the SP group (P0.05). 3. Tunel method: compared with the Sham group, the serum cTnI concentration in the M-SP group was lower than that in the Sham group. Compared with the I / R group, the AI value of myocardial tissue in M-SP group decreased (P0.05), but there was no significant difference in AI value of myocardial tissue in SP group (P0.05). Compared with SP group, the AI value of myocardial tissue in M-SP group decreased (P0.05). 4. Western blot assay results: compared with Sham group, the myocardial tissue AI value of M-SP group decreased (P0.05). The expression of activated caspase-3 in the other three groups was up-regulated (P0.05), the expression of activated caspase-3 in M-SP group was down-regulated (P0.05), the expression of activated caspase-3 in M-SP group was not significantly higher than that in SP group (P0.05), and the expression of activated caspase-3 in M-SP group was lower than that in SP group (P0.05). Conclusion: under the condition of diabetes, the protective effect of sevoflurane on myocardium of rats was ineffective, and the degree of irreversible myocardial damage of MIRI model of T2DM rats after inhibiting the activity of Drp1 was alleviated. The results suggest that the mechanism of myocardial protection failure after sevoflurane treatment in diabetic rats may be related to the activity of Drp1.
【學(xué)位授予單位】：山西醫(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)】：R614
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